R E L A T I O N S H I P S O F IR O N AND C O P P E R T O A S C O R B I C A C I D IN P L A N T S By ROBERT LEON LeBREC A THESIS S u b m i t t e d to t h e S c h o o l of G r a d u a t e S t u d i e s of M i c h i g a n S t a t e C o l l e g e of A g r i c u l t u r e a n d A p p l i e d S c i e n c e i n p a r t i a l f u l f i l l m e n t o f th e r e q u i r e m e n t s f o r t h e d e g r e e of DOCTOR OF PHILOSOPHY D e p a r t m e n t of B otany and P l a n t P ath o lo g y 1951 I ACKNOWLEDGMENTS It i s a p l e a s u r e t o e x p r e s s m y a p p r e c i a t i o n to D r . F . L y l e W yn d f o r h i s a s s i s t a n c e .and v a l u a b l e a d v i c e d u r i n g t h e p l a n n i n g an d e x e c u t i o n o f t h i s i n v e s t i g a t i o n an d t h e p r e p a r a t i o n o f t h e t h e s i s . I a m a l s o g r e a t l y i n d e b t e d to t h e F e r r o E n a m e l C o r p o r a t i o n of C l e v e l a n d a n d to t h e C e r o p h y l L a b o r a t o r i e s , I n c . , of K a n s a s C i t y , M is s o u r i , for t h e i r f in a n c ia l s u p p o rt, w h ich p e r m i t t e d m e to c a r r y o u t th is p r o b l e m . M a n y t h a n k s a r e a l s o d u e to m y f e l l o w s t u d e n t s , M r . G e o r g e * C. V a y o n i s , a n d D r . E r l i n g S t r o m m e , f o r t h e i r h e l p f u l s u g g e s t i o n s a n d e n c o u r a g e m e n t d u r i n g t h e c o u r s e of t h i s s t u d y . R obert Leon L eB rec M ichigan S tate C o lle g e J u n e , 1951 TABLE OF CONTENTS Page 1. I N T R O D U C T I O N ..........................................: .................... A* 1 R e v i e w of L i t e r a t u r e . ; .................................................. 3 1. R e la tio n s h ip B e tw e e n C o p p e r and Iro n and t h e C o n c e n t r a t i o n o f A s c o r b i c A c i d in B i o l o g i c a l M a t e r i a l .................................................... 2. R e la tio n s h ip B e tw e e n C o p p e r and Iro n and t h e S t a b i l i t y of A s c o r b i c A c i d in V i t r o B. II. O b j e c t of t h e P r e s e n t S t u d y ... 6 ............................................. 10 E X P E R I M E N T A L P R O C E D U R E S ............................................. 12 A. E q u i p m e n t ....................................................... . . . . B. III. 3 N u tr ie n t Solution EX PER IM EN T 1 . A. B. ............... 17 P u r p o s e ................................................................... 17 ................................................ C. 1. 18 I r o n a n d C o p p e r L e v e l s in t h e N u t r i e n t S o l u t i o n ............................................. 2. 13 ............................................................................. E x p e rim e n ta l C onditions 1. 12 P lant M aterial 18 .................................................................. 19 E x p e r i m e n t a l R e s u l t s ................................................... 20 O b s e r v a t i o n s o n t h e P l a n t s ...................................... 20 I iv Page a. D e s c r i p t i o n s of t h e P l a n t s a t T i m e of H a r v e s t ......................................... 20 F r e s h W e i g h t of t h e P l a n t s ............................ 21 (1) E f f e c t of I r o n ............................................... 21 (2) E f f e c t of C o p p e r .......................... 21 D r y W e i g h t p e r P l a n t .......................................... 22 (1) E f f e c t of I r o n ............................................... 22 (2) E f f e c t of C o p p e r ........................................ 22 C h e m i c a l A n a l y s e s of t h e P l a n t s ......................... 23 b. c. 2. . . . a. . . . I r o n ...................................................... (1) A m o u n t o f I r o n p e r P l a n t ..................• 23 23 (a) E f f e c t of i n c r e a s i n g th e a m o u n t s of i r o n in t h e s o l u ­ t i o n on t h e i r o n c o n t e n t of t h e p l a n t s .............................................. 23 (b) E f f e c t o f i n c r e a s i n g t h e a m o u n t of c o p p e r in t h e s o l u t i o n on t h e i r o n c o n t e n t of t h e p l a n t s ....................................... 24 (2) A m o u n t o f I r o n p e r 100 G r a m s F r e s h M a t e r i a l ...................... •.................. 25 (a) E f f e c t o f i r o n ..................................... 25 V Page (b) E f f e c t of c o p p e r ............................... 25 (3) A m o u n t o f I r o n p e r 100 G r a m s D ry M aterial b. . . 26 (a) E f f e c t o f i r o n ...................................... 26 (b) E f f e c t of c o p p e r ............................... 26 ........................................................ 27 (1) D e s c r i p t i o n ..................................................... 27 A s c o r b ic A cid (2) A m o u n t o f A s c o r b i c A c i d p e r P l a n t ................................................................... 27 (a) E f f e c t of i r o n ...................................... 27 (b) E f f e c t of c o p p e r 28 ............................... (3) A m o u n t o f A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a tt e r ..................... 29 (a) E f f e c t o f i r o n ...................................... 29 (b) E f f e c t of c o p p e r 29 . • (4) A m o u n t o f A s c o r b i t A c i d p e r 100 G r a m s o f D r y M a t e r i a l ........................ 30 (a) E f f e c t o f i r o n ...................................... 30 (b) E f f e c t of c o p p e r ............................... 30 (5) P e r c e n t a g e o f t h e O r i g i n a l A m o u n t of A s c o r b i c A c i d F o u n d a t E a c h D e t e r m i n a t i o n ............................................. 31 VI Page (a) E f f e c t of i r o n ....................................... 31 ................................ 32 ..................... 33 (b) E f f e c t of c o p p e r c. IV. O th e r C hem ical C o n stitu en ts D. D i s c u s s i o n ofthe F i r s t E x p e r i m e n t .............................. 34 E. S u m m a r y o f t h e F i r s t E x p e r i m e n t ................................... 37 E X P E R I M E N T 2 .................................................................................... 40 A. B. Purpose .......................................................................................... E x p e r i m e n t a l C o n d i t i o n s ....................................... 1. C. P la n ts M aterial 40 .................................................... . . . E x p e r i m e n t a l R e s u l t s ........................................................ 1. O b s e r v a t i o n s on th e P l a n t s ...................................... a. b. c. 2. 40 I r o n a n d C o p p e r L e v e l s in t h e N u t r i e n t S o l u t i o n ................................................ 2. 40 41 42 42 D e s c r i p t i o n of the P l a n t s a t T i m e o f H a r v e s t ........................................................... . 42 F r e s h W e i g h t p e r P l a n t ................................... 42 (1) E f f e c t of I r o n .................................. 42 (2) E f f e c t of Copper .................... 43 ...................................... 43 D r y W eight per P la n t . . , (1) E f f e c t of I r o n ......................................... 43 (2) E f f e c t of C o p p e r .................................. 44 C h e m i c a l C o m p o s i t i o n of t h e P l a n t s ................. 45 vii Page a. I r o n ........................................ 1 .................................... 45 (1) A m o u n t of I r o n p e r P l a n t ..................... 45 (a) E f f e c t of i n c r e a s i n g t h e a m o u n t s of i r o n in t h e s o l u t i o n on t h e a b s o r p t i o n of i r o n b y t h e p l a n t ..................... 45 (b) E f f e c t of i n c r e a s i n g t h e a m o u n t s o f c o p p e r in th e s o l u t i o n s on t h e a b s o r p t i o n of i r o n ............................. ; ................... 45 (2) A m o u n t of I r o n p e r 100 G r a m s of F r e s h M a t e r i a l ................................... 46 (a) E f f e c t of iron . ; ........................ 46 (b) E f f e c t of c o p p e r ........................ 46 (3) A m o u n t of I r o n p e r 100 G r a m s b. of D r y M a t t e r .............................................. 47 (a) E f f e c t of i r o n ............................... 47 (b) E f f e c t of c o p p e r ....................... 47 A s c o r b i c A c i d ........................................................ 48 (1) D e s c r i p t i o n .................................................... 48 (2) A m o u n t of A s c o r b i c A c i d p e r P l a n t . » ............................................................ 48 (a) E f f e c t of i r o n ...................................... 48 V lll Page (b) E f f e c t of c o p p e r ............................... 49 (3) A m o u n t o f A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t t e r ..................... 50 (a) E f f e c t of i r o n ................. ; ................. 50 (b) E f f e c t of c o p p e r ................................ 51 (4) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s of D r y M a t e r i a l . . 52 (a) E f f e c t of i r o n ....................................... 52 (b) E f f e c t of c o p p e r .............. 53 (5) P e r c e n t a g e of t h e O r i g i n a l A m o u n t of A s c o r b i c A c id F o u n d at E a c h D e t e r m i n a t i o n ............................ 53 (a) E f f e c t of i r o n ....................................... 53 (b) E f f e c t of c o p p e r ........... ’................... 54 D. D i s c u s s i o n on t h e S e c o n d E x p e r i m e n t ....................... 55 E. S u m m a r y of t h e S e c o n d E x p e r i m e n t .......................... 58 E X P E R I M E N T 3 ................................................. 61 A. P u r p o s e ......................................................................................... 61 B. E x p e r i m e n t a l C o n d i t i o n s ................................................... 62 1. I r o n a n d C o p p e r L e v e l s in t h e N u t r i e n t S o l u t i o n ................................................................................. 2. P lant M a te rial .................................................................. 62 63 ix Page C. E x p e r i m e n t a l R e s u l t s ................ 1. O b s e r v a t i o n s on t h e P l a n t s ..................... 63 a. D e s c r i p t i o n of t h e P l a n t s ............................... 63 b. F r e s h W e i g h t p e r P l a n t .................................. 64 c. 2. 63 (1) E f f e c t of I r o n ......................................... 64 (2) E f f e c t of C o p p e r .................................. 64 D r y W e i g h t p e r P l a n t ......................................... 65 (1) E f f e c t of I r o n ......................................... 65 (2) E f f e c t of C o p p e r .................................. 65 C h e m i c a l A n a l y s e s of t h e P l a n t s a. Iron ........................ 66 ................................................................. 66 ' (1) A m o u n t s of I r o n p e r P l a n t .................. 66 (a) E f f e c t of i n c r e a s i n g t h e a m o u n t of i r o n in t h e n u t r i e n t s o l u t i o n on t h e i r o n c o n t e n t of t h e p l a n t s ....................................... 66 (b) E f f e c t of i n c r e a s i n g t h e a m o u n t of c o p p e r i n t h e n u t r i e n t s o lu tio n on the a c c u m u l a t i o n of i r o n by th e p l a n t ............................................. 67 Page (2) A m o u n t s of I r o n p e r 100 G r a m s of F r e s h P l a n t M a t e r i a l ..................... 68 (a) E f f e c t of i r o n .......................... 68 (b) E f f e c t of c o p p e r 68 .............................. (3) A m o u n t of I r o n p e r 100 G r a m s of D r y M a t t e r .............................................. 69 (a) E f f e c t of i r o n .................................. 69 ........................... 69 .......................................................................... 70 (1) D e s c r i p t i o n ..................................................... 70 (2) A m o u n t of C o p p e r p e r P l a n t ................ 70 (b) E f f e c t of c o p p e r b. Copper (3) A m o u n t s of C o p p e r p e r 100 G r a m s of F r e s h M a t t e r ............................................ 71 (4) A m o u n t s of C o p p e r p e r 100 G r a m s of D r y M a t t e r c. A sc o rb ic A cid . ............................ ................. 72 73 (1) D e s c r i p t i o n ................ ................................... 73 (2) A m o u n t s of A s c o r b i c A c i d p e r P l a n t .................................................................. 74 (a) E f f e c t of i r o n ....................................... 74 (b) E f f e c t of c o p p e r 75 ............................... xi Page (3) A m o u n t s of A s c o r b i c A c i d p e r 100 G r a m s o f F r e s h M a t t e r ................ 75 (a) E f f e c t of i r o n ....................................... 75 (b) E f f e c t of c o p p e r ................................ 76 (4) P e r c e n t a g e s of t h e O r i g i n a l A m o u n ts of A s c o r b i c A cid VI. Found at E ach D eterm ination . . . . 78 (a) E f f e c t of i r o n ....................................... 78 (b) E f f e c t of c o p p e r ................................ 79 d. O t h e r C h e m i c a l C o n s t i t u e n t s ............................. 81 D. D i s c u s s i o n on t h e T h i r d E x p e r i m e n t ....................... 82 E. S u m m a r y of t h e T h i r d E x p e r i m e n t .................................. 86 E X P E R I M E N T 4 .................................................................................... 90 A. P u r p o s e ........................................................................................... 90 B . E x p e r i m e n t a l C o n d i t i o n s ........................................................ 92 C. 1. T h e N u t r i e n t S o l u t i o n ................................................... 92 2. P lan t M aterial 93 ................................................................. E x p e r i m e n t a l R e s u l t s ........................ 1. O b s e r v a t i o n s on t h e P l a n t s a. b. . •................................. 94 94 D e s c r i p t i o n of t h e P l a n t s a t T i m e of H a r v e s t ..................... •........................................ 94 F r e s h W e i g h t p e r P l a n t ................................... 95 Xll Page c. 2. D r y W e i g h t p e r P l a n t ......................................... 95 C h e m i c a l A n a l y s e s of t h e P l a n t s ......................... 96 a. Iron ................................................................. 96 (1) A m o u n t s of I r o n p e r P l a n t s .................... 96 (2) A m o u n t s of I r o n p e r 100 G r a m s of F r e s h T i s s u e ...... ............................. 96 (3) A m o u n t s of I r o n p e r 100 G r a m s b. of D r y M a t e r i a l ........................................... 97 (a) In the w h e a t t o p s ................... 97 (b) In the w h eat r o o t s ................ 97 M a n g a n e s e ...........................................................• • • 98 (1) A m o u n t s of M a n g a n e s e p e r P l a n t ................................................................... 98 (2) A m o u r t t s of M a n g a n e s e p e r 100 G r a m s of F r e s h M a t e r i a l .................. 98 (3) A m o u n t s of M a n g a n e s e p e r 100 c. G r a m s of D r y M a t t e r ............................. 98 (a) In t h e wTieat t o p s ................... 98 (b) In the w h e a t r o o t s ................ 99 ..................................... 99 A s c o rb ic A cid . . . . (1) A m o u n t s of A s c o r b i c A c i d p e r P l a n t .................................................................. 99 xiii Page (2) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t e r i a l ................. 100 (a) In t h e w h e a t t o p ............................... 100 .................. 100 (b) In t h e w h e a t r o o t (3) A m o u n t of A s c o r b i c A c i d p e r 100 VII. G r a m s of D r y M a t e r i a l ......................... 101 D. D i s c u s s i o n on t h e F o u r t h E x p e r i m e n t ....................... 101 E. S u m m a r y of t h e F o u r t h E x p e r i m e n t ........................... 106 E X P E R I M E N T S ................................................................................... 108 A. P u r p o s e .......................................................................................... 108 13. E x p e r i m e n t a l C o n d i t i o n s ................................................... 109 1. The N u t r i e n t S o l u t i o n ............................................. 109 2. The P lan t M aterial ................................................. 109 . . 110 O b s e r v a t i o n s on t h e P l a n t s ....................................... 110 C. E x p e rim en tal R esults 1. a. D e s c r i p t i o n of t h e P l a n t s a t T i m e of H a r v e s t ............................................................... b. P e r c e n t a g e s of D r y M a t t e r in t h e Leaves 2. 110 ...................................................................... Ill C h e m i c a l A n a l y s e s of t h e P l a n t ............................ Ill a. I ro n . . . . ' ................................................................... Ill I xiv Page (1) A m o u n t s of I r o n in 100 G r a m s o f F r e s h M a t e r i a l ................................... Ill (2) A m o u n t s of I r o n p e r 100 G r a m s of D r y M a t t e r b. . 112 M a n g a n e s e .................................................................. 112 (1) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of F r e s h M a t e r i a l . . . . . . . 112 (2) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of D r y M a t e r i a l ......................... c. A s c o r b ic A cid ........................................................ 113 114 (1) A m o u n t s of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t e r i a l ... 114 100 G r a m s of D r y M a t e r i a l .............. 115 D. D i s c u s s i o n o n t h e F i f t h E x p e r i m e n t ............................ 115 E. S u m m a r y of th e F i f t h E x p e r i m e n t 119 (2) A m o u n t s of A s c o r b i c A c i d p e r VIII. EX PER IM EN T 6 . A. Purpose ............................................ .......................................................................................... B. E x p e r i m e n t a l C o n d itio n s C. . . . . - . • ............. ............................. ,...................... 121 121 122 1. T h e N u t r i e n t S o l u t i o n ............................................. 122 2. T h e P l a n t M a t e r i a l ........................................................ 123 E xperim ental R esu lts ................................................ 124 XV Page 1. O b s e r v a t i o n s on t h e P l a n t s ....................................... a. D e s c r i p t i o n of t h e P l a n t s a t T i m e of H a r v e s t ............................................................... b. 2. 124 124 P e r c e n t a g e s of D r y M a t t e r i n t h e l e a v e s .......................................................................... 125 C h e m i c a l A n a l y s e s of t h e P l a n t s ......................... 126 a . ...................................I r o n ................................................. 126 (1) A m o u n t s of I r o n p e r 100 G r a m s of F r e s h M a t e r i a l ................... •............... 126 (2) A m o u n t s of I r o n p e r 100 G r a m s b. of D r y M a t e r i a l .......................................... 127 M a n g a n e s e .......................................... 127 (1) A m o u n t of M a n g a n e s e p e r 100 G r a m s of F r e s h M a t e r i a l .................. 127 (2) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of D r y M a t t e r ............................ c. A s c o r b i c Acid ..................................... 128 129 (1) A m o u n t s of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t e r i a l . . . 129 100 G r a m s of D r y M a t t e r .................. 130 D i s c u s s i o n on t h e S i x t h E x p e r i m e n t ............................ 131 (2) A m o u n t s of A s c o r b i c A c i d p e r D. I xvi Page S u m m a r y of t h e S i x t h E x p e r i m e n t .................................. 132 E X P E R I M E N T 7 .................................................................................... 135 E. A. P u r p o s e .......................................................................................... 135 B. P r e l i m i n a r y E x p e r i m e n t .................................................... 136 C. E x p e r i m e n t a l C o n d i t i o n s ........................................................ 138 1. T h e N u t r i e n t S o l u t i o n .................................................... 138 2. T h e P l a n t M a t e r i a l ........................................................ 141 E x p e r i m e n t a l R e s u l t s ............................• .............................. 142 D. 1. O b s e r v a t i o n s on t h e P l a n t s . .................... a. D e s c r i p t i o n of t h e P l a n t s ................. 142 b. F r e s h W e i g h t p e r P l a n t .................................. 143 c. P e r c e n t a g e s of D r y M a t t e r in th e Leaves 2. . . . 142 ...................................................................... C h e m i c a l A n a l y s e s of theT o b a c c o P l a n t s a. . . I r o n ................................................................................ 143 145 145 (1) A m o u n t s o f I r o n p e r 100 G r a m s of F r e s h M a t e r i a l ................................... 145 (2) A m o u n t s of I r o n p e r 100 G r a m s b. of D r y M a t t e r .............................................. 146 M a n g a n e s e ................................................................. 146 (1) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of F r e s h M a t e r i a l .................. 146 XVX1 Page (2) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of D r y M a t e r i a l ......................... c. A s c o r b ic A cid 147 ........................................................ 148 (1) D e s c r i p t i o n .................................................... 148 (2) A m o u n t s of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t t e r .............. 149 (a) E f f e c t of I r o n i n t h e p r e s e n c e of c o p p e r .............................................. 149 (b) E f f e c t of i r o n i n t h e a b s e n c e o f c o p p e r ....................... • .................... 152 E. D i s c u s s i o n on t h e S e v e n t h E x p e r i m e n t ..................... 153 F, S u m m a r y of t h e S e v e n t h E x p e r i m e n t ’. . . 156 X. G E N E R A L D I S C U S S I O N .................................................................. 159 XI. G E N E R A L S U M M A R Y A N D C O N C L U S I O N ....................... 1 64 BIBLIO GRAPHY . 166 XII. XIII. ............................................................... . • • • • A P P E N D I C E S , A N A L Y T I C A L P R O C E D U R E S ................ 172 1. INTRODUCTION B e c a u s e of t h e i m p o r t a n c e of a s c o r b i c a c i d in h u m a n d i e t , n u m e r o u s s t u d i e s h a v e b e e n c a r r i e d o u t in o r d e r to o b t a i n i n f o r m a ­ tion c o n c e r n in g its n a tu r a l o c c u r r e n c e . The data obtained by th e se s tu d i e s show the g r e a t v a r i a b i l i t y ' i n the c o n c e n t r a t i o n of a s c o r b i c a c i d in i t s b i o l o g i c a l s o u r c e s . T h e f a c t o r s i n f lu e n c in g the s y n t h e s i s of a s c o r b i c a c i d in p la n ts h a v e b e e n in v e s t i g a t e d d u r in g the p a s t two d e c a d e s . V arious o p in io n s , m a n y of w h ic h a r c c o n t r a d i c t o r y , c o n c e r n i n g th e i m p o r ­ t a n c e of s p e c i e s , v a r i e t i e s , l i g h t , l o c a t i o n , s o i l t y p e a n d f e r t i l i z e r s , on t h e c o n c e n t r a t i o n of t h i s v i t a m i n in p l a n t s a r e e x p r e s s e d in t h e literatu re. T h e d a t a d e a l i n g w i t h t h e e f f e c t of i n o r g a n i c n u t r i e n t s on t h e a s c o r b i c a c i d s t a t u s in p l a n t s a r e e s p e c i a l l y c o n f u s i n g e v e n t h o u g h a v e r y l a r g e n u m b e r of i n v e s t i g a t o r s h a v e s t u d i e d t h i s p h a s e of t h e p r o b l e m . R e v i e w s on t h i s s u b j e c t h a v e b e e n m a d e b y D u t c h e r (1932), H a m n e r and M a y n a r d (1942), and by C a r r o l (1943). S t u d i e s h a v e b e e n c a r r i e d o u t i n t h e f i e l d , g r e e n h o u s e a n d in c o n t r o l l e d n u t r i e n t s o l u t i o n ; a n d t h e e f f e c t of t h e “ m a c r o - e l e m e n t s ” o n t h e a s c o r b i c a c i d s t a t u s of p l a n t s h a v e b e e n d e s c r i b e d b y m a n y authors. H o w e v e r , t h e e f f e c t of t h e “ m i n o r - e l e m e n t s ” on t h e c o n c e n t r a t i o n of a s c o r b i c a c i d in p l a n t s h a s n o t b e e n a s w e l l s t u d ­ i e d a l t h o u g h e l e m e n t s s u c h a s c o p p e r a n d i r o n a r e k n o w n in v i t r o to b e p o w e r f u l c a t a l y s t s o f t h e o x i d a t i o n of a s c o r b i c a c i d . M ost w o r k e rs have a s s u m e d th at any relatio n sh ip betw een e n v i r o n m e n t a l f a c t o r s a n d t h e a m o u n t o f a s c o r b i c a c i d p r e s e n t in t h e p l a n t s r e f l e c t s t h e e f f e c t of t h e s e f a c t o r s on i t s s y n t h e s i s . I t i s p o s s i b l e , h o w e v e r , t h a t t h e s t a b i l i t y r a t h e r t h a n t h e r a t e of f o r ­ m a t i o n of a s c o r b i c a c i d in t h e p l a n t t i s s u e s i s i n f l u e n c e d b y t h e s e f a c to r s .. T h e r e a r e th u s two m a i n p o s s i b l e r e a s o n s f o r the g r e a t v a r i a t i o n s i n t h e c o n c e n t r a t i o n s of t h i s v i t a m i n in s i m i l a r p l a n t tissues. It is not know n w h e t h e r the a s c o r b i c a c id is s t o r e d in the v a c u o l e s o r in t h e c y t o p l a s m of t h e p l a n t c e l l s . R e e d m a n and M c H e n r y (1 938) r e p o r t e d t h a t in c e r t a i n p l a n t t i s s u e s s u c h a s p o ­ t a t o , c a u l i f l o w e r , a c e r t a i n a m o u n t of t h e a s c o r b i c a c i d i s c o m b i n e d w i t h p r o t e i n ; b u t t h e f r a c t i o n of a s c o r b i c a c i d e x t r a c t e d b y t h e m e t h ­ o d s c o m m o n l y u s e d to d e t e r m i n e i t s a m o u n t in t h e p l a n t m a t e r i a l i s w a t e r s o l u b l e a n d i n a f r e e s t a t e in t h e p r o t o p l a s m o r t h e v a c u o l a r sap. T h e s t a b i l i t y a n d s u b s e q u e n t a c c u m u l a t i o n of t h e w a t e r s o l u b l e f r a c t i o n of a s c o r b i c a c i d in t h e p l a n t c e l l m a y t h u s b e d e p e n d e n t o n the f a c t o r s a f f e c tin g i t s s t a b i l i t y in w a t e r s o lu tio n . B e f o r e d e s c r i b i n g t h e e x p e r i m e n t s c a r r i e d o u t , a r e v i e w of t h e l i t e r a t u r e on t h e e f f e c t s of c o p p e r a n d i r o n on t h e c o n c e n t r a t i o n 3 of a s c o r b i c a c i d i n b i o l o g i c a l m a t e r i a l a n d i t s s t a b i l i t y i n v i t r o w ill b e p r e s e n t e d . A. 1. R e v i e w of L i t e r a t u r e R e la tio n s h ip B etw e en C o p p e r and I ro n and the C o n c e n tr a tio n of A s c o r b i c A c i d in B i o l o g i c a l M a t e r i a l H a m d a l l a h (19 3 9 ) w a s t h e f i r s t w o r k e r to t e s t t h e e f f e c t of d e p r i v i n g p l a n t s of i r o n on t h e a s c o r b i c a c i d c o n c e n t r a t i o n s i n t h e tissues. H e f o u n d t h a t b e a n s , s p i n a c h a n d c o r n g r o w n o n a K n o p p ’s iro n f re e solution contained as m u c h o r m o r e a s c o r b i c acid than d id p l a n t s g r o w n on a c o m p l e t e n u t r i e n t s o l u t i o n . R a n d o u in and L e G a llic (1940), a f t e r a n a ly z in g m a n y p la n t s p e c i e s , d i s c o v e r e d th a t s p e c i e s with high p r o t e i n c o n te n t u s u a lly h a v e a h i g h c o n c e n t r a t i o n of i r o n a n d a s c o r b i c a c i d . However these a u t h o r s d i d n o t d u p l i c a t e t h e i r e x p e r i m e n t s w i t h s i n g l e s p e c i e s in o r d e r to t e s t i f t h i s r e l a t i o n s h i p w a s c o n s i s t e n t . T h e a d d i t i o n of i r o n s u l f a t e to t h e s o i l , d e p r e s s e d t h e a m o u n t of a s c o r b i c a c i d in t h e o r a n g e f r u i t s a c c o r d i n g to i n v e s t i g a t i o n s c a r ­ r i e d o u t b y R o y a n d B a h r t (1 940 ) in F l o r i d a . In a v e r y m e t i c u l o u s n u t r i e n t s o l u t i o n , L y o n , B e e s o n a n d E l l i s (1 943) s t u d i e d t h e e f f e c t of d e f i c i e n c i e s of m i n o r e l e m e n t s on t h e v i t a m i n c o n t e n t of t o m a t o e s . T h e y found th a t f r u i t s f r o m the i r o n d e f i c i e n t p l a n t s c o n t a i n e d 30 p e r c e n t m o r e a s c o r b i c a c i d on a f r e s h w e i g h t b a s i s t h a n d i d t h e p l a n t g r o w i n g on a n u t r i e n t solution s u p p lie d w ith iro n . On t h e o t h e r h a n d , t h e a s c o r b i c a c i d c o n t e n t of t h e f r u i t s w a s n o t a f f e c t e d b y l i m i t i n g t h e s u p p l y of m a n g a n e s e , z in c , c o p p e r and m o ly b d e n u m , a lth o u g h the n u m b e r of f r u i t s of e a c h p l a n t w a s t h e n s i g n i f i c a n t l y l e s s e n e d b y a l a c k of these n u trien ts. A s a c o n t i n u a t i o n of t h e i r f i r s t s t u d y L y o n a n d B e e s o n (1948) p u b l i s h e d t h e i r r e s u l t s o n t h e e f f e c t of t o x i c a m o u n t s of m i n o r e l e m e n t s on t o m a t o e s a n d t u r n i p s . T h e ir plants w ere grow n in s a n d s u p p l i e d w i t h n u t r i e n t s o l u t i o n s . T h e c o n c e n t r a t i o n of a s c o r b i c a c i d i n t o m a t o p l a n t s g r o w n on n u t r i e n t s o l u t i o n s u p p l i e d w i t h 50 p a r t s p e r m i l l i o n of c o p p e r w a s f o u n d 60 p e r c e n t g r e a t e r t h a n i n t h e p l a n t s g r o w n w i t h o u t irori. H o w e v e r , t h e a s c o r b i c a c i d c o n c e n t r a t i o n of t u r n i p g r e e n s w a s l e s s e n e d w i t h i n c r e a s e d a m o u n t s of c o p p e r in t h e n u t r i e n t solution. On t h e o t h e r h a n d , no s i g n i f i c a n t d i f f e r e n c e in t h e a s c o r ­ b ic a c i d c o n c e n t r a t i o n of t o m a t o a n d t u r n i p g r e e n w a s o b s e r v e d w i t h i n c r e a s e of t h e a m o u n t of i r o n i n t h e n u t r i e n t s o l u t i o n up to 150 p a r t s p e r m i l l i o n . S i d e r i s a n d Y o u n g (1 948) r e p o r t e d t h a t p i n e a p p l e l e a v e s a c c u m u la te d m o r e a s c o r b ic acid w hen the p lan ts su p p lied with n itr a te n i tr o g e n w e r e d e p r iv e d of iro n , than w hen th ey w e r e s u p ­ plied w ith iro n . H o w ev er, w hen the p la n ts w e r e su p p lied w ith 5 a m m o n i u m n i t r o g e n , t h e c o n c e n t r a t i o n of a s c o r b i c a c i d in t h e l e a v e s w a s t h e s a m e i r r e s p e c t i v e if i r o n w a s p r e s e n t o r n o t in the n u t r i e n t s o lu tio n . In 1 949, t h e s a m e a u t h o r s , s t u d y i n g t h e e f f e c t of d i f f e r e n t i r o n - m a n g a n e s e r a t i o s on t h e a s c o r b i c a c i d c o n t e n t of p i n e a p p l e le a v e s , found a g r e a t e r a s c o r b i c a c i d c o n c e n t r a t i o n in th e p l a n t s d e p r i v e d of i r o n t h a n i n t h o s e s u p p l i e d w i t h 0 . 5 p a r t s p e r m i l l i o n of i r o n in t h e n u t r i e n t s o l u t i o n a n d w h e n a r e l a t i v e l y h i g h a m o u n t of m a n g a n e s e w a s p r e s e n t . T h e e f f e c t of t h e a b s o r p t i o n of i r o n on t h e c o n c e n t r a t i o n of a s c o r b i c a c i d in o a t p l a n t s g r o w n on f i e l d p l o t s i n T e x a s , w a s i n ­ v e s tig a te d by Wynd and N oggle (1950a), who r e p o r t e d th a t . . . t h e c o n c e n t r a t i o n of v i t a m i n C w a s i n v e r s e l y r e ­ l a t e d to t h e c o n c e n t r a t i o n of i r o n f o u n d in t h e p l a n t . T h i s r e l a t i o n s h i p w a s t r u e f o r a l l g r o u p s of c o n t r o l d a t a a n d f o r a m o u n t s of a m m o n i u m s u l f a t e , a m m o n i u m p h o s p h a t e , u r e a o r su lfu r. The data s u p p o rtin g this g e n e r a liz a tio n w e r e obtained f r o m p l a n t s w h i c h s h o w e d w i d e r a n g e s i n t h e a m o u n t of d r y m a t t e r p r o d u c e d , p e r c e n t a g e n i t r o g e n a n d c o n c e n t r a t i o n of i r o n . T h e s o i l s on w h i c h t h e p l a n t s w e r e g r o w n v a r i e d w i d e l y in t h e pH v a l u e . H o w e v e r , t h e s a m e w o r k e r s p u b l i s h e d ( 1 9 5 0 b ) t h a t in r y e p l a n t s g r o w n on f i e l d p l o t s in C a n a d a , t h e c o n c e n t r a t i o n s of a s c o r ­ b ic a c i d w e r e p o s i t i v e l y r e l a t e d t o t h e p e r c e n t a g e of n i t r o g e n , c a l ­ ciu m and iro n . F r o m a p e r s o n a l c o m m u n i c a t i o n w i t h F . L . W ynd it w a s l e a r n e d th a t the p o s it i v e r e l a t i o n s h i p found in th is c a s e b e tw e e n th e c o n c e n t r a t i o n s of i r o n a n d a s c o r b i c a c i d w a s t h e o n l y i n s t a n c e in w h i c h s u c h a r e l a t i o n s h i p o c c u r r e d to t h e s e w o r k e r s . In t h e i r a r t i c l e , t h e a u t h o r s e m p h a s i z e d t h a t t h e “ a c c u m u l a t i o n of a s c o r b i c a c i d in p l a n t t i s s u e p r o b a b l y i s n o t i n t r i n s i c a l l y p o s i t i v e l y c o r r e ­ lated w ith ir o n , b u t r a t h e r w ith a s u b tle s e c o n d a r y r e l a t i o n s h i p betw een iro n and c o p p e r .” T h i s h y p o t h e s i s i s b a s e d on t h e t h o u g h t t h a t t h e c o n c e n t r a t i o n of a s c o r b i c a c i d i n p l a n t s m a y b e t h e i m a g e of i t s s t a b i l i t y i n t h e p l a n t t i s s u e . F r o m in v i t r o s t u d i e s , t h e s t a ­ b i l i t y of a q u e o u s a s c o r b i c a c i d s o l u t i o n i s r e l a t e d to c o p p e r a n d iro n , as w ill be sh o w n in the n e x t p a r a g r a p h . 2. R e la tio n s h ip B e tw e e n C o p p e r and Iro n and the S ta b ility of A s c o r b i c A c i d in V i t r o A s e a r l y a s 1921 , H e s s a n d U n g e r f o u n d t h a t m i l k t r e a t e d a t 6 0 ° C f o r 40 m i n u t e s in a c o p p e r v e s s e l c a u s e d s c u r v y t o g u i n e a pig in f o u r w e e k s . In 1928, S z e n t G y o r g y , m e a s u r e d t h e o x y g e n u p t a k e of s o l u ­ t i o n of h i s p r e p a r a t i o n of a s c o r b i c a c i d in a B a r c r o f t a p p a r a t u s . H e f o u n d t h a t in t h e p r e s e n c e of c o p p e r t h e s o l u t i o n s h o w e d a n i n ­ t e n s e u p t a k e of o x y g e n , b u t h e d i d n o t f in d a n y n o t i c e a b l e e f f e c t of F e +++ a n d M n + + . I n c r e a s i n g c o n c e n t r a t i o n s of c o p p e r a t a pH v a l u e of a b o u t 6 in a n a c e t a t e b u f f e r w e r e s h o w n i n 1933 b y E u l e r , M y r b a c k a n d 7 L a r s s o n , to r e s u l t in i n c r e a s i n g c o n s u m p t i o n s o f o x y g e n i n a W a r ­ burg a p p a r a t u s f r o m an a s c o r b i c a c id solution. T hese authors o b t a i n e d a s m a l l a c t i o n f r o m i r o n o n t h e o x i d a t i o n of t h e a s c o r b i c a c i d in t h e s o l u t i o n . T h e f o l l o w i n g t a b l e p r e s e n t e d b y K e l l i e a n d Z i l v a (1 935) s h o w s t h a t c o p p e r i s a m o r e e f f i c i e n t c a t a l y s t t h a n i r o n in t h e o x i d a t i o n of a s c o r b i c a c i d in w a t e r s o l u t i o n . M i l l i g r a m s of A s c o r b i c A c i d p e r 100 M i l l i l i t e r s of W a t e r Tim e in H ours Q uartz D istilled W ater Q u a rtz D istilled W a t e r 0.01 m g . F e p e r 40 m l . Q u artz D istilled W a t e r 0.01 m g . C u p e r 40 m l . 0 37.0 34.5 34.1 1 35.2 31.8 24.2 2 33 .0 29.6 21.2 3 30.9 28.2 15.2 4 29.4 25.8 8.0 20 25.7 10.3 0. 0 M a w s o n ( 1935) f o u n d t h a t f e r r o u s o r f e r r i c i o n s c a t a l y z e t h e o x i d a t i o n of a s c o r b i c a c i d in a q u e o u s s o l u t i o n , b u t t o a l e s s e r extent than does c o p p e r. H e a l s o o b s e r v e d t h a t a m i x t u r e of c o p p e r a n d i r o n h a s a c a t a l y t i c e f f e c t on t h e a e r o b i c o x i d a t i o n of a s c o r b i c 8 acid g r e a t e r than have e ith e r m e ta l alo n e. He p o in te d out t h a t in s u c h b i o l o g i c a l m e d i a a s s e r u m , t h e a m o u n t of f r e e c o p p e r a n d iro n is so la r g e th a t s o m e in h ib ito rs m u s t p r o te c t the a s c o rb ic acid f r o m bein g ox id ized . He found th a t s o m e p r o te c tiv e actio n was e x e rte d by c y s te in e , c y s tin e and glutathione. He a ls o show ed t h a t i n l e m o n j u i c e t h e r e i s n o s u c h p r o t e c t i v e m e c h a n i s m , b u t if t r a c e s of a n i m a l t i s s u e s a r e a d d e d t o t h e j u i c e , i t s a s c o r b i c a c i d i s p r o t e c t e d f r o m o x i d a t i o n a t a pH v a l u e of 7 . 4 a n d a t a t e m p e r a ­ t u r e of 3 7 ° C. T h e a m o u n t of a s c o r b i c a c i d i n v a r i o u s a n i m a l t i s s u e s w a s o b s e r v e d b y M c F a r l a n e ( 1 9 3 6 ) to r e d u c e a p r o p o r t i o n a l a m o u n t of iron. He a d d e d s o d i u m d i e t h y l d i t h i o c a r b a m a t e to t h e l i s t of i n h i b ­ i t o r s of t h e o x i d a t i o n of a s c o r b i c a c i d a n d h e a l s o f o u n d t h a t if d i p y r i d y l i s a d d e d to s o d i u m d i e t h y l d i t h i o c a r b a m a t e t h e a e r o b i c o x i d a t i o n of a s c o r b i c a c i d in o r a n g e j u i c e i s i n h i b i t e d . B a r r o n , B a r r o n and K l e m p e r e r (1936) s tu d i e d the o x id a tio n of a s c o r b i c a c i d a d d e d to m e d i a s u c h a s b l o o d s e r u m , u r i n e , m i l k a n d f r u i t j u i c e s , b y m e a s u r i n g t h e a m o u n t s of o x y g e n t a k e n up b y t h e s e m e d i a in a W a r b u r g a p p a r a t u s . He found th a t m o s t p lan t e x t r a c t s do n o t c o n ta in i n h i b i t o r y m e c h a n i s m s ; a s e x c e p tio n s w e r e l i s t e d t h e e x t r a c t s f r o m p l a n t s c o n t a i n i n g h i g h c o n c e n t r a t i o n of a s c o rb ic acid su ch as o ra n g e juice, to m a to ju ice and g ra p e fr u it juice. T h i s e x p e r i m e n t a l r e s u l t s u g g e s t s t h e i m p o r t a n c e of t h e S t a b i l i t y of a s c o r b i c a c i d on i t s c o n c e n t r a t i o n f o u n d i n t h e f r e s h tissu e. T h e s e a u t h o r s o b s e r v e d t h a t t h e s t a b i l i t y of a s c o r b i c a c i d , a d d e d to p l a n t e x t r a c t s w h i c h d o n o t p o s s e s s a n i n h i b i t o r y m e c h a n ­ i s m . w a s not d i r e c t l y r e l a t e d to t h e a m o u n t of c o p p e r i n t h e e x ­ tract. They c o n s id e re d h e m o c h ro m o g e n s as p o s sib le c a ta ly s ts for t h e o x i d a t i o n of a s c o r b i c a c i d in t i s s u e s w h i c h c o n t a i n no o x i d i z i n g inhibition m e c h a n is m . B a r r o n , D e M e i o a n d K l e m p e r e r (19 3 6 ) s t u d i e d t h e r a t e of o x i d a t i o n of a s c o r b i c a c i d in v a r i o u s b u f f e r s o l u t i o n s a n d t h e c a t a ­ ly sts w hich a c c e l e r a t e this oxidation. T h e y f o u n d t h a t if t h e a q u e o u s solution w as not c o n ta m in a te d w ith h eav y m e t a l and e s p e c i a l l y w ith c o p p e r , t h e o x i d a t i o n of a s c o r b i c a c i d a t 2 5 ° C f a i l e d to t a k e p l a c e up to a pH v a l u e of 7 . 6 . T h e y f o u n d t h a t in a c i t r a t e b u f f e r of pH 3 . 1 7 , t h e i n c r e a s e in t h e r a t e of o x i d a t i o n of t h e a s c o r b i c a c i d w a s l o g a r i t h m i c a l l y r e l a t e d t o i n c r e a s e in t h e a m o u n t of c o p p e r in the so lu tio n . T h e se w o r k e rs stated that iro n w as a pow erful cell c a t a l y s t w h e n it c o m b i n e s w i t h p o r p h y r i n to f o r m h e m i n . W hen h em in is joined w ith n itro g e n o u s d e r iv a tiv e s su ch a s n ico tin e, pyridine, p ilo c a rp in e , its oxidation re d u c tio n poten tial i n c r e a s e s an d t h e a u t h o r s s u g g e s t a p o s s i b l e r o l e of h e m o c h r o m o g e n in t h e o x i d a t i o n of a s c o r b i c a c i d i n p l a n t t i s s u e s . M a c k a n d K e r t e s z (19 3 6 ) s t u d i e d t h e o x i d a t i o n of k n o w n a m o u n t s of a s c o r b i c a c i d i n t h e a c e t i c e x t r a c t s of p e a p l a n t s . 10 B e c a u s e t h e r a t e of o x i d a t i o n o f th e a s c o r b i c a c i d w a s l e s s in t h e p l a n t e x t r a c t t h a n w h e n t h e s a m e a m o u n t of a s c o r b i c a c i d w a s a d d e d to t h e a s h of t h e s a m e p l a n t m a t e r i a l , t h e y c o n c l u d e d t h a t o n l y a s m a l l f r a c t i o n of t h e c o p p e r n a t u r a l l y o c c u r r i n g i n p e a s w a s in a c a t a l y t i c a l l y a c t i v e f o r m . T h e y a l s o s t a t e d t h a t i r o n ion d o es not c a t a l y z e th e o x id a tio n of a s c o r b i c a c id b u t g r e a t l y i n c r e a s e the c a t a l y s i s by c o p p e r . S t o t z , H a r r e r a n d K i n g (19 3 7 ) i n v e s t i g a t e d t h e o x i d a t i o n of a s c o r b i c a c i d in t h e p r e s s e d j u i c e of s q u a s h a n d c a u l i f l o w e r t o w h i c h k n o w n a m o u n t s o f v a r i o u s f o r m of c o p p e r w e r e a d d e d . They concluded that c o p p e r c o m b in ed w ith p ro te in can oxidize a s c o rb ic acid a t about the s a m e r a te a s in o rg a n ic c o p p e r. A n o t h e r a s p e c t of t h e i m p o r t a n c e of c o p p e r i n r e g a r d to a s c o r b i c a c i d in p l a n t s w a s e m p h a s i z e d b y P o w e r s , L e w i s a n d D a w s o n (19 4 3 ) w h o p u r i f i e d t h e e n z y m e a s c o r b i c a c i d o x i d a s e a n d s h o w e d t h a t i t c o n t a i n e d a t l e a s t 0 . 2 4 p e r c e n t of c o p p e r . B. O b j e c t of t h e P r e s e n t S t u d y T h e m o s t p e r t i n e n t s t u d i e s on t h e e f f e c t of c o p p e r a n d i r o n on t h e c o n c e n t r a t i o n of a s c o r b i c a c i d in p l a n t t i s s u e s h a v e b e e n r e v i e w e d an d found to b e v e r y l i m i t e d in n u m b e r . On the o th e r h a n d , no s t u d y w a s f o u n d i n t h e l i t e r a t u r e on t h e e f f e c t of c o p p e r a n d i r o n o n t h e s t a b i l i t y of a s c o r b i c a c i d i n p l a n t t i s s u e s . Since in v i t r o , t h e r e a c t i o n s of c o p p e r a n d i r o n a r e v e r y w e l l k n o w n a n d h a v e a n o b v i o u s e f f e c t on t h e s t a b i l i t y of a q u e o u s a s c o r b i c a c i d s o l u t i o n , i t s e e m s to b e of i m p o r t a n c e t o f i n d o u t if t h e s a m e f a c ­ t o r s a f fe c t the s ta b i l i t y of a s c o r b i c a cid in p la n t t i s s u e s . T h i s p r o b l e m i s of e s p e c i a l i m p o r t a n c e i n g r a s s b e c a u s e of i t s v a l u e a s a f o r a g e c r o p a n d a l s o b e c a u s e o f t h e i n c r e a s i n g u s e of i m m a t u r e c e r e a l g r a s s e s a s s o u r c e of v i t a m i n s f o r h u m a n consum ption. I t m a y b e n o t e d , t h a t s t u d y i n g t h e e f f e c t o f n u t r i e n t s on t h e p h y s i o l o g y of p l a n t s , m o s t a u t h o r s h a v e i n d i c a t e d t h e c h a n g e s w h i c h m i g h t o c c u r u n d e r t h e c o n d i t i o n s of t h e e x p e r i m e n t , b u t h a v e n o t c o r r e l a t e d t h e e f f e c t s w i t h t h e a m o u n t s of t h e n u t r i e n t a c t u a l l y t a k e n up b y t h e p l a n t s . The fact is known, h o w e v e r, th a t the m e r e p r e s e n c e o f o n e n u t r i e n t in t h e s u b s t r a t e m a y s o m e t i m e s g r e a t l y a f f e c t t h e a b s o r p t i o n of a n o t h e r . In th e p r e s e n t s t u d y t h e e f f e c t s of d i f f e r e n t a m o u n t s of c o p p e r a n d i r o n o n t h e c o n c e n t r a t i o n a n d s t a b i l i t y of a s c o r b i c a c i d w i l l b e c o r r e l a t e d w i t h t h e a m o u n t s of t h e s e n u t r i e n t s s u p p lie d to the p la n ts a n d found in the p l a n t s . II. EX PER IM EN TA L PROCEDURES A. E quipm ent T h e p l a n t s s tu d ie d in the f o llo w in g e x p e r i m e n t s w e r e g r o w n in a r o o m , a r r a n g e d e s p e c i a l l y f o r n u t r i e n t s o l u t i o n s t u d i e s . A c o n c r e te f lo o r, often w a s h e d , r e d u c e d c o n ta m in a tio n f r o m d u st p artic le s. T w o r o w s of t w e n t y g l a z e d e a r t h e n w a r e c o n t a i n e r s of o n e g a l l o n c a p a c i t y w e r e p l a c e d on w o o d e n b e n c h e s . The m echanical a r r a n g e m e n t of t h e c u l t u r e s h a s b e e n d e s c r i b e d b y W y n d ( 1 9 5 1 ) . The pots w e r e filled w ith n u m b e r eight c r u s h e d q u a r t z sand p r e ­ viously w a sh e d w ith tap and d istille d w a te r . A hole, t h r e e - q u a r t e r s of a n i n c h in d i a m e t e r w a s l o c a t e d a t t h e b o t t o m of t h e p o t , w h i c h p e r m i t t e d t h e n u t r i e n t s o l u t i o n to f l o o d t h e p o t f r o m b e l o w . Six­ t e e n l i t e r s of n u t r i e n t s o l u t i o n w e r e s t o r e d in f i v e - g a l l o n s o f t g l a s s c a r b o y s , w h ic h had f i r s t b e e n p a in te d b l a c k to p r e v e n t g r o w th of a l g a e a n d t h e n w i t h a l u m i n u m p a i n t in o r d e r to r e f l e c t a s m u c h h e a t and light a s p o s s ib l e . p o t s (A) a n d ( B ) . E a c h c a r b o y (C) w a s c o n n e c t e d t o t w o T he n u trie n t solution w as a u to m a tic a lly fo rc e d up f r o m t h e c a r b o y i n t o t h e p o t s b y m e a n s of a n a i r p u m p b l o w i n g a i r in to th e c l o s e d s y s t e m c o n n e c te d to all the c a r b o y s . The a ir p r e s s u r e w a s m a i n t a i n e d c o n s t a n t b y a h y d r o s t a t i c c o l u m n (E ) a n d th e h e ig h t to w h ic h th e s o lu tio n r o s e in the c u l t u r e p o ts w a s r e g u l a t e d 13 by t h e a m o u n t of w a t e r in t h e t a n k . The m echan ical a rra n g e m e n t of t h e c u l t u r e s i s i l l u s t r a t e d i n F i g u r e 1. T h e g r e e n h o u s e b e n c h s u p p o r t i n g the c u l t u r e p o ts is in d i­ c a t e d b y ( F ) , t h e c o m m o n a i r - p r e s s u r e l i n e b y (D), a n d t h e h e i g h t of t h e w a t e r i n t h e c o n t a i n e r (E) i s i n d i c a t e d b y (G)-. V ery accu rate a n d d e p e n d a b l e c o n t r o l of t h e r i s e of t h e n u t r i e n t s o l u t i o n in t h e c u ltu re pots w as attain ed by this d ev ice. E v e r y f o u r h o u r s an e l e c t r i c t i m e s w i t c h t u r n e d on t h e a i r p u m p , a n d t h e n u t r i e n t s o l u ­ t i o n w a s f o r c e d up i n t o th e p o t s w h e r e i t r e m a i n e d f o r a b o u t t e n m inutes. W hen the a i r p r e s s u r e w a s r e l e a s e d the s o lu tio n d r a i n e d b a c k into the c a r b o y s . B. N u trie n t Solutions T h e c o m p l e t e n u t r i e n t s o l u t i o n w a s t h a t of S h i v e ( 1915 ) f o r b e s t g r o w t h of w h e a t t o p s : Its final c o m p o s itio n w as: C o m p o s i t i o n of t h e N u t r i e n t S o l u t i o n Salts G ram s per L iter MgSO • 7 H O 5 • 4 2 3.697 Ca(NO ) • 4 3 2 1.228 KH P O 2 4 H O 2 2.450 14 C o n c e n t r a t e d s o l u t i o n s of e a c h s a l t s w e r e m a d e up a s f o l lo w s : C o m p o s i t i o n of t h e S t o c k S o l u t i o n s Salts G r a m s in 16 L i t e r s M gS04 • 7 HzO 5502.0 C a ( N 0 3)2 • 4 H20 1827.3 kh 2p o 4 3646.0 F i f t e e n l i t e r s of t r i p l e - d i s t i l l e d w a t e r w e r e a d d e d to e a c h c a r b o y , a n d t h e n 178 m i l l i l i t e r s of e a c h of t h e s t o c k s o l u t i o n a n d 534 m i l l i l i t e r s of w a t e r w e r e a d d e d , w h i c h b r o u g h t t h e f i n a l v o l ­ u m e to 1 6 l i t e r s . M i c r o e l e m e n t s w e r e s u p p l i e d b y a d d i n g o n e m i l l i l i t e r of a s t o c k s o l u t i o n p e r l i t e r of n u t r i e n t s o l u t i o n . T he sto ck solution of m i c r o e l e m e n t s h a d t h e f o l l o w i n g c o m p o s i t i o n : 15 S t o c k S o l u t i o n of M i c r o E l e m e n t s Salts G ram s per 2 L iters H 3B 0 3 5.720 Z nS 04 • 7 HzO 0.440 M nC l2 2.290 M oQ 3 0.015 T h e f i n a l c o n c e n t r a t i o n s of t h e m i c r o e l e m e n t s in t h e n u ­ t r i e n t so lu tio n w e r e a s follow s: C o n c e n t r a t i o n s of t h e M i c r o E l e m e n t s in t h e N u t r i e n t S o l u t i o n E lem ent P a r t s p e r M illion Boron 0.50 Zinc 0.05 M anganese 0.50 M olybdenum 0.005 C opper and iro n w e r e added s e p a r a te ly at v a rio u s c o n c e n ­ t r a t i o n s a s w ill b e e x p la in e d f o r the d i f f e r e n t e x p e r i m e n t s . 16 T h e i n i t i a l pH of t h e s o l u t i o n s w a s a b o u t 5 . 2 . T h i s pH w a s c h e c k e d a t r e g u l a r i n t e r v a l s d u r i n g t h e g r o w t h p e r i o d of t h e p l a n t s . W a t e r w a s a d d e d p e r i o d i c a l l y to c o m p e n s a t e f o r th a t l o s t by t r a n ­ spiration. O th e r f a c t o r s w h ich w e r e v a r i e d f o r the d if f e r e n t s tu d ie s will be e x p la in e d f o r e a c h e x p e r i m e n t . III. EX PERIM EN T 1 A. Purpose T h e f o l l o w i n g e x p e r i m e n t w a s p l a n n e d in o r d e r to d i s c o v e r w h e t h e r o r n o t i r o n a n d c o p p e r s u p p l i e d to w h e a t p l a n t s m i g h t c a t a l y z e t h e o x i d a t i o n of t h e a s c o r b i c a c i d p r e s e n t in th e p l a n t t i s ­ s u e , in t h e s a m e m a n n e r a s i t h a s b e e n m e n t i o n e d in t h e l i t e r a t u r e in t h e c a s e of in v i t r o e x p e r i m e n t s . T h e p u r p o s e of t h i s f i r s t e x p e r i m e n t w a s to i n v e s t i g a t e t h e r e l a t i o n s h i p s b e t w e e n i n c r e a s i n g a m o u n t s of c o p p e r a n d c o n s t a n t i r o n c o n c e n t r a t i o n in t h e n u t r i e n t s o l u t i o n on t h e c o n c e n t r a t i o n of a s c o r b i c a c i d in t h e f r e s h w h e a t t i s s u e a n d on t h e s t a b i l i t y of t h i s v i t a m i n in t h e d r i e d t i s s u e . In t h e s a m e m a n n e r , it w a s p r o p o s e d to i n v e s t i g a t e t h e s a m e r e l a t i o n s h i p s w i t h i n c r e a s i n g a m o u n t s of i r o n a n d c o n s t a n t c o p p e r c o n c e n t r a t i o n in t h e n u t r i e n t s o l u t i o n . J u s t b e f o r e jointing s ta g e , the a s c o r b i c acid w a s d e t e r m in e d in t h e f r e s h t i s s u e s . T h e r e m a i n d e r of th e p l a n t m a t e r i a l w a s d r i e d a n d a s c o r b i c a c i d w a s d e t e r m i n e d a t i n t e r v a l s to o b t a i n i t s r a t e of oxidation. A n a l y s e s of i r o n a n d o t h e r c o m p o n e n t s of t h e p l a n t m a ­ t e r i a l s s u c h a s c a r b o h y d r a t e , n i t r o g e n and c h lo r o p h y ll, w e r e c a r r i e d out on the p la n t t i s s u e s . Any p o s sib le r e la tio n s h ip s b etw een th e se c o m p o n e n t s a n d t h e a s c o r b i c a c i d s t a t u s of t h e w h e a t p l a n t s w i l l b e d i s c u s s e d in t h e p r e s e n t a t i o n of t h e d a t a . B. 1. E x p e rim e n ta l Conditions I r o n a n d C o p p e r L e v e l s in t h e N u t r i e n t S o l u t i o n A f t e r t h e c o m p l e t e n u t r i e n t s o l u t i o n s w e r e m a d e up, t h e f o l l o w i n g a m o u n t s of i r o n a n d c o p p e r w e r e a d d e d : C o n c e n t r a t i o n of I r o n a n d C o p p e r in t h e N u t r i e n t S o l u t i o n s of t h e F i r s t E x p e r i m e n t Iron S e rie s Copper S eries p p m of F e i n the Solution p p m of C u in the Solution 0.0 1.0 2.0 3.0 4.0 5.0 6.0 7. 0 8.0 9. 0 0.02 0.02 0.02 0.02 0.02 0.02 0.02 0.02 0 .02 0.02 PPm F e in the S o lu tio n 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 p p m of C u in the S o lu tio n 0.00 0.01 0.02 0.03 0.04 0.05 0.06 0.07 0.08 0.09 T h e s e t w o s e r i e s of c u l t u r e s w i l l l a t e r b e m e n t i o n e d a s the “ ir o n s e r i e s " and the “ c o p p e r s e r i e s . " 19 T h e s t o c k s o l u t i o n of i r o n w a s p r e p a r e d b y d i s s o l v i n g 1.6 g r a m s of e l e c t r o l y t i c i r o n in t h r e e m i l l i l i t e r s of c o n c e n t r a t e d s u l ­ furic acid. T h e v o l u m e of t h i s s o l u t i o n w a s b r o u g h t to 500 m i l l i ­ l i t e r s w ith d is tille d w a te r . F i v e m i l l i l i t e r s of t h i s s t o c k s o l u t i o n in 16 l i t e r s of n u t r i e n t s o l u t i o n c o r r e s p o n d t o o n e p a r t p e r m i l l i o n of i r o n . No a t t e m p t w a s m a d e to k e e p t h e i r o n l e v e l c o n s t a n t d u r ­ ing t h e e x p e r i m e n t a l p e r i o d . T h e s t o c k s o l u t i o n of c o p p e r w a s p r e p a r e d b y d i s s o l v i n g 63 m i l l i g r a m s of c o p p e r s u l f a t e ( C u S O ^ • 5 H^O) in d i s t i l l e d w a t e r . T h e v o l u m e of t h i s s o l u t i o n w a s a d j u s t e d to 500 m i l l i l i t e r s . F ive m i l l i l i t e r s of t h i s s t o c k s o l u t i o n in 16 l i t e r s of n u t r i e n t s o l u t i o n c o r r e s p o n d to 0..01 p a r t p e r m i l l i o n of c o p p e r . D u r i n g t h e g r o w t h of t h e p l a n t s , t h e n u t r i e n t s o l u t i o n s w e r e r e n e w e d 13 d a y s a f t e r s e e d i n g . 2. P lan t M aterial T h i r t y s e e d s of w h e a t , “ I l l i n o i s N o . 1 - 1 2 8 , ” h a r v e s t e d in 1 9 4 7 fc w h i c h w a s s u p p l i e d by t h e M i c h i g a n S t a t e C o l l e g e F a r m C r o p s D e p a r t m e n t , w e r e s o w n in e a c h p o t on M a y 24, 1 948. The seeds w e r e e v e n l y s p a c e d a n d c o v e r e d w i t h a b o u t o n e - h a l f i n c h of g r a v e l . D u rin g g e r m in a tio n , the p o ts w e r e c o v e r e d w ith c a r d b o a r d to p r e ­ v e n t e v a p o r a tio n and to s e c u r e e v e n g e r m i n a t i o n . h a r v e s t e d on J u n e 28, 1948, 36 d a y s a f t e r s e e d i n g . The plants w e re All the p la n ts a t t h a t t i m e w e r e s t a r t i n g to j o i n t . T he jointing stag e w as ch o se a s t h e t i m e of h a r v e s t b e c a u s e , a s s h o w n b y K o h l e r ( 1 9 4 4 ) . t h e c o m p o s i t i o n of g r a s s a t t h a t s t a g e i s e s p e c i a l l y h i g h in a s c o r b i c acid as well as m a n y o th e r o rg a n ic c o n s titu e n ts . C. 1. a. E xperim ental R esults O b s e r v a t i o n s on t h e P l a n t s D e s c r i p t i o n o f t h e P l a n t s a t T i m e of H a r v e s t At h a r v e s t t i m e all the p la n ts had f o r m e d the f i r s t joint. T h e p l a n t s of t h e i r o n s e r i e s h a d g e n e r a l l y t h i c k e r s t e m s and g r e e n e r l e a v e s t h a n t h e o n e s of t h e c o p p e r s e r i e s . H o w e v e r . th« d i f f e r e n c e s b e t w e e n t h e two s e r i e s w e r e n o t v e r y s t r i k i n g . P l a n t s g r o w n on t h e n u t r i e n t s o l u t i o n l a c k i n g i r o n s t a r t e d to s h o w c h l o r o s i s a b o u t 20 d a y s a f t e r s e e d i n g . An i n c r e a s e in tl g r o w t h of t h e p l a n t s w a s o b s e r v e d t o b e a b o u t p r o p o r t i o n a l to th< i n c r e m e m t s of i r o n in t h e s o l u t i o n u p to s i x p a r t s p e r m i l l i o n , j t h a t p o i n t t h e g r o w t h s h o w e d a t e n d e n c y to l e s s e n . T h e p l a n t s on t h e c o p p e r s e r i e s , t h o u g h s m a l l e r in s i z e t h a n t h e o n e s of t h e i r o n s e r i e s , w e r e a l l n o r m a l a n d s h o w e d i n ­ c r e a s e d g r o w t h f r o m n o c o p p e r to 0 . 0 6 p a r t s p e r m i l l i o n of co p p in t h e n u t r i e n t s o l u t i o n . T hen g r o w t h w a s l e s s e n e d and the plant 20 at th a t tim e w e r e s ta r t i n g to joint. The jointing stag e w as c h o se n a s t h e t i m e of h a r v e s t b e c a u s e , a s s h o w n b y K o h l e r ( 194 4), t h e c o m p o s i t i o n of g r a s s a t t h a t s t a g e i s e s p e c i a l l y h i g h i n a s c o r b i c acid as w ell as m an y oth er organ ic co n stitu en ts. C. 1. a. E xperim ental R esults O b s e r v a t i o n s on t h e P l a n t s D e s c r i p t i o n of t h e P l a n t s a t T i m e of H a r v e s t At h a r v e s t tim e a ll the p la n ts had f o r m e d the f i r s t joint. T h e p l a n t s of t h e i r o n s e r i e s h a d g e n e r a l l y t h i c k e r s t e m s a n d g r e e n e r l e a v e s t h a n t h e o n e s of t h e c o p p e r s e r i e s . H o w e v e r, the d i f f e r e n c e s b e t w e e n t h e tw o s e r i e s w e r e n o t v e r y s t r i k i n g . P l a n t s g r o w n on t h e n u t r i e n t s o l u t i o n l a c k i n g i r o n s t a r t e d to s h o w c h l o r o s i s a b o u t 20 d a y s a f t e r s e e d i n g . An i n c r e a s e i n t h e g r o w t h of t h e p l a n t s w a s o b s e r v e d t o b e a b o u t p r o p o r t i o n a l t o t h e i n c r e m e m t s of i r o n i n th e s o l u t i o n u p to s i x p a r t s p e r m i l l i o n . A t th a t po in t the g ro w th show ed a te n d e n c y to l e s s e n . T he p la n ts on the c o p p e r s e r i e s , though s m a l l e r in s iz e t h a n t h e o n e s of t h e i r o n s e r i e s , w e r e a l l n o r m a l a n d s h o w e d i n ­ c r e a s e d g r o w t h f r o m no c o p p e r to 0 . 0 6 p a r t s p e r m i l l i o n of c o p p e r in t h e n u t r i e n t s o l u t i o n . T h e n g ro w th w as l e s s e n e d and the plants 21 l o o k e d p o o r e r in t h e f o l l o w i n g c u l t u r e s c o n t a i n i n g i n c r e a s i n g a m o u n t s of c o p p e r i n th e n u t r i e n t s o l u t i o n . b. F r e s h W e i g h t of t h e P l a n t s (1) E f f e c t of I r o n The data indicating the a v e ra g e f r e s h w eight p e r ten plants e x p r e s s e d i n g r a m s a r e a s s e m b l e d i n T a b l e 1, a n d a r e g r a p h e d a g a i n s t t h e a m o u n t of i r o n i n t h e s o l u t i o n i n F i g u r e 2. a r e the a v e r a g e v a lu e s of two d u p lic a te c u l t u r e s . T h e se data If the v a lu e s ob­ t a i n e d f o r t h e p l a n t s g r o w n o n t h r e e p a r t s p e r m i l l i o n of i r o n a r e d i s r e g a r d e d , a n i n c r e a s e of t h e f r e s h w e i g h t up t o s i x p a r t s p e r m i l l i o n of i r o n i s a p p a r e n t . T h e n i t f a l l s off t o r e a c h a p p r o x i m a t e l y the s a m e value a s fo r the p la n ts g ro w n w ithout iro n . (2) E f f e c t of C o p p e r The d ata indicating the a v e r a g e f r e s h w eight p e r ten plants e x p r e s s e d in g r a m s a r e a s s e m b l e d in T ab le 3 and a r e g ra p h e d a g a i n s t t h e a m o u n t s of c o p p e r i n t h e s o l u t i o n i n F i g u r e 45. T h e e f f e c t of c o p p e r o n g r o w t h a s r e f l e c t e d f r o m t h e f r e s h w e i g h t of t h e p l a n t s i s p r o n o u n c e d . The grow th w as in c re a sin g ly s t i m u l a t e d f r o m t h e c u l t u r e w i t h o u t c o p p e r t o t h e o n e w i t h 0 .06 p a r t s p e r m i l l i o n of c o p p e r . Then, a s h a rp decline b rought back I 22 the f r e s h w eight p e r p la n t to a v alu e lo w e r th a n fo r the p la n ts grow n without co p p er. c. D ry W eight p e r P la n t (1) E f f e c t of I r o n The data fo r the a v e ra g e d r y w eight p e r ten plants e x p re s s e d in g r a m s a r e in d ic a te d in T a b le 1 and a r e g r a p h e d a g a i n s t the a m o u n t of i r o n i n t h e s o l u t i o n i n F i g u r e 3. A g e n e r a l u p w a r d te n d e n c y w as p r e s e n t e d b y the d r y w eight of t h e p l a n t s w i t h i n c r e a s i n g i r o n i n t h e s o l u t i o n . T his tendency d i d n o t d e c l i n e a s in t h e c a s e of t h e f r e s h w e i g h t w h e n t h e a m o u n t of i r o n i n t h e s o l u t i o n w a s l a r g e r t h a n s i x p a r t s p e r m i l l i o n . (2) E f f e c t of C o p p e r T h e a v e r a g e d r y w e i g h t s p e r t e n p l a n t s of t h e c o p p e r s e r i e s a r e a r r a n g e d i n T a b l e 3 a n d g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r i n t h e s o l u t i o n i n F i g u r e 46. I n t h i s s e r i e s of c u l t u r e s , t h e d r y w e i g h t of t h e p l a n t s p a r a l ­ le le d the f r e s h w eig h t v a lu e s p e r p la n ts, showing f i r s t an i n c r e a s e up t o 0 . 0 6 p a r t s p e r m i l l i o n of c o p p e r i n t h e n u t r i e n t s o l u t i o n an d then a d e c r e a s e to a value lo w e r than fo r the p lan ts grow n w ithout copper. 23 2. C h e m i c a l A n a l y s e s of t h e P l a n t s In o r d e r to b e ab le to c o m p a r e m o r e e a s i l y th e d a t a f o r i r o n a b s o r p t i o n a n d t h e a m o u n t s of a s c o r b i c a c i d f o u n d a t t h e v a r i o u s d e t e r m i n a t i o n s , t h e s e two c o m p o n e n t s w i l l f i r s t b e d i s c u s s e d . The o th e r p la n t c o m p o n e n ts stu d ied w ill be ta k e n in c o n s id e ra tio n a t the end of t h i s p a r a g r a p h on c h e m i c a l a n a l y s e s . a. Iro n (1) A m o u n t s o f I r o n p e r P l a n t (a) E f f e c t of i n c r e a s i n g t h e a m o u n t s of i r o n i n t h e s o l u t i o n on t h e i r o n c o n t e n t of t h e p l a n t s . T h e m i l l i g r a m s of i r o n o b t a i n e d p e r t e n p l a n t s a r e p r e s e n t e d i n T a b l e 1 a n d g r a p h e d a g a i n s t th e a m o u n t of i r o n i n t h e n u t r i e n t s o l u t i o n i n F i g u r e 4. A n i n c r e a s e i n t h e i r o n c o n t e n t of t h e p l a n t s w a s o b s e r v e d f o l l o w i n g i n c r e a s e of i r o n i n t h e n u t r i e n t s o l u t i o n b u t t h i s i n c r e a s e was not a stra ig h t relationship. On t h e o t h e r h a n d , d i f f e r e n c e s i n t h e a c c u m u l a t i o n of i r o n c o r r e s p o n d e d t o c h a n g e s i n d r y w e i g h t of the d ifferen t c u ltu re s. This m ig h t su g g est th at in the plants m o s t of t h e i r o n w a s t i e d t o t h e c o n s t i t u e n t s f o r m i n g t h e d r y m a t e r i a l . This c o n s id e ra tio n is stren g th en ed by the fa c t th a t the n itro g en p o r t i o n f o l l o w e d v e r y c l o s e l y t h e a m o u n t of i r o n a c c u m u l a t e d . An­ o t h e r c r i t e r i o n of t h e i m p o r t a n c e of i r o n i n t h e p l a n t i s t h e c l o s e 24 c o r r e l a t i o n b e t w e e n t h e a m o u n t of c h l o r o p h y l l a n d t h e t o t a l a m o u n t of i r o n , a s s h o w n i n F i g u r e 13. The c o r re la tio n betw een c h lo ro ­ phyll and ir o n h as b e e n o b s e rv e d by m a n y a u th o rs b u t u s u a lly only a re la tio n s h ip b etw een the s o -c a lle d active iro n and ch lorophy ll is noted. The p r e s e n t data show p o sitiv e ly th at s tra ig h t re la tio n sh ip b e tw e e n total iro n and ch lo ro p h y ll m a y o c c u r. (b) E f f e c t of i n c r e a s i n g t h e a m o u n t of c o p p e r i n t h e s o l u t i o n on t h e i r o n c o n t e n t of t h e p l a n t s . T a b l e 3 i n d i c a t e s t h e m i l l i g r a m s of i r o n a b s o r b e d p e r t e n p l a n t s i n t h e c o p p e r s e r i e s ; t h e s e v a l u e s a r e g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r i n t h e s o l u t i o n i n F i g u r e 47. I n a s m u c h a s t h e s a m e a m o u n t of i r o n h a d b e e n a d d e d to t h e c o m p l e t e n u t r i e n t s o l u t i o n s a t t h e d i f f e r e n t l e v e l s of c o p p e r , i t s h o u l d f o l l o w t h a t a n y d i f f e r e n c e i n t h e a m o u n t of i r o n a c c u m u l a t e d i n t h e p l a n t i s i n t r i n s i c a l l y d u e t o t h e i n f l u e n c e of d i f f e r e n c e s i n t h e c o p p e r c o n t e n t of t h e n u t r i e n t s o l u t i o n . Up t o 0 . 0 6 p a r t s p e r m i l ­ l i o n of c o p p e r i n t h e n u t r i e n t s o l u t i o n c o r r e s p o n d e d t o a n i n c r e a s e i n t h e a m o u n t of i r o n t a k e n up w h i c h p a r a l l e l e d v e r y c l o s e l y t h e a m o u n t of d r y m a t t e r ( F i g u r e 44) a n d c a r b o h y d r a t e ( F i g u r e 43) . It should be noted th a t su c h a r e la tio n w ith the c a r b o h y d ra te f r a c ­ tion had n ot b e e n found in the i r o n s e r i e s . 25 (2) A m o u n t of I r o n p e r 100 G r a m s of F r e s h P l a n t M a t e r i a l (a) E f f e c t of i r o n . T h e m i l l i g r a m s of i r o n p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l a p p e a r i n T a b l e 5 a n d F i g u r e 16. If t h e r e s u l t s o b t a i n e d f o r t h e a m o u n t of i r o n i n e a c h s a m p l e a r e c a lc u la te d on a f r e s h m a t t e r b a s i s i t is p o s s ib le to g e t a n id e a of t h e a c t u a l c o n c e n t r a t i o n of t h a t e l e m e n t i n t h e l i v i n g p l a n t t i s s u e . A s l i g h t u p w a r d t e n d e n c y i n t h e a m o u n t of i r o n i n t h e f r e s h m a t t e r c o r r e s p o n d i n g to i n c r e a s e d a m o u n t of i r o n a d d e d t o t h e n u t r i e n t s o l u t i o n c a n b e n o t i c e d b u t t h i s i n c r e a s e i s n o t a l i n e a r f u n c t i o n of the a m o u n t in the solution. The positive relatio n sh ip with nitro g en w h ic h h a s b e e n found in the d a ta p e r p la n t is a g a in found w hen t h e s e d a t a a r e c a l c u l a t e d on a f r e s h w e i g h t b a s i s b u t i t d o e s n o t h o l d a s closely. T h i s r e l a t i o n s h i p i s p r e s e n t e d i n F i g u r e 25. In t h e s a m e m a n n e r the re la tio n sh ip with chlorophyll is not as c l e a r as before, F i g u r e 38. (b) E f f e c t of c o p p e r . T h e d a t a i n d i c a t i n g t h e m i l l i g r a m s of i r o n p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l of t h e c o p p e r s e r i e s a r e s h o w n i n T a b l e 7 a n d F i g u r e 51. T h e c o n c e n t r a t i o n of i r o n i n t h e f r e s h t i s s u e s h o w e d a s l i g h t d e c r e a s i n g tre n d w ith i n c r e a s i n g c o p p e r in th e solution. This tre n d is w ell c o r r e l a t e d w ith th e d ata fo r d r y w eig h t as shown g ra p h ic a lly in F i g u r e 60 a n d l o o s e l y w i t h t h e t o t a l c a r b o h y d r a t e , F i g u r e 6 l . 26 B o t h of t h e s e r e l a t i o n s h i p s a r e v e r y s t r i k i n g w h e n t h e d a t a f o r t h e a m o u n t of i r o n p e r p l a n t a r e c o m p u t e d . (3) A m o u n t of I r o n p e r 100 G r a m s of D r y M a t e r i a l (a) E f f e c t of i r o n . T h e a m o u n t of i r o n i n m i l l i g r a m s p e r 100 g r a m s of d r y p l a n t m a t e r i a l a r e p r e s e n t e d i n T a b l e 9 a n d F i g ­ u r e 31. T he a m o u n ts obtained f o r the i r o n s e r i e s do n ot show any p r e c i s e tendency. A fte r e x a m in in g all the d a ta obtained, no r e l a ­ t i o n c o u l d b e f o u n d a n d t h e o n e s w h i c h e x i s t e d i n t h e c a s e of t h e a m o u n t of i r o n p e r p l a n t o r i n t h e f r e s h t i s s u e do n o t h o l d t r u e when c a lc u la te d fo r the ir o n in the d r y p la n t m a t e r ia l. (b) E f f e c t of c o p p e r . T h e d a t a f o r t h e a m o u n t of i r o n i n m i l l i g r a m s p e r 100 g r a m s d r y m a t e r i a l i n t h e c o p p e r s e r i e s a r e a s s e m b l e d i n T a b l e 11 a n d F i g u r e 63. W i t h i n c r e a s i n g a m o u n t of c o p p e r i n t h e s o l u t i o n l e s s i r o n w as obtained in the d r y p lan t m a t e r i a l . The relatio n sh ip w hich had b e e n found b e tw e e n ir o n and c a r b o h y d ra te does not hold tr u e when c a l c u l a t e d on a d r y m a t t e r b a s i s a n d n o o t h e r r e l a t i o n s h i p w a s fo u n d . 27 b. A s c o rb ic Acid (1) D e s c r i p t i o n F o r t h e p u r p o s e of s t u d y i n g t h e p o s s i b l e e f f e c t s of i r o n a n d c o p p e r o n t h e t o t a l a m o u n t s of a s c o r b i c a c i d i n t h e w h e a t p la n ts and on its s ta b ility a f t e r d r y in g , four d i f f e r e n t d e t e r m i n a ­ tions w e r e m ad e a t v a rio u s in te rv a ls . The f i r s t d e t e r m in a ti o n on b o t h s e r i e s w a s d o n e a t t h e t i m e of h a r v e s t , t h e s e c o n d a f t e r t h e r e m a i n d e r of t h e p l a n t m a t e r i a l w a s d r i e d i n a f o r c e d a i r o v e n a t 6 0 ° C f o r t h r e e d a y s , t h e t h i r d 31 d a y s l a t e r a n d t h e l a s t 105 d a y s a f t e r d ry in g , the s a m p l e s b e in g k e p t a t r o o m t e m p e r a t u r e . The v a l u e s o b t a i n e d a t e a c h d e t e r m i n a t i o n w e r e c a l c u l a t e d on a b a s i s p e r 10 p l a n t s , p e r 100 g r a m s f r e s h m a t e r i a l a n d p e r 100 g r a m s d ry m a te ria l; each r e s u lt w ill be d is c u s s e d s e p a ra te ly , T h e p e r c e n t a g e of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d left a f t e r e a c h d e te r m in a tio n w as a l s o c a lc u la te d in o r d e r to get a n i d e a of t h e r e l a t i v e r a t e of o x i d a t i o n of t h e a s c o r b i c a c i d i n t h e p la n ts f r o m the d iffe re n t c u ltu r e s . When t h e s e v a lu e s a r e su b ­ t r a c t e d f r o m h u n d r e d t h e p e r c e n t a g e s of l o s s i s o b t a i n e d . (2) A m o u n t of A s c o r b i c A c i d p e r P l a n t (a) E f f e c t of i r o n . T h e d a t a e x p r e s s i n g t h e a m o u n t s of a s c o r b i c a c i d , a t h a r v e s t , a f t e r d r y i n g , 31 d a y s a n d 105 d a y s 28 a f t e r d r y i n g p e r t e n p l a n t s g r o w n a t v a r i o u s l e v e l s of i r o n a r e a s s e m b l e d i n T a b l e 2 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t of i r o n i n t h e s o l u t i o n i n F i g u r e s 5, 6, 7„, a n d 8. T h e t o t a l a m o u n t s of a s c o r b i c a c i d a t h a r v e s t w h e n c a l c u ­ lated p e r p la n t did n o t show any r e la tio n s h ip n e i t h e r w ith the a m o u n t of i r o n i n t h e s o l u t i o n n o r w i t h t h e a m o u n t of i r o n t a k e n up b y t h e p l a n t s . We m ig h t s a y th a t t h e r e w as a slight tendency to d e c r e a s e w i t h i n c r e a s e i r o n i n t h e s o l u t i o n b u t t h i s t r e n d w a s not a t all definite. (b) E f f e c t of c o p p e r . T h e d a t a f o r t h e a m o u n t s of a s c o r b i c a c i d a t h a r v e s t a f t e r d r y i n g , 31 d a y s a n d 105 d a y s a f t e r d r y i n g p e r t e n p l a n t s of t h e c o p p e r s e r i e s a r e p r e s e n t e d i n T a b l e 4 a n d F i g ­ u r e s 48, 49, 50, a n d 51. A d e f i n i t e t r e n d i n t h e a m o u n t of a s c o r b i c a c i d i n t h e p l a n t s g r o w n on t h e c o p p e r s e r i e s w o u l d b e d i f f i c u l t t o i n d i c a t e a l t h o u g h it c a n be s e e n th a t a s the f r e s h w eig h t and a s the d r y w eight, the a m o u n t s of a s c o r b i c a c i d i n c r e a s e d i n t h e p l a n t s f r o m t h e n o c o p ­ p e r l e v e l t o t h e o n e g r o w n o n 0..06 p a r t s p e r m i l l i o n of c o p p e r a n d th en d e c r e a s e d in the s u b se q u e n t c u l t u r e s . The sam e trend p e r ­ s i s t e d b u t l e s s a c c e n t u a t e d f o r t h e v a l u e s of a s c o r b i c a c i d o b t a i n e d j u s t a f t e r d r y i n g a n d 31 d a y s a f t e r d r y i n g . E i g h t y - f i v e t o 95 p e r 29 c e n t of t h e i n i t i a l a m o u n t of a s c o r b i c a c i d h a d b e e n l o s t a f t e r 105 d a y s a n d no t r e n d w a s s h o w n a n y m o r e . (3) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s F r e s h M a t t e r (a) E f f e c t of i r o n . T h e d a t a i n m i l l i g r a m s p e r 100 g r a m s f r e s h m a t t e r of a s c o r b i c a c i d a t h a r v e s t , a f t e r d r y i n g , 31 d a y s a n d 105 d a y s a f t e r d r y i n g , of t h e i r o n s e r i e s a p p e a r i n T a b l e 6 a n d i n F i g u r e s 17, 18, 19, a n d 20. T h e a m o u n t s of a s c o r b i c a c i d a t h a r v e s t d i d n o t b e a r a n y a p p a r e n t r e l a t i o n s h i p w i t h t h e o t h e r c o n s t i t u e n t s of t h e p l a n t . Af­ t e r d r y i n g t h e v a l u e s f o r a s c o r b i c a c i d h a d a t e n d e n c y to b e h i g h e r i n t h e p l a n t g r o w n on h i g h e r i r o n c o n t e n t i n t h e s o l u t i o n s a n d t h i s r e l a t i o n w a s k e p t t i l l 105 d a y s a f t e r d r y i n g . (b) E f f e c t of c o p p e r . T h e m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l g r o w n i n t h e c o p p e r s e r i e s a n d d e t e r m i n e d a t h a r v e s t a f t e r d r y i n g , 31 d a y s a n d 105 d a y s a f t e r d r y i n g a r e r e c o r d e d i n T a b l e 8 a n d F i g u r e s 52, 53, 54, a n d 55. T h e a m o u n t of c o p p e r i n t h e s o l u t i o n d i d n o t h a v e a n y a p ­ p a r e n t e f f e c t on t h e a s c o r b i c a c i d a t h a r v e s t i n t h e f r e s h m a t e r i a l , h o w ev er a f t e r d ry in g the p la n ts w h ic h had b e e n grow n on the h ig h e s t a m o u n t of c o p p e r k e p t t h e m o s t a m o u n t of a s c o r b i c a c i d . 30 (4) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s D r y P l a n t M a t e r i a l (a) E f f e c t o f i r o n . The d ata e x p re s s in g in m illig r a m s p e r 100 g r a m s of d r y p l a n t m a t e r i a l t h e a m o u n t of a s c o r b i c a c i d a t h a r v e s t , a f t e r d r y i n g , 31 d a y s a n d 105 d a y s a f t e r d r y i n g of t h e p l a n t g r o w n o n t h e i r o n s e r i e s a r e p r e s e n t e d i n T a b l e 10 a n d F i g ­ u r e s 32, 33, 34, a n d 35. W h e n c a l c u l a t e d o n a d r y m a t t e r b a s i s t h e a m o u n t of a s c o r b i c a c i d a t h a r v e s t d e c r e a s e d w i t h th e a m o u n t of i r o n i n t h e s o l u t i o n . A f t e r t h e m a t e r i a l h a d b e e n d r i e d t h e a m o u n t of a s c o r b i c a cid left w as a lm o s t the s a m e in all the c u ltu r e s , pointing out the fa c t th at in the p lan ts grow n w ithout ir o n the a s c o rb ic acid w as lost fa ste r. T h is c o n c lu s io n should n o t be g e n e r a liz e d in r e g a r d to t h e e f f e c t of i r o n i n th e p l a n t on t h e a s c o r b i c a c i d p r e s e n t , b e ­ c a u s e t h e a m o u n t of i r o n f o u n d i n t h e d r y m a t t e r w a s n o t p r o p o r ­ t i o n a l t o t h e a m o u n t of i r o n i n t h e s o l u t i o n a n d n o r e l a t i o n s h i p c o u l d be d r a w n b e tw e e n a s c o r b i c acid and th e a m o u n t of i r o n in the p lant. (b) E f f e c t of c o p p e r . T h e a m o u n t i n m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of d r y p l a n t m a t e r i a l g r o w n o n t h e c o p p e r s e r ­ i e s d e t e r m i n e d a t h a r v e s t , j u s t a f t e r d r y i n g , 31 d a y s a n d 105 d a y s a f t e r d r y i n g a r e a s s e m b l e d i n T a b l e 12 a n d g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r i n F i g u r e 64, 65, 66, a n d 67. 31 In th e d r y m a t t e r of the p la n ts g ro w n a t d i f f e r e n t le v e ls of c o p p e r , t h e a m o u n t o f a s c o r b i c a c i d a t h a r v e s t d i d n o t f o l l o w any c l e a r re la tio n sh ip . A f t e r d r y i n g t h e a m o u n t of a s c o r b i c a c i d b e c a m e a l m o s t i n v e r s e l y p r o p o r t i o n a l to t h e a m o u n t of i r o n i n th e d r y m a t t e r a s c a n b e s e e n i n F i g u r e 72, b u t t h i s r e l a t i o n s h i p d id not hold a t the n e x t d e t e r m i n a t i o n s . (5) P e r c e n t a g e s of t h e O r i g i n a l A m o u n t of A s c o r b i c A c i d F o u n d a t E ach D eterm ination (a) E f f e c t of i r o n . T h e v a l u e s of a s c o r b i c a c i d c a l c u l a t e d a s p e r c e n t a g e s of t h e o r i g i n a l a m o u n t s a t e a c h d e t e r m i n a t i o n a r e p r e s e n t e d f o r t h e i r o n s e r i e s in T a b l e 13 a n d F i g u r e 73. T h e p e r c e n t a g e v a l u e s o b t a i n e d a r e i n d e p e n d e n t of t h e b a s i s of c a l c u l a t i o n of t h e a s c o r b i c a c i d a n d a c c o r d i n g l y c a n b e r e l a t e d to t h e o t h e r c h e m i c a l d a t a o b t a i n e d on a p e r p l a n t , p e r 100 g r a m s f r e s h m a t t e r a n d p e r 100 g r a m s d r y m a t t e r b a s i s . U p o n d r y i n g , a s c o r b i c a c i d w a s l o s t a t a s l o w e r r a t e i n th e p l a n t m a t e r i a l s g r o w n on t h e c u l t u r e s s u p p l i e d w i t h h i g h a m o u n t s of i r o n . I n d e e d , o n l y 14 p e r c e n t o f t h e o r i g i n a l a m o u n t o f a s c o r ­ b i c a c i d w e r e l o s t a f t e r d r y i n g f r o m t h e p l a n t m a t e r i a l g r o w n on n i n e p a r t s p e r m i l l i o n o f i r o n , w h e r e a s 38 p e r c e n t w e r e l o s t f r o m t h e p l a n t m a t e r i a l g r o w n o n t h e c u l t u r e w h e r e no i r o n w a s a d d e d . A p o s i t i v e c o r r e l a t i o n b e t w e e n t h e p e r c e n t a g e of t h e o r i g i n a l a m o u n t 32 of a s c o r b i c a c i d f o u n d a f t e r d r y i n g a n d 31 d a y s l a t e r a n d t h e c o n c e n ­ t r a t i o n of i r o n i n t h e s o l u t i o n c a n b e s e e n f r o m F i g u r e 73; a f t e r 105 days this c o r r e l a t i o n s till holds but not a s s tro n g as fo r the o th e r values. W h e n t h e p e r c e n t a g e s a r e g r a p h e d a g a i n s t t h e a m o u n t of i r o n i n t h e f r e s h m a t e r i a l o r in t h e d r y m a t t e r ( F i g u r e 74) n o a p ­ p a r e n t c o r r e l a t i o n is b ro u g h t about, but a p o s itiv e c o r r e l a t i o n , as s e e n i n F i g u r e 27, w a s o b s e r v e d b e t w e e n t h e p e r c e n t a g e o f t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d f o u n d j u s t a f t e r d r y i n g a n d t h e p e r c e n t a g e of d r y m a t t e r in t h e f r e s h t i s s u e . (b) E f f e c t of c o p p e r . T h e d a t a f o r t h e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d f o u n d a t e a c h d e t e r m i n a t i o n in t h e p l a n t s g r o w n on t h e c o p p e r c u l t u r e s a r e r e p o r t e d in T a b l e 14 a n d F i g u r e 74. An u p w a r d t r e n d in th e p e r c e n t a g e o f a s c o r b i c a c i d l e f t a f t e r d r y i n g c a n b e d e t e c t e d w i t h i n c r e a s e of c o p p e r i n t h e s o l u t i o n b u t t h i s t e n d e n c y i s n o t k e p t f o r th e p e r c e n t a g e l e f t 31 d a y s a f t e r drying. A f t e r 105 d a y s a l m o s t a l l t h e a s c o r b i c a c i d h a s d i s a p p e a r e d , o n l y 5 . 4 to 14 p e r c e n t w e r e l e f t , w h e n t h e s e v a l u e s a r e g r a p h e d a g a i n s t t h e a m o u n t s of c o p p e r in t h e s o l u t i o n s , a s l i g h t d e c r e a s e i s s h o w n f r o m no c o p p e r t o 0 . 0 2 p a r t s p e r m i l l i o n of c o p p e r a n d th en an i n c r e a s e to 0,08 p a r t s p e r m il lio n i s a p p a r e n t. 33 F r o m t h e o b s e r v a t i o n of F i g u r e 76, i t c a n b e s e e n t h a t t h e r a t e o f l o s s d i d n o t s h o w a n y p r o p o r t i o n a l i t y w i t h t h e a m o u n t of i r o n in th e d r y m a t t e r . H o w e v e r , to a s m a l l e x t e n t i t s h o w e d a n e g a t i v e r e l a t i o n a f t e r 105 d a y s a f t e r d r y i n g a s s h o w n i n F i g u r e 76. No o th e r c o r r e l a t i o n s w e r e found w ith an y of the c o m p o n e n ts a n a l y z e d a n d c a l c u l a t e d o n a p e r p l a n t , p e r 100 g r a m s f r e s h m a t ­ t e r a n d p e r 100 g r a m s d r y m a t t e r . c. O ther C hem ical C onstituents T h e n itr o g e n , c h l o r o p h y ll and c a r b o h y d r a t e c o n te n ts of the p l a n t s h a v e b e e n c a l c u l a t e d in the s a m e m a n n e r a s f o r i r o n and a s c o rb ic acid. No f u r t h e r d i s c u s s i o n w i l l b e u n d e r t a k e n a b o u t th e s e c o n s titu e n ts s in c e any r e la tio n s h ip b etw een th e m and ir o n and a s c o r b ic a c id have a lr e a d y b e e n d i s c u s s e d . T he follow ing ta b le w ill m e n tio n the p l a c e s w h e r e the d a ta c a n b e fou n d : 34 A m ounts p e r T en P lants C onstituents A m ounts per 100 F r e s h M aterial A m ounts per 100 g. D r y M aterial T ables F igures T ables F igures T ables F igures N itrogen 1 & 3 10 & 41 5 & 7 22 & 57 9 & 11 28 & 69 C hlorophyll 1 & 3 1 1 & 42 5 & 7 23 & 58 9 & 11 29 h 70 C arbo­ hydrate 1 & 3 9 & 40 5 & 7 21 & 56 9 & 11 36 & 68 D. D i s c u s s i o n of t h e F i r s t E x p e r i m e n t If w e o n l y h a d t h e d a t a on th e t o t a l a m o u n t of a s c o r b i c a c i d f o und a t h a r v e s t , no s i g n i f i c a n t e f f e c t of t h e a m o u n t s of i r o n a n d c o p p e r a d d e d to t h e s o l u t i o n c o u l d b e r e p o r t e d o n t h e a s c o r b i c a c i d in w h e a t p l a n t s . I n d e e d f r o m o n e c u l t u r e to a n o t h e r , t h e v a l ­ u e s of a s c o r b i c a c i d v a r i e d w i d e l y w i t h n o c o n s i s t e n t r e l a t i o n s h i p n e i t h e r w i t h t h e a m o u n t of c o p p e r a n d i r o n in t h e n u t r i e n t s o l u t i o n n o r w i t h t h e a m o u n t s of o t h e r c o m p o n e n t s ( i r o n , n i t r o g e n , c a r b o ­ h y d r a t e , c h lo ro p h y ll) a n a ly z e d in the p la n t m a t e r i a l . H o w e v e r , a n e w o u t l o o k on t h e p r o b l e m e m e r g e s w h e n t h e d a t a on t h e a m o u n t of a s c o r b i c a c i d l e f t a f t e r d r y i n g a r e s t u d i e d . It c a n t h e n b e c o n c l u d e d t h a t t h e o v e r - a l l e f f e c t of a g r e a t e r a m o u n t 35 of i r o n o r c o p p e r i n t h e n u t r i e n t s o l u t i o n c o r r e s p o n d e d t o a g r e a t e r s t a b i l i t y of t h e a s c o r b i c a c i d in t h e w h e a t p l a n t s d u r i n g d r y i n g . T h e s e r e l a t i o n s h i p s b e c o m e a p p a r e n t w h e n t h e p e r c e n t a g e of t h e o r i g i n a l a m o u n t s of a s c o r b i c a c i d l e f t a t t h e s u c c e s s i v e d e t e r m i ­ n a t i o n s m a d e on t h e m a t e r i a l l e f t a f t e r d r y i n g a r e c a l c u l a t e d . The p la n ts f r o m the ir o n s e r i e s show ed the m o s t s ig n ifi­ c a n t i n c r e a s e i n t h e s t a b i l i t y of t h e i r a s c o r b i c a c i d w i t h i n c r e a s e of i r o n i n t h e n u t r i e n t s o l u t i o n s a n d t h i s r e l a t i o n s h i p h e l d t r u e a t t h e various d e te rm in a tio n s a fte r drying. On t h e o t h e r h a n d , a p o s i t i v e r e l a t i o n s h i p b e t w e e n i n c r e a s e of c o p p e r in t h e n u t r i e n t s o l u t i o n a n d s t a b i l i t y of a s c o r b i c a c i d in t h e p l a n t s e x i s t e d j u s t a f t e r d r y i n g b u t d i s a p p e a r e d 31 d a y s a f t e r d r y i n g . I t w o u l d s e e m l i k e l y t h a t s i n c e a g r e a t e r s t a b i l i t y of a s c o r ­ b ic a c i d o c c u r r e d in t h e p l a n t s g r o w n on h i g h e r a m o u n t s of i r o n a n d c o p p e r , t h i s s t a b i l i t y w o u l d b e c o r r e l a t e d to t h e a m o u n t of i r o n and c o p p e r in the t i s s u e . Of t h e s e tw o e l e m e n t s o n l y i r o n w a s d e ­ t e r m i n e d a n d in t h e c o n d i t i o n s of t h e e x p e r i m e n t t h e r e w a s no p r o ­ p o r t i o n a l i n c r e a s e of i r o n in t h e t i s s u e w i t h i n c r e a s e of t h e a m o u n t s in t h e n u t r i e n t s o l u t i o n s a n d s u b s e q u e n t l y no r e l a t i o n c o u l d b e f o u n d b e tw e e n the p l a n t - i r o n and the a s c o r b i c a c id s ta b ility . I t w a s n o t i c e d t h a t i n t h e c o p p e r s e r i e s , w h e r e t h e a m o u n t s of iro n added in the n u tr ie n t s o lu tio n s w e r e s i m i l a r fo r the w hole s e r i e s , t h e f i n a l c o n c e n t r a t i o n of t h i s e l e m e n t in t h e p l a n t v a r i e d w i d e l y 36 b u t no c l e a r r e l a t i o n s h i p c o u l d b e d r a w n b e t w e e n th e a m o u n t p r e s ­ ent and the a s c o r b i c acid s ta b ility . F r o m t h e s e c o n c l u s io n s , two s u g g e s t i o n s w ill be a d v a n c e d to e x p l a i n t h e c o r r e l a t i o n b e t w e e n t h e i n c r e a s e of s t a b i l i t y of a s c o r b i c a c i d in t h e p l a n t m a t e r i a l a f t e r d r y i n g a n d t h e i n c r e a s e of i r o n in t h e n u t r i e n t s o l u t i o n . It is c o n c e iv a b le th a t only a f r a c ­ t i o n of t h e t o t a l i r o n p r e s e n t i n t h e p l a n t i s r e s p o n s i b l e f o r th e i n c r e a s e in the s ta b i l i t y ; th is f r a c t i o n w ould th e n be p r o p o r t i o n a l to t h e a m o u n t of i r o n p r e s e n t i n t h e n u t r i e n t s o l u t i o n . Indeed it is known th a t fo r o t h e r p r o c e s s e s s u c h as c h l o r o p h y ll f o r m a t i o n only a p o r t i o n of i r o n i s a c t i v e a s h a s b e e n r e p o r t e d b y J a c o b s o n ( 1 9 4 5 ) . T h e o t h e r s u g g e s t i o n i s t h a t b y i n c r e a s i n g th e a m o u n t of i r o n in the n u t r i e n t s o lu tio n , a n o t h e r f a c t o r is p r o p o r t i o n a l l y a f f e c te d w h i c h h a s a d i r e c t b e a r i n g on t h e a s c o r b i c a c i d s y s t e m of t h e p l a n t ; a m o r e c o m p l e t e f r a c t i o n a t i o n of t h e p l a n t c o m p o n e n t s s h o u l d t h e n b e c o n d u c t e d in o r d e r t o find t h i s p o s s i b l e f a c t o r . A s c a n be s e e n f r o m the r e s u l t s o b ta in e d in th is e x p e r i ­ m e n t and i f w e a s s u m e t h a t n o a s c o r b i c a c i d i s s y n t h e s i z e d a f t e r t h e h a r v e s t i n g of t h e p l a n t , i t c a n b e c o n c l u d e d t h a t t h e a p p a r e n t b e h a v i o u r of a s c o r b i c a c i d i n d r y i n g p l a n t m a t e r i a l i s q u i t e d i f ­ fe re n t fro m its b ehaviour in vitro. I t s h o u l d b e n o t e d t h a t t h e a c t u a l d i f f e r e n c e s i n a m o u n t s of i r o n a b s o r b e d b y the p l a n t s g r o w n on v a r i o u s l e v e l s of i r o n in the 37 n u t r i e n t s o lu tio n w e r e n o t g r e a t and not p r o p o r t i o n a l to the a m o u n t s a d d e d in t h e n u t r i e n t s o l u t i o n . L y o n a n d B e e s o n (1 9 4 8 )) on t h e o t h e r h a n d , w e r e a b l e t o o b t a i n p r o p o r t i o n a l i n c r e a s e of i r o n i n t h e p l a n t s g r o w n on i n c r e a s i n g a m o u n t s of i r o n . It is s u g g e s te d th at the g r a ­ v e l i t s e l f on w h i c h t h e p l a n t s w e r e g r o w n m i g h t c o n t a i n i m p u r i t i e s w hich should be tak en out b e f o re s t a r t i n g a new e x p e r im e n t . A p o s s i b l e r e a s o n f o r t h e d i f f e r e n c e in t h e a m o u n t of i r o n t a k e n up by t h e p l a n t s on t h e c o p p e r s e r i e s m i g h t a l s o l i e in t h e i m p u r i t i e s of t h e g r a v e l . It i s s u g g e s t e d t h a t a n e w e x p e r i m e n t u s i n g t h e s a m e p l a n t m a t e r i a l g r o w n o v e r a w i d e r r a n g e of i r o n a n d c o p p e r c o n c e n t r a ­ t i o n s in t h e s u b s t r a t e s h o u l d b e u n d e r t a k e n to f in d o u t if t h e s a m e t e n d e n c i e s w ould be d u p lic a te d and hold t r u e a t the h i g h e r c o n c e n ­ t r a t i o n s of t h e s e e l e m e n t s . E. 1. S u m m a r y of t h e F i r s t E x p e r i m e n t W h e a t p l a n t s w e r e g r o w n in g r a v e l on t w o s e r i e s of c u l t u r e s ; t h e b a s i c m e d i u m of e a c h c u l t u r e w a s a c o m p l e t e t h r e e s a lts n u tr ie n t solution. In t h e f i r s t s e r i e s , e a c h c u l t u r e w a s d i f ­ f e r e n t i a t e d f r o m t h e o t h e r b y a n i n c r e m e n t of i r o n of o n e p a r t p e r m i l l i o n , t h e w h o l e s e r i e s r a n g i n g f r o m no i r o n to n i n e p a r t s p e r m illion. In t h e s e c o n d s e r i e s , e a c h c u l t u r e d i f f e r e n t i a t e d f r o m t h e 38 o t h e r b y a n i n c r e m e n t o f c o p p e r of 0 .01 p a r t s p e r m i l l i o n , t h e w h o l e s e r i e s r a n g i n g f r o m no c o p p e r to 0-.09 p a r t s p e r m i l l i o n . 2. The plan ts w e r e h a rv e s te d at jointing stag e . A scorbic a c i d a n d c h l o r o p h y l l w e r e t h e n d e t e r m i n e d on t h e f r e s h t i s s u e s . I r o n , n i t r o g e n a n d c a r b o h y d r a t e w e r e d e t e r m i n e d on o v e n d r y s a m p l e s of t h e t i s s u e s . A s c o rb ic acid was again d e te r m in e d ju st a f t e r d r y i n g , 31 d a y s a n d 105 d a y s a f t e r d r y i n g . The data w e re c a l c u l a t e d o n a b a s i s p e r p l a n t , p e r 100 g r a m s f r e s h m a t t e r a n d p e r 100 g r a m s d r y m a t t e r . F r e s h a n d d r y w e i g h t of t h e c u l t u r e s w ere recorded. 3. In t h e i r o n s e r i e s , t h e f r e s h w e i g h t of t h e p l a n t s i n ­ c r e a s e d w i t h h i g h e r i r o n c o n c e n t r a t i o n up to s i x p a r t s p e r m i l l i o n of i r o n in t h e s o l u t i o n . The d r y w eight show ed an u p w ard te n d en cy w i t h i n c r e a s e i r o n up to t h e l a s t c u l t u r e . In t h e c o p p e r s e r i e s , t h e f r e s h and d r y w eig h t p a r a ll e le d e a c h o th e r , in c r e a s in g w ith h ig h e r c o p p e r c o n c e n t r a t i o n in t h e s o l u t i o n up to 0 , 0 6 p a r t p e r m i l l i o n of c o p p e r a n d t h e n d e c r e a s i n g u n t i l t h e l a s t c u l t u r e of t h e s e r i e s . 4. A t h a r v e s t , n o c o r r e l a t i o n b e t w e e n t h e a m o u n t of a s c o r ­ b i c a c i d c o u l d b e f o u n d w i t h t h e a m o u n t of i r o n in t h e n u t r i e n t s o ­ l u t i o n o r i n t h e p l a n t c a l c u l a t e d on a b a s i s p e r p l a n t , p e r 100 g r a m s d r y m a t t e r o r p e r 100 g r a m s of f r e s h t i s s u e . 39 5. T he r a te a t w hich a s c o r b i c acid w a s lo s t d u rin g and a f t e r d r y i n g in the p la n ts g r o w n on th e i r o n s e r i e s w a s i n v e r s e l y c o r r e l a t e d w i t h t h e a m o u n t of i r o n in t h e s o l u t i o n on w h i c h t h e plants w e r e grow n. No r e l a t io n s h i p , h o w e v e r , co u ld be found b e ­ t w e e n t h e a m o u n t of i r o n i n t h e f r e s h t i s s u e a n d t h e r a t e a t w h i c h a s c o rb ic acid w as lost. 6. T h e a m o u n t of i r o n f o u n d i n t h e p l a n t s g r o w n o n th e c o p p e r s e r i e s w a s p o s i t i v e l y c o r r e l a t e d w i t h t h e d r y w e i g h t of t h e plants. The r e la tio n s h ip b e tw e e n iro n and d r y m a t t e r held tru e when th e d a ta w e r e c a l c u l a t e d on a f r e s h t i s s u e b a s i s b u t d i s a p p e a r e d on a d r y w e i g h t b a s i s . 7. A t h a r v e s t , no c o r r e l a t i o n b e t w e e n t h e a m o u n t of a s c o r ­ b ic a c i d c o u l d b e d r a w n w i t h t h e a m o u n t of c o p p e r in t h e n u t r i e n t s o l u t i o n o r t h e a m o u n t of i r o n in t h e f r e s h t i s s u e of t h e c o p p e r series. 8. T h e r a t e a t w h ic h a s c o r b i c a c id w a s l o s t d u r in g and a f t e r d r y i n g w a s i n v e r s e l y c o r r e l a t e d w i t h t h e a m o u n t of c o p p e r in t h e s o l u t i o n on w h i c h t h e p l a n t s g r e w . IV. EX PER IM EN T 2 A. Purpose I t w a s s u g g e s t e d in t h e d i s c u s s i o n of t h e f i r s t e x p e r i m e n t t h a t t h e p o s i t i v e r e l a t i o n s h i p f o u n d b e t w e e n t h e s t a b i l i t y of a s c o r ­ b i c a c i d a n d t h e c o n c e n t r a t i o n o f i r o n and c o p p e r in t h e s o l u t i o n b e s u b j e c t e d to a n e w t r i a l . In o r d e r t o c h e c k th e r e s u l t s o b t a i n e d d u r i n g t h e f i r s t e x ­ p e r i m e n t a n d a t t h e s a m e t i m e to e x p l o r e t h e e f f e c t s of h i g h e r c o n c e n t r a t i o n s of c o p p e r a n d i r o n in t h e s o l u t i o n , i t w a s t h o u g h t to s t a r t t h e s e r i e s of c u l t u r e a t t h e s a m e l e v e l a s f o r t h e f i r s t e x ­ p e r i m e n t , i . e . , no i r o n o r no c o p p e r b u t to a d d i n t h e f o l l o w i n g c u l t u r e s l a r g e r i n c r e m e n t s of t h e s e e l e m e n t s . In th is m a n n e r by o v e rla p p in g w ith the p r e v io u s e x p e r i m e n t and u sin g the s a m e p la n t m a t e r i a l and the s a m e e x p e r im e n t a l m e th o d s , the r e s u l t s ob tain ed w o u l d b e a c o m p l e m e n t t o th e f i r s t o b s e r v a t i o n s . B. 1. E x p e rim e n ta l C onditions I r o n a n d C o p p e r L e v e l s in th e N u t r i e n t S o lu tio n T o c o m p l e t e t h r e e s a l t s n u t r i e n t s o l u t i o n s , a s u s e d in t h e f i r s t e x p e r i m e n t , w e r e a d d e d t h e f o l l o w i n g a m o u n t s of i r o n a n d copper: 41 C o n c e n t r a t i o n of I r o n a n d C o p p e r in t h e N u t r i e n t S o l u t i o n s of t h e S e c o n d E x p e r i m e n t Iron S e rie s C opper S eries p p m of F e in th e S o l u t i o n p p m of C u in the Solution p p m of F e in the Solution p p m of C u in the Solution 0.0 2.0 4.0 6.0 8. 0 10.0 12.0 14.0 16.0 18.0 0.02 0.02 0.02 0.02 0.02 0.02 0.02 0.02 0.02 0.02 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 0.00 0.02 0.04 0.06 0.08 0.10 0.12 0.14 0.16 0.18 I r o n a n d c o p p e r w e r e a d d e d in t h e s a m e f o r m a s i n t h e f i r s t e x p e r i m e n t f r o m a s t o c k s o l u t i o n of f e r r i c s u l f a t e a n d c u p r i c sulfate. 2* P l a n t M a t e r i a l T h i r t y s e e d s of w h e a t I l l i n o i s N o . 1 - 1 2 8 w e r e s e e d e d i n t h e q u a r t z g r a v e l i n e a c h p o t on O c t o b e r 25, 1948, a n d w e r e h a r v e s t e d 19 d a y s l a t e r j u s t b e f o r e j o i n t i n g s t a g e on N o v e m b e r 13, 1948. 42 C. 1. a. E x p e rim en tal R esults O b s e r v a t i o n s on t h e P l a n t s D e s c r i p t i o n o f t h e P l a n t s a t T i m e of H a r v e s t W hen the p la n ts w e r e h a r v e s te d , they had not y et s ta r t e d to j o i n t . A s w a s o b s e r v e d a t t h e e n d of t h e f i r s t e x p e r i m e n t , t h e p l a n t s on t h e i r o n s e r i e s w e r e s o m e w h a t b e t t e r t h a n t h o s e g r o w n on t h e c o p p e r s e r i e s , s h o w i n g t h i c k e r s t e m a n d g r e e n e r l e a v e s . On t h e i r o n s e r i e s , t h e h e a l t h i e s t l o o k i n g p l a n t s w e r e g r o w n on t h e c u l t u r e c o n t a i n i n g s i x p a r t s p e r m i l l i o n of i r o n . T h e p l a n t s g r o w i n g on t h e v a r i o u s l e v e l s of c o p p e r w e r e a ll h e a l t h y a n d t h e b e s t g r o w t h w a s o b t a i n e d on t h e h i g h e s t l e v e l of c o p p e r , i . e . , 0 . 1 8 p a r t s p e r m i l l i o n , c o n t r a r y to w h a t w a s o b ­ s e r v e d d u rin g the f i r s t e x p e r im e n t w h e r e b e s t p la n ts w e r e o b tain ed a t 0 . 0 6 p a r t s p e r m i l l i o n of c o p p e r . b. F r e s h W e i g h t of t h e P l a n t s (1) E f f e c t of I r o n T he d a ta for the a v e r a g e f r e s h w eight p e r ten p lan ts e x p r e s s e d in g r a m s a r e g i v e n i n T a b l e 15 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t of i r o n i n t h e n u t r i e n t s o l u t i o n i n F i g u r e 82. 43 A c c o r d i n g to t h e s e v a l u e s , the h i g h e s t w e ig h t p e r p la n t w a s o b t a i n e d o n t h e h i g h e s t c o n c e n t r a t i o n of i r o n in t h e n u t r i e n t solution. H o w e v e r t h i s v a l u e i s s o m e w h a t o u t of t h e g e n e r a l l i n e of t h e o t h e r r e s u l t s a n d if i t i s t a k e n o u t , t h e c u r v e t h e n s h o w s a m a x i m u m a t s ix p a r t s p e r m i l l i o n w h ic h c o r r e s p o n d s to th e o b ­ s e r v a t i o n m a d e on t h e a p p e a r a n c e of t h e p l a n t s a t h a r v e s t . (2) E f f e c t of C o p p e r T h e d a t a i n d i c a t i n g t h e a v e r a g e of t h e f r e s h w e i g h t p e r t e n p l a n t s e x p r e s s e d in g r a m s a r e a s s e m b l e d i n T a b l e 16 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r i n t h e n u t r i e n t s o l u t i o n i n F i g u r e 119. A g e n e r a l i n c r e a s e in t h e w e i g h t of t h e p l a n t s w a s s h o w n f r o m t h e c u l t u r e w i t h o u t c o p p e r to t h e o n e w i t h 0*18 p a r t s p e r m i l l i o n of c o p p e r . T h is te n d e n c y w a s d i f f e r e n t f r o m the one o b ­ t a i n e d d u r i n g t h e f i r s t e x p e r i m e n t , w h e r e t h e b e s t g r o w t h o f th e p l a n t s o c c u r r e d a t 0 . 0 6 p a r t s p e r m i l l i o n of c o p p e r a n d t h e n d e ­ creased* c. D ry W eight p e r P la n ts (1) E f f e c t o f I r o n The d a ta f o r the a v e r a g e d r y w eig h t p e r ten p la n ts e x p r e s s e d 44 in g r a m s a r e s h o w n in T a b l e 15 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t of i r o n in t h e s o l u t i o n i n F i g u r e 83. T h e v a r i a t i o n s in t h e d r y w e i g h t of t h e p l a n t s s h o w e d t h e s a m e te n d en cy a s the f r e s h w eight and this te n d e n c y w as even m o r e pronounced. If t h e v a l u e f o r t h e p l a n t s g r o w n o n t h e s o l u t i o n w i t h 18 p a r t s p e r m i l l i o n i s n o t c o n s i d e r e d , t h e n a m a x i m u m of d r y w e i g h t i s o b t a i n e d a t a c o n c e n t r a t i o n o f s i x p a r t s p e r m i l l i o n of ir o n in the n u t r i e n t s o lu tio n . (2) E f f e c t of C o p p e r T he a v e r a g e d r y w e ig h t p e r ten p la n ts of th is s e r i e s a r e a r r a n g e d in T a b l e 16 a n d g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r in t h e s o l u t i o n in F i g u r e 120. T h e d r y w e i g h t of t h e p l a n t s of t h e c o p p e r s e r i e s d i d n o t p a r a l l e l t h e f r e s h w e i g h t a n d s h o w e d tw o m a x i m a , o n e a t 0 . 0 6 p a r t s p e r m i l l i o n a n d t h e o t h e r a t 0 . 1 4 p a r t s p e r m i l l i o n of c o p p e r . r e a s o n f o r t h e i r r e g u l a r i t i e s in t h e d r y w e i g h t of t h e p l a n t s i s r a t h e r o b s c u r e b u t a s we w ill s e e l a t e r it w a s fo llo w e d by the a m o u n t of i r o n i n t h e p l a n t s . The 45 2. a. C h e m i c a l C o m p o s i t i o n of t h e P l a n t s Iron (1) A m o u n t of I r o n p e r P l a n t (a) E f f e c t of i n c r e a s i n g t h e a m o u n t s of i r o n i n t h e s o l u t i o n on t h e a b s o r p t i o n o f i r o n b y t h e p l a n t . T h e m i l l i g r a m s of i r o n a b ­ s o r b e d p e r t e n p l a n t s a r e p r e s e n t e d i n T a b l e 15 a n d g r a p h e d a g a i n s t t h e a m o u n t of i r o n i n s o l u t i o n in F i g u r e 77. T o a n i n c r e a s e of i r o n i n t h e s o l u t i o n c o r r e s p o n d e d a n i n ­ c r e a s e of a m o u n t of i r o n a b s o r b e d p e r p l a n t . T his re s p o n s e w as s i m i l a r to t h e o n e o b t a i n e d d u r i n g t h e f i r s t e x p e r i m e n t . A posi­ tive c o r r e l a t i o n b e tw e e n i r o n and d r y w e ig h t of the p la n t w a s a l s o o b ta in e d a s d u r in g the f i r s t e x p e r i m e n t . The other relatio n s d e a l­ ing w ith a s c o r b i c a c id w ill be c o n s i d e r e d l a t e r . (b) E f f e c t of i n c r e a s i n g t h e a m o u n t s of c o p p e r i n t h e s o l u ­ t i o n s on t h e a b s o r p t i o n of i r o n . T h e a m o u n t s of i r o n , e x p r e s s e d i n m i l l i g r a m s , a b s o r b e d p e r t e n p l a n t s of t h e c o p p e r s e r i e s a r e r e p o r t e d in T a b l e 16 a n d g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r in t h e s o l u t i o n i n F i g u r e 114. A l t h o u g h t h e a m o u n t s of i r o n a b s o r b e d p e r p l a n t d i d n o t s h o w a n y c l e a r r e l a t i o n s h i p w i t h t h e a m o u n t s of c o p p e r i n t h e s o l u t i o n , i t s v a r i a t i o n s w e r e f o l l o w e d b y t h e d r y w e i g h t of t h e 46 plants. T h i s d a t a g i v e s a g r e a t w e i g h t t o t h e c l o s e r e l a t i o n s h i p of i r o n to d r y m a t t e r , w h i c h h a d b e e n o b t a i n e d d u r i n g t h e f i r s t e x ­ p e r i m e n t on b o t h i r o n a n d c o p p e r s e r i e s . R e la tio n s h ip s with a s c o r b i c a c id w ill be d i s c u s s e d l a t e r . (2) A m o u n t of I r o n p e r 100 G r a m s of F r e s h M a t e r i a l (a) E f f e c t of i r o n . The m illig ra m s of f r e s h p l a n t m a t e r i a l a p p e a r in T a b l e 17 a n d of i r o n p e r 100 gram s F i g u r e 89. V e r y c l e a r l y t h e c o n c e n t r a t i o n s of i r o n i n c r e a s e d i n th e f r e s h t i s s u e of t h e p l a n t s g r o w n w i t h i n c r e a s i n g a m o u n t s o f i r o n in t h e s o l u t i o n . T h i s r e l a t i o n s h i p i s s i m i l a r to t h e o n e o b t a i n e d d u r i n g th e f i r s t e x p e r i m e n t b u t i s m u c h m o r e s i g n i f i c a n t . (b) E f f e c t of c o p p e r . T h e d a ta in d ic a tin g the m i l l i g r a m s of i r o n p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l of t h e c o p p e r s e r i e s are s h o w n in T a b l e 18 a n d F i g u r e 126. As in t h e f i r s t e x p e r i m e n t t h e a m o u n t s of i r o n p e r 100 g r a m s of f r e s h m a t e r i a l w e r e q u i t e v a r i a b l e f r o m o n e c u l t u r e to the o t h e r a n d a s t r o n g p o s i t i v e r e l a t i o n s h i p c o r r e l a t e d i t w i t h d r y weight. (3) A m o u n t of I r o n p e r 100 G r a m s of D r y M a t t e r (a) E f f e c t of i r o n . T h e d a t a f o r t h e a m o u n t of i r o n in m i l ­ l i g r a m s p e r 100 g r a m s of d r y m a t e r i a l i n t h e i r o n s e r i e s a r e a s ­ s e m b l e d i n T a b l e 19 a n d in F i g u r e 106. W i t h i n c r e a s i n g i n c r e m e n t s of i r o n in t h e n u t r i e n t s o l u t i o n w e r e o b t a i n e d i n c r e a s i n g a m o u n t of i r o n in t h e d r y m a t t e r . At t h e s a m e t i m e , t h e p e r c e n t a g e of w a t e r in t h e t i s s u e s w a s i n c r e a s ­ i n g a s c a n b e s e e n i n t h e f i g u r e 111 g r a p h i n g t h e g r a m s of f r e s h m a t e r i a l c o r r e s p o n d i n g t o 100 g r a m s o f d r y m a t t e r f o r t h e d i f f e r ­ e n t c u l t u r e s of t h e i r o n s e r i e s , a g a i n s t t h e a m o u n t of i r o n in t h e solution. (b) E f f e c t of c o p p e r . T h e a m o u n t of i r o n in m i l l i g r a m s p e r 100 g r a m s o f d r y p l a n t m a t e r i a l a r e p r e s e n t e d in T a b l e 20 a n d F i g u r e 135. A s t h e a m o u n t o f c o p p e r v a r i e s in t h e s o l u t i o n , t h e a m o u n t of i r o n f o u n d i n t h e d r y p l a n t m a t e r i a l c h a n g e d b u t d i d n o t f o l l o w any c le a r relatio n sh ip , D u r in g the f i r s t e x p e r i m e n t , l e s s i r o n w a s found in th e t i s s u e s g r o w n on h i g h e r c o p p e r l e v e l s , b u t t h i s t e n d e n c y did not show a g a in d u r in g th is e x p e r i m e n t . 48 b. A s c o rb ic Acid (1) D e s c r i p t i o n A s f o r t h e f i r s t e x p e r i m e n t , a s t u d y of t h e s t a b i l i t y of t h e a s c o r b i c a c id in the p la n t a f t e r h a r v e s t w a s u n d e r ta k e n . It w a s j u d g e d f r o m t h e r e s u l t s of t h e f i r s t e x p e r i m e n t t h a t t h e s u c c e s s i v e d e t e r m i n a t i o n s s h o u l d b e d o n e o v e r a s h o r t e r p e r i o d of t i m e . The f i r s t d e te r m in a tio n w as done a t h a r v e s t, the second ju s t a f t e r d r y in g in a f o r c e d a i r oven a t 6 0 ° C f o r t h r e e d a y s , th e th ir d s e v e n d a y s l a t e r a n d the l a s t f o u r t e e n d a y s a f t e r d r y in g . T h e s a m p l e s of p l a n t m a t e r i a l a f t e r d r y i n g w e r e k e p t a t c o n s t a n t t e m p e r a t u r e in a n i n c u b a t o r a t 3 0 ° C . (2) A m o u n t of A s c o r b i c A c i d p e r P l a n t (a) E f f e c t of i r o n . T h e d a t a f o r t h e a m o u n t s of a s c o r b i c acid a t h a r v e s t , a f t e r d ry in g , se v e n d ay s and f o u rte e n d a y s a f te r d r y i n g p e r t e n p l a n t s of t h e i r o n s e r i e s , a r e p r e s e n t e d in T a b l e 15 a n d F i g u r e s 78, 79, 80, a n d 81. T h e a m o u n t of a s c o r b i c a c i d p e r p l a n t , a t h a r v e s t , w a s i n ­ v e r s e l y p r o p o r t i o n a l t o t h e a m o u n t of i r o n t a k e n up b y t h e p l a n t , e x c e p t f o r t h e v a l u e of a s c o r b i c a c i d o b t a i n e d p e r p l a n t g r o w n w i t h ­ out iro n . T h i s v a l u e w a s t h e l o w e s t o n e a n d o u t of t h e g e n e r a l line of the o th e r v a lu e s. 49 A v e r y s p e c i a l b e h a v i o u r w a s s h o w n b y t h e a m o u n t of a s c o r b i c a c id left a f t e r d ry in g ; in d e e d the r e l a t i o n s h i p o b s e r v e d a t h a r v e s t b e t w e e n t h e a m o u n t of a s c o r b i c a c i d p e r p l a n t a n d t h e a m o u n t of i r o n in t h e c o r r e s p o n d i n g n u t r i e n t s o l u t i o n s h i f t e d to th e o p p o s i t e s i g n : m o r e a s c o r b i c a c i d r e m a i n e d i n th e p l a n t s g r o w n on t h e h i g h e s t l e v e l s of i r o n . This re la tio n s h ip held tru e with a m o u n t of a s c o r b i c a c id found s e v e n and f o u r t e e n d a y s a f t e r drying.S ig n ifican t p o sitiv e c o r r e l a t i o n s w e r e o btained b e tw e e n the a m o u n t of a s c o r b i c a c i d r e m a i n i n g a f t e r d r y i n g , s e v e n d a y s a n d f o u r t e e n d a y s l a t e r a n d th e a m o u n t of i r o n i n t h e p l a n t s , b u t in a l l th e c a s e s t h e v a l u e f o r t h e p l a n t s g r o w n w i t h o u t i r o n s h o u l d b e c o n s i d e r e d a p a r t ( F i g u r e s 86, 87, a n d 88 ). (b) E f f e c t of c o p p e r . T h e d a t a e x p r e s s i n g t h e a m o u n t of a s c o r b ic acid a t h a r v e s t , a f t e r d ry in g , s e v e n d ay s and f o u rte e n d a y s a f t e r d r y i n g p e r t e n p l a n t g r o w n a t v a r i o u s l e v e l s of c o p p e r a r e a s s e m b l e d in T a b l e 16 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r in t h e s o l u t i o n in F i g u r e s 115, 11 6, 117, a n d 118. W h e n t h e a m o u n t s of a s c o r b i c a c i d a t h a r v e s t a n d a t th e d i f f e r e n t d e t e r m i n a t i o n s a f t e r d r y i n g a r e o b s e r v e d , no a p p a r e n t r e l a t i o n s h i p w i t h t h e a m o u n t of c o p p e r i n t h e s o l u t i o n c a n b e d r a w n . 50 H o w e v e r , a p o s it i v e c o r r e l a t i o n is a p p a r e n t b e tw e e n the v a l u e s of a s c o r b i c a c i d o f t h e s e p l a n t s a t h a r v e s t a n d t h e a m o u n t s of i r o n f o u n d i n t h e p l a n t s . T h i s r e l a t i o n s h i p c a n b e s e e n in F i g ­ u r e 122. A f t e r d r y i n g , w h e n t h e a m o u n t of a s c o r b i c a c i d l e f t i s p l o t t e d a g a i n s t t h e a m o u n t of i r o n i n t h e p l a n t a n e g a t i v e c o r r e l a ­ tion a p p e a r s . B ut a fte r fo u rte e n d ay s this re la tio n h as d is a p p e a re d . (3) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t t e r (a) E f f e c t of i r o n . T h e a v e r a g e m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l g r o w n i n t h e i r o n s e ­ r ie s and d e t e r m in e d at h a r v e s t , a f t e r d ry in g , s e v e n d a y s and f o u r ­ t e e n d a y s a f t e r d r y i n g a r e r e c o r d e d i n T a b l e 17 a n d F i g u r e s 90,‘ 91, 92,' a n d 93. W h e n t h e r e s u l t s f o r a s c o r b i c a c i d a r e c a l c u l a t e d on a b a s i s p e r 100 g r a m s of f r e s h t i s s u e , t h e c o r r e l a t i o n s w h i c h a p ­ p e a r e d b e tw e e n i r o n and a s c o r b i c a c id p e r p la n t s ta y e d a l m o s t unchanged. I n d e e d , t h e a m o u n t s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h m a t t e r a t h a r v e s t w a s i n v e r s e l y p r o p o r t i o n a l to t h e c o n ­ c e n t r a t i o n s o f i r o n in t h e s o l u t i o n a n d i n t h e p l a n t , F i g u r e 90 a n d 95. T h e s h i f t i n g f r o m a n e g a t i v e to a p o s i t i v e c o r r e l a t i o n b e t w e e n a s c o r b i c a c i d a n d t h e c o n c e n t r a t i o n of i r o n in t h e f r e s h t i s s u e , o c c u r r e d a fte r d ry in g and m o r e c le a r ly seven days a f te r drying, 51 a s h a d b e e n o b s e r v e d b e f o r e , w h e n t h e d a t a w e r e c a l c u l a t e d on a per plant b a s is . T h e s e r e l a t i o n s m a y b e s e e n i n F i g u r e s 97, 98, an d 99. A s tr o n g p o s i t i v e c o r r e l a t i o n w a s found a s s e e n in F i g u r e 96 b e t w e e n t h e c o n c e n t r a t i o n of d r y m a t t e r i n t h e f r e s h t i s s u e a n d th e a m o u n t of a s c o r b i c a c i d in t h a t t i s s u e a t h a r v e s t . (b) E f f e c t of c o p p e r . T h e d a t a i n m i l l i g r a m s p e r 100 g r a m s of f r e s h m a t t e r of a s c o r b i c a c i d a t h a r v e s t , a f t e r d r y i n g , s e v e n d a y s a n d f o u r t e e n d a y s a f t e r d r y i n g , of t h e c o p p e r s e r i e s a p p e a r in T a b l e 18 a n d in F i g u r e s 127, 128, 129, a n d 130. No a p p a r e n t r e l a t io n s h i p w as in d ic a te d b e tw e e n the a m o u n t of c o p p e r i n t h e s o l u t i o n a n d t h e a m o u n t of a s c o r b i c a c i d f o u n d a t h a r v e s t i n 100 g r a m s of f r e s h t i s s u e . H ow ever a significant p o s ­ i t i v e r e l a t i o n s h i p i s o b t a i n e d , w h e n t h e a m o u n t of a s c o r b i c a c i d a t h a r v e s t i s g r a p h e d a g a i n s t t h e a m o u n t of i r o n in 100 g r a m s of f r e s h t i s s u e ; th is c o r r e l a t i o n had a l r e a d y b e e n o b ta in e d w h e n the v a l u e s w e r e c a l c u l a t e d on a p e r p l a n t b a s i s . A n e g a tiv e c o r r e l a t i o n a p p e a r s b e tw e e n the a s c o r b i c acid l e f t a f t e r d r y i n g a n d t h e a m o u n t of i r o n i n t h e t i s s u e , t h i s c o r r e l a ­ t i o n b e c o m e s n o t s o a p p a r e n t a f t e r s e v e n o r f o u r t e e n d a y s of d r y i n g , F i g u r e s 102, 103, a n d 104. 52 In b o th c o p p e r a n d i r o n s c r i e s a p o s i t i v e c o r r e l a t i o n w a s o b t a i n e d w i t h t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d a t h a r v e s t a n d the d r y m a t e r i a l in th e f r e s h t i s s u e . (4) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s of D r y M a t e r i a l (a) E f f e c t of i r o n . T h e a m o u n t s in m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s o f d r y p l a n t m a t e r i a l g r o w n o n t h e i r o n s e r i e s , d e te rm in e d at h a r v e s t, ju s t a fte r drying, sev e n days and fo u rte e n d a y s a f t e r d r y i n g a r e a s s e m b l e d in T a b l e 19 a n d g r a p h e d a g a i n s t th e a m o u n t o f i r o n in t h e s o l u t i o n i n F i g u r e s 107, 108, 109, a n d 110. T o a n i n c r e a s e i n t h e a m o u n t of i r o n i n t h e s o l u t i o n c o r ­ r e s p o n d e d a t h a r v e s t a d e c r e a s e of a s c o r b i c a c i d in t h e d r y m a ­ t e r i a l a n d a n e g a t i v e c o r r e l a t i o n b e t w e e n t h e a m o u n t of i r o n in t h e d r y m a t t e r a n d t h e a s c o r b i c a c i d c a n b e s h o w n i n F i g u r e 112. The v a lu e s for the p la n ts grow n w ithout iro n should be c o n s id e r e d as a s p e c ia l c a s e , in d eed a t h a r v e s t and a f te r d ry in g th e se v a lu e s b e h a v e d i n t h e o p p o s i t e w a y to a l l t h e o t h e r v a l u e s . S e v e n d a y s a f t e r h a r v e s t , the p l a n t m a t e r i a l w ith hig h a m o u n t o f i r o n h a d t h e g r e a t e s t a m o u n t of a s c o r b i c a c i d l e f t a s i s i n d i c a t e d i n F i g u r e 113 a n d t h e s a m e r e l a t i o n s h i p w a s k e p t fo u rteen days a fte r drying. 53 (b) E f f e c t of c o p p e r . T h e d a t a e x p r e s s i n g in m i l l i g r a m s p e r 100 g r a m s of d r y m a t e r i a l , t h e a m o u n t of a s c o r b i c a c i d a t h a r v e s t , a f t e r d r y i n g , s e v e n a n d f o u r t e e n d a y s a f t e r d r y i n g of t h e p l a n t s g r o w n on t h e c o p p e r s e r i e s a r e p r e s e n t e d i n T a b l e 20 a n d F i g u r e s 136, 137, 1 38, a n d 139. T h e a m o u n t s o f c o p p e r i n th e n u t r i e n t s o l u t i o n d i d n o t s h o w an y c o r r e l a t i o n w i t h t h e a m o u n t o f a s c o r b i c a c i d a t h a r v e s t c a l ­ c u l a t e d on a d r y w e i g h t b a s i s . H ow ever a slight positive c o r r e l a ­ ti o n w a s f o u n d b e t w e e n t h e a m o u n t of i r o n in t h e d r y m a t t e r a n d the a s c o r b i c a c i d a t h a r v e s t , t h i s r e l a t i o n s h i p i s p r e s e n t e d in F i g u r e 132. Seven d a y s a f t e r d r y in g no a p p a r e n t r e l a t i o n s h i p c a n be d r a w n n e i t h e r w ith c o p p e r n o r w ith i r o n a s c a n be s e e n in F i g ­ u r e s 134 a n d 138. (5) P e r c e n t a g e of t h e O r i g i n a l A m o u n t of A s c o r b i c A c i d F o u n d a t E ach D eterm ination (a) E f f e c t of i r o n , T h e d a t a s h o w i n g t h e p e r c e n t a g e of t h e o r i g i n a l a m o u n t s of a s c o r b i c a c i d f o u n d i n t h e i r o n s e r i e s a f t e r drying, se v e n days and fourteen days a f te r d ry in g a r e p r e s e n te d in T a b l e 21 a n d F i g u r e 141. F r o m F i g u r e 141 i t i s a p p a r e n t t h a t , w i t h t h e e x c e p t i o n of th e “ n o i r o n ” v a l u e , t h e p e r c e n t a g e s o f t h e o r i g i n a l a m o u n t s of a s c o r b i c a c i d w e r e a t e a c h d e t e r m i n a t i o n p o s i t i v e l y c o r r e l a t e d to 54 t h e a m o u n t of i r o n i n t h e s o l u t i o n . - F r o m F i g u r e s 86 a n d 87, i t c a n b e s e e n t h a t t h i s p o s it i v e r e l a t i o n s h i p h o ld s t r u e w h e n the a m o u n t of i r o n p e r ten p la n ts is c o n s i d e r e d . T h e g r e a t e s t r a t e of o x i d a t i o n o f t h e a s c o r b i c a c i d o c c u r r e d d u r in g the d r y in g p r o c e s s and a t a h i g h e r r a t e in the p la n ts c o n ­ taining le s s iron: In t h e p l a n t g r o w n on t h e t w o p a r t s p e r m i l l i o n l e v e l , 65 p e r c e n t of t h e a s c o r b i c a c i d w e r e l o s t , w h e n o n l y 49 p e r c e n t w e r e l o s t i n t h e p l a n t s g r o w n on t h e 16 p a r t s p e r m i l l i o n l e v e l . (b) E f f e c t of c o p p e r . T a b l e 22 a n d F i g u r e 142 r e p o r t t h e v a l u e s of t h e p e r c e n t a g e s o f t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d f o und in t h e c o p p e r s e r i e s a f t e r d r y i n g , s e v e n d a y s a n d f o u r t e e n days a fte r d rying. A n o b s e r v a t i o n of t h e F i g u r e 142 d o e s n o t l e a d to a n y a p ­ p a r e n t r e l a t i o n s h i p b e t w e e n t h e p e r c e n t a g e of a s c o r b i c a c i d l e f t , a n d t h e a m o u n t of c o p p e r i n th e s o l u t i o n , b u t if t h e p e r c e n t a g e s v a l ­ u e s a r e g r a p h e d a g a i n s t t h e a m o u n t of i r o n in 100 g r a m s of f r e s h ti s s u e , a v e r y s ig n if ic a n t n e g a tiv e c o r r e l a t i o n a p p e a r s a s c a n be s e e n i n F i g u r e 105. In t h i s c a s e t h e i n f l u e n c e of i r o n on t h e c a t a l ­ y s i s of a s c o r b i c a c i d in t h e d r y i n g m a t e r i a l i s s h o w n v e r y d e f i n i t e l y . 55 D. D i s c u s s i o n on the S e c o n d E x p e r i m e n t To r e p r o d u c e the e n v ir o n m e n ta l f a c t o r s is a l m o s t i m p o s s ib le a n d i s t h e m a j o r c a u s e to t h e g r e a t d i f f i c u l t y i n d u p l i c a t i n g q u a n ­ t i t a t i v e r e s u l t s in b i o l o g i c a l e x p e r i m e n t a t i o n . B e c a u s e of n e c e s s i t y this s e c o n d e x p e r i m e n t w a s n o t s t a r t e d a t the s a m e t i m e of the y e a r a s the f i r s t one and the p la n t s w e r e h a r v e s t e d at a y o u n g e r s ta g e ; t h e s e r e a s o n s would e x p la in th e g r e a t d i f f e r e n c e s b e tw e e n th e q u a n t i t a t i v e r e s u l t s o b t a i n e d d u r i n g t h e t w o e x p e r i m e n t s , b u t a l s o w i l l g i v e m o r e s i g n i f i c a n c e to t h e c o m p a r a b l e t e n d e n c i e s . As f a r as iro n is c o n c e rn e d , this seco n d e x p e r im e n t was m o r e c o n c l u s i v e t h a n th e f i r s t o n e , b e c a u s e a c c o r d i n g to t h e e x ­ p e r i m e n t a l r e s u l t s t h e a b s o r p t i o n of t h i s e l e m e n t w a s d i r e c t l y p r o p o r t i o n a l to t h e a m o u n t of i r o n a d d e d in t h e s o l u t i o n . This t r e n d w a s t h e s a m e a s t h a t r e p o r t e d b y L y o n a n d B e e s o n (1 948) in t u r n i p s a n d t o m a t o e s g r o w n a t d i f f e r e n t l e v e l s o f i r o n , t h e s e w o r k e r s i n d e e d o b s e r v e d a s i g n i f i c a n t d i f f e r e n c e in t h e i r o n c o n ­ t e n t of t h e v e g e t a t i v e p l a n t p a r t s a t t h e s u c c e s s i v e i r o n t r e a t m e n t s . T h e s e a u t h o r s a l s o r e p o r t e d t h e r a t h e r l i m i t e d p o w e r of p l a n t s t o a c c u m u l a t e i r o n , i n f a c t t o a t h i r t y - f o l d i n c r e a s e in t h e c o n c e n ­ t r a t i o n of i r o n s u p p l i e d to t u r n i p s w a s a s s o c i a t e d l e s s t h a n a t w o ­ f o l d i n c r e a s e i n t h e i r o n c o n c e n t r a t i o n of t h e l e a v e s . The results of t h e p r e s e n t e x p e r i m e n t s h o w e d t h a t b y a c o m p a r i s o n of t h e 56 r e s u l t s o b ta in e d f r o m w h e a t p la n ts s u p p lie d w ith two p a r t s p e r m i l l i o n a n d t h o s e s u p p l i e d w i t h 18 p a r t s p e r m i l l i o n of i r o n , l e s s t h a n a t w o - f o l d i n c r e a s e in i r o n c o n c e n t r a t i o n o c c u r r e d . T h e a m o u n t s o f a s c o r b i c a c i d f o u n d a t h a r v e s t in t h e p l a n t s g r o w n on t h e i r o n s e r i e s , e x c e p t f o r t h e o n e g r o w n w i t h o u t i r o n , w e r e i n v e r s e l y p r o p o r t i o n a l to the a m o u n t of i r o n in the p la n t t i s ­ sue. T his tenden cy w hich w as not show n a t the f i r s t e x p e r im e n t s h o u l d h o w e v e r b e c o n s i d e r e d a s s p e c i a l l y i m p o r t a n t b e c a u s e of i t s s i m i l a r i t y w i t h t h e o n e o b t a i n e d b y W y n d ( 1 9 5 0 a ) in o a t s g r o w n in T e x a s . O n t h e o t h e r h a n d , in t h e c o p p e r s e r i e s , w h e r e t h e a m o u n t of i r o n w a s c o n s t a n t in t h e s o l u t i o n , t h e a m o u n t of a s c o r b i c a c i d f o u n d a t h a r v e s t s h o w e d a p o s i t i v e r e l a t i o n w i t h t h e a m o u n t of iro n in the f r e s h t i s s u e . T h e s e two o p p o s it e t e n d e n c i e s s u g g e s t t h a t i r o n a n d c o p p e r h a v e a s p e c i a l i n t e r r e l a t i o n s h i p e f f e c t on t h e a s c o r b i c a c i d e q u i l i b r i u m in t h e p l a n t s . A m ong the r e s u l t s w h ich show ed the s a m e tr e n d s d u rin g both e x p e r i m e n t s w a s the p o s itiv e r e l a t i o n s h i p b e tw e e n the a m o u n t of i r o n a b s o r b e d b y t h e p l a n t s a n d t h e y i e l d of d r y m a t t e r of t h e s e plants. A l t h o u g h t h i s s t u d y i s n o t d i r e c t l y r e l a t e d to t h i s p h a s e of t h e e f f e c t of i r o n , i t s h o u l d b e e m p h a s i z e d t h a t t h i s r e l a t i o n s h i p o c c u r r e d u n d e r d i f f e r e n t e n v i r o n m e n t a l c o n d itio n s and a l s o in b o th co p p e r and ir o n s e r i e s . 57 The m o s t in te r e s tin g r e s u l t obtained d u rin g this second e x p e r i m e n t w a s t h e d u p l i c a t i o n of t h e t e n d e n c y o b s e r v e d i n t h e f i r s t e x p e r i m e n t that the p la n ts f r o m the iro n s e r i e s show ed a s i g n i f i c a n t i n c r e a s e of t h e s t a b i l i t y of t h e i r a s c o r b i c a c i d w i t h i n ­ c r e a s e of i r o n i n t h e s o l u t i o n . It w a s a l s o o b s e r v e d t h a t a p o s ­ i t i v e c o r r e l a t i o n o c c u r r e d b e t w e e n i n c r e a s e of c o p p e r in t h e s o l u ­ t i o n a n d s t a b i l i t y of a s c o r b i c a c i d i n t h e p l a n t s d u r i n g d r y i n g . I r o n w a s f o u n d , in t h i s e x p e r i m e n t , to b e d i r e c t l y r e l a t e d to t h e a s c o r b i c a c i d i n tw o d i f f e r e n t a n d e v e n o p p o s i t e w a y s . In th e i r o n s e r i e s a n e g a t i v e c o r r e l a t i o n w a s f o u n d b e t w e e n t h e p e r ­ c e n t a g e of l o s s o f a s c o r b i c a c i d d u r i n g d r y i n g a n d t h e a c t u a l a m o u n t of i r o n i n th e f r e s h t i s s u e . O n t h e o t h e r h a n d , in t h e c o p ­ p e r s e r i e s a p o s itiv e c o r r e l a t i o n w a s o b ta in e d w h en the p e r c e n t a g e of l o s s of a s c o r b i c a c i d w a s c o m p a r e d to t h e a m o u n t of i r o n in t h e fre sh tissue. T h e s e two o p p o s ite r e s u l t s do n ot c o n t r a d i c t t h e m s e l v e s , b e c a u s e t h e y w e r e o b t a i n e d i n tw o d i f f e r e n t s e t s of c o n d i t i o n s , i n d e e d in t h e f i r s t t h e a m o u n t of i r o n i n t h e s o l u t i o n w a s i n c r e a s ­ in g a n d t h e c o p p e r m a i n t a i n e d c o n s t a n t a n d i n t h e s e c o n d t h e a m o u n t o f i r o n w a s l e f t c o n s t a n t i n t h e s o l u t i o n b u t t h e l e v e l s of c o p p e r w ere in creased . I t i s w o r t h p o i n t i n g o u t t h a t a t h a r v e s t in t h e i r o n s e r i e s t h e a m o u n t of a s c o r b i c a c i d f o u n d w a s i n v e r s e l y p r o p o r t i o n a l t o 58 th e a m o u n t of i r o n in t h e f r e s h t i s s u e a n d t h a t d u r i n g d r y i n g th e g r e a t e r s t a b i l i t y of t h e a s c o r b i c a c i d o c c u r r e d i n t h e p l a n t s w i t h th e h i g h e s t a m o u n t of i r o n . On t h e o p p o s i t e , in t h e c o p p e r s e r i e s , th e a m o u n t of a s c o r b i c a c i d w a s d i r e c t l y p r o p o r t i o n a l to the a m o u n t of i r o n in t h e p l a n t a n d d u r i n g d r y i n g t h e g r e a t e r s t a b i l i t y of t h e a s c o r b i c a c i d o c c u r r e d in t h e p l a n t w i t h t h e l o w e s t a m o u n t of i r o n in t h e t i s s u e . B e c a u s e of t h e c o n s i s t e n c y of t h e t r e n d of t h e i n f l u e n c e of h i g h e r l e v e l s of i r o n a n d c o p p e r on t h e s t a b i l i t y of a s c o r b i c a c i d d u r i n g th e tw o f i r s t e x p e r i m e n t s , it w o u l d b e v e r y d e s i r a b l e to k n o w if t h e s e t e n d e n c i e s p e r s i s t i n t h e r a n g e of t o x i c i t y of t h e s e elem ents. T h e r e would be a l s o a need fo r a m o r e c o m p le te a n a l ­ y s i s of t h e e f f e c t s c a u s e d b y th e d i f f e r e n t t r e a t m e n t s on t h e a m o u n t of t h e o t h e r e l e m e n t s a b s o r b e d a n d in p a r t i c u l a r on t h e a m o u n t s of c o p p e r t a k e n up b y t h e p l a n t s . E. 1. S u m m a r y of t h e S e c o n d E x p e r i m e n t W h e a t p l a n t s w e r e g r o w n in g r a v e l on t w o s e r i e s of c u l ­ tu re s whose b asic m e d iu m w as a com p lete th re e salt n u trien t so lu ­ tion w h e r e i r o n and c o p p e r w e r e the only v a ry in g f a c t o r s . The i r o n s e r i e s r a n g e d f r o m no i r o n to 18 p a r t s p e r m i l l i o n , e a c h c u l ­ t u r e v a r y i n g f r o m t h e o n e b e l o w b y a n i n c r e m e n t of tw o p a r t s p e r m illion. T h e c o p p e r s e r i e s r a n g e d f r o m n o c o p p e r to 0 . 1 8 p a r t s 59 p e r m illio n , e a c h c u ltu r e v a ry in g f r o m the one below by an i n c r e ­ m e n t of 0 . 0 2 p a r t s p e r m i l l i o n o f c o p p e r . This e x p e r im e n t w as d e s i g n e d t o o v e r l a p w i t h t h e f i r s t e x p e r i m e n t a n d to o b s e r v e if t h e s a m e t e n d e n c i e s w o u l d p e r s i s t f o r a w i d e r r a n g e of c o n c e n ­ t r a t i o n s of t h e s e t w o e l e m e n t s on t h e s y n t h e s i s a n d s t a b i l i t y of a s c o rb ic acid. 2. The plants w e r e h a r v e s te d ju s t b e fo re jointing stage. A s c o r b i c a cid w a s d e t e r m i n e d on the f r e s h t i s s u e s . d e t e r m i n e d on o v e n d r y s a m p l e s of t h e t i s s u e s . Iro n was A s c o rb ic acid w as d e te r m in e d ag ain j u s t a fte r dry in g , sev en d a y s a f te r drying and f o u r te e n d a y s a f t e r d ry in g , the d r i e d m a t e r i a l b eing k e p t a t co n sta n t te m p e r a t u r e at 30° C. T h e d a t a w e r e c a l c u l a t e d on a b a s i s p e r p l a n t , p e r 100 g r a m s f r e s h m a t t e r a n d p e r 100 g r a m s dry m atter. 3. F r e s h a n d d r y w e i g h t of t h e c u l t u r e s w e r e r e c o r d e d . T h e f r e s h w e i g h t of t h e p l a n t s in t h e i r o n s e r i e s f o l ­ lowed the s a m e te n d e n c y a s d u r in g the f i r s t e x p e r i m e n t : It i n ­ c r e a s e d w i t h h i g h e r i r o n c o n c e n t r a t i o n u p to s i x p a r t s p e r m i l l i o n i n t h e s o l u t i o n ; t h e d r y w e i g h t p a r a l l e l e d t h e t r e n d of t h e f r e s h w eight. I n t h e c o p p e r s e r i e s t h e f r e s h w e i g h t i n c r e a s e d up to t h e l a s t c u l t u r e of 0 . 1 8 p a r t s p e r m i l l i o n o f c o p p e r b u t t h e d r y w e i g h t v a r i e d w i d e l y a n d w a s f o u n d to c o r r e l a t e p o s i t i v e l y w i t h t h e i r o n a b s o rb e d p e r plant. 60 4. A s t h e i r o n c o n c e n t r a t i o n s i n c r e a s e d in t h e s o l u t i o n , a c o r r e s p o n d i n g i n c r e a s e of i r o n a b s o r b e d b y t h e p l a n t w a s o b ­ s e r v e d w h i c h w a s a l s o f o u n d in t h e c o n c e n t r a t i o n of t h i s e l e m e n t in t h e f r e s h a n d d r y t i s s u e s . 5. T h e a m o u n t o f a s c o r b i c a c i d f o u n d a t h a r v e s t in t h e p l a n t s of t h e i r o n s e r i e s , e x c e p t f o r t h e o n e g r o w n w i t h o u t i r o n , w a s i n v e r s e l y p r o p o r t i o n a l to th e a m o u n t of i r o n i n t h e s o l u t i o n a n d a l s o t o t h e a m o u n t of i r o n a b s o r b e d b y t h e p l a n t s . 6. T h e r a t e of l o s s of a s c o r b i c a c i d i n t h e p l a n t s of t h e i r o n s e r i e s d u r i n g d r y i n g w a s i n v e r s e l y c o r r e l a t e d to t h e a m o u n t of i r o n in t h e p l a n t a n d in t h e s o l u t i o n . 7. In th e c o p p e r s c r i e s , a p o s i t i v e c o r r e l a t i o n c o u l d b e d r a w n b e t w e e n t h e a m o u n t of a s c o r b i c a c i d a t h a r v e s t a n d t h e t o t a l a m o u n t of i r o n p e r p l a n t a n d a l s o t h e p e r c e n t a g e of i r o n i n the d r y m a t t e r . 8. T h e r a t e of l o s s of a s c o r b i c a c i d w a s i r r e g u l a r i n t h e p l a n t s of t h e c o p p e r s e r i e s a n d c o u l d n o t b e c o r r e l a t e d to t h e a m o u n t of c o p p e r i n t h e s o l u t i o n b u t w a s p o s i t i v e l y r e l a t e d to t h e c o n c e n t r a t i o n of i r o n i n t h e f r e s h t i s s u e . V. EX PERIM EN T 3 A. Purpose I t w a s f o u n d i n t h e tw o f i r s t e x p e r i m e n t s t h a t to a n i n c r e a s e of i r o n a n d c o p p e r i n t h e n u t r i e n t s o l u t i o n s c o r r e s p o n d e d a n i n ­ c r e a s e i n t h e s t a b i l i t y o f t h e a s c o r b i c a c i d d u r i n g t h e d r y i n g of t h e plants. S ince th e s e te n d e n c ie s p e r s i s te d o v e r the ra n g e of iro n a n d c o p p e r w h e r e t h e s e e l e m e n t s s h o w e d a s t i m u l a t i n g e f f e c t on g r o w t h , i t w a s p r o p o s e d to i n v e s t i g a t e t h e e f f e c t of t h e s e s a m e e l e ­ m e n t s up to t o x i c l e v e l s . I t w a s a l s o d e e m e d u s e f u l t o f o l l o w t h e e f f e c t of i n c r e a s i n g a m o u n t s of i r o n a n d c o p p e r up to t h e t o x i c l e v e l s on t h e a b s o r p t i o n of t h e o t h e r m a j o r e l e m e n t s , n i t r o g e n , p h o s p h o r u s , p o t a s s i u m , c a lc iu m , and m a g n e s iu m . T h e s y m p t o m s of t o x i c i t y a n d t h e p o s ­ s i b l e e f f e c t s on t h e a s c o r b i c a c i d w o u l d t h e n b e c o n n e c t e d w i t h t h e o b s e r v e d s t a t u s of t h e o t h e r e l e m e n t s . T h e u s e of l a r g e r i n c r e m e n t s f r o m o n e c u l t u r e to t h e o t h e r would p e r m i t a g a in an o v e r la p p in g of the p r e v i o u s c o n c e n t r a t i o n s and a l s o r e a c h toxic l e v e l s of c o p p e r and iro n . T he s a m e p la n ts and s a m e b a s ic n u t r i e n t so lu tio n would a l s o a llo w a c o m p a r i s o n of th e e ffe c t o b ta in e d w ith th o s e p r e v i o u s l y estab lish ed . 62 B. 1. E x p e rim e n ta l C onditions I r o n a n d C o p p e r L e v e l s in t h e N u t r i e n t S o l u t i o n T h e f o l l o w i n g a m o u n t s of i r o n a n d c o p p e r w e r e a d d e d to the s a m e c o m p l e t e t h r e e s a l t s s o lu ti o n s a s u s e d in the two f i r s t experim ents: C o n c e n t r a t i o n of I r o n an d C o p p e r in th e N u t r i e n t S o lu tio n s of t h e T h i r d E x p e r i m e n t Iron S e rie s Copper S eries p p m of F e i n th e S o l u t i o n p p m of C u in the S olution 0. 0 3.0 6. 0 9. 0 12.0 15.0 18.0 21.0 24.0 27.0 0.02 0.02 0.02 0.02 0.02 0.02 0,02 0.02 0.02 0.02 p p m of F e i n the Solution 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 1.0 p p m of C u i n the S olution 0. 0 0. 3 0.6 0.9 1.2 1.5 1.8 2.1 2,4 2.7 I r o n a n d c o p p e r w e r e a d d e d in t h e s a m e f o r m a s i n t h e t w o f i r s t e x p e r i m e n t s f r o m a s t o c k s o l u t i o n of f e r r i c s u l f a t e a n d c u p r i c sulfate. 63 2. P lan t M aterial On M a r c h 2, 1949, f o r t y s e e d s of w h e a t I l l i n o i s N o . 1 - 1 2 8 w e r e s o w n in t h e q u a r t z g r a v e l in e a c h p o t . The co leoptiles ap ­ p e a r e d on M a r c h 6, f o u r d a y s a f t e r p l a n t i n g . The p la n ts w e r e h a r v e s t e d on A p r i l 15, f o r t y - f i v e d a y s a f t e r p l a n t i n g . C. 1. a. E xperim ental R esults O b s e r v a t i o n s on t h e P l a n t s D e s c r i p t i o n of t h e P l a n t s On M a r c h 16, w h e n th e p l a n t s w e r e 15 d a y s o l d , n o t i c e a b l e d i f f e r e n c e s c o u l d b e o b s e r v e d b e t w e e n t h e v a r i o u s c u l t u r e s on b o t h series. In t h e c o p p e r s e r i e s a t o x i c i t y c o u l d b e n o t i c e d a t 0 . 6 p a r t s p e r m i l l i o n w h i c h i n c r e a s e d w i t h h i g h e r l e v e l s of c o p p e r . In t h e c u l t u r e s w h e r e t h e l e v e l of c o p p e r e x c e e d e d 0 . 6 p a r t s p e r m i l l i o n , the f i r s t le a f f o r m e d r e m a i n e d g r e e n , but th e new l e a v e s b e c a m e m o r e a n d m o r e c h l o r o t i c w i t h i n c r e a s i n g l e v e l s of c o p p e r . At h a r v e s t t h e p l a n t s g r o w n on 2 . 7 p a r t s p e r m i l l i o n of c o p p e r w e r e v e r y m u c h s t u n t e d i n g r o w t h a n d w e r e s t a r t i n g to p e r i s h . C h l o r o s i s s t a r t e d to a p p e a r 15 d a y s a f t e r s e e d i n g o n t h e p l a n t s g r o w n on t h e c u l t u r e s w h e r e n o i r o n h a d b e e n s u p p l i e d . In 64 th e r e m a i n i n g of t h e i r o n s e r i e s , t h e v i s i b l e q u a l i t y of t h e p l a n t s s h o w n b y s i z e a n d c o l o r of l e a v e s i n c r e a s e d w i t h h i g h e r c o n c e n ­ t r a t i o n s of i r o n up t o 15 p a r t s p e r m i l l i o n of i r o n i n t h e n u t r i e n t solution. On t h e h i g h e r l e v e l s of i r o n , t h e g e n e r a l a p p e a r a n c e of the p la n ts did n o t c h a n g e . b. F r e s h W eight p e r P la n t (1) E f f e c t of I r o n The d a ta in d ic a tin g the f r e s h w e ig h t p e r ten p la n ts e x p r e s s e d in g r a m s a r e g i v e n in T a b l e 23 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t of i r o n in t h e n u t r i e n t s o l u t i o n i n F i g u r e 151. T h e p l a n t s g r o w n on th e n u t r i e n t s o l u t i o n w h e r e n o i r o n h a d b e e n a d d e d w e r e m u c h s m a l l e r t h a n t h o s e of t h e r e m a i n i n g of t h e series. T h e f r e s h w e i g h t of t h e o t h e r p l a n t s s h o w e d a g e n e r a l i n ­ c r e a s e w i t h i n c r e a s e of i r o n i n t h e n u t r i e n t s o l u t i o n b u t t h e d i f f e r ­ e n c e s i n w e i g h t b e t w e e n tw o c o n s e c u t i v e c u l t u r e s w e r e n o t v e r y significant. (2) E f f e c t of C o p p e r T he d a t a f o r th e a v e r a g e f r e s h w e ig h t p e r ten p l a n t s of the c o p p e r s e r i e s e x p r e s s e d i n g r a m s a r e a s s e m b l e d i n T a b l e 25 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t s of c o p p e r i n t h e n u t r i e n t s o l u t i o n i n F i g u r e 193.. 65 T h e t e n d e n c y s h o w n b y t h e f r e s h w e i g h t s of t h e p l a n t s of the c o p p e r s e r i e s w a s m u c h m o r e s i g n i f i c a n t th a n the one o b ta in e d w ith iro n . Up to 0 . 6 p a r t s p e r m i l l i o n , no d e f i n i t e c h a n g e s o c c u r r e d i n t h e w e i g h t of t h e p l a n t s b u t t h e n w i t h i n c r e a s i n g c o n c e n t r a t i o n of c o p p e r, an a b r u p t d e c lin e w as o b s e rv e d . Indeed the plants grow n in t h e s o l u t i o n w i t h 1.8 p a r t s p e r m i l l i o n of c o p p e r w e i g h e d a b o u t 50 p e r c e n t l e s s t h a n t h o s e g r o w n on 0 . 6 p a r t s p e r m i l l i o n a n d a t 2 . 7 p a r t s p e r m i l l i o n t h e y w e i g h e d a b o u t 80 p e r c e n t l e s s t h a n t h o s e g r o w n on 0 . 6 p a r t s p e r m i l l i o n . c. D ry W eight p e r P la n t (1) E f f e c t of I r o n T h e a v e r a g e d r y w e i g h t p e r 10 p l a n t s o f t h e i r o n s e r i e s a r e a r r a n g e d in T a b l e 24 a n d g r a p h e d a g a i n s t th e a m o u n t s of i r o n in t h e n u t r i e n t s o l u t i o n in F i g u r e 144. F r o m n o n e to s i x p a r t s p e r m i l l i o n of i r o n i n t h e n u t r i e n t s o l u t i o n , t h e d r y w e i g h t of t h e p l a n t s i n c r e a s e d , b u t w i t h h i g h e r c o n c e n t r a t i o n the d r y w e ig h t s ta y e d a b o u t c o n s ta n t and no s p e c ia l tre n d could be defined. (2) E f f e c t of C o p p e r The d a ta fo r the a v e r a g e d r y w eight p e r ten p lan ts e x p r e s s e d 66 in g r a m s a r e s h o w n in T a b l e 26 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t s of c o p p e r i n n u t r i e n t s o l u t i o n i n F i g u r e 185. T o a n i n c r e a s e o f c o p p e r i n t h e n u t r i e n t s o l u t i o n of 0 . 6 to 2.7 p a r t s p e r m i l l i o n c o r r e s p o n d e d a d e c r e a s e in th e d r y w e ig h t of t h e w h e a t p l a n t s ; t h e s e l e v e l s w e r e t o x i c t o t h e p l a n t s a n d w e r e s h o w n to p r o d u c e c h l o r o s i s i n t h e n e w l e a v e s f o r m e d . . I n a g e n e r a l way, th e d r y w e ig h t fo llo w e d the s a m e te n d e n c y a s the f r e s h w eig h t. 2. C h e m i c a l A n a l y s e s of t h e P l a n t s T h e e f f e c t s of i n c r e a s i n g t h e a m o u n t o f i r o n a n d c o p p e r in t h e n u t r i e n t s o l u t i o n o n t h e c h e m i c a l c o m p o s i t i o n of t h e w h e a t p l a n t s w a s d e t e r m i n e d b y t h e a n a l y s e s of t h e e x p e r i m e n t a l s a m ­ p les fo r ir o n and c o p p e r and the o th e r m a j o r c h e m i c a l c o m p o n e n ts: nitrogen, phosphorus, p o ta ssiu m , c alciu m , and m a g n e siu m , as w ell a s a s c o rb ic acid. a. Iron (1) A m o u n t s o f I r o n p e r P l a n t (a) E f f e c t of i n c r e a s i n g t h e a m o u n t of i r o n in t h e n u t r i e n t s o l u t i o n o n t h e i r o n c o n t e n t of t h e p l a n t s . T h e a m o u n ts of i r o n e x ­ p r e s s e d i n m i l l i g r a m s , a c c u m u l a t e d p e r p l a n t s of t h e i r o n s e r i e s a r e r e p o r t e d i n T a b l e 24 a n d g r a p h e d a g a i n s t t h e a m o u n t of i r o n i n F i g u r e 145. 67 A s in t h e tw o f i r s t e x p e r i m e n t s , to a n i n c r e a s e of i r o n in t h e s o l u t i o n c o r r e s p o n d e d a n i n c r e a s e i n t h e t o t a l a m o u n t of i r o n f ound p e r p l a n t . T h is i n c r e a s e did not follow a s t r a i g h t c u r v e but w a s g r e a t e r in t h e l o w c o n c e n t r a t i o n s of i r o n in t h e n u t r i e n t s o l u ­ t i o n up to n i n e p a r t s p e r m i l l i o n a n d t h e n f l a t t e n e d . W hen the v a l u e s f o r t h e t o t a l a m o u n t o f i r o n p e r p l a n t w e r e c o m p a r e d to t h e v a l u e s f o r t h e t o t a l a m o u n t s of t h e o t h e r c o m p o n e n t s , p o s i t i v e c o r ­ r e l a t io n s w ith the follow ing c o m p o n e n ts w e r e o b s e r v e d : phosphorus, n itro g e n , p o t a s s i u m , m a g n e s i u m , and a l s o the d r y w e ig h t and the f re s h w eight. The o th e r r e la tio n s d ealing w ith a s c o r b ic acid w ill be c o n s id e re d l a te r . T h e se n u m e ro u s p o sitiv e c o r re la tio n s show th at an i n c r e a s e of i r o n p r e s e n t in the s o lu tio n s t i m u l a t e d g r o w th a n d t h e a b s o r p t i o n of t h e o t h e r e l e m e n t s . No l e v e l s h a d b e e n r e a c h e d w h e r e t h e a m o u n t of i r o n s e e m e d d e t r i m e n t a l . (b) E f f e c t of i n c r e a s i n g t h e a m o u n t of c o p p e r in t h e n u t r i e n t s o l u t i o n on t h e a c c u m u l a t i o n of i r o n by t h e p l a n t . The m illig ra m s of i r o n a c c u m u l a t e d p e r t e n p l a n t s a r e p r e s e n t e d i n T a b l e 26 a n d g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r in th e n u t r i e n t s o l u t i o n in F i g u r e 186. A b o v e 0 . 6 p a r t s p e r m i l l i o n of c o p p e r i n t h e s o l u t i o n , t o a n i n c r e a s e of c o p p e r c o r r e s p o n d e d a d e c r e a s e i n t h e t o t a l a m o u n t of iro n a b s o rb e d p e r plant. T h is d e c r e a s e w a s fo llo w e d by a ll the 68 o t h e r c o m p o n e n t s a n a l y z e d f o r i n t h e p l a n t w i t h t h e e x c e p t i o n of calcium . T h is o v e r - a l l effe c t of c o p p e r is u n d e r s to o d by the fact t h a t a b o v e 0 . 6 p a r t s p e r m i l l i o n in t h e n u t r i e n t s o l u t i o n , t h e c o n ­ c e n t r a t i o n s w e r e m o r e a n d m o r e t o x i c to t h e p l a n t s a n d c h e c k e d grow th c o rre s p o n d in g ly . (2) A m o u n t s of I r o n p e r 100 G r a m s of F r e s h M a t e r i a l (a) E f f e c t of i r o n . T h e d a t a i n d i c a t i n g th e m i l l i g r a m s of i r o n p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l of t h e i r o n s e r i e s a r e s h o w n i n T a b l e 28 a n d F i g u r e 159. T h e c o n c e n t r a t i o n s of i r o n i n t h e p l a n t t i s s u e i n c r e a s e d w i t h h i g h e r a m o u n t s of i r o n in t h e s o l u t i o n . T his tendency w as s i m i l a r t o t h e o n e o b t a i n e d d u r i n g th e tw o f i r s t e x p e r i m e n t s , b u t a b o v e 18 p a r t s p e r m i l l i o n o f i r o n in t h e n u t r i e n t s o l u t i o n to t h e f o l l o w i n g i n c r e a s e s of i r o n in t h e n u t r i e n t s o l u t i o n c o r r e s p o n d e d a d e c r e a s e in t h e c o n c e n t r a t i o n of i r o n i n th e p l a n t m a t e r i a l . Pos­ i t i v e c o r r e l a t i o n s b e t w e e n t h e c o n c e n t r a t i o n s of i r o n i n t h e f r e s h t i s s u e and n itro g e n , p o ta s s iu m , and d r y m a t t e r w e r e o b s e r v e d . (b) E f f e c t of c o p p e r . T h e m i l l i g r a m s of i r o n p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l of t h e c o p p e r s e r i e s a p p e a r in T a b l e 30 a n d a r e g r a p h e d a g a i n s t t h e a m o u n t of c o p p e r i n t h e s o l u t i o n i n F i g u r e 201. 69 W i t h t h e i n c r e a s e s of c o p p e r in t h e n u t r i e n t s o l u t i o n u p to 1.5 p a r t s p e r m i l l i o n c o r r e s p o n d e d a d e c r e a s e in t h e a m o u n t s of i r o n in t h e t i s s u e . A f t e r t h i s l e v e l t h e c o n c e n t r a t i o n of i r o n i n t h e t i s s u e s t a r t e d to i n c r e a s e up to 2 . 7 p a r t s p e r m i l l i o n of c o p p e r in the n u t r i e n t s o lu tio n . T h i s t e n d e n c y w a s n o t f o l l o w e d b y a n y of the o th e r c o m p o n e n t s and fo r the f i r s t tim e no a p p a r e n t c o r r e l a ­ t i o n w a s o b t a i n e d b e t w e e n t h e i r o n of t h e t i s s u e a n d t h e a m o u n t of d ry w eight. (3) A m o u n t of I r o n p e r 100 G r a m s of D r y M a t t e r (a) E f f e c t o f i r o n . T h e a m o u n t s of i r o n in m i l l i g r a m s p e r 100 g r a m s of d r y p l a n t m a t e r i a l a r e p r e s e n t e d in T a b l e 31 a n d F i g u r e 170. N o t m u c h d i f f e r e n c e s w e r e o b s e r v e d in t h e c o n c e n t r a t i o n of i r o n f o u n d in t h e d r y t i s s u e s of t h e p l a n t s g r o w n on t h e v a r i o u s l e v e l s of i r o n , h o w e v e r a s l i g h t u p w a r d t e n d e n c y c a n b e d e t e c t e d up to 24 p a r t s p e r m i l l i o n o f i r o n . T he v a lu e f o r the “ no i r o n ” p l a n t s i s o u t of t h e g e n e r a l l i n e a n d o u g h t to b e d i s r e g a r d e d f o r this c o n s id e ra tio n . No a p p a r e n t r e la tio n w ith o th e r c o m p o n e n ts w e r e found. (b) E f f e c t of c o p p e r . T h e d a t a f o r t h e a m o u n t s of i r o n p e r 100 g r a m s of t h e d r y m a t e r i a l in t h e c o p p e r s e r i e s a r e a s s e m b l e d i n T a b l e 32 a n d in F i g u r e 208. 70 T h e a m o u n t s o f i r o n f o u n d in t h e d r y t i s s u e , w a s i n v e r s e l y r e l a t e d t o t h e a m o u n t of c o p p e r in t h e n u t r i e n t s o l u t i o n . T his r e ­ la tio n s h ip w a s o b s e r v e d d u r in g the f i r s t e x p e r i m e n t b u t did not a p p e a r a t the second. No e v id e n t c o r r e l a t i o n could be found w ith the o th e r c o m p o n e n ts. b. Copper (1) D e s c r i p t i o n I t w a s of i n t e r e s t f o r t h i s s t u d y t o o b t a i n i n f o r m a t i o n on t h e a c c u m u l a t i v e p o w e r of t h e w h e a t p l a n t s t o w a r d t h e c o p p e r p r e s e n t in i n c r e a s i n g a m o u n t s in t h e s o l u t i o n . A nalyses for copper w ere m a d e o n l y on t h e p l a n t s of t h e c o p p e r s e r i e s w h e r e s i g n i f i c a n t d i f f e r e n c e s in t h e a m o u n t s of c o p p e r b e t w e e n p l a n t s c o u l d b e o b ­ tained. (2) A m o u n t of C o p p e r p e r P l a n t T h e m i l l i g r a m s o f c o p p e r a b s o r b e d p e r p l a n t g r o w n on n u t r i e n t s o l u t i o n s c o n t a i n i n g i n c r e a s i n g a m o u n t s of t h i s e l e m e n t a r e p r e s e n t e d in T a b l e 25 a n d g r a p h e d a g a i n s t t h e a m o u n t of c o p ­ p e r in t h e s o l u t i o n i n F i g u r e 192. Up to 0 . 6 p a r t s p e r m i l l i o n of c o p p e r i n t h e n u t r i e n t s o l u ­ t i o n c o r r e s p o n d e d a s h a r p i n c r e a s e i n th e t o t a l a m o u n t o f c o p p e r a c c u m u la te d p e r plant. A f t e r t h i s l e v e l , w h i c h c o r r e s p o n d e d to 71 t h e b e g i n n i n g of t o x i c i t y s y m p t o m s , , t h e t o t a l a m o u n t of c o p p e r found p e r p l a n t d e c r e a s e d s h a r p l y in an i n v e r s e r e l a t i o n s h i p w ith t h e l e v e l s o f c o p p e r in t h e n u t r i e n t s o l u t i o n . T h is r e s u l t m ig h t be e x p l a i n e d b y t h e e f f e c t of t o x i c l e v e l s of c o p p e r on t h e t o t a l g r o w t h , w h i c h t h e n d i d n o t a l l o w t h e p l a n t to a b s o r b m o r e of t h i s e l e m e n t . W h e n t h e d a t a f o r t h e o t h e r c o m p o n e n t s a r e c a l c u l a t e d on a p e r p la n t b a s i s , the s a m e d e c r e a s i n g te n d e n c y is o b s e r v e d . (3) A m o u n t s of C o p p e r p e r 100 G r a m s of F r e s h M a t t e r T h e a m o u n t s of c o p p e r i n m i l l i g r a m s p e r 100 g r a m s of f r e s h t i s s u e of t h e c o p p e r s e r i e s a r e r e p o r t e d in T a b l e 30 a n d F i g u r e 202. A s tr o n g p o s itiv e c o r r e l a t i o n w a s found b e tw e e n the c o n c e n ­ t r a t i o n s of c o p p e r in t h e n u t r i e n t s o l u t i o n a n d t h e a m o u n t s o f c o p p e r f o u n d in t h e f r e s h t i s s u e . I n d e e d t h e c o n c e n t r a t i o n of c o p p e r i n t h e f r e s h t i s s u e of p la n ts g ro w n on the 2.7 p a r t s p e r m i l l i o n of c o p p e r w a s 3 . 4 t i m e s g r e a t e r t h a n i t s c o n c e n t r a t i o n in t h e p l a n t t i s s u e grow n w ithout co p p er. I t i s i n t e r e s t i n g to n o t e t h a t a b o v e 1.2 p a r t s p e r m i l l i o n o f c o p p e r in the n u trie n t solution c o r r e s p o n d e d an a p p a r e n t and g e n ­ e r a l c h a n g e of r a t e of a c c u m u l a t i o n of t h e f o l l o w i n g e l e m e n t s : p h o s p h o r u s , c a l c i u m , a n d m a g n e s i u m ( F i g u r e s 195, 198, a n d 199). 72 (4) A m o u n t s of C o p p e r p e r 100 G r a m s of D r y M a t t e r T h e d a t a f o r t h e a m o u n t s of c o p p e r in m i l l i g r a m s p e r 100 g r a m s o f d r y m a t e r i a l in t h e c o p p e r s e r i e s a r e a s s e m b l e d i n T a b l e 32 a n d g r a p h e d a g a i n s t t h e a m o u n t o f c o p p e r in t h e s o l u t i o n i n F i g ­ u r e 214. T h e s a m e i n c r e a s i n g t e n d e n c y i n t h e c o n c e n t r a t i o n of c o p p e r a s in t h e f r e s h m a t t e r , o c c u r r e d i n t h e d r y t i s s u e of p l a n t s g r o w n on i n c r e a s i n g l e v e l s of c o p p e r . H ow ever, when th ese values a r e c o n s i d e r e d , i t i s o b s e r v e d t h a t t h e r a t e of a c c u m u l a t i o n o f c o p p e r on a d r y m a t t e r b a s i s w a s n o t a s g r e a t a t t h e h i g h e r l e v e l s of su p p ly a s at the lo w e r l e v e l s . This s a m e o b s e rv a tio n had been m a d e b y L y o n a n d B e e s o n (1 94 8) f o r o t h e r m i c r o e l e m e n t s s u c h a s m o ly b d e n u m , m a n g a n e s e and zinc. A s w a s o b s e r v e d w i t h t h e d a t a on a f r e s h w e i g h t b a s i s , a c h a n g e in t h e r a t e of a b s o r p t i o n of p h o s p h o r u s , c a l c i u m a n d m a g ­ n e s iu m and a l s o p o t a s s i u m and m a g n e s i u m w as a p p a r e n t , w hen the v a l u e s w e r e c a l c u l a t e d on a d r y m a t t e r b a s i s , f o r t h e p l a n t s g r o w n a b o v e t h e 1.2 p a r t s p e r m i l l i o n o f c o p p e r l e v e l ( F i g u r e s 209, 2 1 2 , 2 1 0 , a n d 21 3). ,c. A s c o rb ic A cid (1) D e s c r i p t i o n It w a s o b s e r v e d d u r in g the two f i r s t e x p e r i m e n t s t h a t the a m o u n t s of a s c o r b i c a c i d d e c r e a s e d m o s t l y d u r i n g t h e d r y i n g p r o ­ cess. F o r t h i s r e a s o n , i t w a s d e c i d e d t o f o l l o w t h e l o s s of a s c o r b i c a c i d d u r i n g t h e d r y i n g of t h e p l a n t m a t e r i a l . The o riginal am ounts of a s c o r b i c a c i d a t h a r v e s t w e r e f i r s t d e t e r m i n e d , t h e n t h e p l a n t s w e r e l e f t in t r a y s in t h e l a b o r a t o r y a t r o o m t e m p e r a t u r e a n d a f t e r i n t e r v a l s , p o r t i o n s of t h e d r y i n g s a m p l e s w e r e t a k e n a n d a n a l y z e d . F o r the ir o n s e r i e s , the f i r s t d e te r m in a tio n s w e r e done at h a r v e s t, t h e s e c o n d f o u r t e e n h o u r s l a t e r , t h e t h i r d 43 h o u r s a f t e r h a r v e s t , t h e n 62, 110, a n d 157 h o u r s a f t e r h a r v e s t . F o r the c o p p e r s e r i e s t h e f i r s t d e t e r m i n a t i o n s w e r e d o n e a t h a r v e s t , t h e s e c o n d 13 h o u r s l a t e r , t h e t h i r d 30 h o u r s a f t e r h a r v e s t , t h e n 52, 100, a n d 147 h o u r s after harvest. T h e a m o u n t s of a s c o r b i c a c i d f o u n d a t h a r v e s t w e r e c a l ­ c u l a t e d on a p e r p l a n t , p e r 100 g r a m s f r e s h m a t t e r a n d 100 g r a m s d r y m a t t e r b a s i s i n o r d e r t o b e a b l e to c o m p a r e t h e s e r e s u l t s w i t h t h o s e of t h e o t h e r c o m p o n e n t s a n a l y z e d . On t h e o t h e r h a n d , t h e a m o u n t s of a s c o r b i c a c i d l e f t a t t h e v a r i o u s i n t e r v a l s of d r y i n g w e r e o n l y c a l c u l a t e d o n a 100 g r a m s f r e s h w e i g h t b a s i s b e c a u s e t h e r a t e of o x i d a t i o n of t h e a s c o r b i c a c i d i s i n d e p e n d e n t of t h e w a y the am o u n t is c a lc u la te d and c o m p a r a b l e r e s u l t s a r e ob tain ed when t h e d a t a a r e c o m p u t e d a s t h e p e r c e n t a g e of t h e o r i g i n a l a m o u n t found at h a r v e s t . A s p e c i a l p a r a g r a p h w ill c o n s i d e r th e p e r c e n t ­ a g e s o f t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d l e f t a t e a c h d e t e r m i n a ­ tion. (2) A m o u n t s of A s c o r b i c A c i d p e r P l a n t (a) E f f e c t of iro n . T h e d a t a e x p r e s s i n g t h e a m o u n t s of a s c o r b i c a c i d a t h a r v e s t p e r t e n p l a n t s g r o w n a t v a r i o u s l e v e l s of i r o n a r e a s s e m b l e d in T a b l e 23 a n d a r e g r a p h e d a g a i n s t th e a m o u n t of i r o n in t h e s o l u t i o n i n F i g u r e 143. T h e g e n e r a l t e n d e n c y of t h e t o t a l a m o u n t of a s c o r b i c a c i d in t h e p l a n t s , g r o w n on i n c r e a s i n g a m o u n t s of i r o n , w a s to i n c r e a s e ; a l t h o u g h t h i s t r e n d w a s a p p a r e n t , th e d i f f e r e n c e s b e t w e e n a v e r a g e v a l u e s of a s c o r b i c a c i d w e r e o n l y of f e w m i l l i g r a m s p e r t e n p l a n t s . T h is p o s itiv e r e la t io n s h i p do n ot co in c id e w ith the r e s u l t s o b tained d u r i n g t h e s e c o n d e x p e r i m e n t w h e r e , on t h e o p p o s i t e , a n e g a t i v e r e la tio n s h ip had b een obtained. W h e n t h e v a l u e s of a s c o r b i c a c i d a t h a r v e s t a r e c o m p a r e d to the o th er c o m p o n e n ts a p o sitiv e re la tio n can be o b s e rv e d with t h e t o t a l f r e s h a n d d r y w e i g h t a s w e l l a s w i t h t h e t o t a l a m o u n t of iro n , p h o s p h o ru s , n itro g e n and p o t a s s i u m p e r plant. (b) E f f e c t of c o p p e r . T h e d a t a f o r t h e a m o u n t of a s c o r b i c a c id a t h a r v e s t p e r ten p la n ts of th e c o p p e r s e r i e s a r e p r e s e n t e d i n T a b l e 25 a n d F i g u r e 184. T h e m o r e c o p p e r w a s a d d e d to the n u t r i e n t s o lu tio n , the le s s total a s c o r b i c acid w as obtained p e r plant. The m ain cause of t h i s d e c r e a s e in t h e a m o u n t of a s c o r b i c a c i d w a s t h e d e c r e a s i n g e f f e c t of c o p p e r on t h e g r o w t h of t h e p l a n t s . P ositive co rrelatio n s b e t w e e n a s c o r b i c a c i d a n d o t h e r c o m p o n e n t s of t h e p l a n t s w e r e o b ­ t a i n e d w i t h t h e f r e s h a n d d r y w e i g h t , t h e t o t a l a m o u n t s of i r o n , p h o s p h o r u s , p o t a s s i u m , m a g n e s i u m , a n d in t h e p l a n t s g r o w n on s o l u t i o n s c o n t a i n i n g m o r e t h a n 0 . 6 p a r t s p e r m i l l i o n of c o p p e r w i t h t h e t o t a l a m o u n t of c o p p e r i t s e l f . (3) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t t e r (a) E f f e c t of i r o n . T h e d a t a e x p r e s s e d in m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h m a t t e r a t h a r v e s t a p p e a r in T a b l e 28 a n d t h e a m o u n t s of a s c o r b i c a c i d a t h a r v e s t a n d a t t h e v a r i o u s t i m e s d u r i n g d r y i n g a r e g r a p h e d a g a i n s t t h e a m o u n t s of i r o n i n t h e s o l u t i o n in F i g u r e 152 a n d f r o m F i g u r e s 160 t o 1 64 inclusive. T h e c o n c e n t r a t i o n s of a s c o r b i c a c i d in t h e f r e s h t i s s u e a t h a r v e s t s h o w e d a d e c r e a s i n g t e n d e n c y w i t h i n c r e a s i n g a m o u n t s of i r o n in the n u t r i e n t so lu tio n . A lthough l e s s a p p a re n t, this te n d e n c y 76 w a s s i m i l a r to t h e o n e r e c o r d e d d u r i n g t h e s e c o n d e x p e r i m e n t . On the o th e r hand, a p o s itiv e c o r r e l a t i o n i s o b s e r v e d b e tw e e n the c o n ­ c e n t r a t i o n of a s c o r b i c a c i d f o u n d in t h e d r y i n g m a t e r i a l 15 h o u r s a f t e r h a r v e s t a n d t h e a m o u n t of i r o n in t h e n u t r i e n t so lu tio n.. W h e n t h e v a l u e s f o r th e a m o u n t s of a s c o r b i c a c i d f o u n d a t h a r v e s t a r e p l o t t e d a g a i n s t th e a m o u n t s of i r o n i n t h e f r e s h t i s s u e a n e g a t i v e c o r r e l a t i o n i s o b s e r v e d ( F i g u r e 165). On t h e o t h e r h a n d , w h e n t h e v a l u e s f o r t h e a m o u n t o f a s c o r b i c a c i d f o und a t t h e f i r s t d e t e r m i n a t i o n 14 h o u r s a f t e r h a r v e s t a r e p l o t t e d a g a i n s t t h e a m o u n t of i r o n in t h e f r e s h t i s s u e a p o s i t i v e c o r r e l a t i o n a p p e a r s . This c h a n g e f r o m a n e g a tiv e to a p o s itiv e c o r r e l a t i o n d u r in g th e f i r s t h o u r s of d r y i n g h a d a l r e a d y b e e n o b s e r v e d d u r i n g t h e s e c o n d e x ­ p erim ent. (b) T h e s e r e l a t i o n s m a y b e s e e n i n F i g u r e 166. E f f e c t of c o p p e r . T h e a v e r a g e m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l g r o w n in t h e c o p p e r s e r i e s a n d d e t e r m i n e d a t h a r v e s t a r e r e c o r d e d i n T a b l e 30 a n d t h e a m o u n t s of a s c o r b i c a c i d a t h a r v e s t a n d a t t h e v a r i o u s t i m e s d u r i n g d r y i n g a r e g r a p h e d a g a i n s t th e a m o u n t of c o p p e r i n t h e n u ­ t r i e n t s o l u t i o n in F i g u r e 194 a n d f r o m F i g u r e s 203 to 2 0 7 , i n c l u s i v e . Up to a l e v e l of 0 . 9 p a r t s p e r m i l l i o n of c o p p e r i n t h e n u ­ t r i e n t s o l u t i o n , t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d i n t h e c o r r e ­ sponding f r e s h m a t e r i a l a t h a r v e s t had a te n d e n c y to d e c r e a s e ; t h e n t h i s t e n d e n c y c h a n g e d to a p o s i t i v e d i r e c t i o n w i t h h i g h e r a m o u n t s of c o p p e r i n t h e s o l u t i o n . T hese tendencies a re co m p arab le to t h o s e o b s e r v e d f o r t h e c o n c e n t r a t i o n s of p h o s p h o r u s , c a l c i u m , a n d m a g n e s i u m in t h e f r e s h m a t e r i a l ( F i g u r e s 195, 198, a n d 199). In t h e p l a n t t i s s u e g r o w n on t h e l o w c o p p e r c o n c e n t r a t i o n s , the a s c o r b i c a c id w a s l o s t a t a f a s t e r r a t e th a n in the t i s s u e g ro w n on t h e h i g h c o p p e r l e v e l d u r i n g t h e 13 f i r s t h o u r s a f t e r h a r v e s t . A p o s itiv e c o r r e l a t i o n is o b s e r v e d , w hen the c o n c e n tr a tio n s of a s c o r ­ b ic a c i d a t t h a t d e t e r m i n a t i o n a r e g r a p h e d a g a i n s t th e a m o u n t s of c o p p e r in t h e s o l u t i o n ( F i g u r e 2 03) . A f t e r 30 h o u r s , t h e v a l u e s f o r a s c o r b i c a c i d in a l l t h e c u l ­ t u r e s b e c a m e a l m o s t s i m i l a r ( F i g u r e 204). W h e n t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d a t h a r v e s t a n d 13 h o u r s a f t e r h a r v e s t a r e p l o t t e d a g a i n s t t h e a m o u n t s o f c o p p e r in t h e f r e s h t i s s u e , a s in F i g u r e s 167 a n d 168, i t i s f o u n d t h a t to t h e h i g h e r a m o u n t of c o p p e r c o r r e s p o n d a t b o t h d e t e r m i n a t i o n s t h e h i g h e r a m o u n t of a s c o r b i c a c i d : A positive re la tio n sh ip w as ob­ ta in e d in b o th c a s e s b e tw e e n a s c o r b i c a c id and c o p p e r . These r e ­ s u l t s c a n b e c o m p a r e d t o t h e o n e o b t a i n e d b y L y o n a n d B e e s o n (1948) w i t h t o m a t o e s w h e r e s i g n i f i c a n t l y h i g h e r a m o u n t s of a s c o r b i c a c i d r e s u l t e d f r o m i n c r e a s e i n t h e c o p p e r c o n c e n t r a t i o n of t h e n u t r i e n t m edium . 78 I n t h e p l a n t s of t h e c o p p e r s e r i e s , no a p p a r e n t c o r r e l a t i o n b e t w e e n t h e c o n c e n t r a t i o n of a s c o r b i c a c i d a n d t h e a m o u n t of i r o n in t h e t i s s u e c a n b e d r a w n a s i n t h e p l a n t t i s s u e s f r o m t h e s e c o n d e x p e r i m e n t ( F i g u r e 169). (4) P e r c e n t a g e s o f t h e O r i g i n a l A m o u n t s of A s c o r b i c A c i d F o u n d at E a ch D eterm in atio n (a) E f f e c t of i r o n . T h e v a l u e s f o r t h e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d f o u n d i n t h e i r o n s e r i e s 14, 43 , 62, 110, a n d 157 h o u r s a f t e r h a r v e s t , a r e r e p o r t e d in T a b l e 33, g r a p h e d a g a i n s t t h e a m o u n t of i r o n i n th e n u t r i e n t s o l u t i o n in F i g ­ u r e 2 29, a n d g r a p h e d a g a i n s t t h e a m o u n t of i r o n in 100 g r a m s of f r e s h m a t t e r in F i g u r e s 176, 177, 178, a n d 179. A s in t h e tw o f i r s t e x p e r i m e n t s , t h e a m o u n t of a s c o r b i c a c i d l o s t d u r i n g d r y i n g w a s s m a l l e r in t h e p l a n t s g r o w n on t h e s o l u ­ tion w ith a h ig h e r a m o u n t of iro n . In f a c t , a f t e r 14 h o u r s , 5 3 . 6 p e r c e n t of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d w a s l o s t i n t h e p l a n t g r o w n on t h r e e p a r t s p e r m i l l i o n o f i r o n , w h i l e o n l y 22 p e r c e n t w e r e l o s t f r o m t h e p l a n t s g r o w n on 24 p a r t s p e r m i l l i o n of i r o n . A p o s it i v e c o r r e l a t i o n is o b ta in e d w h en th e a m o u n t s o f i r o n in the f r e s h t i s s u e a r e g r a p h e d a g a i n s t t h e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d . T h i s c o r r e l a t i o n i s v e r y a p p a r e n t 14 h o u r s a n d 43 h o u r s a f t e r h a r v e s t a n d t h e n s t a r t s to d i s a p p e a r a f t e r 79 62 h o u r s . A t t h e f o u r t h d e t e r m i n a t i o n m a d e 110 h o u r s a f t e r h a r ­ v e s t , n o c o r r e l a t i o n b e t w e e n t h e p e r c e n t a g e of a s c o r b i c a c i d a n d t h e a m o u n t of i r o n i n t h e f r e s h t i s s u e e x i s t e d a n y m o r e . These r e l a t i o n s h i p s c a n b e f o l l o w e d i n F i g u r e s 176, 177, 178, a n d 179. I t w a s o b s e r v e d t h a t d u r i n g t h e d r y i n g of t h e p l a n t s a t r o o m t e m p e r a t u r e , th e l o s s of w a t e r f r o m t h e p l a n t m a t e r i a l w a s of t h e s a m e r e l a t i v e m a g n i t u d e a s t h e l o s s i n a s c o r b i c a c i d . This r e l a t i o n s h i p c a n b e n o t e d w h e n F i g u r e 22 8 a n d F i g u r e 229 a r e com pared. W h e n th e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t s of w a t e r l e f t a f t e r 43 h o u r s a r e g r a p h e d a g a i n s t t h e a m o u n t of i r o n i n t h e f r e s h tis s u e , a positive c o r r e la tio n is obtained as seen fro m F i g ­ u r e 181. T h i s r e l a t i o n s h i p i s a p p a r e n t 43 h o u r s a f t e r h a r v e s t a n d s t a r t s t o d i s a p p e a r a f t e r 62 h o u r s a s i n t h e c a s e of t h e r e l a t i o n ­ s h i p b e t w e e n i r o n in t h e f r e s h t i s s u e a n d t h e p e r c e n t a g e l o s s of a s c o r b i c a c i d ( F i g u r e s 180, 181, 182, a n d 183). (b) E f f e c t of c o p p e r . T h e d a t a f o r t h e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d f o u n d a t e a c h d e t e r m i n a t i o n i n t h e p l a n t s g r o w n on t h e c o p p e r c u l t u r e s a r e l i s t e d in T a b l e 34 a n d F i g u r e 227. D e t e r m i n a t i o n s of a s c o r b i c a c i d w e r e m a d e a t h a r ­ v e s t , 13, 30, 52, 100, a n d 147 h o u r s a f t e r h a r v e s t . D u r i n g t h e f i r s t 13 h o u r s a f t e r h a r v e s t , t h e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t o f a s c o r b i c a c i d l o s t w a s i n v e r s e l y 80 p r o p o r t i o n a l to t h e a m o u n t of c o p p e r i n t h e n u t r i e n t s o l u t i o n . In­ d e e d , 4 3 . 4 p e r c e n t of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d h a d b e e n l o s t f r o m t h e “ no c o p p e r ” c u l t u r e t h o u g h o n l y t e n p e r c e n t w a s l o s t in the p la n t g r o w n on the n u t r i e n t s o lu tio n s u p p lie d w ith 2 , 7 p a r t s p e r m i l l i o n of c o p p e r . It w a s a ls o n o te d th a t the g e n e r a l r a t e o f l o s s of a s c o r b i c a c i d in t h e p l a n t s g r o w n on t h e c o p p e r s e r i e s w a s s l o w e r d u r i n g t h e f i r s t 15 h o u r s t h a n in t h e p l a n t s of the iro n s e r i e s . W h e n t h e p e r c e n t a g e v a l u e s o b t a i n e d f o r th e p l a n t s w h i c h w e r e l e f t 30 h o u r s a r e c o n s i d e r e d , i t i s f o u nd t h a t t h e p o s i t i v e r e l a t i o n b e t w e e n a s c o r b i c a c i d s t a b i l i t y a n d t h e a m o u n t of c o p p e r in t h e n u t r i e n t s o l u t i o n d i s a p p e a r e d a n d e v e n s h o w e d a s l i g h t n e g ­ ative tendency. A t t h e n e x t d e t e r m i n a t i o n s 52, 100, a n d 147 h o u r s a f t e r h a r v e s t , no r e l a t i o n s h i p w a s n o t e d a n y m o r e , b e t w e e n c o p p e r in th e n u t r i e n t s o l u t i o n a n d t h e p e r c e n t a g e of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d . W h e n t h e a m o u n t s of c o p p e r in 100 g r a m s of f r e s h m a t e r i a l a r e p l o t t e d a g a i n s t t h e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d l e f t a f t e r 13 h o u r s of d r y i n g a s i g n i f i c a n t p o s i t i v e r e l a t i o n s h i p a p p e a r s , b u t w h ic h l o s e s i t s s ig n i f i c a n c e w hen the v a l u e s a t 30 h o u r s a r e g r a p h e d . i n F i g u r e 219 a n d 2 2 0 . T h ese re latio n sh ip s can be seen 81 N o r e l a t i o n s h i p i s a p p a r e n t a f t e r 13 a n d 30 h o u r s b e t w e e n t h e r a t e of l o s s of a s c o r b i c a c i d a n d t h e a m o u n t o f i r o n f o u n d in t h e f r e s h t i s s u e a s s h o w n in F i g u r e s 221 a n d 2 2 2 . I t w a s n o t e d t h a t d u r i n g t h e d r y i n g of t h e p l a n t s i n t h e c a s e of th e i r o n s e r i e s a n d of t h e c o p p e r s e r i e s , t h e l o s s of w a t e r f r o m t h e p l a n t m a t e r i a l w a s of t h e s a m e r e l a t i v e m a g n i t u d e a s t h e l o s s in a s c o r b i c a c i d . This re la tio n s h ip can be o b s e rv e d when F ig u r e s 226 a n d 22 7 a r e c o m p a r e d . A l t h o u g h t h e r e l a t i v e m a g n i t u d e s of the l o s s e s w e r e s i m i l a r , t h e i r r a t e s w e r e s p e c if ic . T h e r a t e of w a t e r l o s s f r o m t h e p l a n t s g r o w n on t h e c o p p e r s e r i e s w a s p r o ­ p o r t i o n a l t o t h e a m o u n t of c o p p e r in t h e n u t r i e n t s o l u t i o n s on w h i c h t h e y g r e w , a s s h o w n in F i g u r e 2 2 6 . This ra te w as a lso p ro p o rtio n a l to t h e a m o u n t of c o p p e r in t h e f r e s h t i s s u e ( F i g u r e s 215 a n d 2 1 6 ) . No r e l a t i o n s h i p c o u l d b e o b t a i n e d , h o w e v e r , b e t w e e n t h e r a t e of w a t e r l o s s a n d t h e a m o u n t o f i r o n i n t h e f r e s h t i s s u e of t h e p l a n t s o f t h e c o p p e r s e r i e s a s s e e n i n F i g u r e s 2 1 7 a n d 218. d. O ther C hem ical C onstituents T h e n itro g e n , p o ta s s iu m , m a g n e s iu m , c a lc iu m , and p h o s ­ p h o r u s c o n t e n t s of t h e p l a n t s h a v e b e e n c a l c u l a t e d on a b a s i s p e r t e n p l a n t s , p e r 100 g r a m s of f r e s h m a t e r i a l a n d p e r 100 g r a m s o f d ry plant m a te r ia l. No f u r t h e r d i s c u s s i o n w i l l b e u n d e r t a k e n about th e se co n stitu e n ts sin ce any re la tio n sh ip b etw een th em and iro n , c o p p e r and a s c o r b ic acid have a l r e a d y b e e n d is c u s s e d . T he follow ing ta b le w ill m e n tio n the p l a c e s w h e r e the d a ta c a n b e fo u nd: A m ounts per Ten Plants C onstituents A m ounts p e r 100 g. F r e s h M aterial A m ounts p e r 100 g. D r y M aterial T ables F igures T ables F igures T ables F igures N itrogen 24 and 26 148 and 189 27 and 29 155 and 197 31 and 32 173 and 211 Phosphorus 24 an d 26 146 and 187 27 and 29 153 and 195 31 and 32 171 and 209 P otassium 24 an d 26 147 and 188 27 and 29 154 and 196 31 and 32 172 and 210 C alcium 23 an d 25 149 and 190 27 and 29 156 and 198 31 and 32 174 and 212 M agnesium 23 and 25 150 and 191 27 and 29 157 and 199 31 and 32 175 and 213 D. D i s c u s s i o n on t h e T h i r d E x p e r i m e n t T h e m a i n o b j e c t of t h i s t h i r d e x p e r i m e n t w a s to s t u d y t h e e f f e c t s of i n c r e a s i n g a m o u n t s of c o p p e r a n d i r o n up to t o x i c l e v e l s , 83 on t h e s y n t h e s i s a n d s t a b i l i t y o f a s c o r b i c a c i d i n w h e a t p l a n t s . T h is e x p e r i m e n t w a s only p a r t l y s u c c e s s f u l b e c a u s e if it w a s p o s ­ s i b l e to r e a c h t h e l e v e l of c o p p e r t o x i c i t y , t h e p l a n t s of t h e i r o n s e r i e s , on t h e o t h e r h a n d , d i d n o t s h o w a n y t o x i c s y m p t o m s w h e n g r o w n o n t h e h i g h e s t a m o u n t s of i r o n s u p p l i e d . A s f a r a s t h e e f f e c t s of c o p p e r a n d i r o n on t h e a s c o r b i c a c i d s t a b i l i t y of d r y i n g p l a n t s a r e c o n c e r n e d , t h e r e s u l t s o b t a i n e d c o n ­ f ir m e d th o se r e p o r te d d u rin g the f i r s t and se c o n d e x p e r im e n ts . I n d e e d , t o h i g h c o n c e n t r a t i o n s o r i r o n in t h e s o l u t i o n c o r r e s p o n d e d a g r e a t e r s t a b i l i t y of i t s a s c o r b i c a c i d a n d in t h e c o p p e r s e r i e s t o h i g h c o p p e r c o n c e n t r a t i o n s c o r r e s p o n d e d a l o w r a t e of l o s s of a s c o rb ic acid. The fact that this o b s e rv a tio n w as noted w ith p lan ts h a r v e s t e d a t d i f f e r e n t t i m e s of t h e y e a r a n d d r i e d u n d e r d i f f e r e n t c o n d i t i o n s g i v e to i t a g r e a t s i g n i f i c a n c e . If t h e c o n c e n t r a t i o n s of i r o n in t h e t i s s u e s of t h e p l a n t s of t h e i r o n s e r i e s w e r e p o s i t i v e l y r e l a t e d t o t h e s t a b i l i t y of t h e a s c o r b ic acid d u rin g this th ird e x p e rim e n t, it should h o w ev er be r e c a l l e d t h a t t h i s c o r r e l a t i o n w a s n o t a s a p p a r e n t in t h e t w o f i r s t experim ents. I t s h o u l d a l s o b e e m p h a s i z e d t h a t i n t h e p l a n t s of th e s e c o n d e x p e r i m e n t in th e c o p p e r s e r i e s , a n e g a t i v e c o r r e l a t i o n w a s o b t a i n e d b e t w e e n t h e i r o n c o n c e n t r a t i o n of t h e f r e s h t i s s u e a n d th e a s c o r b i c a c id s ta b ility ; in the t h i r d e x p e r i m e n t no c l e a r r e l a ­ tionship w as th en a p p a re n t. The facts ju st m entioned su g g est that 84 i r o n p l a y s a n i m p o r t a n t r o l e i n t h e s t a b i l i t y of a s c o r b i c a c i d in t h e p l a n t , b u t i t s m o d e of a c t i o n i s n o t k n o w n . B e c a u s e no c o n ­ s is te n t re la tio n a p p e a r s betw een the a c tu a l c o n c e n tra tio n of iro n in t h e t i s s u e a n d t h e a s c o r b i c a c i d t h e s p e c i f i c r o l e of i r o n i s r a ­ ther obscure. A n o th e r fa c t th a t should be s t r e s s e d is th at the c o n c e n t r a ­ t i o n of i r o n i n t h e f r e s h t i s s u e d u r i n g t h e t h r e e e x p e r i m e n t s w h e r e v a r i o u s l e v e ls of th is e l e m e n t w e r e u s e d , did not sh o w m u c h v a r i ­ a t i o n s in i t s c o n c e n t r a t i o n i n t h e t i s s u e s , a l t h o u g h t h e t o t a l a m o u n t of i r o n p e r p l a n t v a r i e d g r e a t l y . T h e a b s o r p t i o n of t h e m a j o r e l e m e n t s , c a l c i u m , m a g n e s i u m , p o t a s s i u m , p h o s p h o r u s , and n it r o g e n , w a s show n to b e a ffe c te d , a s s e e n f r o m t h e i r c o n c e n t r a t i o n i n the p l a n t t i s s u e , b y t h e v a r i o u s c o n c e n t r a t i o n s of i r o n a n d c o p p e r in t h e s o l u t i o n s . It w a s th o u g h t to c o r r e l a t e t h e a m o u n t s of t h e s e e l e m e n t s a n d t h e a m o u n t of a s c o rb ic acid at h a r v e s t. In the c o p p e r s e r i e s th e p l a n t s w ith h i g h a m o u n t of a s c o r b i c a c i d c o r r e s p o n d e d to t h o s e w i t h h i g h c o n ­ c e n t r a t i o n of p h o s p h o r u s , c a l c i u m , a n d m a g n e s i u m . On t h e o t h e r h a n d , t h e s a m e t r e n d s d i d n o t a p p e a r i n t h e i r o n s e r i e s w h e r e to a h i g h c o n c e n t r a t i o n of m a g n e s i u m , p o t a s s i u m a n d n i t r o g e n , a n d t o a l o w c o n c e n t r a t i o n of c a l c i u m , c o r r e s p o n d e d a h i g h c o n c e n t r a ­ t i o n of a s c o r b i c a c i d . R e la tio n s h ip b e tw e e n a s c o r b i c acid and m a c r o e l e m e n t s in the p la n ts w e r e w o r k e d out by Wynd and Noggle (1950), who r e p o r t e d t h a t in r y e , g r o w n on a f ie ld in C a n a d a , the c o n c e n t r a t i o n s of a s c o r b i c a c i d w a s f o u n d to b e p o s i t i v e l y r e l a t e d t o t h e p e r c e n t a g e of n i t r o g e n , c a l c i u m , m a g n e s i u m , a n d i r o n . M a n y a u t h o r s h a v e s t u d i e d t h e e f f e c t of m a c r o - e l e m e n t s on t h e a c c u m u l a t i o n of a s c o r b i c a c i d , i n p l a n t t i s s u e b y v a r y i n g t h e l e v e l s a t w h i c h t h e y w e r e s u p p l i e d to t h e p l a n t s . W atson and N o g g l e (19 4 7 ) n o t e d t h a t t h e m i n u s p o t a s s i u m a n d m i n u s m a g n e ­ s iu m t r e a t m e n t s gave l a r g e i n c r e a s e s in th e a s c o r b i c a c id c o n ­ c e n t r a t i o n of o a t l e a v e s . B e r n s t e i n , H a m n e r a n d P a r k s (1 945 ) f o u n d t h a t t u r n i p g r e e n s g r o w n in s a n d c u l t u r e s w i t h o u t p o t a s s i u m p r o d u c e d p l a n t s l o w in a s c o r b i c a c i d . S o m e r s and K elly (1951), i n f i l t r a t i n g s o l u t i o n s of v a r i o u s i n o r g a n i c s a l t s in i l l u m i n a t e d l e a f d i s c s of t u r n i p s a n d b r o c c o l i , f o u n d t h a t c a l c i u m r e d u c e s t h e a c c u m u l a t i o n of a s c o r b i c a c i d . W hen the r e s u l t s r e p o r t e d by t h e s e a u t h o r s a r e c o m p a r e d to th o s e o b ta in e d d u r in g the p r e s e n t i n v e s t i ­ g a t i o n , i t d o e s n o t s e e m t h a t a v a r i a t i o n in t h e s u p p l y of a n e l e ­ m e n t a f f e c t i n t h e s a m e w a y t h e a c c u m u l a t i o n of a s c o r b i c a c i d in th e p l a n t a s a s i m i l a r v a r i a t i o n s of t h e c o n c e n t r a t i o n o f t h e s a m e e l e m e n t in t h e p l a n t t i s s u e . A n i n t e r e s t i n g e f f e c t of t h e h i g h l e v e l s of c o p p e r w a s m a d e c l e a r b y t h e s i g n i f i c a n t i n c r e a s e d a c c u m u l a t i o n of t h e e l e m e n t s p h o s p h o r u s , c a l c i u m , a n d m a g n e s i u m i n t h e p l a n t t i s s u e g r o w n on s u b s t r a t e c o n t a i n i n g m o r e t h a n 1.2 p a r t s p e r m i l l i o n of c o p p e r . 86 O n e of t h e t o x i c e f f e c t s o f e x c e s s c o p p e r m i g h t b e d u e to t h e f o r m ­ a t i o n of s o m e c o p p e r p r o t e i n w h i c h t h r o u g h c h a n g e in t h e i r p r o p ­ e r t i e s w o u l d d i s t u r b s o m u c h t h e p e r m e a b i l i t y of t h e p r o t o p l a s m t h a t a b n o r m a l a c c u m u l a t i o n of s a l t m a y o c c u r a n d b e d e t r i m e n t a l to v i t a l f u n c t i o n s of t h e c e l l . It is known th a t c o p p e r b in d s r e a d i l y w ith p r o t e i n s a s show n by T h o m p s o n , K o c h e r and F r i t z s c h e (1948). It w a s o b s e r v e d t h a t d u r i n g t h e d r y i n g of t h e v a r i o u s s a m p l e s f r o m the c o p p e r s e r i e s , the w a t e r w a s l o s t a t a g r e a t e r r a t e in the s a m ­ p l e s f r o m th e h i g h e r c o p p e r l e v e l s . This sp ecific ev ap o ratio n ra te m i g h t b e e x p l a i n e d b y a c h a n g e in t h e p e r m e a b i l i t y of t h e p r o t o ­ p l a s m d u e to t h e p r e s e n c e of c o p p e r . E. 1, S u m m a r y of th e T h i r d E x p e r i m e n t In t h e s a m e m a n n e r a s f o r t h e f i r s t t w o e x p e r i m e n t s , w h e a t p l a n t s w e r e g r o w n o n g r a v e l on tw o s e r i e s of c u l t u r e s o l u ­ t i o n s v a r y i n g in t h e i r a m o u n t s of c o p p e r a n d i r o n . The iron s e ­ r i e s r a n g e d f r o m n o i r o n t o 27 p a r t s p e r m i l l i o n , e a c h c u l t u r e v a r y i n g f r o m t h e o n e b e l o w b y a n i n c r e m e n t of t h r e e p a r t s p e r m i l l i o n of i r o n . T h e c o p p e r s e r i e s r a n g e d f r o m n o c o p p e r to 2 . 7 p a r t s p e r m i l l i o n , e a c h c u l t u r e v a r y i n g b y a n i n c r e m e n t of 0.3 p a r t s p e r m i l l i o n of c o p p e r . T h i s e x p e r i m e n t w a s d e s i g n e d to o v e r l a p w i t h t h e p r e c e d i n g o n e s a n d to o b s e r v e if t h e s a m e t e n d e n c i e s 87 in t h e a c c u m u l a t i o n a n d s t a b i l i t y of a s c o r b i c a c i d i n t h e p l a n t t i s ­ s u e s would p e r s i s t a t l e v e l s w h e r e the c o p p e r and i r o n w ould be toxic.' 2. The p lan ts w e r e h a r v e s te d a t jointing sta g e . A scorbic a c i d w a s d e t e r m i n e d on t h e f r e s h t i s s u e s a n d a t v a r i o u s i n t e r ­ v a l s d u r i n g t h e d r y i n g of t h e p l a n t s a t r o o m t e m p e r a t u r e . Iron, nitrogen, p h o sp h o ru s, p o ta s s iu m , c a lc iu m , and m a g n e s iu m w e re d e t e r m i n e d on o v e n d r y s a m p l e s . F r e s h and d r y w eight, a s well a s t h e r a t e of w a t e r l o s s d u r i n g d r y i n g , w e r e r e c o r d e d . 3. T h e f r e s h w e i g h t s o f t h e p l a n t s of t h e i r o n s e r i e s w e r e f o u n d to i n c r e a s e s l i g h t l y w i t h i n c r e a s e in t h e i r o n l e v e l o f t h e s o l u t i o n a n d no t o x i c l e v e l w a s r e a c h e d . In t h e c o p p e r s e r i e s a n i n c r e a s e in w e i g h t w a s o b t a i n e d in t h e p l a n t s up to 0 . 6 p a r t s p e r m i l l i o n in t h e s o l u t i o n a n d a b o v e t h i s l e v e l a t o x i c e f f e c t w a s r e ­ v e a l e d b y t h e s t u n t i n g of th e p l a n t s . In b o t h i r o n a n d c o p p e r s e r i e s , t h e d r y w e i g h t p a r a l l e l e d t h e f r e s h w e i g h t s of t h e p l a n t s . 4. T o a n i n c r e a s e of i r o n in th e s o l u t i o n c o r r e s p o n d e d an i n c r e a s e in t h e t o t a l a m o u n t s of i r o n in t h e p l a n t s ; b u t on a u n i t w e i g h t of f r e s h a n d d r y t i s s u e , a d e c r e a s e in th e c o n c e n t r a t i o n of i r o n w a s o b s e r v e d a b o v e t h e l e v e l of 18 p a r t s p e r m i l l i o n of i r o n in t h e s u b s t r a t e . In t h e c o p p e r s e r i e s , t h e t o t a l a m o u n t s of i r o n 88 a b s o r b e d p e r p la n t d e c r e a s e d w ith i n c r e a s e s in th e a m o u n t s of c o p p e r in t h e n u t r i e n t s o l u t i o n , h o w e v e r th e c o n c e n t r a t i o n s of i r o n in t h e f r e s h m a t t e r s h o w e d a m i n i m u m f o r t h e p l a n t s g r o w n on 1.5 p a r t s p e r m i l l i o n of c o p p e r , f o l l o w e d b y a n i n c r e a s e to 2 . 7 p a rts p er m illion. T h e i r o n c o n t e n t o f t h e d r y m a t t e r of t h e p l a n t s f r o m th e c o p p e r s e r i e s w a s i n v e r s e l y r e l a t e d to t h e a m o u n t s of c o p p e r in t h e n u t r i e n t s o l u t i o n . 5. In t h e p l a n t s of t h e c o p p e r s e r i e s t h e c o n c e n t r a t i o n s of c o p p e r in t h e d r y a n d f r e s h m a t t e r i n c r e a s e d c o r r e s p o n d i n g l y to t h e h i g h e r l e v e l s of c o p p e r in th e s o l u t i o n s . 6. T h e c o n c e n t r a t i o n s of a s c o r b i c a c i d in t h e f r e s h m a t e ­ r i a l a t h a r v e s t s h o w e d a n e g a t i v e r e l a t i o n to t h e a m o u n t of i r o n in the s o lu tio n . In t h e c o p p e r s e r i e s , t h e a m o u n t s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h m a t t e r a t h a r v e s t s h o w e d a d e c r e a s e up to 0. 9 p a r t s p e r m i l l i o n in t h e s o l u t i o n a n d t h e n i n c r e a s e d w i t h h i g h e r c o p p e r l e v e l s to h i g h e r c o n c e n t r a t i o n s of a s c o r b i c a c i d t h a n in t h e lo w c o p p e r l e v e l s . 7. A p o sitiv e c o r r e la tio n w as obtained betw een the a m o u n ts of i r o n i n t h e f r e s h t i s s u e a n d t h e s t a b i l i t y of t h e a s c o r b i c a c i d in t h e d r y i n g t i s s u e a f t e r 15 a n d 43 h o u r s of d r y i n g . In the c o p p e r s e r i e s a p o sitiv e c o r r e l a t i o n w as shown b e tw e e n the c o n c e n tra tio n s 89 of c o p p e r i n t h e t i s s u e a n d th e s t a b i l i t y of i t s a s c o r b i c a c i d a f t e r 15 h o u r s of d r y i n g . VI. EX PERIM EN T 4 A. Purpose In t h e t h r e e f i r s t e x p e r i m e n t s t h e t o p s of w h e a t p l a n t s g r o w n on w i d e l y d i f f e r e n t c o n c e n t r a t i o n s o f i r o n w e r e o b s e r v e d to a c c u m u ­ l a t e v e r y l i m i t e d a n d r a t h e r c o n s t a n t a m o u n t s of i r o n . In a s i m i l a r w a y , f r o m t h e d a t a of v a r i o u s w o r k e r s , i t c a n b e s e e n t h a t t h e a c ­ c u m u l a t i v e p o w e r of p l a n t s f o r i r o n i s r a t h e r l i m i t e d . For exam ­ p l e , B i d d u l p h ( 1 9 4 7 ) , s t u d y i n g th e i r o n a c c u m u l a t i o n i n l e a v e s of R e d K i d n e y b e a n s g r o w n on n u t r i e n t s o l u t i o n w i t h v a r i o u s f o r m s a n d l e v e l s of i r o n , f o u n d t h a t t h e c o n c e n t r a t i o n of t h i s e l e m e n t in th e s t e m , h e a r t - s h a p e d l e a v e s a n d t r i f o l i a t e l e a v e s r e m a i n e d c o m ­ p a r a t i v e l y u n i f o r m u n d e r th e v a r i o u s c o n d i t i o n s . S i d e r i s (1 94 9) f o u n d in p i n e a p p l e t h a t , a l t h o u g h i r o n a n d m a n g a n e s e i n c r e a s e in t h e t i s s u e s w i t h h i g h e r c o n c e n t r a t i o n s of t h e s e e l e m e n t s in t h e c u l t u r e s o l u t i o n s , t h e i r o n a c c u m u l a t e s i n t h e l e a v e s a n d s t e m to a m u c h l e s s e r e x te n t than m a n g a n e s e . L y o n a n d B e e s o n (1 94 8) o b s e r v e d t h e s a m e r e l a t i o n s h i p in t u r n i p s a n d t o m a t o e s . B e s i d e s t h e r e s t r i c t e d p o w e r of p l a n t s to a c c u m u l a t e i r o n i n s t e m s a n d l e a v e s , a n o t h e r r e a s o n f o r t h e r e l a t i v e l y lo w c o n c e n ­ t r a t i o n of t h i s e l e m e n t , e v e n i n t h e p l a n t t i s s u e s g r o w n o n s o l u t i o n s w h e r e h i g h a m o u n t s of i r o n h a v e b e e n a d d e d , m i g h t b e a n a c t u a l 91 u n a v a i l a b i l i t y of i r o n to th e r o o t s . I n d e e d , a g r e a t d i f f i c u l t y in s u p p l y i n g i r o n in w a t e r c u l t u r e m i g h t b e d u e , a s h a s b e e n p o i n t e d out b y B i d d u l p h ( 1 9 4 7 ), t o a r a p i d f o r m a t i o n of i n s o l u b l e c o m p l e x w i t h c e r t a i n s a l t s of t h e n u t r i e n t m e d i a . It is a ls o known, th a t at h ig h p H , i r o n e a s i l y p r e c i p i t a t e s a s t h e h y d r o x i d e a n d i f p h o s p h a t e ions a r e p r e s e n t , a s th e p h o s p h a te s a lt . In m o s t of t h e e x p e r i m e n t s p re v io u sly r e p o r te d , the iro n le v e ls r e p r e s e n t the to ta l iro n added when the s o lu tio n w a s m a d e up. U n l e s s s p e c i f i c a l l y e m p h a s i z e d by the a u t h o r s , i t i s r e a s o n a b l e to b e l i e v e t h a t i n t h e l i t e r a t u r e t h e l e v e l of t h i s e l e m e n t in s o l u b l e f o r m in t h e m e d i a i s a t a m u c h low er c o n c e n t r a t i o n th an e x p e c te d f r o m the in itia l a m o u n t a d d e d . In o r d e r to o b t a i n s u b s t a n t i a l a n d s i g n i f i c a n t i n f o r m a t i o n on th e r e l a t i o n s h i p of i r o n to a s c o r b i c a c i d in p l a n t s , i t s e e m e d o f t h e g r e a t e s t i n t e r e s t to o b t a i n p l a n t s w i t h v a r i o u s a n d w i d e l y d i f f e r e n t a m o u n t s of i r o n in t h e i r t i s s u e s . T he follow ing s te p s w e r e then p r o p o s e d to i n v e s t i g a t e t h e i r o n a c c u m u l a t i o n in w h e a t p l a n t s b y s u p p l y i n g s o l u t i o n s of c o n t r o l l e d i r o n l e v e l s : In o r d e r to p r e v e n t t h e p r e c i p i t a t i o n of t h e i r o n , a p H v a l u e of 4, l o w e r t h a n p r e v i o u s l y u s e d , w a s m a i n t a i n e d in t h e n u t r i e n t s o l u t i o n s a n d t h e i r o n s a l t w a s a d d e d to a c a r b o u y n o t c o n t a i n i n g t h e p h o s p h a t e i o n s . The c o m p l e t e n u t r i e n t s , w a s th e n s u p p lie d to the p l a n t s by c o n n e c tin g , a t i n t e r v a l s , t h e c u l t u r e - p o t s t o t h e i r o n c o n t a i n i n g - c a r b o u y a n d to the p h o s p h a te co n ta in in g c a r b o u y . A c c o r d i n g to B i d d u l p h ( 1 9 4 7 ) , f e r r i c c i t r a t e s e e m t o g i v e s a t i s f a c t i o n in s u p p l y i n g i r o n to p l a n t s a t p H 4 a n d t h i s f o r m of i r o n w a s u t i l i z e d . A t s h o r t i n t e r v a l s , the i r o n c o n c e n t r a t i o n s of t h e s o l u t i o n s w e r e c h e c k e d a n d m o r e i r o n added when n e c e s s a r y . At the end of the g r o w th p e r io d , the p la n ts w e r e h a r v e s t e d and v a r i o u s i n f o r m a t i o n o b ta in e d on t h e i r c h e m i c a l c o m p o s i t i o n s , a s done in the p r e v i o u s e x p e r i m e n t s . It s e e m e d u s e ­ ful to i n v e s t i g a t e a t t h e s a m e t i m e t h e r e l a t i v e a c c u m u l a t i o n of i r o n in t h e r o o t s , b e c a u s e a p o s s i b l e e x p l a n a t i o n o f t h e r a t h e r c o n s t a n t c o n c e n t r a t i o n s of i r o n in t h e t o p s m i g h t b e d u e to t h e p r e c i p i t a t i o n of t h i s e l e m e n t in t h e r o o t s a n d i t s s u b s e q u e n t u n a v a i l a b i l i t y t o t h e r e s t of t h e p l a n t . D a t a on t h e a s c o r b i c a c i d s t a t u s i n t h e r o o t s w e r e a l s o c o l l e c t e d b e c a u s e o f t h e r a t h e r l i m i t e d a m o u n t of i n f o r m a t i o n on t h i s c o m p o u n d in t h a t p a r t of t h e p l a n t s . B. E x p e rim e n ta l C onditions 1. The N u trie n t Solution T h e n u t r i e n t s o lu tio n f o r e a c h c u l t u r e w a s c o n ta in e d in two c a r b o y s of a v o l u m e of 1 8 l i t e r s . In o n e c a r b o y w a s a d d e d t h e p o t a s s i u m d i h y d r o g e n p h o s p h a t e and the c a l c i u m n i t r a t e a t the s a m e c o n c e n t r a t i o n s a s fo r the p r e v ­ io u s e x p e r i m e n t s and in the s e c o n d c a r b o y w a s p la c e d the m a g n e ­ s iu m s u lf a te and the m i n o r e l e m e n t s in c lu d in g 0.02 p a r t s p e r m i l ­ li o n of c o p p e r a n d v a r i o u s l e v e l s of i r o n a s d e s c r i b e d l a t e r . 93 T h e c a r b o y c o n ta in in g the p o t a s s i u m d ih y d ro g e n p h o sp h a te a n d t h e c a l c i u m n i t r a t e w a s c o n n e c t e d to t h e p u m p i n g d e v i c e d u r i n g the d a y and the o t h e r c a r b o y d u r in g the n ight. T h e only c o n t r o ll e d v a ry in g f a c t o r w a s th e ir o n s u p p lie d a t the follow ing le v e ls ; 0 . 0 , 0 . 5 , 1.0, 5 . 0 a n d 10 p a r t s p e r m i l l i o n . T w o p a i r s of c u l t u r e s w e r e g r o w n on e a c h l e v e l of i r o n . T h e pH of t h e s o l u t i o n w a s a d j u s t e d a t 4 a n d w a s c h e c k e d e v e r y f o u r d a y s a n d b r o u g h t b a c k t o pH 4 b y a d d i t i o n o f 0 . 2 5 N su lfu ric acid. T h e s t o c k s o l u t i o n of i r o n w a s p r e p a r e d b y d i s s o l v i n g 8 . 6 g r a m s of f e r r i c c i t r a t e w i t h d i s t i l l e d w a t e r to a v o l u m e of 500 m illiliters. T h is s o lu tio n w as then f il t e r e d and a n a ly z e d for its iro n content a g a in s t a s ta n d a rd iro n solution. T h e i r o n in t h e s o l u t i o n s w a s d e t e r m i n e d a t i n t e r v a l s a n d a d d i t i o n a l a m o u n t s w e r e a d d e d to b r i n g t h e n u t r i e n t s o l u t i o n to th e desired iro n -lev el. 2. P lan t M aterial F i f t y s e e d s of w h e a t I l l i n o i s N o. 1 - 1 2 8 w e r e s e e d e d i n e a c h p o t e v e n l y s p a c e d a n d c o v e r e d w i t h o n e - h a l f i n c h of g r a v e l on M a r c h 12, 1950, a n d w e r e h a r v e s t e d 38 d a y s l a t e r a t t h e b e g i n n i n g of th e j o i n t i n g s t a g e o n A p r i l 19, 195 0. A t the t i m e of h a r v e s t t h r e e s a m ­ p l e s of t h r e e g r a m s of l e a f b l a d e s w e r e t a k e n a t r a n d o m i n t h e 94 four c u l t u r e p o ts of e a c h lev el and t h e i r a s c o r b i c a cid c o n te n t d e ­ term ined. T w en ty -fiv e p la n ts w e r e then h a r v e s te d f ro m each pot a n d t h e f r e s h w e i g h t of t h e p l a n t t o p s w e r e r e c o r d e d . The plants w e r e t h e n p l a c e d in an a i r f o r c e d o v e n a t 6 0 ° C to b e d r i e d . The r o o t s of t h e p l a n t s w e r e w a s h e d f r e e of g r a v e l a n d t h e w a t e r b l o t t e d f r o m the t i s s u e s . T w o s a m p l e s of t w e n t y g r a m s of r o o t s w e r e u s e d fo r the d e t e r m i n a t i o n of a s c o r b i c a c id . T h e r e m a i n i n g o f th e r o o t m a t e r i a l w a s p l a c e d in a n o v e n a t 6 0 ° C to b e d r i e d a t t h e s a m e tim e as the tops. The d rie d m a te r ia l w as kept for fu rth e r C. 1. a. an alysis. E xperim ental R esults O b s e r v a t i o n s on t h e P l a n t s D e s c r i p t i o n of t h e P l a n t s a t T i m e of H a r v e s t A ll the p la n ts at h a r v e s t t i m e sh o w e d the f i r s t jo in t and m an y had the second joint. T h e p l a n t s g r o w n on t h e “ no i r o n ” c u l t u r e s w e r e s m a l l e r t h a n t h e o t h e r p l a n t s of t h e s e r i e s b u t d i d n o t s h o w a n y c h l o r o s i s . N o t m u c h d i f f e r e n c e c o u l d b e v i s u a l l y n o t e d b e t w e e n e a c h of t h e o t h e r l e v e l s , a l t h o u g h t h e p l a n t s g r o w n on t h e n u t r i e n t s o l u t i o n w i t h o n e p a r t p e r m i l l i o n of i r o n l o o k e d s o m e w h a t b i g g e r a n d h e a l t h i e r than all the o t h e r s . 95 b. F r e s h W eight p e r P la n t T h e d a t a i n d i c a t i n g t h e a v e r a g e of t h e f r e s h w e i g h t of t h e t o p s of t e n p l a n t s e x p r e s s e d in g r a m s a r e a s s e m b l e d in T a b l e 37 an d g r a p h e d a g a i n s t t h e a m o u n t of i r o n in t h e n u t r i e n t s o l u t i o n in F i g u r e 236. An i n c r e a s e i n t h e f r e s h w e i g h t of t h e p l a n t s w a s f o u n d f r o m t h e p l a n t s g r o w n on t h e n o - i r o n c u l t u r e s to a m a x i m u m f o r t h o s e g r o w n on t h e o n e p a r t p e r m i l l i o n i r o n l e v e l ; t h e w e i g h t s of t h e p l a n t s of th e n e x t c u l t u r e s w e r e s o m e w h a t l o w e r a l t h o u g h n o t o x i c sym ptom was apparent. c. D ry W eight p e r P la n t T h e a v e r a g e d r y w e i g h t p e r t e n p l a n t s a r e a r r a n g e d in T a b l e 37 a n d g r a p h e d a g a i n s t t h e a m o u n t of i r o n in t h e s o l u t i o n in F i g u r e 2 3 7 . T h e d r y w e i g h t s of t h e p l a n t s p a r a l l e l e d t h o s e of t h e f r e s h w e i g h t r e a c h i n g a m a x i m u m f o r t h e p l a n t s g r o w n on o n e p a r t p e r m i l l i o n of i r o n . 96 2. a. C h e m i c a l A n a l y s e s of t h e P l a n t s Iron (1) A m o u n t s of I r o n p e r P l a n t T h e a m o u n t s o f i r o n e x p r e s s e d in m i l l i g r a m s a c c u m u l a t e d in t h e a e r i a l p a r t of t h e p l a n t s a r e p r e s e n t e d i n T a b l e 37 a n d i n F i g u r e 233. No c o r r e l a t i o n c o u l d b e d r a w n b e t w e e n t h e a m o u n t of i r o n f o u n d p e r p l a n t t o p a n d t h e l e v e l of i r o n i n t h e s o l u t i o n s . T h e l e a s t a m o u n t of i r o n w a s f o u n d i n t h e p l a n t s g r o w n on t h e n u t r i e n t s o l u t i o n w h e r e no i r o n h a s b e e n a d d e d , h o w e v e r , t h e t o t a l a m o u n t of i r o n f o u n d i n t h e s e p l a n t s w a s a l m o s t a s h i g h a s in t h e p l a n t s g r o w n on t h e t e n p a r t s p e r m i l l i o n i r o n l e v e l . This f a c t m i g h t b e e x p l a i n e d b y t h e lo w pH of t h e s o l u t i o n w h i c h p e r ­ m i t t e d t h e i r o n i m p u r i t i e s of t h e g r a v e l to b e d i s s o l v e d a n d r e n ­ d e r e d m o r e i r o n a v a i l a b l e to th e p l a n t s . T h is would a l s o e x p la in w h y no c h l o r o s i s w a s o b s e r v e d in t h e n o - i r o n c u l t u r e s . (2) A m o u n t s of I r o n p e r 100 G r a m s of F r e s h T i s s u e T h e m i l l i g r a m s of i r o n p e r 100 g r a m s of f r e s h p l a n t t i s ­ s u e s a p p e a r i n T a b l e 38 a n d in F i g u r e 2 3 8 . I 97 T h e c o n c e n t r a t i o n s of i r o n in t h e f r e s h m a t t e r i n c r e a s e d v e r y s l i g h t l y f r o m t h e c u l t u r e s w i t h no i r o n t o t h o s e w i t h f i v e p a r t s p e r m i l l i o n of i r o n b u t t h e d i f f e r e n c e s c a n n o t b e c o n s i d e r e d as significant. (3) A m o u n t of I r o n p e r 100 G r a m s of D r y M a t e r i a l (a) In t h e w h e a t t o p s . T h e d a t a f o r t h e a m o u n t of i r o n p e r 100 g r a m s of d r y p l a n t m a t e r i a l a r e a s s e m b l e d in T a b l e 39 a n d in F i g u r e 243. No s p e c i a l t r e n d in th e c o n c e n t r a t i o n of i r o n in t h e d r y m a ­ t e r i a l c a n b e d e f i n e d f r o m t h e o b s e r v a t i o n of t h e d a t a m e n t i o n e d above. (b) In t h e w h e a t r o o t s . T h e a m o u n t s o f i r o n in m i l l i g r a m s p e r 100 g r a m s o f d r y r o o t m a t e r i a l a r e p r e s e n t e d in T a b l e 39 a n d F i g u r e 246. A v e r y s i g n i f i c a n t i n c r e a s e w a s f o u n d in t h e c o n c e n t r a t i o n of i r o n in t h e r o o t s of t h e p l a n t s g r o w n f r o m t h e no i r o n l e v e l to t h o s e g r o w n on t h e t e n p a r t s p e r m i l l i o n of i r o n l e v e l . Indeed a 699 p e r c e n t i n c r e a s e w a s o b t a i n e d b e t w e e n t h e c o n c e n t r a t i o n s of i r o n f o u n d i n t h e r o o t s g r o w n on t h o s e t w o c o n d i t i o n s . The values o b ta in e d f o r the d i f f e r e n t i r o n c o n c e n t r a t i o n s in th e d r y r o o t t i s s u e s a r e p o s i t i v e l y r e l a t e d t o t h e i r o n l e v e l s of t h e n u t r i e n t s o l u t i o n s . 98 b. M anganese (1) A m o u n t s of M a n g a n e s e p e r P l a n t The d a ta fo r the a m o u n ts of m a n g a n e s e p e r ten p la n ts a r e a s s e m b l e d in T a b l e 37 a n d in F i g u r e 2 3 4 . T h e a m o u n t s of m a n g a n e s e f o u n d in t h e p l a n t t o p s g r o w n on t h e d i f f e r e n t l e v e l s of i r o n w a s a b o u t t h e s a m e f o r a l l th e c u l ­ t u r e s a n d n o a p p a r e n t d e p r e s s i n g e f f e c t of t h e h i g h e r l e v e l s of iron was o b served. (2) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of F r e s h M a t e r i a l T h e a m o u n t s of m a n g a n e s e in m i l l i g r a m s p e r 100 g r a m s of f r e s h t i s s u e a r e r e p o r t e d i n T a b l e 38 a n d F i g u r e 239 . W h e n t h e c o n c e n t r a t i o n s of m a n g a n e s e in t h e f r e s h t i s s u e a r e c o n s i d e r e d an a p p a r e n t le s s e n i n g is o b s e r v e d f r o m the p la n ts g r o w n on n o i r o n to t h o s e g r o w n on t e n p a r t s p e r m i l l i o n of i r o n . (3) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of D r y M a t t e r (a) In t h e w h e a t t o p s . T h e d a t a f o r t h e a m o u n t s of m a n ­ g a n e s e i n m i l l i g r a m s p e r 100 g r a m s of d r y m a t e r i a l a r e a s s e m b l e d in T a b l e 39 a n d F i g u r e 2 4 4 . In t h e s a m e m a n n e r a s f o r t h e c o n c e n t r a t i o n o f m a n g a n e s e in t h e f r e s h t i s s u e , a s i g n i f i c a n t d e c r e a s e of i t s c o n c e n t r a t i o n in th e d r y m a t e r i a l w a s o b s e r v e d in t h e p l a n t s t o p s i n r e l a t i o n t o a n i n c r e a s e i n t h e i r o n l e v e l in t h e n u t r i e n t s o l u t i o n . (b) In the w h e a t r o o t s . T h e m i l l i g r a m s of m a n g a n e s e p e r 100 g r a m s of d r y r o o t m a t e r i a l a p p e a r in T a b l e 39 a n d i n F i g u r e 247. T h e o p p o s i t e t r e n d to t h e o n e f o u n d i n th e p l a n t t o p s w a s o b s e r v e d i n t h e w h e a t r o o t s a s f a r a s t h e c o n c e n t r a t i o n s of m a n ­ g a n e s e in r e l a t i o n to t h e a m o u n t of i r o n in t h e n u t r i e n t s o l u t i o n are concerned. In f a c t t h e c o n c e n t r a t i o n s of m a n g a n e s e in t h e r o o t s g r o w n on t e n p a r t s p e r m i l l i o n of i r o n w a s a l i t t l e m o r e t h a n t w i c e i t s c o n c e n t r a t i o n i n t h e r o o t s g r o w n on no i r o n . c. A s c o rb ic A cid (1) A m o u n t s of A s c o r b i c A c i d p e r P l a n t T h e d a t a f o r t h e a v e r a g e a m o u n t s of a s c o r b i c a c i d e x p r e s s e d in m i l l i g r a m s p e r t e n p l a n t s a r e p r e s e n t e d in T a b l e 37 a n d F i g u r e 235. T h e h i g h e s t a m o u n t of a s c o r b i c a c i d w a s f o u n d b o t h in th e p l a n t s g r o w n on t h e n u t r i e n t s o l u t i o n d e p r i v e d of i r o n a n d o n t h e s o l u t i o n w i t h f i v e p a r t s p e r m i l l i o n of i r o n . T he v a lu e s fo r the c u l t u r e s g r o w n on 0 . 5 , o n e a n d t e n p a r t s p e r m i l l i o n of i r o n w e r e s o m e w h a t lo w e r and did n ot show s ig n i f i c a n t d i f f e r e n c e s b e tw e e n 100 each, o t h e r . T h e v a r i a t i o n s in t h e t o t a l a m o u n t s of a s c o r b i c a c i d c o u l d n o t b e r e l a t e d t o a n y of t h e o t h e r c o m p o n e n t s a n a l y z e d . (2) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t e r i a l (a) I n t h e w h e a t t o p . T h e c o n c e n t r a t i o n s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h m a t e r i a l f r o m t h e a e r i a l p a r t of w h e a t a r e p r e s e n t e d i n T a b l e 38 a n d F i g u r e 240. T h e h i g h e s t c o n c e n t r a t i o n of a s c o r b i c a c i d w a s f o u n d i n t h e plants f r o m the “ no i r o n ” c u ltu re s . T he d if f e r e n c e s b e tw e e n the c o n c e n t r a t i o n s of a s c o r b i c a c i d i n t h e f r e s h m a t e r i a l of t h e p l a n t s t o p s of t h e o t h e r c u l t u r e s a r e s i g n i f i c a n t b u t c o u l d n o t b e c o r r e l a t e d w ith the co m p o n en ts analy zed . (b) I n t h e w h e a t r o o t . T h e m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h r o o t m a t e r i a l s a r e g i v e n i n T a b l e 38 a n d F i g ­ u r e 242. I t w a s o b s e r v e d t h a t t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d i n t h e w h e a t r o o t s w e r e v e r y low; i n f a c t , t h e a v e r a g e c o n c e n t r a t i o n of a s c o r b i c a c i d i n t h e w h e a t t o p s of a l l t h e c u l t u r e s w a s 16 t i m e s hig h er than the a v e r a g e c o n c e n tra tio n in the w heat r o o ts . The dif­ f e r e n c e s b e t w e e n t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d i n t h e r o o t s g r o w n a t t h e v a r i o u s l e v e l s of i r o n w e r e o n l y of f e w m i l l i g r a m s and cannot be co n s id e re d significant. 101 (3) A m o u n t of A s c o r b i c A c i d p e r 100 G r a m s of D r y M a t e r i a l T h e d a t a i n d i c a t i n g t h e m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of d r y m a t t e r a r e s h o w n in T a b l e 39 an d F i g u r e 245. T h e c o n c e n t r a t i o n s of a s c o r b i c a c i d i n t h e d r y m a t t e r of t h e w h e a t t o p s s h o w s t h e s a m e t r e n d s a s t h o s e found in the f r e s h t i s s u e and the s a m e o b s e rv a tio n s c a n be d ra w n . D. D i s c u s s i o n on t h e F o u r t h E x p e r i m e n t A l t h o u g h t h e c o n c e n t r a t i o n s of i r o n w e r e k e p t a t t h e d e s i r e d l e v e l s d u r i n g t h e t i m e of t h e e x p e r i m e n t , no s i g n i f i c a n t g r a d i e n t w a s f o u n d i n t h e c o n c e n t r a t i o n s of t h i s e l e m e n t i n t h e t i s s u e f r o m t h e a e r i a l p a r t s of t h e p l a n t s . The r e s t r i c t e d a c c u m u la tiv e po w er of w h e a t p l a n t s f o r i r o n i s h e n c e c o n f i r m e d b y t h i s e x p e r i m e n t . On th e o t h e r h a n d , t h e p l a n t r o o t s b e h a v e q u i t e d i f f e r e n t l y a n d a p o s i t i v e r e l a t i o n s h i p w a s o b s e r v e d b e t w e e n th e a c c u m u l a t i o n of i r o n in the r o o ts and its c o n c e n tr a tio n in the n u tr ie n t m e d ia . T h e f a c t t h a t t h e a m o u n t s of i r o n w e r e f ound t o v a r y w i d e l y in t h e r o o t s b u t d i d n o t c o r r e l a t e w i t h t h e a m o u n t s i n t h e t o p s w o u l d t e n d to s h o w t h a t a f t e r i t s a b s o r p t i o n , i r o n i s p r e c i p i t a t e d i n t h e r o o t s a n d s o r e n d e r e d u n a v a i l a b l e t o t h e r e s t of t h e p l a n t . T h e p o s s i b l e a d s o r p t i o n of c o l l o i d a l f e r r i c h y d r o x i d e on t h e e x t e r n a l p a r t of t h e r o o t s s u r f a c e s m i g h t a l s o b e a n e x p l a n a t i o n 102 t o t h e g r e a t a m o u n t of t h i s e l e m e n t f ound i n t h e r o o t s g r o w n on t h e high iro n -le v e l m ed iu m . S i d e r i s ( 195 0), s t u d y i n g i r o n a b s o r p t i o n i n p i n e a p p l e , w i t h r a d io a c tiv e i r o n ^ , found high co unts for th e r o o t m a t e r i a l in c o m p a r i s o n t o th e l e a v e s a n d s u g g e s t e d t h a t m o s t of t h e i r o n 59 in the r o o t s w a s n o t a b s o r b e d i n t o th e t i s s u e s b u t p r e s u m a b l y p r e c i p i ­ t a t e d in t h e e x o d e r m a l l a y e r of c e l l s . A g r e a t a c c u m u l a t i o n of i r o n i n t h e r o o t s of R e d K i d n e y b e a n s w a s a l s o n o t e d b y B i d d u l p h (1947) i n c o m p a r i s o n to t h e c o n c e n t r a t i o n of t h i s e l e m e n t i n t h e s t e m a n d leaves. I t i s i n t e r e s t i n g t o n o te t h a t , a l t h o u g h t h e p l a n t s of t h e n o - i r o n c u l t u r e s w e r e s m a l l e r in s i z e t h a n t h o s e of t h e o t h e r c u l ­ t u r e s , t h e y did n o t s h o w a n y c h l o r o s i s . T h is fa c t would s u g g e s t th a t at pH 4 the s o - c a l l e d “ active i r o n " n e e d e d in v e r y s m a l l am o u n ts f o r ch lo ro p h y ll f o rm a tio n is av ailab le f r o m the i m p u r i t i e s of t h e s o l u t i o n b u t i r o n i s y e t a l i m i t i n g f a c t o r f o r o t h e r g r o w t h p r o c e s s e s r e q u i r i n g a g r e a t e r a m o u n t of i t . In a s e r i e s of e x p e r i ­ m e n t s , S t r o m m e (1951), g r o w i n g w h e a t a t d i f f e r e n t p H s , o b s e r v e d a l s o t h a t a t pH 4 n o a p p a r e n t c h l o r o s i s d e v e l o p e d i n th e p l a n t s grow n w ithout iro n . The s a m e r e s u l t was obtained by Biddulph (1947) i n R e d K i d n e y b e a n s . T h i s a u t h o r s u g g e s t e d t h a t a t p H 4, ir o n “ r e m a i n s m o b ile and capable of m o v e m e n t to the new ly f o r m e d s t e m t i p s a n d l e a v e s an d t h a t a n e x t r e m e l y lo w i r o n s u p p l y w i l l 103 suffice p ro v id in g conditions a r e s u c h th a t the iro n w ill not b e c o m e i m m o b i l i z e d w i t h i n t h e t i s s u e s ” (p. 93). M a n g a n e se should be c o n s id e re d am ong the f a c t o r s w hich m i g h t b e r e s p o n s i b l e f o r a g r a d i e n t i n t h e c o n c e n t r a t i o n of i r o n i n the p la n ts. I n d e e d , t h e t r a n s l o c a t i o n of i r o n i n p i n e a p p l e h a s b e e n r e p o r t e d b y S i d e r i s (1950) to b e a f f e c t e d b y t h e p r e s e n c e of m a n ­ g a n e s e ; in the s a m e m a n n e r , w hen the d a ta r e p o r t e d by S o m e r s a n d S h i v e (1942) a r e c o n s u l t e d , i t c a n b e o b s e r v e d t h a t t o a h i g h e r i r o n - l e v e l in the n u trie n t solution c o r r e s p o n d e d a lo w e r a b s o r p ­ t i o n of m a n g a n e s e i n t h e s o y b e a n t h a n a t l o w e r i r o n - l e v e l s . I n t h e p r e s e n t e x p e r i m e n t , t h e l e v e l of m a n g a n e s e s u p p l i e d to a l l t h e c u l t u r e s w a s c o n s t a n t , b u t th e a c c u m u l a t i o n o f t h i s e l e ­ m e n t in the p la n t p a r t s w as found to v a r y w idely. In th e d r ie d m a t e r i a l f r o m the p lant tops, a negative re la tio n s h ip w a s obtained w i t h t h e l e v e l s of i r o n i n t h e s o l u t i o n s , b u t on t h e c o n t r a r y a p o s i ­ t i v e r e l a t i o n s h i p w a s f o u n d i n t h e roots.. The fact also, that leaves and s te m s showed a m u c h g r e a te r p o w e r of a c c u m u l a t i o n f o r m a n g a n e s e t h a n t h e r o o t s w a s m a d e c l e a r b y t h e d a t a of t h e p r e s e n t s t u d y . I r o n i n t e r f e r e n c e i n t h e t r a n s l o ­ c a t i o n of m a n g a n e s e c a n h a r d l y e x p l a i n t h e f a c t r e p o r t e d a b o v e , b e c a u s e , a l t h o u g h t h e r o o t s h a d h i g h a m o u n t s of i r o n i n c o m p a r i s o n to th e s t e m s no c o r r e l a t i o n could b e d r a w n b e tw e e n i r o n and m a n ­ g a n e s e c o n c e n t r a t i o n s i n b o t h r o o t s an d s t e m s . It should b e noted 104 t h a t n o i n f o r m a t i o n on t h e i n f l u e n c e of th e c o n c e n t r a t i o n o f i r o n in t h e r o o t s on t h e m a n g a n e s e c o n c e n t r a t i o n w a s f o u n d i n t h e l i t e r a ­ ture. T o o b t a i n m o r e i n f o r m a t i o n on t h e m a n g a n e s e s t a t u s i n t h e p l a n t s i s of i n t e r e s t b e c a u s e t h i s e l e m e n t h a s b e e n c l a i m e d to be involved in the a s c o r b i c a c id s y n th e s is . I n d e e d , R u d r a (1939) r e p o r t e d t h a t w h e n a s o l u t i o n of m a n n o s e c o n t a i n i n g s m a l l a m o u n t s of m a n g a n e s e w a s i n j e c t e d i n r a t l i v e r , i n c r e a s e i n a s c o r b i c a c i d c o n te n t could b e d e te c te d . H e s t e r (1941) c l a i m e d t h a t a n a d d i t i o n of s o l u b l e m a n g a n e s e to s o i l s d e f i c i e n t in t h i s e l e m e n t r e s u l t e d in an i n c r e a s e d a s c o r b i c a c id c o n c e n tra tio n in the to m a to e s g ro w n on t h i s s o i l ; b u t L y o n a n d B e e s o n (1948) c o u l d n o t o b t a i n a n y s i g ­ n i f i c a n t e f f e c t of m a n g a n e s e o n t h e a s c o r b i c a c i d c o n t e n t of t o m a ­ t o e s g r o w n on v a r i o u s s o l u t i o n . F r o m t h e p r e s e n t e x p e r i m e n t no a p p a r e n t r e l a t i o n b e t w e e n m a n g a n e s e c o n c e n t r a t i o n i n t h e t i s s u e and i t s a s c o r b i c a c i d c o n ­ t e n t c a n b e d r a w n ; i t s h o u ld , h o w e v e r , b e p o i n t e d o u t t h a t i n w h e a t top the h ig h e s t a s c o r b i c a c id c o n c e n tr a tio n w a s found in the p la n ts of t h e “ n o - i r o n ” l e v e l a n d w h i c h c o n t a i n e d t h e h i g h e s t c o n c e n t r a ­ t i o n of m a n g a n e s e i n t h e p l a n t m a t e r i a l . V e ry significant d iffe re n c e s w e r e found b e tw e e n the co n ­ c e n t r a t i o n s of a s c o r b i c a c i d i n t h e p l a n t t o p s a n d t h e o n e s i n t h e roots. T h i s r e s u l t c o n f i r m e d t h e d a t a r e p o r t e d b y R e i d (1937 ), who 105 f o u n d t h a t i n c o w p e a , 75 t o 85 p e r c e n t of t h e a s c o r b i c a c i d i s f o u n d in the le a f b la d e s . R e i d ' s r e s u l t s a r e g i v e n i n t h e f o ll o w i n g t a b l e : L o c a l i z a t i o n of A s c o r b i c A c i d in t h e C o w p e a P l a n t M gms per G ram P ercentage of T o t a l Q uantity Total Mgms Leaves B lades 0 . 818 2 1 .4 8 6 P etioles Leaf buds B lo s s o m buds Stem s R oots 0 .1 7 7 0 . 612 0 .560 0.141 0.12 5 1.7 16 0.164 0.084 1. 494 1. 788 80 ) ) ) ) ) 20 2 6.7 3 2 I t w a s s u g g e s t e d t h e n b y R e i d t h a t h i g h e r a c c u m u l a t i o n of a s c o r b i c a c id c o r r e s p o n d e d to t i s s u e s w ith h ig h e r c e l l u l a r a c t i v i ­ ty , h o w e v e r , i n p l a n t s i t i s r e a s o n a b l e to s a y t h a t r o o t t i s s u e e x e r t as m u c h activity as the a e r ia l p a rt. The b e s t explanation for the g r e a t d i f f e r e n c e i n c o n c e n t r a t i o n s of a s c o r b i c a c i d b e t w e e n l e a f and ro o t is th at a s c o rb ic acid is m o s tly sy n th esized in the g r e e n a e r i a l p a r t of t h e p l a n t . 106 E. S u m m a r y of t h e F o u r t h E x p e r i m e n t 1. W h e a t p l a n t s w e r e g r o w n o n n u t r i e n t s o l u t i o n s of c o n ­ t r o l l e d i r o n l e v e l ; t h e i r o n b e i n g s u p p l i e d i n th e c i t r a t e f o r m f r o m a s o l u t i o n f r e e of p h o s p h o r u s a n d a t a pH of 4 i n o r d e r to l e s s e n its precipitation. solutions w ere: 2. ing s ta g e . T h e c o n c e n t r a t i o n s of t h e i r o n i n t h e v a r i o u s n o n e , 0. 5, 1.5 a n d t e n p a r t s p e r m i l l i o n . T h e p l a n t s w e r e h a r v e s t e d a t th e b e g i n n i n g of t h e j o i n t ­ A s c o r b i c a c i d w a s d e t e r m i n e d on t h e f r e s h t i s s u e s . The d e t e r m i n a t i o n s of i r o n a n d m a n g a n e s e w e r e c a r r i e d on t h e o v e n dried sam p les. 3. F r e s h and d r y w eig h t w e r e r e c o r d e d . T h e f r e s h an d d r y w e i g h t of t h e p l a n t s s h o w e d a m a x i m u m f o r t h o s e g r o w n on one p a r t p e r m i l l i o n of i r o n . N either chlorosis n o r t o x i c s y m p t o m s w e r e a p p a r e n t on t h e p l a n t s f r o m t h e e x t r e m e cultures. 4. T h e h i g h e s t a m o u n t of i r o n a c c u m u l a t e d i n t h e a e r i a l p a r t of t h e p l a n t s w a s i n t h o s e f r o m t h e c u l t u r e s s u p p l i e d w i t h one p a r t p e r m i l l i o n of i r o n , c o r r e s p o n d i n g to t h e h i g h e s t f r e s h a n d d r y weight. On t h e o t h e r h a n d , no a p p a r e n t t r e n d i n t h e c o n c e n t r a t i o n of i r o n i n t h e t i s s u e of t h e t o p s c o u l d b e n o t e d . In t h e d r y r o o t m a t e r i a l , t h e c o n c e n t r a t i o n of i r o n w a s p o s i t i v e l y r e l a t e d t o t h e a m o u n t of i r o n i n t h e s o l u t i o n . i 107 5. No c l e a r r e l a t i o n s h i p c o u l d b e d r a w n b e t w e e n m a n ­ g a n e s e and ir o n o r m a n g a n e s e and a s c o r b i c acid . The m o s t s t r i k ­ in g r e s u l t o b t a i n e d f r o m t h e m a n g a n e s e d a t a w a s t h e c o m p a r a t i v e l y h i g h e r a c c u m u l a t i v e p o w e r of s t e m s a n d l e a v e s f o r t h i s e l e m e n t than the p lan t r o o ts . 6. No r e l a t i o n s h i p c o u l d b e d r a w n b e t w e e n a s c o r b i c a c i d a n d t h e c o n c e n t r a t i o n s of i r o n o r m a n g a n e s e i n t h e s o l u t i o n s o r in the p la n t t i s s u e s . On t h e o t h e r h a n d , a v e r y s i g n i f i c a n t d i f f e r ­ e n c e in t h e c o n c e n t r a t i o n of a s c o r b i c a c i d w a s f o u n d b e t w e e n t h e l e a v e s a n d t h e r o o t s , t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d in th e ro o ts bein g a l m o s t nil. VII. EX PERIM EN T 5 A. Purpose All the p re v io u s e x p e rim e n ts had b e e n conducted w ith w h eat p l a n t s a n d t h e i r r e s p o n s e s t o w a r d i r o n an d c o p p e r s t u d i e d i n r e l a ­ tion to a s c o r b ic acid. It w as thought n e c e s s a r y to study so m e o th e r p la n t m a t e r i a l to find out if the r e s u l t s a l r e a d y obtained could b e generalized. T h e f i r s t o b j e c t of t h e s t u d y w a s t o f i n d o u t h o w d i f f e r e n t l e v e l s of i r o n m i g h t a f f e c t t h e c o n c e n t r a t i o n of a s c o r b i c a c i d f o u n d a t h a r v e s t in s u ch o th e r p lan ts a s to m a to e s , f o r w hich the n u t r i ­ tional r e q u ir e m e n ts have a lr e a d y b e e n m u c h studied. B e c a u s e of t h e i n t e r r e l a t i o n s h i p b e t w e e n i r o n a n d m a n g a n e s e t h e s t a t u s of t h e l a t t e r i n t h e p l a n t w a s a l s o s t u d i e d . In o r d e r to k e e p th e i r o n le v e ls a s c o n s ta n t a s p o s s i b l e the n u t r i e n t s o l u t i o n w a s k e p t a t a pH v a l u e of 4 a n d t h e tw o c a r b o y s s y s t e m w a s u s e d a s i n t h e p r e c e d i n g e x p e r i m e n t , to p r e v e n t t h e i r o n to b e i n t h e s a m e s o l u t i o n a s t h e p h o s p h a t e s a l t . I t w a s a l s o p r o p o s e d to s t u d y t h e p o s s i b l e g r a d i e n t in t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d in t h e l e a v e s f r o m th e to p a n d t h e b a s e of t h e p l a n t s a n d r e l a t e a n y d i f f e r e n c e f o u n d w i t h t h e d a t a av aila b le . It w as thought th a t b y studying v a r ia tio n s in the d iffe re n t 109 p a r t s of t h e s a m e p l a n t s , o n e of t h e g r e a t a d v a n t a g e w a s t h a t t h e e f f e c t of u n c o n t r o l l a b l e e n v i r o n m e n t a l f a c t o r s w o u l d t h e n b e r e ­ d u c e d to t h e m i n i m u r r u B. E x p e r im e n ta l Conditions 1. T he N u t r ie n t Solution The s a m e n u tr ie n t so lu tio n a s in the l a s t e x p e r i m e n t w ith w h e a t w as u s e d to g ro w the to m a to p la n ts . T h e s a m e l e v e l s of i r o n w e r e c h o s e n and a s b e f o r e the c a r b o y containing the p o ta s s iu m a c id p h o sp h ate and the c a lc iu m n i tr a te w a s co n n ected to the p u m p ­ in g d e v i c e d u r i n g t h e d a y an d t h e o t h e r c a r b o y d u r i n g t h e n i g h t . T h e pH of t h e n u t r i e n t s o l u t i o n w a s a l s o a d j u s t e d a t a pH of 4. S i n c e a p o s s i b i l i t y of i r o n c o n t a m i n a t i o n h a d b e e n a t t r i b u t e d to th e q u a r t z g r a v e l , a h a n d m a g n e t w a s p a s s e d t h r o u g h t h e l a t t e r d u r i n g th e w a s h i n g ; i t w a s p o s s i b l e t o g e t r i d i n t h i s m a n n e r of s m a l l p a r t i c l e s of i r o n w h i c h m i g h t h a v e r e l e a s e d e n o u g h i r o n f o r g r o w t h a t a low pH. 2. T h e P l a n t M a t e r i a l The to m a to plan ts u sed fo r th is e x p e r im e n t had f i r s t b een sown in a fla t filled w ith soil. W h e n t h e y w e r e o n e m o n t h old, on M a y 5, 1950, f o u r p l a n t s w e r e t r a n s p l a n t e d p e r p o t s . A s i n t h e 110 p r e v i o u s e x p e r i m e n t t w o p a i r s of c u l t u r e s w e r e g r o w n on e a c h iro n level. B e f o r e tr a n s p la n ta tio n to the q u a r t z g r a v e l the r o o ts of t h e t o m a t o p l a n t s h a d b e e n w a s h e d i n t a p a n d d i s t i l l e d w a t e r . . A t t h e t i m e of h a r v e s t , 45 d a y s a f t e r t r a n s p l a n t i n g , on J u n e 19, 1950, t w o k i n d s of s a m p l e s w e r e t a k e n . F r o m e a c h c u l ­ t u r e a c o m p o s i t e s a m p l e of t h e f o u r l e a v e s f r o m t h e t i p of e a c h p lant w as tak en w hich l a t e r w ill be m en tio n ed a s “ t e r m i n a l l e a v e s " a n d f o u r l e a v e s f r o m t h e b a s e of e a c h p l a n t s of e a c h c u l t u r e w h i c h l a t e r w ill be r e f e r r e d as “ b a s a l l e a v e s ." C. E x perim ental R esults 1. O b s e r v a t i o n s on t h e P l a n t s a. D e s c r i p t i o n of t h e P l a n t s a t T i m e of H a r v e s t A t t i m e of h a r v e s t n o t m u c h d i f f e r e n c e c o u l d v i s u a l l y b e s e e n b e t w e e n t h e p l a n t s g r o w n on t h e v a r i o u s l e v e l s of i r o n . The p l a n t s g r o w n on t h e “ n o i r o n " l e v e l w e r e of s m a l l e r s i z e t h a n t h o s e g r o w n on t h e o t h e r i r o n l e v e l s b u t d i d n o t s h o w a n y c h l o r o s i s . T h e h i g h e s t p l a n t s w e r e t h o s e g r o w n on t h e f i v e p a r t s p e r m i l l i o n iro n level. T h e a v e r a g e h e i g h t of a l l t h e p l a n t s a t h a r v e s t w a s about seventy c e n tim e te rs. Ill b. P e r c e n t a g e s of D r y M a t t e r i n t h e L e a v e s T h e v a l u e s f o r t h e p e r c e n t a g e s of d r y m a t t e r in t h e b a s a l a n d t e r m i n a l l e a v e s of t h e t o m a t o p l a n t s a r e p r e s e n t e d in T a b l e 40 a n d F i g u r e 250. V a r i a t i o n s in t h e p e r c e n t a g e s of d r y m a t t e r of t h e l e a v e s w e r e o b s e r v e d f r o m o n e l e v e l of i r o n to t h e o t h e r b u t no d e f i n i t e tre n d can be noted. C o n s i s t e n t l y a n d s i g n i f i c a n t l y , a h i g h e r p e r c e n t a g e of d r y m a t t e r w a s f o und in t h e t e r m i n a l l e a v e s t h a n i n t h e b a s a l l e a v e s . 2. a. C h e m i c a l A n a l y s e s of t h e P l a n t Iron (1) A m o u n t s of I r o n p e r 100 G r a m s of F r e s h P l a n t M a t e r i a l T h e a m o u n t s of i r o n p e r 100 g r a m s of f r e s h p l a n t m a t e r i a l in t h e b a s a l a n d t e r m i n a l l e a v e s a p p e a r in T a b l e 40 a n d in F i g u r e 248 . In t h e b a s a l l e a v e s a n e g a t i v e r e l a t i o n s h i p w a s o b s e r v e d b e t w e e n t h e c o n c e n t r a t i o n of i r o n i n t h e f r e s h t i s s u e s a n d i t s c o n ­ c e n t r a t i o n in t h e n u t r i e n t s o l u t i o n . In t h e t e r m i n a l l e a v e s , n o s i g n i f i c a n t t r e n d i n t h e i r o n c o n ­ c e n tr a tio n w as noted. In a l l t h e c u l t u r e s t h e i r o n c o n c e n t r a t i o n s in t h e f r e s h t i s ­ s u e of t h e b a s a l l e a v e s w e r e s i g n i f i c a n t l y h i g h e r t h a n in t h e t e r ­ m inal leaves. (2) A m o u n t s of I r o n p e r 100 G r a m s of D r y M a t t e r T h e c o n c e n t r a t i o n s of i r o n i n m i l l i g r a m s p e r 100 g r a m s of d r y m a t t e r in t h e b a s a l a n d t e r m i n a l l e a v e s a r e p r e s e n t e d in T a b l e 41 a n d in F i g u r e 252. T he n eg ativ e r e la tio n s h ip found b e tw e e n the i r o n c o n c e n t r a ­ t i o n s in t h e f r e s h t i s s u e of t h e b a s a l l e a v e s a n d t h e c o n c e n t r a t i o n s of i r o n in t h e n u t r i e n t s o l u t i o n h o l d t r u e b u t t o a l e s s e r e x t e n t f o r t h e c o n c e n t r a t i o n s of i r o n in t h e d r y m a t t e r . No g r e a t v a r i a t i o n s w e r e r e p o r t e d i n t h e a m o u n t of i r o n f o u n d in t h e d r y m a t e r i a l of t h e t e r m i n a l l e a v e s . In a l l t h e c u l t u r e s , t h e i r o n c o n c e n t r a t i o n s in t h e d r y m a ­ t e r i a l of t h e b a s a l l e a v e s w e r e s i g n i f i c a n t l y h i g h e r t h a n t h o s e of the te r m in a l leav es. b. M anganese (1) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of F r e s h M a t e r i a l T h e a m o u n t s of m a n g a n e s e i n m i l l i g r a m s p e r 100 g r a m s of f r e s h t i s s u e of t h e b a s a l a n d t e r m i n a l l e a v e s a r e r e p o r t e d in T a b l e 113 V a r i a t i o n s in t h e c o n c e n t r a t i o n s o f t h e m a n g a n e s e w a s o b ­ s e r v e d i n t h e f r e s h t i s s u e of b o t h b a s a l a n d t e r m i n a l l e a v e s b u t no t r e n d c o u l d b e r e l a t e d to t h e v a r i o u s l e v e l s of t h e i r o n in t h e n u ­ tr ie n t solutions. In a l l t h e c a s e s , e x c e p t f o r t h e p l a n t s g r o w n on t e n p a r t s p e r m i l l i o n of i r o n , t h e c o n c e n t r a t i o n of m a n g a n e s e i n t h e f r e s h t i s s u e w a s h i g h e r i n t h e b a s a l t h a n in t h e t e r m i n a l l e a v e s . (2) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of D r y M a t e r i a l T h e c o n c e n t r a t i o n s o f m a n g a n e s e in m i l l i g r a m s p e r 100 g r a m s of d r y m a t t e r i n t h e b a s a l a n d t e r m i n a l l e a v e s a r e a s s e m b l e d in T a b l e 41 a n d F i g u r e 25 3. N o a p p a r e n t r e l a t i o n s h i p b e t w e e n t h e a m o u n t s of i r o n i n t h e n u t r i e n t s o l u t i o n s a n d t h e c o n c e n t r a t i o n of m a n g a n e s e i n t h e d r y t i s s u e of t h e b a s a l l e a v e s w a s o b s e r v e d . H o w e v e r , i n t h e t e r m i n a l l e a v e s , i f t h e o n e s g r o w n on “ n o i r o n ” a r e let a s id e , a p o s itiv e r e la tio n s h ip is o b s e r v e d b etw een t h e m a n g a n e s e c o n c e n t r a t i o n s i n t h e d r y t i s s u e a n d t h e l e v e l s of i r o n in th e n u t r i e n t s o lu tio n . In a l l th e c a s e s , m o r e m a n g a n e s e w as found in the d r i e d b a s a l l e a v e s than in the d r i e d t e r m i n a l l e a v e s . 114 c. A s c o r b ic A cid (1) A m o u n t s of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t e r i a l T h e m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h m a t e r i a l i n t h e b a s a l a n d t e r m i n a l l e a v e s of t h e t o m a t o p l a n t s a r e p r e s e n t e d i n T a b l e 40 a n d F i g u r e 251. In t h e b a s a l l e a v e s , t h e a m o u n t s of a s c o r b i c a c i d d i d n o t s h o w a n y r e l a t i o n s h i p w i t h t h e c o n c e n t r a t i o n s of i r o n i n t h e s o l u ­ t i o n on w h i c h t h e p l a n t s g r e w . On the o t h e r han d , in the t e r m i n a l leav es a p o sitiv e c o r r e la tio n w as o b s e rv e d with the c o n c e n tra tio n of i r o n i n t h e s o l u t i o n . If t h e d a t a f o r t h e a m o u n t s of a s c o r b i c a c i d i n t h e f r e s h m a t t e r of t h e b a s a l a n d t e r m i n a l l e a v e s a r e c o n s i d e r e d s e p a r a t e l y no c o r r e l a t i o n i s a p p a r e n t w i t h t h e d r y m a t t e r , i r o n o r m a n g a n e s e p e r u n i t of f r e s h w e i g h t . H o w ev er, when the r e s u l t s a r e pooled a n e g a t i v e r e l a t i o n s h i p a p p e a r s b e tw e e n the i r o n in th e f r e s h t i s s u e a n d t h e a s c o r b i c a c i d ( F i g u r e 257) w h i l e a p o s i t i v e c o r r e l a t i o n i s s h o w n b e t w e e n a s c o r b i c a c i d a n d t h e p e r c e n t a g e of d r y m a t t e r ( F i g u r e 259). No c o r r e l a ti o n w as shown b etw een m a n g a n e s e and a s c o r b i c a c i d in t h e f r e s h t i s s u e ( F i g u r e 258). 115 (2) A m o u n t s of A s c o r b i c A c i d p e r 100 G r a m s of D r y M a t e r i a l T h e m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of d r y m a t e r i a l in th e b a s a l a n d t e r m i n a l l e a v e s a r e g i v e n in T a b l e 41 a n d F i g u r e 254. W h e n t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d a r e c a l c u l a t e d on a d r y m a t t e r b a s i s , no r e l a t i o n c a n b e f o u n d in th e b a s a l l e a v e s w ith the o th e r d a ta , b u t a p o s itiv e c o r r e l a t i o n is a p p a r e n t b e tw e e n t h e c o n c e n t r a t i o n of a s c o r b i c a c i d in t h e d r y m a t e r i a l of t h e t e r ­ m i n a l l e a v e s a n d t h e l e v e l of i r o n on w h i c h t h e p l a n t s g r e w . H o w e v e r , a s c a n b e s e e n i n F i g u r e s 255 a n d 256, no r e l a ­ t i o n s h i p c a n b e d r a w n b e t w e e n t h e c o n c e n t r a t i o n s of i r o n o r m a n ­ g a n e s e a n d t h e a m o u n t s of a s c o r b i c a c i d in 100 g r a m s of d r y m ateria l. D. D i s c u s s i o n on t h e F i f t h E x p e r i m e n t A s i g n i f i c a n t i n c r e a s e in t h e a m o u n t s of a s c o r b i c a c i d in t h e t e r m i n a l l e a v e s of t h e t o m a t o p l a n t s w a s r e p o r t e d to c o r r e s p o n d to an i n c r e a s e in t h e l e v e l of i r o n in th e s o l u t i o n . H o w e v e r, this r e s u l t m i g h t b e m i s l e a d i n g a s to t h e e f f e c t of i r o n on t h e a s c o r b i c a c i d s t a t u s in t h e p l a n t t i s s u e : Indeed, a negative relatio n sh ip was f o u n d b e t w e e n t h e l e v e l s of i r o n in th e n u t r i e n t s o l u t i o n a n d t h e c o n c e n t r a t i o n s of i r o n in t h e f r e s h t i s s u e . In c o n s e q u e n c e , in th e p r e s e n t e x p e r i m e n t t h e p o s s i b l e e f f e c t o f t h e i r o n on t h e a s c o r b i c a c i d c o n t e n t of t h e p l a n t a t h a r v e s t w a s a d e p r e s s i n g o n e . This r e l a t io n s h i p is not a p p a r e n t, w hen the d a ta fo r the b a s a l le a v e s a r e c o n s id e r e d , although w hen a ll the d a ta a r e pooled, an o v e r - a l l n eg ativ e re la tio n s h ip can be se e n b etw een iro n c o n c e n tra tio n and a s c o r b i c a c i d in t h e f r e s h m a t t e r . A p o sitiv e c o r r e la tio n betw een the a s c o r b ic acid c o n c e n t r a ­ t i o n in t h e f r e s h l e a v e s a n d t h e d r y m a t t e r w a s o b t a i n e d w h e n a l l the d ata fo r b a s a l and t e r m i n a l le a v e s w e r e pooled. A sim ilar p o s i t i v e r e l a t i o n s h i p h a s b e e n f o u n d in t o m a t o b y A b e r g ( 1 9 4 8 ) , a n d in o a t by Wynd (1946). A h i g h c o r r e l a t i o n b e t w e e n a c c u m u l a t i o n of d r y m a t t e r and a s c o r b i c a c id is a ls o r e p o r t e d by S o m e r s and K elly (1 95 1) in t u r n i p a n d b r o c c o l i l e a f d i s c s . It s h o u ld be n oted th a t hig h a s c o r b i c a c id v a l u e s w e r e found i n th e t e r m i n a l l e a v e s a n d t h a t t h e s e l e a v e s h a d a h i g h d r y w e i g h t content. If t h e p r e s e n t d a t a a r e s e g r e g a t e d i n t o b a s a l a n d t e r m i n a l l e a v e s t h e r e l a t i o n s h i p b e t w e e n a s c o r b i c a c i d a n d d r y w e i g h t no lo n g er holds tru e . It is s u g g e s te d f r o m th is f a c t th at the h ig h e r a c t i v i t y of t h e y o u n g t e r m i n a l l e a v e s c o r r e s p o n d e d t o a t i s s u e w i t h a r e l a t i v e l y h i g h d r y w e i g h t w h i c h h a d a g r e a t e r p o w e r of s y n t h e s i s f o r a s c o r b i c a c id n o t n e c e s s a r i l y r e l a t e d to the d r y m a t t e r . This i d e a w a s p u t f o r t h b y R e id (1937), who o b s e r v e d t h a t h i g h e r c o n ­ c e n t r a t i o n s of a s c o r b i c a c i d w e r e f o u n d i n p a r t s of t i s s u e w i t h g r e a t e r a c tiv ity , a s show n in th e follow ing table: L o c a l i z a t i o n of A s c o r b i c A c i d i n R o o t s a n d S t e m s M illig ram s per G ram R oots T i p s l / 2 ” lo n g A djoining s e g m e n t 1” R em aining portion 0.475 0.171 0.115 Stem T ip s 1” M iddle Basal 0.169 0.018 0.056 A p o s i t i v e r e l a t i o n s h i p b e t w e e n t h e a g e of t h e t i s s u e a n d i ts a s c o r b i c a c id c o n te n t w as o b s e r v e d by B e r n s t e i n , H a m n e r and P a r k s (19 4 5 ) a s p r e s e n t e d in t h e f o l l o w i n g t a b l e : E f f e c t of L e a f A g e on A s c o r b i c A c i d C o n c e n t r a t i o n ( m g . / l O O g. f r e s h w e i g h t ) . F i v e P l a n t s H a r v e s t e d f r o m Sand C u ltu re B ala n c e d N u trie n t. P o s i t i o n of L e a v e s O ldest M iddle Youngest A v e r a g e of 5 P l a n t s 192.8 204.3 227.9 On the o th e r hand, a s ig n if ic a n t g r a d i e n t in the c o n c e n t r a ­ t i o n s of i r o n a n d m a n g a n e s e w a s a l s o f o u n d b e t w e e n t h e t e r m i n a l a n d b a s a l l e a v e s of t h e t o m a t o p l a n t s ; t h i s r e l a t i o n w a s c o n s i s t e n t l y found in all the c u l t u r e s . A s i m i l a r t e n d e n c y of a h i g h e r a m o u n t of i r o n a n d m a n g a n e s e i n th e b a s a l t h a n in t h e t e r m i n a l l e a v e s w a s r e p o r t e d b y L y o n , B e e s o n a n d E l l i s (1 943) a s s h o w n in t h e t a b l e g i v e n b e lo w : I r o n an d M a n g a n e s e A n a l y s e s of T o m a t o P l a n t s G r o w n in C o m p l e t e C u l t u r e S o l u t i o n Plant P a rt P p m of I r o n i n the D r y M a t t e r P p m of M a n g a n e s e in t h e D r y M a t t e r T o p o n e - t h i r d of th e p l a n t 175 + 14.9 M i d d l e o n e - t h i r d of t h e plant 199 + 9.0 248.3 + 29.95 L o w e r o n e - t h i r d of t h e plant 229 + 10.8 398.0 + 23.79 69.7 + 9 .08 It is o b s e rv e d that the b a s a l le a v e s w hich had a r e la tiv e ly h i g h i r o n a n d m a n g a n e s e c o n t e n t w e r e f o u n d t o h a v e a r a t h e r low a s c o r b i c a cid c o n c e n tr a tio n ; the o p p o s ite b e in g t r u e fo r the t e r ­ m inal leaves. I t c a n n o t b e s a i d t h a t t h e i n t e r n a l c o n c e n t r a t i o n of t h e s e e l e m e n t s a f fe c te d d i r e c t l y the a s c o r b i c acid s t a t u s b e c a u s e no c o n s is te n t re la tio n s h ip could be d ra w n b e tw e e n th e s e f a c to r s . 119 E. 1. S u m m a r y of t h e F i f t h E x p e r i m e n t T o m a t o p l a n t s w e r e g r o w n on n u t r i e n t s o l u t i o n s of c o n ­ t r o l l e d i r o n l e v e l s , a t pH 4, in t h e s a m e m a n n e r a s t h e w h e a t p l a n t s of t h e f o u r t h e x p e r i m e n t . 2. T e r m i n a l and b a s a l l e a v e s w e r e h a r v e s t e d w hen the p l a n t s w e r e s t a r t i n g to f l o w e r . the f r e s h tis s u e s . A s c o r b i c a c i d w a s d e t e r m i n e d on T h e d e t e r m i n a t i o n s of i r o n a n d m a n g a n e s e w e r e c a r r i e d on c o r r e s p o n d i n g o v e n d r i e d s a m p l e s . T h e p e r c e n t a g e s of dry m atter w ere recorded. 3. An i n c r e a s e i n g r o w t h of t h e p l a n t s w a s o b s e r v e d c o r ­ r e s p o n d i n g to a n i n c r e a s e in t h e c o n c e n t r a t i o n of i r o n in t h e s o l u ­ t i o n u p to f i v e p a r t s p e r m i l l i o n . A l t h o u g h , s m a l l e r in s i z e , t h e p l a n t s d e p r i v e d of i r o n d i d n o t s h o w a n y c h l o r o t i c s y m p t o m s . The p e r c e n t a g e of d r y m a t t e r w a s c o n s i s t e n t l y h i g h e r in t e r m i n a l t h a n b asal leaves. 4. T h e c o n c e n t r a t i o n of i r o n i n t h e f r e s h t i s s u e s h o w e d a n e g a t i v e r e l a t i o n s h i p w i t h t h e a m o u n t o f i r o n in t h e n u t r i e n t s o l u ­ t i o n b u t t h i s t e n d e n c y d i d n o t h o l d on a d r y m a t t e r b a s i s . In a l l c a s e s , a s i g n i f i c a n t l y h i g h e r a m o u n t of i r o n w a s f o u n d i n t h e b a s a l leaves. 120 5. T h e m a n g a n e s e v a l u e s of t h e b a s a l l e a v e s d i d n o t s h o w a n y s p e c i a l t r e n d b u t i n t h e t e r m i n a l l e a v e s t h e a m o u n t s of t h i s e l e m e n t w a s f o und to i n c r e a s e w i t h h i g h e r i r o n l e v e l s i n t h e n u ­ tr ie n t solution. 6. T h e a m o u n t s of a s c o r b i c a c i d in t h e b a s a l l e a v e s v a r i e d o n l y s l i g h t l y w i t h t h e i r o n l e v e l s of t h e n u t r i e n t s o l u t i o n b u t i t s c o n c e n t r a t i o n in t h e t e r m i n a l l e a v e s s h o w e d a p o s i t i v e r e l a t i o n to th e i r o n in t h e n u t r i e n t s o l u t i o n . In t h e f r e s h m a t t e r of t h e p l a n t s a n eg ativ e r e la tio n s h ip w as found b e tw e e n ir o n and a s c o r b i c acid and a l s o a p o s iti v e r e l a t i o n s h i p b e tw e e n d r y m a t t e r and a s c o r b i c a c i d w h e n a l l th e d a t a f r o m b a s a l a n d t e r m i n a l l e a v e s w e r e p o o l e d . T h e s e r e l a t i o n s d id n o t h o l d t r u e , h o w e v e r , w h e n t h e v a l u e s w e r e c a l c u l a t e d on a d r y b a s i s . VIII. EX PER IM EN T 6 A. Purpose It w a s e s t a b l i s h e d in the p r e c e d i n g e x p e r i m e n t w ith t o m a t o p l a n t s t h a t d e f i n i t e g r a d i e n t s in t h e c o n c e n t r a t i o n of a s c o r b i c a c i d , i r o n , a n d m a n g a n e s e e x i s t e d i n t h e d i f f e r e n t p a r t s of t h e p l a n t . Not e n o u g h d a t a w e r e t h e n a v a i l a b l e to f in d o u t if t h e i n t e r n a l c o n c e n ­ t r a t i o n of e a c h of t h e a b o v e c o m p o n e n t s h a d a n i n f l u e n c e o n t h e o t h e r . It w a s then p la n n e d to g r o w t o m a t o p la n ts on v a r i o u s l e v e ls of i r o n a n d c o p p e r a n d to c a r r y c h e m i c a l a n a l y s e s o n t h e p l a n t s a t d i f f e r e n t t i m e s of t h e g r o w i n g s e a s o n a n d f r o m d i f f e r e n t p a r t s of th e p l a n t . T h i s w a y of a t t a c k p e r m i t t e d t o o b t a i n i n f o r m a t i o n on t h e c o n s i s t e n c y of th e g r a d i e n t s in t h e c o n c e n t r a t i o n s of t h e v a r i o u s c o m p o n en ts d u rin g grow th. B e c a u s e of t h e r a t h e r u n e x p e c t e d i r o n c o n c e n t r a t i o n f o u n d in t h e p l a n t s of t h e f i f t h e x p e r i m e n t g r o w n w i t h o u t i r o n , i t w a s s u g g e s t e d t o p l a n t t h e s e t o m a t o e s on q u a r t z s a n d i n s t e a d of s o i l a n d t o t r a n s p l a n t t h e m l a t e r to t h e c u l t u r e p o t s . T h e p r o c e d u r e of s u p p l y i n g t h e n u t r i e n t s o l u t i o n i n t w o c a r b o y s w a s not u s e d b e c a u s e it did not give e n t i r e s a tis f a c tio n as fa r as iro n availability is concerned. F e r r i c c i t r a te w as supplied 122 d i r e c t l y to t h e c o m p l e t e s o l u t i o n w h e r e c i t r i c a c i d w a s a d d e d to m a i n t a i n the i r o n in so lu tio n . B. E x p e r im e n ta l C onditions 1. T he N u tr ie n t Solution T h e s a m e t h r e e s a l t s s o l u t i o n s o l u t i o n a s u s e d in t h e f i r s t e x p e r i ­ m e n t w i t h w h e a t p l a n t s w a s m a d e up w i t h v a r i o u s c o n c e n t r a t i o n s of i r o n a n d t w o l e v e l s of c o p p e r a s f o l l o w s : C o n c e n t r a t i o n s of I r o n in t h e N u t r i e n t S o l u t i o n s of t h e S i x t h E x p e r i m e n t No C o p p e r S e r i e s No i r o n 0 . 0 4 p p m . of C o p p e r S e r i e s 0.0 p p m F e 0.5 p p m F e 0.5 1.0 1.0 . 5.0 5.0 10.0 10.0 T h e s e tw o s e r i e s of c u l t u r e s w i l l b e r e f e r r e d to l a t e r a s t h e “ n o c o p p e r - s e r i e s ” a n d t h e “ 0 . 0 4 p a r t s p e r m i l l i o n of c o p p e r series.” 123 F e r r i c c i t r a t e w a s ad d ed f r o m a sto c k so lu tio n w h ic h had been s ta n d a rd iz e d fo r its iro n content. T h e i r o n in t h e s o l u t i o n w a s d e t e r m i n e d a t i n t e r v a l s a n d a d d i t i o n a l a m o u n t s of t h e s t o c k s o lu tio n w e r e added to b r in g the so lu tio n to th e d e s i r e d ir o n le v e l. In o r d e r to k e e p t h e i r o n i n s o l u t i o n , 20 m i l l i l i t e r s of 20 p e r c e n t c i t r i c a c i d w a s a d d e d p e r c a r b o y s o f 16 l i t e r s of s o l u t i o n . T h e c o m p l e t e n u t r i e n t s o l u t i o n w a s f i r s t m a d e up on N o v e m b e r 1, 1950, a n d r e n e w e d o n N o v e m b e r 29, 1 9 50, b e c a u s e of t h e g r o w t h of m o ld in the s o lu tio n s . The se c o n d tim e the s o lu tio n s w e r e m a d e up, 25 m i l l i l i t e r s of 20 p e r c e n t c i t r i c a c i d w e r e a d d e d . D uring t h e w h o l e e x p e r i m e n t t h e p H of t h e n u t r i e n t s o l u t i o n s v a r i e d f r o m 5.3 t o 6.0. B e f o r e s t a r t i n g the e x p e r i m e n t , th e q u a r t z g r a v e l w a s p a s s e d t h r o u g h an e l e c t r o - m a g n e t t o g e t r i d of i r o n d u s t s a n d p a r t i c l e s s t i l l f o u n d in th e g r a v e l . T o e a c h l e v e l of i r o n c o r r e s p o n d e d a d u p l i c a t e p a i r of c u l ­ tures. 2. The P lan t M aterial On S e p t e m b e r 28, 1 9 50, s e e d s of t o m a t o e s , v a r i e t y R u t g e r s F e r r y M o r s e D e t r o i t N o. 290 s t o c k D 7 0 6 8 w e r e s o w n i n v e r m i c u lite. F iv e p la n ts w e r e tr a n s p l a n t e d in th e q u a r tz g r a v e l p e r c u l­ t u r e p o t s w h e n t h e p l a n t s w e r e 35 d a y s o l d on N o v e m b e r 2, 1950. 124 T h e d a y a f t e r t r a n s p l a n t i n g , t h e c o t y l e d o n s of e a c h p l a n t w e r e c u t off. W h e n t h e p l a n t s w e r e 58 d a y s o l d , o n e p l a n t p e r p o t w a s h a r ­ vested. A t t h a t t i m e , s a m p l e s of t h e w h o l e p l a n t s w e r e u s e d to d e t e r m in e the a s c o r b i c acid, iro n , and d r y m a t t e r co n ten t. The s e c o n d h a r v e s t w a s m a d e w h e n t h e p l a n t s w e r e 78 d a y s o ld a n d then two p la n ts p e r p o ts w e r e ta k e n . B e c a u s e the p lan ts w e r e r a t h e r l a r g e , only the le a v e s w e r e u s e d fo r the d i f f e r e n t a n a l y s e s of a s c o r b i c acid, iro n , and m a n g a n e s e . F o r th e se s a m p le s a dif­ fe re n tia tio n w as m a d e b etw een the four u p p e r le a v e s r e f e r r e d la te r a s “ t e r m i n a l l e a v e s ” and the o n e s b e lo w a s “ b a s a l l e a v e s . ” In the s a m e m a n n e r , a th ird h a r v e s t w as m a d e w hen the p la n ts w e r e 97 d a y s old on J a n u a r y 3, 1951. C. 1. a. E xperim ental R esults O b s e r v a t i o n s on t h e P l a n t s D e s c r i p t i o n of t h e P l a n t s a t T i m e of H a r v e s t D u r i n g t h e w h o l e g r o w t h p e r i o d , no c h l o r o s i s s y m p t o m s a p ­ p e a r e d on t h e p l a n t s g r o w n o n t h e “ n o - i r o n l e v e l s ” of b o t h s e r i e s . In g e n e r a l t h e p l a n t s g r o w n on t h e “ 0 . 0 4 p a r t s p e r m i l l i o n of c o p ­ p e r s e r i e s ” w e r e of g r e a t e r s i z e t h a n t h o s e g r o w n on t h e s o l u t i o n s i n w h i c h no c o p p e r h a d b e e n a d d e d . A t all t i m e s , all th e p la n ts looked h e a lth y and s ta y e d v e r y g r e e n . J u s t b e f o r e the l a s t 125 h a r v e s t , s o m e p l a n t s s t a r t e d to f l o w e r ; t h e f l o w e r s w e r e t h e n p i n c h e d off. b. P e r c e n t a g e s of D r y M a t t e r i n t h e L e a v e s T h e a m o u n t s of d r y m a t t e r a t t h e d i f f e r e n t h a r v e s t s , e x ­ p r e s s e d a s p e r c e n t a g e s of t h e f r e s h m a t t e r of t h e l e a v e s of b o t h c o p p e r s e r i e s a r e g i v e n i n T a b l e 45 a n d F i g u r e s 260 t o 265, i n ­ clusive . In g e n e r a l t h e p l a n t s g r o w n o n t h e s o l u t i o n w h e r e 0 . 0 4 p a r t s p e r m i l l i o n of c o p p e r h a d b e e n a d d e d , g a v e a h i g h e r y i e l d of d r y m atter. T h is d i f f e r e n c e w a s m o s t a p p a r e n t w h en the p la n ts w e r e y o u n g , i n d e e d t h e a v e r a g e v a l u e of a l l t h e p l a n t s of t h e 0 . 0 4 p p m C u s e r i e s w h e n 58 d a y s old w a s 16 p e r c e n t g r e a t e r t h a n t h e a v e r ­ a g e v a l u e of th e p l a n t s g r o w n on n o c o p p e r . T his d ifferen ce betw een b o th s e r i e s s o m e w h a t d i s a p p e a r e d a t the l a t e r h a r v e s t s . In a l m o s t a l l t h e c a s e s w h e n b o t h b a s a l a n d t e r m i n a l l e a v e s w e r e s a m p l e d , the g r e a t e s t a m o u n t of d r y m a t t e r w a s found in th e te rm in a l leaves. A t e n d e n c y w h i c h i s s h o w n i n t h e d a t a on d r y w e i g h t s i s a n i n c r e a s e of t h e d r y m a t t e r in t h e l e a v e s w i t h a g e a n d in b o t h c o p ­ per series. s e r i e s . ' 1' T h is t e n d e n c y w a s m o r e a p p a r e n t in th e “ no c o p p e r 126 2. a. C h e m i c a l A n a l y s e s of t h e P l a n t s Iron (1) A m o u n t s of I r o n p e r 100 G r a m s of F r e s h M a t e r i a l T h e c o n c e n t r a t i o n s of i r o n i n m i l l i g r a m s p e r 100 g r a m s of f r e s h m a t t e r in t h e l e a v e s h a r v e s t e d w h e n t h e p l a n t s w e r e 58, 78 an d 97 d a y s o l d , f r o m t h e no c o p p e r a n d th e 0 . 0 4 p a r t s p e r m i l ­ l i o n of c o p p e r s e r i e s a r e l i s t e d in T a b l e 42 a n d F i g u r e s 2 6 6 to 271, i n c l u s i v e . In a l l t h e s a m p l e s , t h e c o n c e n t r a t i o n s of i r o n in t h e f r e s h m a t t e r did not show any c l e a r t e n d e n c ie s and no s ig n if ic a n t c o r ­ r e l a t i o n c a n b e d r a w n b e t w e e n t h e a m o u n t of i r o n in t h e l e a v e s a n d i t s c o n c e n t r a t i o n in t h e n u t r i e n t s o l u t i o n . When c o m p a r i n g both the “ n o - c o p p e r ” a n d the “ 0.04 p a r t s p e r m i l l i o n of c o p p e r ” s e r i e s , i t i s o b s e r v e d t h a t t h e a c c u m u l a ­ t i o n of i r o n in th e l e a v e s g e n e r a l l y g r e a t e r in t h e p l a n t s g r o w n on t h e “ no c o p p e r ” s e r i e s t h a n on t h e “ 0 . 0 4 p a r t s p e r m i l l i o n c o p p e r series.” T h i s o b s e r v a t i o n i s m o s t a p p a r e n t in t h e 58 d a y s o ld p l a n t s , l e s s i n t h e 78 d a y s o ld a n d h a s d i s a p p e a r e d in t h e 97 d a y s old p la n ts . In o p p o s i t i o n w i t h t h e r e s u l t s o b t a i n e d i n t h e p r e v i o u s t o ­ m a t o e x p e r i m e n t , a t e n d e n c y w a s s h o w n b y t h e t e r m i n a l l e a v e s to 127 h a v e a h i g h e r c o n c e n t r a t i o n of i r o n , h o w e v e r t h e d i f f e r e n c e s b e ­ tw een b a s a l and te r m in a l le a v e s w e r e not v e ry sig n ifican t. (2) A m o u n t s of I r o n p e r 100 G r a m s of D r y M a t e r i a l T h e d a t a f o r t h e a m o u n t of i r o n p e r 100 g r a m s of d r y p l a n t m a t e r i a l e x p r e s s e d in m i l l i g r a m s f o r the l e a v e s h a r v e s t e d f r o m p l a n t s 58, 78 a n d 97 d a y s o l d f r o m t h e “ n o c o p p e r ” a n d t h e “ 0 . 0 4 p a r t s p e r m i l l i o n of c o p p e r ” s e r i e s a r e a s s e m b l e d i n T a b l e 46 a n d F i g u r e s 2 7 2 to 2 77, i n c l u s i v e . T he s a m e o b s e r v a t i o n s a s th o s e r e p o r t e d f o r the c o n c e n ­ t r a t i o n s of i r o n i n t h e f r e s h m a t t e r c a n a l s o b e d r a w n f o r t h e d a t a o f i r o n in th e d r y m a t t e r . H e n c e no f u r t h e r d i s c u s s i o n w i l l b e carried . b. M anganese (1) A m o u n t of M a n g a n e s e p e r 100 G r a m s o f F r e s h M a t e r i a l T h e c o n c e n t r a t i o n s of m a n g a n e s e i n m i l l i g r a m s p e r 100 g r a m s of f r e s h m a t e r i a l in t h e l e a v e s of 78 a n d 97 d a y s o l d t o m a t o p l a n t s g r o w n on t h e “ n o c o p p e r ” a n d t h e “ 0 . 0 4 p a r t s p e r m i l l i o n o f c o p p e r ” s e r i e s a r e r e p o r t e d in T a b l e 43 a n d F i g u r e s 27 8 to 281, in c lu s iv e . 128 T o i n c r e a s e a m o u n t s of i r o n i n t h e n u t r i e n t s o l u t i o n s c o r ­ r e s p o n d e d a s i g n i f i c a n t d e c r e a s e i n t h e a c c u m u l a t i o n of m a n g a n e s e in t h e l e a v e s . In a g r e e m e n t w i t h t h e p r e v i o u s e x p e r i m e n t , a g r e a t e r c o n ­ c e n t r a t i o n of m a n g a n e s e w a s f o u n d i n t h e b a s a l l e a v e s of t h e t o ­ m a to p la n ts than in the t e r m i n a l le a v e s . This relationship was m o s t s i g n i f i c a n t i n t h e 97 d a y s o ld p l a n t s . In t h i s e x p e r i m e n t t h e c o p p e r c o n t e n t of t h e s o l u t i o n h a d a p p a r e n t l y no e f f e c t on t h e a c c u m u l a t i o n of m a n g a n e s e in t h e leaves. (2) A m o u n t s of M a n g a n e s e p e r 100 G r a m s of D r y M a t t e r T h e v a l u e s f o r t h e a m o u n t of m a n g a n e s e i n t h e d r y m a t t e r c o r r e s p o n d i n g to t h o s e g i v e n i n th e p r e c e d i n g p a r a g r a p h f o r t h e f r e s h m a t t e r a r e p r e s e n t e d in T a b l e 47 a n d F i g u r e s 282 t o 285, inclusive. The s a m e o b s e rv a tio n s as th o se m a d e for th e m a n g a n e s e c o n c e n t r a t i o n s in t h e f r e s h m a t t e r c a n b e d r a w n f r o m t h e d a t a on t h e d r y m a t t e r , in c o n s e q u e n c e no f u r t h e r d i s c u s s i o n w i l l b e d o n e . 129 c. A s c o r b ic A cid (1) A m o u n t s o f A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t e r i a l T h e c o n c e n t r a t i o n s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h m a t e r i a l g r o w n on d i f f e r e n t l e v e l s of i r o n w i t h o u t o r w i t h c o p p e r a r e l i s t e d i n T a b l e 44 a n d F i g u r e s 2 8 6 to 291. i n c l u s i v e . T h e c o n c e n t r a t i o n s of a s c o r b i c a c i d v a r i e d t o a s i g n i f i c a n t e x t e n t b e t w e e n s a m p l e s g r o w n on v a r i o u s i r o n l e v e l s b u t n o s i g ­ n i f i c a n t r e l a t i o n s h i p w a s a p p a r e n t b e t w e e n t h e i r o n l e v e l s in t h e n u t r i e n t s o l u t i o n s a n d t h e a s c o r b i c a c i d in th e p l a n t . In a l l c a s e s a n d a t a l l t h e h a r v e s t s , t h e a m o u n t s of a s c o r b i c a c i d w e r e m u c h g r e a t e r in t h e t e r m i n a l l e a v e s t h a n in t h e b a s a l l e a v e s . In o r d e r to f i n d a n y p o s s i b l e r e l a t i o n s h i p b e t w e e n t h e p l a n t s c o m p o n e n ts a n a ly z e d and the a s c o r b i c a c id at the d if f e r e n t h a r ­ v e s t s , t h e v a l u e s of a s c o r b i c a c i d in t h e f r e s h t i s s u e w e r e g r a p h e d r e s p e c t i v e l y a g a i n s t t h e a m o u n t s o f i r o n , of m a n g a n e s e a n d t h e p e r c e n t a g e s of d r y m a t t e r a s s e e n in F i g u r e s 2 9 8 to 3 05, i n c l u s i v e . A p o s i t i v e c o r r e l a t i o n b e t w e e n i r o n a n d a s c o r b i c a c i d in t h e t e r m i n a l l e a v e s of t h e 97 d a y s old p l a n t s of b o t h s e r i e s i s s h o w n in F i g u r e 3 0 1 . W hen the v a lu e s f o r t e r m i n a l and b a s a l le a v e s a r e pooled a p o s itiv e r e la tio n s h ip is o b s e r v e d betw een a s c o r b i c a c i d in t h e f r e s h t i s s u e a n d t h e p e r c e n t a g e s of d r y m a t t e r of t h e 78 d a y s old p l a n t s ( F i g u r e 3 0 4 ) . A positive rela tio n sh ip 130 i s a l s o o b s e r v e d in F i g u r e 30 5 b e t w e e n t h e p e r c e n t a g e s o f d r y m a t t e r of t h e b a s a l l e a v e s of t h e 97 d a y s old p l a n t s a n d t h e i r a s c o rb ic acid c o n c e n tra tio n s. B e s i d e s t h e s e few o b s e r v a t i o n s , no c o n s i s t e n t r e l a t i o n s h i p w a s o b s e r v e d a n d it w a s n o t p o s s i b l e to r e l a t e t h e v a r i a t i o n s o b ­ s e r v e d in th e a s c o r b i c a c i d c o n c e n t r a t i o n s a n d t h e f a c t o r s s t u d i e d . (2) A m o u n t s of A s c o r b i c A c i d p e r 100 G r a m s of D r y M a t t e r T h e m i l l i g r a m s o f a s c o r b i c a c i d p e r 100 g r a m s of d r y p l a n t m a t e r i a l g r o w n on d i f f e r e n t l e v e l s of i r o n w i t h o u t o r w i t h c o p p e r a r e r e c o r d e d in T a b l e 48 a n d F i g u r e s 2 9 2 to 297. W h e n t h e v a l u e s f o r a s c o r b i c a c i d a r e c a l c u l a t e d on a d r y m a t t e r b a s i s , ab o u t the s a m e te n d e n c i e s a s show n in its c o n c e n ­ t r a t i o n in t h e f r e s h m a t t e r a p p e a r e d . Should b e m e n tio n e d how­ e v e r , t h a t , on a d r y w e i g h t b a s i s , m o r e a s c o r b i c a c i d w a s f o und in t h e 58 d a y s o l d p l a n t s g r o w n w i t h o u t c o p p e r t h a n on t h o s e g r o w n w i t h 0 . 0 4 p a r t s p e r m i l l i o n of c o p p e r , b u t t h i s r e l a t i o n d o e s n o t hold t r u e at o ld e r a g e s . T h e o t h e r o b s e r v a t i o n s r e p o r t e d f o r t h e a s c o r b i c a c i d in th e f r e s h m a t t e r h o l d t r u e f o r t h e v a l u e s i n t h e d r y m a t t e r . 131 D. D i s c u s s i o n on th e S ix th E x p e r i m e n t In t h i s e x p e r i m e n t , u n d e r v a r i o u s s t a g e s o f g r o w t h a n d f r o m t h e b a s e to t h e t i p of t h e p l a n t s , g r a d i e n t s of i n c r e a s i n g c o n ­ c e n t r a t i o n s of a s c o r b i c a c i d a n d d r y m a t t e r a n d d e c r e a s i n g c o n ­ c e n t r a t i o n s of m a n g a n e s e w e r e f o u n d a n d c o n f i r m e d t h e f i n d i n g s e s t a b l i s h e d a n d d i s c u s s e d a t t h e e n d of t h e f i f t h e x p e r i m e n t . A s in t h e p r e c e d i n g e x p e r i m e n t , t h e c o n c e n t r a t i o n of a s c o r ­ b ic a c id in th e f r e s h p la n t t i s s u e w a s found to b e p o s i t i v e l y r e l a t e d to t h e a m o u n t of d r y m a t t e r o f t h e t i s s u e . T his relatio n sh ip s e e m s to b e h i g h l y s i g n i f i c a n t b e c a u s e i t h o l d s t r u e in m o s t c a s e s w h e n the d a ta fo r t e r m i n a l and b a s a l le a v e s a r e p o oled o r s e g r e g a t e d . On t h e o t h e r h a n d , t h e e f f e c t of i r o n a n d m a n g a n e s e c o n c e n t r a t i o n s i n t h e t i s s u e on t h e a s c o r b i c a c i d c o n t e n t w a s n o t c o n s i s t e n t a n d no g e n e r a l i z a t i o n c a n be m a d e . A l e s s e n i n g in t h e a c c u m u l a t i o n of i r o n in t h e p l a n t t i s s u e w a s n o t e d a s a s p e c i a l e f f e c t of t h e p r e s e n c e of c o p p e r in t h e n u ­ t r ie n t solution. T h is r e s u l t c o n f i r m s th o s e r e p o r t e d by M a q u e n n e a n d D e m o u s s y (1 920 ) a n d a l s o b y W i l l i s ( 1 9 3 6 ) . It w a s a l s o o b s e r v e d i n t h e p r e s e n t e x p e r i m e n t t h a t t o h i g h i r o n l e v e l s in t h e s o l u t i o n c o r r e s p o n d e d lo w a m o u n t s of m a n g a n e s e a c c u m u l a t e d in the t i s s u e s . T h is o b s e rv a tio n a g r e e s w ith a s i m ­ i l a r r e l a t io n s h i p o b s e r v e d in w h e a t tops d u rin g the f o u rth e x p e r im e n t. 132 In t h e l i t e r a t u r e , t h e u s u a l i n t e r a c t i o n b e t w e e n i r o n a n d m a n g a n e s e r e p o r t e d , i s a d e p r e s s i n g e f f e c t of h i g h a m o u n t s o f m a n g a n e s e on t h e a c c u m u l a t i o n of i r o n , f a c t w h i c h w a s o b s e r v e d b y J o h n s o n ( 1917) i n p i n e a p p l e , b y H o p k i n s (1 930) i n C h l o r e l l a , a n d m o r e r e c e n t l y b y S i d e r i s (1950). D u r i n g t h e p r e s e n t s t u d y , f e r r i c c i t r a t e w a s s u p p l i e d to t h e n u t r i e n t s o l u t i o n s a n d c i t r i c a c i d w a s a d d e d to p r e v e n t t h e p r e c i p ­ i t a t i o n of i r o n ; t h i s m e t h o d g a v e r a t h e r g o o d r e s u l t s a s c a n b e o b ­ s e r v e d f r o m t h e a m o u n t s of i r o n t a k e n up b y t h e p l a n t s . O n e of t h e d i s a d v a n t a g e s h o w e v e r i s t h a t t h e p r e s e n c e of c i t r i c i o n f a v o u r t h e g r o w t h of m o l d in t h e s o l u t i o n , w h i c h t h e n m u s t b e c h a n g e d . The s a m e m e t h o d of p r o v i d i n g i r o n in c o m b i n a t i o n w i t h c i t r i c a c i d h a s b e e n r e p o r t e d v e r y r e c e n t l y to b e u s e d b y B i t c o v e r a n d S i e l i n g (1 951) w h o n o t e d t h a t a s o l u t i o n c o n t a i n i n g t e n p a r t s p e r m i l l i o n , w i t h a 4 t o 1 m o l a r r a t i o c i t r i c a c i d to i r o n , g a v e t h e b e s t g r o w t h of S p i r o d e l a . E. 1. S u m m a r y of t h e S i x t h E x p e r i m e n t T o m a t o p l a n t s w e r e g r o w n o n n u t r i e n t s o l u t i o n s of c o n ­ tro lle d i r o n - l e v e l s w ith and w ithout c o p p e r. The iro n w as supplied in t h e s a m e c a r b o y s a s t h e o t h e r n u t r i e n t s in t h e f o r m of f e r r i c c i t r a te s u p p le m e n te d w ith c itric acid . 133 2. W h o l e p l a n t s w e r e h a r v e s t e d w h e n 58 d a y s o l d . T er­ m i n a l a n d b a s a l l e a v e s w e r e c o l l e c t e d on 78 a n d 97 d a y s old plants. A s c o r b i c a c i d w a s d e t e r m i n e d on t h e f r e s h t i s s u e s . The d e t e r m i n a t i o n s of i r o n a n d m a n g a n e s e w e r e c a r r i e d on c o r r e s p o n d ­ in g o v e n d r i e d s a m p l e s . T h e p e r c e n t a g e s of d r y m a t t e r w e r e recorded. 3. L a r g e r p l a n t s w e r e g r o w n on t h e n u t r i e n t s o l u t i o n s s u p ­ plied w ith c o p p e r; th e s e p la n ts a l s o c o n tain ed a g r e a t e r p e r c e n ta g e of d r y m a t t e r . A g r e a t e r a m o u n t of d r y m a t t e r w a s found in a ll th e t e r m i n a l t h a n in t h e b a s a l l e a v e s . In a l l t h e c a s e s , t h e a m o u n t s of d r y m a t t e r in t h e t i s s u e s i n c r e a s e d w i t h a g e . 4. T h e c o n c e n t r a t i o n of i r o n in t h e t i s s u e d i d n o t s h o w a n y s i g n i f i c a n t r e l a t i o n s h i p to t h e l e v e l of i r o n in t h e s o l u t i o n . A d e p r e s s i n g e f f e c t on th e i n t a k e of i r o n in t h e p r e s e n c e of c o p p e r in t h e s o l u t i o n w a s o b s e r v e d . No s i g n i f i c a n t g r a d i e n t in t h e c o n ­ c e n t r a t i o n of t h i s e l e m e n t w a s n o t e d in t h e d i f f e r e n t p a r t of t h e plants. 5. A n e g a t i v e r e l a t i o n s h i p b e t w e e n t h e c o n c e n t r a t i o n of m a n g a n e s e i n t h e t i s s u e a n d t h e l e v e l of i r o n in t h e s o l u t i o n w a s reported. served. N o e f f e c t of c o p p e r on t h e m a n g a n e s e u p t a k e w a s o b ­ G e n e r a l l y a h i g h e r a m o u n t of t h i s e l e m e n t w a s f o u n d i n th e b a s a l t h a n i n t h e t e r m i n a l l e a v e s . 6. No s i g n i f i c a n t r e l a t i o n s h i p c o u l d b e d r a w n b e t w e e n t h e i r o n c o n t e n t of t h e s o l u t i o n a n d t h e c o n c e n t r a t i o n o f a s c o r b i c a c i d in the p la n t t i s s u e . H ow ever a p o sitiv e c o r r e la tio n betw een the i r o n in the t i s s u e and its a s c o r b i c a c id c o n c e n t r a t i o n w a s o b s e r v e d in t h e t e r m i n a l l e a v e s w h e n t h e p l a n t s w e r e 97 d a y s old^ A con­ s i s t e n t p o s i t i v e r e l a t i o n s h i p b e t w e e n t h e p e r c e n t a g e of d r y m a t t e r a n d t h e c o n c e n t r a t i o n of a s c o r b i c a c i d i n t h e p l a n t t i s s u e w a s o b ­ s e rv e d at each d e term in a tio n . IX. EX PER IM EN T 7 A. Purpose T h e p r e c e d i n g tw o e x p e r i m e n t s e s t a b l i s h e d t h e e x i s t e n c e of a s i g n i f i c a n t g r a d i e n t i n th e c o n c e n t r a t i o n s of a s c o r b i c a c i d , in t h e t e r m i n a l a n d b a s a l l e a v e s of t h e t o m a t o p l a n t s . H o w e v e r , no c o n s i s t e n t r e l a t i o n s h i p b e t w e e n t h e s e v a r i o u s c o n c e n t r a t i o n s of a s c o rb ic acid and o th e r f a c to r s could be d ra w n , ex cep t w ith the d ry m a t t e r c o n t e n t of t h e t i s s u e s . T h e h i g h e r c o n c e n t r a t i o n of a s c o r b i c a c i d i n t h e t e r m i n a l l e a v e s , m i g h t b e e x p l a i n e d b y a g r e a t e r r a t e of s y n t h e s i s o r b y a g r e a t e r s t a b i l i t y of t h e a s c o r b i c a c i d i n t h a t p l a n t t i s s u e . A s t u d y of t h e s t a b i l i t y o f t h e a s c o r b i c a c i d in t h e d r y i n g t i s s u e f r o m d i f f e r e n t p a r t s of t o b a c c o p l a n t s w a s t h e n i n v e s t i g a t e d . The to b a c c o p la n ts w e r e c h o s e n a s the e x p e r i m e n t a l m a t e r i a l b e c a u s e of t h e l a r g e l e a v e s w h i c h g i v e a r a t h e r u n i f o r m m a t e r i a l a n d w h i c h e a s i l y c a n b e s p l i t i n tw o c o m p a r a t i v e s a m p l e s a l o n g the m i d r i b . O n o n e h a l f of t h e l e a f t h e a s c o r b i c a c i d d e t e r m i n a t i o n w a s c a r r i e d on t h e o t h e r h a l f t h e a s h a n a l y s e s w e r e d o n e . I t w a s p r o p o s e d t o f o l l o w t h e o x i d a t i o n of t h e a s c o r b i c a c i d in c o m p a r a t i v e s a m p l e s f r o m t h r e e d i f f e r e n t p a r t s of th e to b a c c o p l a n t s , g r o w n o n s u b s t r a t e d i f f e r i n g in t h e i r i r o n a n d c o p p e r c o n t e n t . R e l a t i o n s h i p s b e t w e e n t h e r a t e of o x i d a t i o n a n d t h e c h e m i c a l c o m ­ p o s i t i o n of t h e t i s s u e s w e r e t h e n i n v e s t i g a t e d . In a p r e l i m i n a r y e x p e r i m e n t , t h e v a r i a t i o n s i n c o n c e n t r a ­ t i o n s of a s c o r b i c a c i d , i r o n a n d m a n g a n e s e w a s f i r s t d e t e r m i n e d f o r e a c h l e a v e s of a t o b a c c o p l a n t . B. P re lim in a ry E xperim ent I n o r d e r to f i n d o u t t h e p o s s i b l e v a r i a t i o n s in a s c o r b i c a c i d , i r o n , and m a n g a n e s e in to b a c c o l e a v e s , the follow ing e x p e r i m e n t w a s c a r r i e d out: A l l t h e l e a v e s of a t o b a c c o p l a n t , g r o w n on s o i l in t h e g r e e n h o u s e , w e r e c u t in h a l f a l o n g t h e m i d r i b . O n e h a l f of e a c h le a f w a s u s e d fo r the a s c o r b i c a c id d e t e r m i n a t i o n and the o th e r h a l f w a s u s e d f o r a n a l y s e s of i r o n a n d m a n g a n e s e ; a t t h e s a m e t i m e t h e p e r c e n t a g e s of d r y m a t t e r of e a c h l e a f w e r e r e c o r d e d . T h e d a t a c o m p i l e d on t h i s t o b a c c o p l a n t a r e l i s t e d i n T a b l e s 49 a n d 50 a n d g r a p h e d in F i g u r e s 306 to 313, i n c l u s i v e . F r o m t h e b a s e t o t h e t o p of t h i s t o b a c c o p l a n t , t e n l e a v e s w e r e c u t and a n a ly z e d . A s s e e n f r o m the d a ta , d e fin ite g r a d i e n t s i n t h e c o n c e n t r a t i o n s of i r o n , m a n g a n e s e , a n d a s c o r b i c a c i d w e r e found, and a r e d i s c u s s e d below . T h e c o n c e n t r a t i o n of i r o n i n t h e f r e s h m a t e r i a l w a s t h e h i g h e s t in th e b o tto m leaf, th e n d e c r e a s e d to a m i n i m u m in the 137 f i f t h l e a f , 29 c e n t i m e t e r s f r o m t h e s o i l , a n d t h e n i n c r e a s e d a g a i n t i l l t h e l a s t l e a f 62 c e n t i m e t e r s f r o m t h e s o i l ( F i g u r e 3 06) . T h e a m o u n t s of m a n g a n e s e w e r e f o u n d to b e of v e r y low concentrations: u r e 301). o n l y o n e - t e n t h of t h e c o n c e n t r a t i o n of i r o n ( F i g ­ T h e g r a d i e n t in t h e c o n c e n t r a t i o n s of m a n g a n e s e w a s a p p a r e n t l y i n c r e a s i n g f r o m th e b o tto m le a f to the tip le a f ( F i g u r e 31 2 ) , a l t h o u g h f e w l e a v e s g a v e v a l u e s w h i c h w e r e o u t of t h e g e n ­ e ra l trend. T h e s e v a r i a t i o n s in t h e v a l u e s f r o m t h e g e n e r a l t r e n d m a y be e x p la in e d b y the r e l a t i v e s m a l l c o n c e n t r a t i o n found, w h ic h i t s e l f i s m o r e d i f f i c u l t to d e t e c t q u a n t i t a t i v e l y w i t h a c c u r a c y . T h e c o n c e n t r a t i o n of a s c o r b i c a c i d i n e a c h l e a f s h o w e d a d e f i n i t e g r a d i e n t i n t h e c o n c e n t r a t i o n s f r o m b o t t o m to t o p ( F i g u r e s 308 a n d 3 1 3 ) . T h e l e a s t c o n c e n t r a t i o n w a s f o u n d i n th e b o t t o m l e a f a n d t h e n i n c r e a s e d up t o a m a x i m u m a t t h e s i x t h l e a f , 34 c e n t i m e t e r s fro m the soil. In t h e u p p e r l e a v e s (3 9 c e n t i m e t e r s and above) the c o n c e n t r a t i o n s had a te n d e n c y to d e c r e a s e b u t the d i f f e r e n c e s in a m o u n t s b e t w e e n s u c c e s s i v e l e a v e s w e r e n o t a s g r e a t in m a g n i t u d e t h a n t h o s e b e t w e e n t h e l o w e r l e a v e s , No s ig n ific a n t c o r r e l a t i o n s could b e d ra w n e i t h e r b e tw e e n t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d a n d i r o n , o r m a n g a n e s e , in th e f r e s h m a t t e r . H ow ever, a slight negative re la tio n sh ip was o b s e r v e d b e t w e e n t h e v a l u e s of m a n g a n e s e a b o v e 0 . 2 m i l l i g r a m s p e r 100 g r a m s of f r e s h t i s s u e a n d t h e c o r r e s p o n d i n g c o n c e n t r a t i o n s of a s c o r b i c a c i d ( F i g u r e 3 0 9) . A lth o u g h no s ig n i f i c a n t c o r r e l a t i o n c a n be b r o u g h t o u t b e tw e e n a s c o r b i c a c id and i r o n , i t s h o u ld be noted th a t a s lig h t n e g a tiv e t r e n d r e l a t e d th e s e two c o m p o n e n ts ( F i g u r e 310). A s i g n i f i c a n t r e l a t i o n s h i p b e t w e e n t h e a m o u n t of a s c o r b i c a c i d a n d t h e p e r c e n t a g e s of d r y m a t t e r w a s o b s e r v e d i n t h e l e a v e s . I n d e e d w h e n t h e v a l u e s of a s c o r b i c a c i d i n t h e f r e s h m a t t e r w e r e g r a p h e d a g a i n s t t h e p e r c e n t a g e s of d r y m a t t e r , a p a r a b o l i c c u r v e w a s o b t a i n e d , a s s e e n in F i g u r e 311. F r o m the ab o v e r e p o r t e d d a ta , it w a s c o n c lu d e d th a t to ­ b a c c o p l a n t s w o u l d b e a d e q u a t e to p r o v i d e p l a n t t i s s u e s of v a r y i n g a s c o rb ic acid, iro n , and m a n g a n e s e c o n c e n tr a tio n s . C. E x p e rim e n ta l C onditions 1. T he N u tr ie n t Solution T h e n u t r i e n t s o l u t i o n r e c o m m e n d e d b y S p e n c e r (1 941 ) w a s used. T h e f i n a l c o m p o s i t i o n of t h e s o l u t i o n w a s a s f o l l o w s : 139 C o m p o s i t i o n of t h e N u t r i e n t S o l u t i o n Salt G r a m s p e r 16 L i t e r s M gS04 • 7 HzO C a ( N 0 3) 2 • 4 H z O kh 2p o 7.9 20.8 14.0 4 3. 5 ( n h 4) 2 s o 4 T h e f i n a l v o l u m e i n e a c h c a r b o y w a s m a d e up t o 16 l i t e r s w ith d istille d w a te r. M i c r o e l e m e n t s w e r e s u p p l i e d b y a d d i n g 10 m i l l i l i t e r s p e r c a r b o y of a s t o c k s o l u t i o n h a v i n g t h e f o l l o w i n g c o m ­ position: C o m p o s i t i o n of t h e S t o c k S o l u t i o n of M i c r o e l e m e n t s Salt G ram s per L iter H 3B 0 3 4.568 M nS04 2.462 T h e f i n a l c o n c e n t r a t i o n s of t h e m i c r o - e l e m e n t s in t h e n u ­ t r i e n t so lu tio n s w e r e a s follow s: 140 C o n c e n t r a t i o n of M i c r o - e l e m e n t s i n t h e N u t r i e n t S o l u t i o n s E lem ent P p m of t h e E l e m e n t B oron 0. 5 M anganese 0. 5 I n t h e s a m e m a n n e r a s in t h e s e c o n d e x p e r i m e n t w i t h t o ­ m a t o p l a n t s , t w o l e v e l s of c o p p e r w e r e u s e d : one w h e r e no c o p p e r w a s a d d e d a n d t h e o t h e r w i t h 0 . 0 8 p a r t s p e r m i l l i o n of c o p p e r . T h e s e t w o s e r i e s w i l l l a t e r b e r e f e r r e d to a s t h e “ n o c o p p e r s e ­ r i e s " and the “ c o p p e r s e r i e s . ” In t h e s e tw o s e r i e s , e x i s t e d t h r e e d i f f e r e n t c u l t u r e s i n r e s p e c t to i r o n : o n e w h e r e no i r o n w a s a d d e d , a s e c o n d w h e r e a c o n c e n t r a t i o n of f o u r p a r t s p e r m i l l i o n w a s m a d e up b y a d d i n g f e r r i c s u l f a t e a n d t h e t h i r d w h e r e t h e i r o n w a s s u p ­ p l i e d f r o m a g l a s s y f r i t , u s e d i n s t e a d of t h e q u a r t z g r a v e l . This i r o n - f r i t N o. A 6 3 0 0 A w a s s u p p l i e d b y t h e F e r r o E n a m e l C o m p a n y in C l e v e l a n d . T h e c u l t u r e s m e n t i o n e d a b o v e w i l l b e r e f e r r e d to l a t e r a s the “ No i r o n , ” the “ I r o n - g r a v e l ” and the “ I r o n - F r i t ” cu ltu res. 141 2. The P lan t M aterial S e e d s of N i c o t i a n a t o b a c u m v a r i e t y H a v a n a N o. 38, w e r e p l a n t e d in s o i l on D e c e m b e r 7, 1950. The s e e d s w e r e supplied t h r o u g h t h e c o u r t e s y o f D r . J a m e s J o h n s o n o f t h e U n i v e r s i t y of W isconsin. W h e n t h e p l a n t s w e r e 34 d a y s o l d , on J a n u a r y 10, t h e y w e r e t r a n s p l a n t e d in t h e c u l t u r e p o t s . D u rin g the t r a n s p l a n t i n g th e r o o t s w e r e w a s h e d w i t h t a p a n d d i s t i l l e d w a t e r . A t t h e t i m e of h a r v e s t 59 d a y s a f t e r t r a n s p l a n t i n g , on M a r c h 10, 1951, t h r e e k i n d s of s a m p l e s w e r e t a k e n : the te r m in a l, m e d ia n an d b a s a l l e a v e s , a s d e s c r i b e d b e l o w . F r o m e a c h c u l t u r e the f o u r t e r m i n a l l e a v e s w e r e ta k e n to c o n s t i t u t e a c o m p o s i t e s a m p l e of w h a t w i l l b e m e n t i o n e d l a t e r a s “ term in al leav es.” T h e n e x t t h r e e l e a v e s of e a c h p l a n t s w e r e c o l l e c t e d a n d p u t t o g e t h e r to f o r m t h e s a m p l e s o f “ m e d i a n l e a v e s . ” A t l a s t , g o in g f u r t h e r d o w n on t h e p l a n t s , t h e t h r e e n e x t le a v e s w e r e used for the “ b a s a l le a v e s ” s a m p le s . E a c h s a m p l e w a s su b d iv id ed into six s u b s a m p l e s by cu ttin g each leaf into six p a r t s and d is tr ib u tin g each p a r t into ea c h s u b ­ s a m p l e s o t h a t a c o m p o s i t e of a l l t h e l e a v e s p a r t s w o u l d c o n s t i ­ tute e a c h s u b s a m p le . O n e of t h e s u b s a m p l e s o f e a c h c u l t u r e w a s i m m e d i a t e l y u s e d to d e t e r m i n e t h e a m o u n t s of a s c o r b i c a c i d a t h a r v e s t . An­ o t h e r s u b s a m p l e of e a c h c u l t u r e w a s w a s h e d w i t h d i s t i l l e d w a t e r a n d p u t to d r y i n a f o r c e d a i r o v e n a n d w a s l a t e r u t i l i z e d f o r t h e iro n and m a n g a n e s e a n a ly s e s . The r e m n a n t s u b s a m p le s w e r e left i n t r a y s t o d r y a t r o o m t e m p e r a t u r e in t h e l a b o r a t o r y a n d w e r e u s e d t o d e t e r m i n e t h e a s c o r b i c a c i d a t v a r i o u s t i m e of d r y i n g . D. 1. a. E x p erim en tal R esults O b s e r v a t i o n s on t h e P l a n t s D e s c r i p t i o n of th e P l a n t s F o u r t e e n d a y s a f t e r t r a n s p l a n t i n g , t h e p l a n t s g r o w n on t h e n u t r i e n t s o l u t i o n w i t h o u t i r o n a n d w i t h 0 . 0 8 p a r t s p e r m i l l i o n of c o p p e r s t a r t e d to s h o w c h l o r o s i s , w h i l e t h e p l a n t s of t h e “ n o i r o n ” c u l t u r e i n t h e “ no c o p p e r s e r i e s ” d i d n o t s h o w a n y c h l o r o s i s . At h a r v e s t t h e p l a n t s of t h e “ n o i r o n ” c u l t u r e i n t h e “ c o p p e r s e r i e s ” w e r e v e r y s t u n t e d a n d c h l o r o t i c , w h i l e in t h e “ no c o p p e r s e r i e s ” no a p p a r e n t c h l o r o s i s w a s s h o w i n g a n d t h e p l a n t s w e r e a l m o s t a s big a s the one s u p p lie d w ith iro n . T h e p l a n t s of t h e “ I r o n - g r a v e f ” a n d “ I r o n - f r i t ” w e r e a t h a r v e s t i n v e r y h e a l t h y c o n d i t i o n a n d t h e p l a n t of b o t h of t h e s e c u l t u r e s w e r e s o m e w h a t l a r g e r in the “ c o p p e r s e r i e s . ” 143 b. F r e s h W eight p e r P la n t T h e a v e r a g e f r e s h w e i g h t s in g r a m s p e r p l a n t g r o w n on t h e v a r i o u s c u l t u r e s of b o t h s e r i e s a r e g i v e n in t h e f o l l o w i n g t a b l e : A v e r a g e F r e s h W eig h ts p e r P l a n t in G r a m s C ultures “ C o p p e r S e r i e s ’’ “ No C o p p e r S e r i e s ” “ Iron-F r it” 592.9 522.9 “ Iron-gravel ” 581.0 537.0 “ No i r o n ” 332.0 500.0 It c a n b e s e e n t h a t a t t h e e x c e p t i o n of t h e “ No i r o n ” c u l ­ t u r e , t h e p l a n t s on t h e c o p p e r s e r i e s w e r e s i g n i f i c a n t l y h e a v i e r t h a n t h e p l a n t s g r o w n on t h e s o l u t i o n s w h e r e no c o p p e r h a d b e e n added. T h i s o b s e r v a t i o n a g r e e s w i t h t h e o n e m a d e on t h e a p p e a r ­ a n c e of t h e p l a n t s a t h a r v e s t . c. P e r c e n t a g e s of D r y M a t t e r i n t h e L e a v e s T h e p e r c e n t a g e s of d r y m a t t e r i n t h e t e r m i n a l , m e d i a n a n d b a s a l l e a v e s of t o b a c c o p l a n t s g r o w n on v a r i o u s c u l t u r e s d i f f e r i n g in i r o n c o n t e n t of b o t h “ c o p p e r ” a n d “ n o c o p p e r ” s e r i e s a r e l i s t e d in T a b l e 51 a n d F i g u r e s 314 t o 319» i n c l u s i v e . 144 In b o t h s e r i e s t h e p e r c e n t a g e s of d r y m a t t e r s h o w e d t h e s a m e t e n d e n c i e s a s f a r a s i t s c o n c e n t r a t i o n in t h e d i f f e r e n t k i n d of l e a v e s i s c o n c e r n e d : the t e r m i n a l l e a v e s h a v in g the g r e a t e s t p e r c e n t a g e s of d r y m a t t e r , th e b a s a l o n e s th e l e a s t and the m e d i a n l e a v e s b e i n g i n b e t w e e n i n t h e i r a m o u n t s of d r y m a t t e r p e r u n i t w eight. C o m p a r in g the c o p p e r s e r i e s w ith the “ no c o p p e r ” s e r i e s , i t i s f o u n d t h a t t h e p e r c e n t a g e s of d r y m a t t e r w e r e g e n e r a l l y h i g h e r in b o t h “ i r o n - f r i t ” a n d “ i r o n - g r a v e l ” c u l t u r e s of t h e c o p p e r s e r i e s t h a n of t h e “ n o c o p p e r s e r i e s . ” H o w e v e r, the o p p o s ite w as ob­ s e r v e d i n t h e c a s e of t h e “ no i r o n ” c u l t u r e s , w h e r e a g r e a t e r p e r ­ c e n t a g e of d r y m a t t e r w a s f o u n d in t h e t e r m i n a l a n d m e d i a n l e a v e s of t h e “ no c o p p e r ” s e r i e s t h a n of t h e “ c o p p e r ” s e r i e s . It should b e r e c a l l e d t h a t t h e p l a n t s of t h e c o p p e r s e r i e s g r o w n w i t h o u t i r o n w e r e v e r y c h l o r o t i c a n d s t u n t e d a t t h e t i m e of h a r v e s t , w h i l e t h e p l a n t s of t h e o t h e r s e r i e s on t h e s a m e c u l t u r e d i d n o t a p p e a r a b ­ n o r m a l e x c e p t f o r t h e i r s m a l l e r g r o w t h in c o m p a r i s o n t o t h e plan ts supplied w ith iro n . W hen c o m p a r i n g the d i f f e r e n t c u l t u r e s , i t i s n o te d th a t the h i g h e s t p e r c e n t a g e of d r y m a t t e r w a s o b t a i n e d i n t h e f r i t g r o w n p la n ts and the l e a s t in the c u l t u r e s d e p r i v e d of i r o n . 2. a. C h e m i c a l A n a l y s e s of t h e T o b a c c o P l a n t s Iron ( l ) A m o u n t s of I r o n p e r 100 G r a m s o f F r e s h M a t e r i a l T h e c o n c e n t r a t i o n s of i r o n in m i l l i g r a m s p e r 100 g r a m s of f r e s h m a t t e r in t h e t e r m i n a l , m e d i a n a n d b a s a l l e a v e s of t o b a c c o p l a n t s g r o w n o n t h e “ i r o n - f r i t , ” “ i r o n - g r a v e l ” a n d “ no i r o n ” c u l ­ t u r e s of b o t h “ n o c o p p e r ” a n d “ c o p p e r ” s e r i e s a r e r e c o r d e d in T a b l e 51 a n d F i g u r e s 320 to 3 25, i n c l u s i v e . C o n s i s t e n t l y in the c u l t u r e s s u p p lie d w ith i r o n , i.e ., the f r i t a n d t h e g r a v e l c u l t u r e s of b o t h c o p p e r s e r i e s , t h e m e d i a n l e a v e s c o n t a i n e d t h e h i g h e s t c o n c e n t r a t i o n of i r o n . H o w e v e r , it s h o u l d b e r e m i n d e d t h a t in t h e t o b a c c o p l a n t g r o w n on s o i l a n d u s e d f o r t h e p r e l i m i n a r y e x p e r i m e n t t h e l e a s t c o n c e n t r a t i o n of i r o n h a d b e e n f ound in t h e m e d i a n l e a v e s . W hen the plan ts w e r e d e p r i v e d of i r o n , in b o t h s e r i e s , w i t h o r w i t h o u t c o p p e r , th e g r e a t ­ e s t c o n c e n t r a t i o n of i r o n w a s f o un d i n t h e b a s a l l e a v e s an d t h e l e a s t in t h e t e r m i n a l l e a v e s . T h e a c c u m u l a t i o n of i r o n in a l l th e l e a v e s s e e m to h a v e b e e n d i r e c t l y i n f l u e n c e d b y t h e p r e s e n c e of c o p p e r i n t h e n u t r i e n t s o l u t i o n ; i n d e e d , i n a l l t h e l e a v e s of a l l t h e p l a n t s g r o w n on t h e 146 s o l u t i o n s d e p r i v e d of c o p p e r m o r e i r o n w a s f o u n d t h a n i n t h e s e r i e s supplied w ith c o p p e r. (2) A m o u n t s of I r o n p e r 100 G r a m s of D r y M a t t e r T h e d a t a f o r t h e a m o u n t s of i r o n p e r 100 g r a m s of d r y p l a n t m a t e r i a l of t h e v a r i o u s l e a v e s of t o b a c c o p l a n t s a r e r e p o r t e d i n T a b l e 54 a n d F i g u r e s 32 6 t o 3 31, i n c l u s i v e . T h e s a m e te n d e n c i e s a s th o s e o b s e r v e d in the c o n c e n t r a ­ t i o n s of i r o n in t h e f r e s h t i s s u e h o l d t r u e f o r t h e c o n c e n t r a t i o n s of i r o n in t h e d r y m a t e r i a l . I t s h o u l d a l s o b e m e n t i o n e d t h a t in a l l t h e c a s e s t h e c o n c e n t r a t i o n s of i r o n in t h e d r y m a t t e r w a s t h e l e a s t in t h e t e r m i n a l l e a v e s . The o b s e rv a tio n m a d e about the d e ­ p r e s s i n g e f f e c t of c o p p e r on t h e a c c u m u l a t i o n of i r o n in t h e l e a v e s i s e v e n m o r e a p p a r e n t on a d r y m a t t e r b a s i s . b. M anganese (1) A m o u n t s of m a n g a n e s e p e r 100 G r a m s of F r e s h M a t e r i a l T h e c o n c e n t r a t i o n s of m a n g a n e s e i n m i l l i g r a m s p e r 100 g r a m s of f r e s h m a t e r i a l in t h e t e r m i n a l , m e d i a n a n d b a s a l l e a v e s of t o b a c c o p l a n t s g r o w n o n t h e “ i r o n - f r i t , ” “ i r o n - g r a v e l ” a n d t h e “ no i r o n ” c u l t u r e s o f b o t h “ n o c o p p e r ” a n d “ c o p p e r ” s e r i e s a r e a s s e m b l e d i n T a b l e 51 a n d F i g u r e s 332 to 3 3 7 , i n c l u s i v e . In b o t h c u l t u r e s p r o v i d e d w i t h i r o n a n d w i t h o r w i t h o u t c o p p e r , a g r a d i e n t i n t h e c o n c e n t r a t i o n s of m a n g a n e s e w a s o b ­ s e r v e d f r o m t h e b a s a l to t h e t e r m i n a l l e a v e s , t h e b a s a l c o n t a i n i n g th e l e a s t and the t e r m i n a l the m o s t . On t h e o p p o s i t e , i n b o t h of t h e “ n o i r o n ” c u l t u r e s a n d e s p e c i a l l y i n t h e o n e of t h e “ c o p p e r s e r i e s ” a s i g n i f i c a n t h i g h e r c o n c e n t r a t i o n of m a n g a n e s e w a s f o u n d i n t h e b a s a l l e a v e s , in c o m p a r i s o n to t h e t e r m i n a l o n e s w hich co n ta in e d the le a s t. In t h e s e c u l t u r e s t h e p l a n t s f r o m th e “ c o p p e r s e r i e s ” h a d a m u c h h i g h e r c o n c e n t r a t i o n of m a n g a n e s e t h a n t h o s e of t h e “ no c o p p e r ” s e r i e s . This re la tio n s h ip betw een b o t h c o p p e r s e r i e s d i d n o t e x i s t in t h e c u l t u r e s s u p p l i e d w i t h i r o n . (2) A m o u n t s of M a n g a n e s e p e r 100 G r a m s o f D r y M a t e r i a l T h e v a l u e s f o r t h e a m o u n t s of m a n g a n e s e in t h e d r y m a t t e r c o r r e s p o n d i n g to t h o s e g i v e n in th e p r e c e d i n g p a r a g r a p h f o r th e f r e s h m a t t e r a r e p r e s e n t e d in T a b l e 54 a n d F i g u r e s 338 to 34 3, inclusive. O n a d r y b a s i s , n o s i g n i f i c a n t g r a d i e n t in th e c o n c e n t r a t i o n s of m a n g a n e s e in t h e v a r i o u s l e a v e s of t o b a c c o p l a n t s c o u l d b e o b ­ s e r v e d a t t h e e x c e p t i o n of t h o s e of t h e “ no i r o n ” c u l t u r e s . It w as f o u n d , a s on a f r e s h w e i g h t b a s i s t h a t i n t h e “ n o i r o n ” c u l t u r e s , t h e h i g h e s t c o n c e n t r a t i o n s of m a n g a n e s e w a s o b t a i n e d i n t h e b a s a l l e a v e s w h ile the l e a s t w a s found in the t e r m i n a l l e a v e s . The leaves 148 f r o m t h e p l a n t s of t h e c o p p e r s e r i e s g r o w n on t h e " n o i r o n " c u l t u r e h a d a m u c h g r e a t e r a m o u n t of m a n g a n e s e p e r u n i t w e i g h t of t i s s u e th a n the s a m e c u l t u r e in the " n o c o p p e r " s e r i e s . c. A s c o r b ic A cid (1) D e s c r i p t i o n T h e w a y b y w h i c h t h e l o s s of a s c o r b i c a c i d i n t h e f r e s h a n d d r y i n g p l a n t m a t e r i a l h a d b e e n f o l l o w e d i n t h e t h i r d e x p e r i m e n t on w h e a t p r o v e d to b e s a t i s f a c t o r y a n d to y i e l d r e s u l t s i n a r a t h e r s h o r t p e r i o d of t i m e . T h e s a m e p r o c e d u r e w a s u s e d f o r t h e s t u d y of a s c o r b i c a c i d l o s s in t h e t o b a c c o l e a v e s d u r i n g d r y i n g . As d escribed b e f o r e , e a c h k i n d of l e a v e s , t h e t e r m i n a l , m e d i a n , a n d b a s a l , w e r e d i v i d e d in c o m p a r a b l e c o m p o s i t e s u b s a m p l e s w h i c h w e r e l e f t to d r y in the l a b o r a t o r y . F o r t h e s a m p l e s of t h e " n o c o p p e r s e r i e s , " t h e f i r s t d e t e r m i n a t i o n w a s d o n e a t h a r v e s t , t h e s e c o n d 24 h o u r s l a t e r , t h e t h i r d 48 h o u r s a f t e r h a r v e s t a n d t h e n 72, 120 a n d 144 hours after h arv est. F o r t h e s a m p l e s of t h e " c o p p e r s e r i e s " t h e f i r s t d e t e r m i n a t i o n w a s c a r r i e d a t h a r v e s t , t h e s e c o n d 24 h o u r s l a t e r , t h e t h i r d 4 8 h o u r s a f t e r h a r v e s t a n d t h e n 96, 120 a n d 192 h o u r s after h arvest. T h e a m o u n t s of a s c o r b i c a c i d f o u n d a t e a c h d e t e r m i n a t i o n w e r e c a l c u l a t e d o n a 100 g r a m s f r e s h m a t t e r b a s i s a n d t h e p e r c e n t a g e of t h e o r i g i n a l a m o u n t l e f t a t e a c h d e t e r m i n a t i o n w a s a l s o c o m ­ puted. (2) A m o u n t s of A s c o r b i c A c i d p e r 100 G r a m s of F r e s h M a t t e r (a) E f f e c t of i r o n i n t h e p r e s e n c e of c o p p e r . The data ex­ p r e s s e d i n m i l l i g r a m s o f a s c o r b i c a c i d p e r 100 g r a m s of f r e s h m a t t e r a t h a r v e s t a n d a t t h e v a r i o u s t i m e s o f d r y i n g a p p e a r in T a b l e 53 a n d F i g u r e s 3 4 4 t o 3 4 6 , i n c l u s i v e . In t h e p l a n t s g r o w n on b o t h c u l t u r e s s u p p l i e d w i t h i r o n th e c o n c e n t r a t i o n s of a s c o r b i c a c i d w a s g r e a t e r t h a n in t h e “ n o i r o n ” c u l t u r e a n d in a l l t h e c a s e s t h e f o l l o w i n g s e q u e n c e i n c o n c e n t r a ­ t i o n s of a s c o r b i c a c i d in t h e l e a v e s w a s a p p a r e n t : m edian T erm inal basal. No c o r r e l a t i o n w a s o b t a i n e d b e t w e e n t h e a m o u n t s of i r o n in t h e f r e s h m a t e r i a l a n d t h e a m o u n t s of a s c o r b i c a c i d f o u n d a t h a r v e s t ( s e e F i g u r e 350) . On t h e o t h e r h a n d , a p o s i t i v e c o r r e l a t i o n b e t w e e n m a n g a n e s e a n d a s c o r b i c a c i d w a s f o u n d , a s s e e n i n F i g u r e 352, w h e n t h e d a t a fo r b o th c u ltu r e s su pplied with iro n w e r e pooled. On t h e o p p o s i t e , a n e g a tiv e r e l a t i o n s h i p is o b s e r v e d in the no i r o n c u l t u r e , b u t th is f a c t s h o u l d n o t b e g e n e r a l i z e d b e c a u s e of t h e f e w d a t a a v a i l a b l e for this c u ltu re . 150 D u r i n g t h e f i r s t 48 h o u r s of d r y i n g , a g r e a t a m o u n t of th e o r i g i n a l v a l u e of a s c o r b i c a c i d w a s l o s t . T h e r a t e of l o s s w i l l b e d i s c u s s e d l a t e r in the p a r a g r a p h w h e r e the p e r c e n t a g e s of the o r i g i n a l a m o u n t s of a s c o r b i c a c i d a t e a c h d e t e r m i n a t i o n a r e r e ­ ported. (b) E f f e c t of i r o n i n t h e a b s e n c e of c o p p e r . The average v a l u e s i n m i l l i g r a m s of a s c o r b i c a c i d p e r 100 g r a m s of f r e s h t i s s u e a t h a r v e s t a n d a t t h e v a r i o u s t i m e s of d r y i n g a r e r e c o r d e d i n T a b l e 52 a n d F i g u r e s 347 to 349. In m o s t of t h e l e a v e s of t h e p l a n t s s u p p l i e d w i t h i r o n , th e c o n c e n t r a t i o n s of a s c o r b i c a c i d w e r e h i g h e r t h a n in t h e c u l t u r e d e p r i v e d of i r o n . In a l l t h e c a s e s a n d i n th e s a m e m a n n e r a s in the “ c o p p e r ” s e r i e s the s e q u e n c e : term inal m edian basal w a s f o u n d in t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d in t h e l e a v e s . N o c o r r e l a t i o n w a s o b t a i n e d b e t w e e n t h e a m o u n t of i r o n i n t h e f r e s h m a t e r i a l a n d t h e a m o u n t of a s c o r b i c a c i d f o u n d a t h a r ­ v e s t ( F i g u r e 351) . W hen the r e s u l t s fo r th e “ c o p p e r ” and the “ no c o p p e r ” s e r i e s a r e p o o l e d ( F i g u r e 351) a n d t h e d a t a f o r e a c h c a t e g o r y of le a f c o n s id e re d s e p a r a te l y , p o s itiv e r e l a tio n s h ip s a r e then o b ­ s e r v e d b etw een iro n and a s c o rb ic acid; a re la tio n s h ip is e s p e c ia lly a p p a r e n t i n t h e c a s e of t h e t e r m i n a l l e a v e s . 151 A s f a r a s m a n g a n e s e is c o n c e r n e d in the “ no c o p p e r ” s e r i e s ( F i g u r e 35 3), a s l i g h t p o s i t i v e r e l a t i o n s h i p i s o b t a i n e d . W hen the d a ta f o r the “ c o p p e r ” and the “ no c o p p e r ” s e r i e s a r e pooled, a m o r e s ig n ific a n t p o s itiv e r e la tio n s h ip a p p e a r s ( F ig u r e 35 3 ) . A s ig n if ic a n t p o s itiv e c o r r e l a t i o n w a s o b ta in e d w h en the d a ta f o r the p e r c e n t a g e s of d r y m a t t e r in the l e a v e s of the p la n ts f r o m a l l t h e c u l t u r e s a n d of b o t h c o p p e r s e r i e s a n d t h e a m o u n t of a s c o rb ic acid w e r e pooled. A t h a r v e s t m o r e a s c o r b i c a c id w a s found in a ll the c a t e ­ g o r i e s of l e a v e s of t h e “ no c o p p e r ” s e r i e s a t t h e e x c e p t i o n of t h e m e d i a n l e a v e s of t h e “ i r o n - g r a v e l ” c u l t u r e a n d t h e b a s a l l e a v e s of t h e “ no i r o n ” c u l t u r e t h a n in t h e c o r r e s p o n d i n g l e a v e s of t h e “ copper s e rie s .” D uring drying , le s s a s c o rb ic acid w as g e n e ra lly d e t e c t e d in t h e l e a v e s f r o m t h e p l a n t s of t h e “ n o c o p p e r ” s e r i e s a t t h e e x c e p t i o n of t h o s e f r o m t h e c u l t u r e d e p r i v e d of i r o n , a s c a n b e o b s e r v e d i n F i g u r e s 355 to 360, i n c l u s i v e . (3) P e r c e n t a g e s of t h e O r i g i n a l A m o u n t s of A s c o r b i c A c i d F o u n d at E a c h D eterm ination (a) E f f e c t of i r o n in t h e p r e s e n c e of c o p p e r . The values for t h e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t s of a s c o r b i c a c i d f o u n d in t h e t e r m i n a l , m e d i a n a n d b a s a l l e a v e s of t h e v a r i o u s c u l t u r e s of 152 t h e “ c o p p e r ” s e r i e s a r e l i s t e d i n T a b l e 56 a n d g r a p h e d a g a i n s t t h e t i m e of t h e v a r i o u s d e t e r m i n a t i o n s i n F i g u r e s 361 to 3 63, i n ­ clusive. A p o s i t i v e r e l a t i o n s h i p b e t w e e n t h e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t s of a s c o r b i c a c i d a n d t h e a m o u n t s of i r o n p e r u n i t w e i g h t of f r e s h m a t t e r i s a p p a r e n t i n F i g u r e s 370 t o 373, i n ­ c l u s i v e , a n d h o l d t r u e d u r i n g a l l t h e p r o c e s s of d r y i n g . These r e s u l t s a g r e e w ith th o se o b tain ed in the p r e v io u s w h e a t e x p e r i ­ m e n t , w h e r e to a g r e a t e r a m o u n t of i r o n in t h e f r e s h t i s s u e c o r ­ r e s p o n d e d a g r e a t e r s t a b i l i t y of t h e a s c o r b i c a c i d . (b) E f f e c t of i r o n i n t h e a b s e n c e of c o p p e r . The data for th e p e r c e n t a g e s of t h e o r i g i n a l a m o u n t s of a s c o r b i c a c i d f o u n d a t e a c h d e t e r m i n a t i o n s i n t h e d i f f e r e n t l e a v e s of t h e “ n o c o p p e r ” s e r i e s a r e r e p o r t e d in T a b l e 55 a n d g r a p h e d a g a i n s t t h e t i m e of t h e v a r i o u s d e t e r m i n a t i o n s i n F i g u r e s 364 to 3 66, i n c l u s i v e . I n t h e c a s e of t h e “ no c o p p e r ” s e r i e s , a s l i g h t p o s i t i v e r e l a t i o n s h i p w a s f o u n d b e t w e e n t h e c o n c e n t r a t i o n of i r o n in t h e p l a n t a n d t h e p e r c e n t a g e s of a s c o r b i c a c i d ( F i g u r e s 374 t o 377) le f t a f t e r 24 h o u r s of d r y i n g , h o w e v e r t h i s r e l a t i o n s h i p d i s a p p e a r e d at the next d e te r m in a tio n s . In o r d e r to c o m p a r e t h e r a t e of o x i d a t i o n of a s c o r b i c a c i d in t h e c o p p e r a n d t h e “ no c o p p e r ” s e r i e s , t h e d i f f e r e n c e s b e t w e e n the p e r c e n t a g e s found fo r e a c h s e r i e s a t e a c h d e t e r m in a ti o n w e r e c a l c u l a t e d a n d a r e p r e s e n t e d i n T a b l e 57 a n d F i g u r e s 36 7 t o 369. F r o m t h e s e d a t a i t c a n b e s e e n t h a t i n g e n e r a l t h e r a t e of o x i d a ­ t i o n w a s g r e a t e r i n t h e l e a v e s of t h e p l a n t s of t h e c o p p e r s e r i e s s u p p l i e d w i t h i r o n i n c o m p a r i s o n t o t h e c o r r e s p o n d i n g p l a n t s of t h e no c o p p e r s e r i e s . On th e o t h e r hand, in the p la n ts d e p r i v e d of i r o n of t h e “ c o p p e r ” s e r i e s , t h e r a t e of o x i d a t i o n of t h e a s c o r ­ bic a c id w a s s m a l l e r th a n in th e c o r r e s p o n d i n g p la n ts of the “ no copper s e rie s .” E. D i s c u s s i o n on t h e S e v e n t h E x p e r i m e n t D u r i n g t h i s e x p e r i m e n t on t o b a c c o p l a n t s , m a n y of t h e f a c t s o b s e r v e d on t h e e f f e c t s of i r o n a n d c o p p e r n u t r i t i o n on s o m e a s p e c t s of t h e p h y s i o l o g y of t h e p l a n t s , w e r e f o u n d to b e i n a g r e e m e n t w i t h th o s e a l r e a d y d i s c l o s e d d u rin g the p r e v i o u s e x p e r i m e n t s w ith o th e r plant m a te r ia ls . G r e a t e r w e i g h t s h o u l d h e n c e b e g i v e n to t h o s e re la tio n s h ip s w hich o c c u r r e d under th ese v a rio u s conditions. F o r e x a m p l e , t h e d e p r e s s i n g e f f e c t of t h e p r e s e n c e of c o p ­ p e r i n t h e n u t r i e n t s o l u t i o n on t h e a c c u m u l a t i o n of i r o n in t h e l e a v e s , e f fe c t w h ic h h ad a l r e a d y b e e n d i s c u s s e d in the s ix th e x p e r i m e n t , w a s s h o w n a g a i n to e x i s t i n t h e c a s e of t o b a c c o p l a n t s . It w as o b s e rv e d th a t when the p lan ts w e r e d e p riv e d of iro n , a h i g h e r c o n c e n t r a t i o n of m a n g a n e s e w a s f o u n d i n t h e l e a v e s f r o m 154 t h e p l a n t s of t h e c o p p e r s e r i e s t h a n f r o m t h o s e o f t h e “ n o c o p p e r ” series. T h i s f a c t c o u l d b e e x p l a i n e d b y t h e d e p r e s s i n g e f f e c t of c o p p e r o n i r o n - a c c u m u l a t i o n , w h i c h in t u r n h a s a d e p r e s s i n g e f ­ f e c t on m a n g a n e s e a c c u m u l a t i o n . This explanation, ho w ev er, is not c o m p le te ly s a t i s f a c t o r y b e c a u s e it w a s o b s e r v e d th a t this f a c t o n l y o c c u r r e d f o r t h e p l a n t s d e p r i v e d of i r o n ; a n o t h e r e x ­ p l a n a t i o n i s t h a t , t h e p l a n t s d e p r i v e d of i r o n b e i n g l a r g e r i n t h e “ no c o p p e r ” s e r i e s th a n in the “ c o p p e r ” s e r i e s , the s m a l l e r c o n ­ c e n t r a t i o n s of m a n g a n e s e i n t h e f o r m e r m i g h t b e d u e t o a d i l u t i o n effect. A s in t h e c a s e of t h e t o m a t o l e a v e s , a p o s i t i v e r e l a t i o n s h i p w a s o b t a i n e d b e t w e e n t h e a m o u n t s of a s c o r b i c a c i d a t h a r v e s t a n d t h e p e r c e n t a g e s of d r y m a t t e r i n t h e t i s s u e s . T his relatio n sh ip w a s d i s c u s s e d a t t h e e n d of t h e f i f t h e x p e r i m e n t , a n d m o r e e v i d e n c e fo r its g e n e r a l o c c u r r e n c e is g iven by th e r e s u l t s r e p o r t e d d u r in g the p r e s e n t e x p e r i m e n t . A s f a r a s the i r o n - a s c o r b i c a c id r e l a t i o n s h i p is c o n s i d e r e d , i t s h o u l d b e p o i n t e d o u t t h a t on o n e h a n d , w h e n t h e r e s u l t s f o r e a c h c a t e g o r y of l e a f a r e t a k e n s e p a r a t e l y , a p o s i t i v e r e l a t i o n s h i p i s o b t a i n e d b e t w e e n t h e c o n c e n t r a t i o n s of a s c o r b i c a c i d a t h a r v e s t a n d t h e c o n c e n t r a t i o n s of i r o n i n t h e f r e s h t i s s u e s ; o n t h e o t h e r h a n d , w hen th e s a m e r e s u l t s a r e pooled no s p e c i a l c o r r e l a t i o n c a n be draw n. It c a n t h e n b e c o n c l u d e d t h a t t h e i r o n - a s c o r b i c a c i d 155 r e l a t i o n s h i p had a c e r t a i n i m p o r t a n c e on th e a s c o r b i c a c id s t a t u s in t h e l i v i n g p l a n t s , b u t a m o r e f u n d a m e n t a l f a c t o r s e e m t o b e d i r e c t l y r e l a t e d w ith the a g e and the a c tiv ity of the t i s s u e s , b e ­ c a u s e of t h e g r e a t i n f l u e n c e of t h e p o s i t i o n o f t h e l e a v e s ( i . e . , a t t h e b a s e , t h e m i d d l e , o r t h e t i p of t h e p l a n t ) on t h e i r a s c o r b i c acid contents. T he a s c o r b i c a c id , in the d r y in g p la n t t i s s u e s w a s found to b e m o r e s t a b l e i n t h e p l a n t c o n t a i n i n g a h i g h t h a n a lo w c o n c e n ­ t r a t i o n of i r o n . This g r e a te r stab ility w as m o r e a p p a re n t b efo re f if t y p e r c e n t of t h e o r i g i n a l a m o u n t of a s c o r b i c a c i d w e r e l o s t . It sh o u ld be r e c a l l e d t h a t th is o b s e r v a t i o n had a l r e a d y b e e n m a d e in the t h i r d e x p e r i m e n t w ith w h e a t p l a n t s . An i n t e r a c t io n b e tw e e n c o p p e r a n d i r o n , r e l a t e d to a s c o r b i c a c i d s t a b i l i t y i s a p p a r e n t f r o m th e p r e s e n t d a t a , i n d e e d t h e r e l a t i o n s h i p i r o n - a s c o r b i c a c i d s t a b i l i t y a p p e a r e d in t h e p l a n t s s u p p l i e d w i t h 0 . 0 8 p a r t s p e r m i l l i o n of c o p ­ p e r w h i l e i t w a s l e s s c o n s i s t e n t i n t h e p l a n t s d e p r i v e d of c o p p e r . On t h e o t h e r h a n d , a n d i n a g r e e m e n t w i t h t h e r e s u l t s o b ­ t a i n e d in t h e w h e a t e x p e r i m e n t s , i t w a s f o u n d t h a t t h e a s c o r b i c acid f r o m the p la n ts supplied w ith c o p p e r, w as lo st a t a slo w e r r a t e t h a n t h o s e d e p r i v e d of c o p p e r . W hat co u ld b e s a id to e x p lain th e p r e s e n t f a c ts would only b e s p e c u l a t i o n , b u t a t l e a s t , t h e p o s s i b l e r o l e of i r o n a n d c o p p e r on t h e i n c r e a s e i n s t a b i l i t y of a s c o r b i c a c i d i n t h e d r y i n g p l a n t t i s s u e s sh o u ld be e m p h a s iz e d . F. 1. S u m m a r y of the S e v e n th E x p e r i m e n t T o b a c c o p l a n t s w e r e g r o w n on n u t r i e n t s o l u t i o n s , u n d e r t h r e e d i f f e r e n t c o n d i t i o n s of i r o n a v a i l a b i l i t y : on g r a v e l w i t h o u t i r o n , o n a n i r o n - f r i t , on g r a v e l s u p p l i e d w i t h f o u r p a r t s p e r m i l ­ l i o n of f e r r i c s u l f a t e i n t h e n u t r i e n t s o l u t i o n . T h e s e c o n d i t i o n s of i r o n a v a i l a b i l i t y w e r e s e t up a t tw o c o p p e r le v e ls : one s e r i e s w ith ­ o u t c o p p e r , t h e o t h e r w i t h 0 . 0 8 p a r t s p e r m i l l i o n of c o p p e r . 2. T e rm in a l, m ed ian , and b a s a l leav es w e re co llected w h e n t h e p l a n t s w e r e h a r v e s t e d o n 93 d a y s o ld p l a n t s . A scorbic a c i d w a s d e t e r m i n e d on t h e f r e s h t i s s u e s a n d a t v a r i o u s i n t e r v a l s d u r i n g t h e d r y i n g of t h e p l a n t s a t r o o m t e m p e r a t u r e , The d e te rm in a ­ t i o n s of i r o n a n d m a n g a n e s e w e r e c a r r i e d on c o r r e s p o n d i n g o v e n dried sam p les. 3. T h e p e r c e n t a g e s of d r y m a t t e r w e r e r e c o r d e d . A d e c r e a s i n g g r a d i e n t in t h e p e r c e n t a g e s of d r y m a t t e r w a s f o u n d f r o m t h e t e r m i n a l t o t h e b a s a l l e a v e s a n d e x c e p t on t h e “ n o i r o n ” c u l t u r e s t h e p e r c e n t a g e of d r y m a t t e r w a s g r e a t e r f o r th e p l a n t s f r o m the c o p p e r s e r i e s th an f o r t h o s e f r o m the “ no c o p ­ p e r ” series. 157 4. T h e a m o u n t s of i r o n p e r 100 g r a m s of f r e s h m a t e r i a l w e r e found th e h i g h e s t in th e m e d i a n l e a v e s of th e c u l t u r e s s u p p lie d w i t h i r o n of b o t h c o p p e r c u l t u r e s ; t h e p l a n t s f r o m t h e “ n o i r o n ” c u l t u r e s s h o w e d a n i n c r e a s i n g g r a d i e n t i n t h e p e r c e n t a g e s of i r o n f r o m t e r m i n a l to b a s a l l e a v e s . A g r e a t e r c o n c e n t r a t i o n of i r o n w a s found in a ll th e l e a v e s f r o m the no c o p p e r s e r i e s . The sam e r e s u l t s c a l c u l a t e d on a p e r c e n t d r y m a t e r i a l b a s i s g a v e a p p r o x ­ im a te ly the s a m e te n d e n c ie s a s r e p o rte d above. 5. T h e a m o u n t s of m a n g a n e s e p e r 100 g r a m s of f r e s h m a ­ t e r i a l of p l a n t t i s s u e s h o w e d a n i n c r e a s e f r o m t h e b a s a l to th e t e r m i n a l l e a v e s of t h e p l a n t s g r o w n on t h e c u l t u r e s s u p p l i e d w i t h iron. W h e n no i r o n w a s p r e s e n t i n th e n u t r i e n t s o l u t i o n , t h e o p p o ­ s i t e g r a d i e n t in t h e c o n c e n t r a t i o n w a s f o u n d , i . e . , t h e c o n c e n t r a ­ t i o n s in t h e b a s a l l e a v e s w a s t h e g r e a t e s t a n d i n t h e t e r m i n a l l e a v e s the l e a s t . W h e n t h e p l a n t s w e r e d e p r i v e d of i r o n , a h i g h e r c o n c e n ­ t r a t i o n of m a n g a n e s e w a s f o u n d i n t h e l e a v e s of t h e p l a n t s f r o m t h e 0 . 0 8 p a r t s p e r m i l l i o n c o p p e r s e r i e s t h a n of t h e s e of t h e no copper series. 6. In a l l t h e p l a n t s , f r o m a l l t h e c u l t u r e s , t h e g r e a t e s t c o n c e n t r a t i o n s of a s c o r b i c a c i d a t h a r v e s t w e r e f o u n d i n t h e t e r ­ m i n a l l e a v e s a n d t h e l e a s t in t h e b a s a l l e a v e s . In b o t h c o p p e r s e r i e s , th e p la n ts f r o m th e c u l t u r e s s u p p lie d w ith i r o n had the 158 m o s t a s c o r b i c a c i d a n d w h e n t h e d a t a of b o t h s e r i e s a r e p o o l e d a n d e a c h c a t e g o r y of l e a f c o n s i d e r e d a p o s i t i v e c o r r e l a t i o n i s f o u n d b e t w e e n th e i r o n a n d t h e a s c o r b i c a c i d . At harvest, m ore a s c o r b i c a c i d w a s f o u n d , in m o s t c a s e s in t h e l e a v e s f r o m th e p l a n t s of t h e “ no c o p p e r ” s e r i e s , b u t d u r i n g d r y i n g l e s s a s c o r b i c a c id w a s found in th is s a m e s e r i e s . 7. T h e p e r c e n t a g e s o f t h e o r i g i n a l a m o u n t s of a s c o r b i c a c id found a t e a c h d e t e r m i n a t i o n , d u r in g d r y in g show ed a p o s itiv e r e l a t i o n s h i p w i t h t h e c o n c e n t r a t i o n of i r o n i n t h e f r e s h t i s s u e s of the p la n ts f r o m the “ c o p p e r ” s e r i e s . In t h e p l a n t s f r o m t h e “ no c o p p e r ” s e r i e s , the s a m e r e la tio n s h ip w as not so a p p a r e n t and d i s a p p e a r e d a f t e r 24 h o u r s of d r y i n g . A g r e a t e r r a t e of o xidation of t h e a s c o r b i c a c i d w a s f o u n d i n th e l e a v e s f r o m t h e p l a n t s of t h e “ c o p p e r ” s e r i e s s u p p l i e d w i t h i r o n t h a n in t h o s e of t h e “ no c o p ­ p e r ” series. B u t , w h e n t h e p l a n t s w e r e d e p r i v e d of i r o n , t h e r a t e of o x i d a t i o n w a s s m a l l e r in th e l e a v e s f r o m t h e “ c o p p e r ” s e r i e s t h a n f r o m t h o s e of t h e “ no c o p p e r ” s e r i e s . X. G E N E R A L DISCUSSION T h e c o m p a r a t i v e s t u d y of t h e e f f e c t s of v a r i o u s c o n c e n t r a ­ t i o n s of c o p p e r a n d i r o n , in t h e n u t r i e n t m e d i a a n d i n t h e p l a n t t i s ­ s u e s on t h e c o n c e n t r a t i o n a n d s t a b i l i t y of a s c o r b i c a c i d i n p l a n t s b r o u g h t a b o u t c e r t a i n f a c t s w h i c h s e e m w o r t h y of b e i n g e m p h a s i z e d a t t h e e n d of t h i s i n v e s t i g a t i o n . Q uantitative c o m p a riso n s betw een the d a ta o b ta in e d by the d i f f e r e n t e x p e r i m e n t s h a v e b e e n av o id e d b e c a u s e t h e p l a n t s , a l t h o u g h s u p p l i e d w i t h n u t r i e n t s o l u t i o n s of c o n t r o l l e d c o m p o s i t i o n s , w e r e g r o w n a t d i f f e r e n t p e r i o d s of t h e year. C o n s e q u e n tly the c l i m a t i c f a c t o r s know n to a ffe c t th e a b ­ s o r p t i o n of t h e i n o r g a n i c n u t r i e n t a n d t h e s y n t h e s i s of a s c o r b i c a c id v a r ie d b etw een wide lim its . On t h e o t h e r h a n d , a f e w g e n ­ e r a l q u a l i t a t i v e t r e n d s w e r e found in the v a r i o u s e x p e r i m e n t s . A s a g e n e r a l c o n c lu sio n , it can be sa id that a lth o u g h c o p p e r a n d i r o n a r e p o w e r f u l c a t a l y s t s of t h e o x i d a t i o n of a s c o r b i c a c i d i n v i t r o , t h e p r e s e n c e of t h e s e e l e m e n t s in t h e p l a n t s d i d n o t i n ­ c r e a s e t h e r a t e of o x i d a t i o n of t h e a s c o r b i c a c i d in t h e d r y i n g p l a n t tissues. S u r p r i s i n g l y , i t w a s g e n e r a l l y f o u n d t h a t t h e s t a b i l i t y of t h e a s c o r b i c a c i d in t h e s e t i s s u e s w a s p o s i t i v e l y r e l a t e d to t h e a m o u n t s of i r o n and c o p p e r in the n u t r i e n t so lu tio n . A t the s a m e t i m e , i t w a s s u g g e s t e d t h a t a p o s s i b l e i n t e r a c t i o n b e t w e e n i r o n an d 160 c o p p e r w a s involved in the a s c o r b i c a c id s y s t e m of p la n ts . A p o s itiv e r e l a t i o n s h i p b e tw e e n the a s c o r b i c a c id s t a b i l i t y and the i r o n c o n t e n t of t h e p l a n t t i s s u e w a s m o r e a p p a r e n t i n t h e p l a n t s s u p p l i e d w i t h v a r i o u s a m o u n t s of i r o n t h a n in t h o s e s u p p l i e d w i t h v a r i o u s a m o u n t s of c o p p e r . T h e m e c h a n i s m i n v o l v e d in t h e r e l a t i o n b e t w e e n i r o n , c o p ­ p e r , and a s c o r b i c a c id in p l a n t a r e n o t know n, i t i s p o s s i b l e , h o w ­ e v e r , t h a t c o p p e r an d i r o n m a y f o r m c o m p l e x e s o f o r g a n i c n a t u r e a f t e r t h e i r a b s o r p tio n by the p la n t. It h a s b e e n sh o w n in the l i t ­ e r a t u r e t h a t the p r o te in f r a c t i o n s e a s i l y c o m p l e x w ith i r o n and copper. A c h a n g e in t h e p r o p e r t i e s of i r o n i s i l l u s t r a t e d b y t h e o x i d a t i o n - r e d u c t i o n p o t e n t i a l of F e ++ —> F e +** - e w h i c h i s - 0 7 7 v o l t s , w h i l e t h e p o t e n t i a l of t h e s a m e r e a c t i o n i s - 1 . 1 4 v o l t s w h e n th e i r o n i s c o m p l e x e d w i t h o r t h o p h e n a n t h r o l e n e . I t i s e v i d e n t t h a t i t i s n o t p o s s i b l e to p r e d i c t t h e a c t i o n of m e t a l s s u c h a s c o p p e r and i r o n in o x i d a t i o n - r e d u c t i o n r e a c t i o n s in r e l a t i o n to t h e a s c o r b i c a c i d s t a t u s of p l a n t s u n t i l d e t a i l e d know ledge is av ailab le c o n c e rn in g the p a r t i c u l a r m e ta llic co m p o u n d s involved. B u t t h e c h a n g e s of s o m e p r o p e r t i e s of t h e m e t a l s w h e n t r a n s f o r m e d in to c o m p le x o r g a n ic c o m p o u n d m a y e x p la in the i n ­ c r e a s e d s t a b i l i t y of t h e a s c o r b i c a c i d o b t a i n e d w i t h i n c r e a s e s of i r o n a n d c o p p e r c o n c e n t r a t i o n s in t h e p l a n t t i s s u e s . 161 D u r i n g t h i s i n v e s t i g a t i o n , a s i g n i f i c a n t i n c r e a s e i n th e g r a d i e n t of a s c o r b i c a c i d w a s o b s e r v e d f r o m t h e o ld , b a s a l l e a v e s to t h e y o u n g , t e r m i n a l l e a v e s of t o b a c c o a n d t o m a t o p l a n t s . Two m a i n f a c t o r s m a y b e t h e c a u s e of t h i s c h a n g e in c o n c e n t r a t i o n : (1) a g r e a t e r s t a b i l i t y o f t h e a s c o r b i c a c i d i n t h e h i g h e r p a r t of t h e p l a n t , a n d (2) a g r e a t e r s y n t h e t i c p o w e r of t h a t p a r t of t h e p l a n t . I n o r d e r t o t e s t t h e s t a b i l i t y of a s c o r b i c a c i d i n t h e p l a n t t i s s u e s , d e t e r m i n a t i o n s w e r e c a r r i e d o u t a t d i f f e r e n t i n t e r v a l s d u r i n g th e d r y i n g of t h e t i s s u e s . I t w a s a s s u m e d t h a t t h e s t a b i l i t y of th e a s c o r b i c a c i d w o u l d t h e n b e r e l a t i v e l y c o m p a r a b l e t o t h a t of l i v i n g tissue. It should, h o w e v e r , be r e a l i z e d how d i f f e r e n t the c o n d i­ t i o n s c a n b e in b o t h c a s e s a n d h o w l i m i t e d t h i s a s s u m p t i o n i s ; b u t no p r e s e n t t e c h n i q u e o f f e r s a n y i n f o r m a t i o n a s to t h e s t a b i l i t y of a s c o r b i c a c i d in l i v i n g t i s s u e s , b e c a u s e i t s a c t i v e s y n t h e s i s w i l l m a s k t h e a c t u a l s t a t u s of i t s s t a b i l i t y . It w a s found f r o m s ta b ility s t u d i e s p e r f o r m e d d u r i n g t h e d r y i n g of d i f f e r e n t p l a n t p a r t s c o n ­ t a i n i n g v a r y i n g a m o u n t s of a s c o r b i c a c i d , t h a t t h e r a t e of o x i d a ­ t i o n w a s a b o u t a s g r e a t a n d s o m e t i m e s g r e a t e r in t h e t e r m i n a l l e a v e s t h a n in t h e b a s a l l e a v e s . The f ir s t facto r m entioned as a c a u s e f o r g r e a t e r a c c u m u l a t i o n of a s c o r b i c a c i d i n t h e t e r m i n a l p a r t s of t h e p l a n t s w a s t h u s e l i m i n a t e d . On the o th e r hand it w a s o b s e r v e d c o n s i s t e n t l y t h a t th e a m o u n t s of a s c o r b i c a c id found a t h a r v e s t w e r e p o s i t i v e l y r e l a t e d to t h e p e r c e n t a g e of d r y m a t t e r in 162 the t i s s u e . I t i s , t h e r e f o r e , p o s s i b l e to s u g g e s t t h a t t h e a s c o r b i c a c i d c o n t e n t of p l a n t t i s s u e s m a y b e p o s i t i v e l y r e l a t e d to t h e a c ­ t i v i t y of t h e c e l l s . T h e p e r c e n t a g e of d r y w e i g h t h a s b e e n s h o w n b y R e i d ( 1 9 4 1 a ) to b e c o r r e l a t e d , i n y o u n g t i s s u e s , to a g r e a t e r a c ­ t i v i t y of t h e c e l l s . T h i s a s p e c t o f t h e p r o b l e m w o u l d t h e n b e in a g r e e m e n t w i t h t h e r e s u l t s of R e i d ( 1 9 4 1 b ) , w h o f o u n d a c o r r e l a t i o n b e t w e e n d r y w e i g h t , a s c o r b i c a c i d , a n d t h e a c t i v i t y of t h e c e l l s in c o w p e a s e e d l i n g s ; a n d w i t h t h o s e of S h a w a n d P a s c o e ( 1 9 4 9 ) , w ho r e p o r t e d a r a p i d s y n t h e s i s of a s c o r b i c a c i d i n t h e b r o a d b e a n d u r ­ i n g t h e f i r s t d a y s of g e r m i n a t i o n a n d f o u n d t h e g r e a t e s t c o n c e n t r a ­ t i o n of a s c o r b i c a c i d in t h e m o s t a c t i v e p a r t s of t h e s e e d l i n g s . F r o m t h i s i n f o r m a t i o n , i t i s r a t h e r d i f f i c u l t to g i v e a n y e x p l a n a t i o n of t h e r e s u l t s r e p o r t e d e a r l i e r in t h e i n t r o d u c t i o n , w h i c h s h o w e d a g r e a t e r c o n c e n t r a t i o n of a s c o r b i c a c i d r e l a t e d to a l o w e r c o n c e n t r a t i o n of i r o n in t h e t i s s u e s . It is p o ssib le , how­ e v e r , t h a t u n d e r s p e c i f i c c o n d i t i o n s , a l u x u r i o u s g r o w t h d u e to t h e g r e a t a c t i v i t y of d i v i d i n g a n d e n l a r g i n g c e l l w i l l c o r r e s p o n d to a d i l u t i o n of t h e t o t a l i r o n p r e s e n t . T h e l a r g e a m o u n t of a s c o r b i c a c i d f o u n d c o u l d b e a c a u s e o r a n e f f e c t of t h e g r e a t a c t i v i t y of this tiss u e . F r o m t h e r e s u l t s of a n a l y s e s , i t w o u l d t h e n s e e m t h a t t o a s m a l l a m o u n t of i r o n c o r r e s p o n d e d a g r e a t a m o u n t of a s c o r b i c acid , but th is c o r r e l a t i o n would not be a tr u e c a u s e o r e ffe c t r e l a ­ tionship. 163 A l t h o u g h t h e s t u d y of t h e r e l a t i o n s h i p b e t w e e n a s c o r b i c a c i d , i r o n , a n d c o p p e r w a s t h e m a i n p u r p o s e of t h i s i n v e s t i g a t i o n , so m e in te ra c tio n betw een co p p e r, iro n and m a n g a n e se w e re also observed. At th is point, a d iff e r e n tia tio n should b e m a d e b e tw e e n t h e i n t e r a c t i o n s of i o n s in t h e n u t r i e n t s o l u t i o n f r o m t h e i r i n t e r ­ a c t i o n s i n th e p l a n t t i s s u e . The o x id a tio n -re d u c tio n potential, w h i c h w a s s h o w n b y E r k a m a (1 949) to h a v e a d i r e c t e f f e c t on t h e a b s o r p t i o n of h e a v y m e t a l s , i s a t a s t a t e of e q u i l i b r i u m i n the n u t r i e n t s o l u t i o n ; w h i l e a n e q u i l i b r i u m n e v e r e x i s t s in t h e r e d o x s y s t e m s of t h e l i v i n g c e l l s , t h u s t h e s a m e i n t e r a c t i o n s w i l l n o t b e apparent. It w a s f o u n d t h a t , in t o m a t o a n d t o b a c c o p l a n t s , t h e p r e s e n c e of c o p p e r i n t h e s o l u t i o n e x e r t e d a d e p r e s s i n g e f f e c t o n th e a c c u m u l a t i o n of i r o n in t h e l e a v e s a n d t h a t a h i g h c o n c e n t r a t i o n of i r o n i n t h e s o l u t i o n c o r r e s p o n d e d to a low a c c u m u l a t i o n of m a n ­ g a n e s e in th e p l a n t s . Such an i n t e r a c t i o n , h o w e v e r did n o t e x i s t i n th e l e a v e s , w h e r e h i g h a m o u n t s o f i r o n w e r e d e t e c t e d i n th e p r e s e n c e of w i d e l y v a r y i n g a m o u n t s of m a n g a n e s e . X.I. G E N E R A L S U M M A R Y AND C O N C L U S I O N 1. W h e a t , t o m a t o a n d t o b a c c o p l a n t s w e r e g r o w n on n u ­ t r i e n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s c o n c e n t r a t i o n s of c o p p e r and i r o n in o r d e r to f i n d o u t th e p o s s i b l e e f f e c t o f t h e s e n u t r i e n t on t h e c o n c e n t r a t i o n a n d on t h e s t a b i l i t y of a s c o r b i c a c i d in p l a n t tissues. 2. In m o s t c a s e s , a g r e a t e r s t a b i l i t y o f a s c o r b i c a c i d in th e d r y i n g p l a n t t i s s u e s w a s a s s o c i a t e d w i t h a g r e a t e r c o n c e n t r a t i o n of c o p p e r a n d i r o n in t h e n u t r i e n t s o l u t i o n . B u t no g e n e r a l c o r r e l a ­ t i o n b e t w e e n t h e c o n c e n t r a t i o n of a s c o r b i c a c i d a t h a r v e s t a n d th e c o n c e n t r a t i o n s of c o p p e r a n d i r o n in t h e n u t r i e n t s o l u t i o n w a s a p ­ p a r e n t f r o m the d a ta o b ta in e d . 3. The p o s s ib ility of an in te r a c tio n b e tw e e n c o p p e r and ir o n w a s s u g g e s t e d b e c a u s e a p o s itiv e r e l a t i o n s h i p b e tw e e n the a m o u n t of i r o n in t h e p l a n t a n d th e s t a b i l i t y of a s c o r b i c a c i d w a s m o r e a p ­ p a r e n t in p l a n t s s u p p l i e d w i t h v a r i o u s a m o u n t s of i r o n t h a n w i t h v a r i o u s a m o u n t s of c o p p e r i n t h e n u t r i e n t s o l u t i o n . 4. A g r e a t e r c o n c e n t r a t i o n o f a s c o r b i c a c i d w a s f o u n d in the young a c tiv e t i s s u e s th a n in the old m a t u r e t i s s u e s . The la r g e r c o n c e n t r a t i o n of a s c o r b i c a c i d w a s n o t r e l a t e d to t h e s t a b i l i t y o f 165 t h i s v i t a m i n d u r i n g th e d r y i n g of t h e t i s s u e . It c o u ld b e c o n c lu d e d , t h e r e f o r e , t h a t t h e p o w e r of s y n t h e s i s o f t h e t i s s u e s to f o r m a s c o r ­ b i c a c i d i s th e m o s t i m p o r t a n t f a c t o r g o v e r n i n g i t s c o n c e n t r a t i o n in t h e l i v i n g p l a n t t i s s u e s . 5. T h e p r e s e n c e of c o p p e r i n t h e n u t r i e n t s o l u t i o n w a s f o u n d to e x e r t a d e p r e s s i n g e f f e c t on t h e a c c u m u l a t i o n of i r o n i n to m ato and to b acco p la n ts. I t w a s a l s o f o u n d t h a t to a h i g h c o n ­ c e n t r a t i o n of i r o n i n t h e s o l u t i o n c o r r e s p o n d e d a l o w a c c u m u l a t i o n of m a n g a n e s e i n t h e p l a n t s . H o w e v e r , n e i t h e r of t h e s e c o n d i t i o n s c o u l d b e r e l a t e d to t h e c o n c e n t r a t i o n of a s c o r b i c a c i d i n t h e f r e s h t i s s u e s o r to i t s s t a b i l i t y d u r i n g d r y i n g . XJLI. BIBLIO GRAPHY A b e r g , B . , a n d I. E k d a h l . E f f e c t s of N i t r o g e n f e r t i l i z a t i o n on t h e a s c o r b i c a c i d c o n t e n t of g r e e n p l a n t s . P h y s . P l a n t . 1 : 2 9 0 329. 1948. B a r r o n , E . S. G , , A . G. B a r r o n , a n d F . K l e m p e r e r . S t u d i e s on b i o l o g i c a l o x i d a t i o n s . VII. T h e o x i d a t i o n of a s c o r b i c a c i d i n b i o l o g i c a l f l u i d s . J o u r . B i o l . C h e m . 1 1 6 : 5 6 3 - 5 7 3 . 1936. B a r r o n , E . S. G . , R . H. D e M e i o , a n d F . K l e m p e r e r . S t u d i e s on b i o l o g i c a l o x i d a t i o n s . V. C o p p e r a n d h e m o c h r o m o g e n s a s c a t a l y s t s f o r t h e o x i d a t i o n of a s c o r b i c a c i d . T h e m e c h a n i s of t h e o x i d a t i o n . J o u r . B i o l . C h e m . 1 1 2 : 6 2 5 - 6 4 0 . 1936. B e r n s t e i n , L . , K. C . H a m n e r , a n d R . Q. P a r k s . T h e i n f l u e n c e of m i n e r a l n u tritio n , so il f e r tility , and c l i m a t e on c a r o t e n e a n d a s c o r b i c a c i d c o n t e n t of t u r n i p g r e e n s . P l a n t P h y s i o l . 2 0 : 5 4 0 - 5 7 2 . 1945. B e s s e y , O. A . A m e t h o d f o r t h e d e t e r m i n a t i o n of s m a l l q u a n t i t i e s of a s c o r b i c a c i d a n d d e h y d r o a s c o r b i c a c i d i n t u r b i d o r c o l o r e d s o l u t i o n s i n t h e p r e s e n c e of o t h e r r e d u c i n g s u b ­ s t a n c e s . J . B i o l . C h e m . 1 2 6 : 7 7 1 - 7 8 4 . 1938. B i d d u l p h , O. S t u d i e s of c h l o r o s i s u s i n g r a d i o i r o n a n d r a d i o p h o s ­ p h o r u s . P r o c e e d i n g s of t h e A u b u r n c o n f e r e n c e on th e u s e of r a d i o a c t i v e i s o t o p e s . i n a g r i c u l t u r a l r e s e a r c h . A l a b a m a P o l y t e c h n i c I n s t i t u t e . 9 0 - 1 0 2 . 1947. B i t c o v e r , E . H . , a n d D. H. S i e l i n g . E f f e c t of v a r i o u s f a c t o r s on t h e u t i l i z a t i o n of n i t r o g e n a n d i r o n b y s p i r o d e l a P o l y r h i z a ( L . ) S c h l e i d P l a n t P h y s i o l . £ 6 : 2 9 0 - 3 0 3 . 1951. C a r r o l l , G. H. T h e r o l e o f a s c o r b i c a c i d in p l a n t n u t r i t i o n . B o t . R e v . 9 : 4 1 - 4 8 . 1943. The C o m a r , C. L . , E . J . B e n n e , a n d E . K. B u t e y n . C a l i b r a t i o n of a p h o t o e l e c t r i c c o l o r i m e t e r f o r t h e d e t e r m i n a t i o n of c h l o r o p h y ll . Ind. E n g . C h e m . A n a l. E d . 1 5 :5 2 4 -5 2 6 . 1943. 167 D u t c h e r , R_ A . F a c t o r s i n f l u e n c i n g t h e v i t a m i n c o n t e n t of f o o d s . P e n n . A g r . E x p t . S t a . B u l l . 2 7 5 : 1 - 2 4 . 1 9 32. E u l e r , H. V . , K. M y r b a c k , a n d H. L a r s s o n . S a u e r s t o f f a u f n a h m e d u r c h V i t a m i n C - h a l t i g e O r g a n e un d u r c h G l u c o - R e d u k t o n H o p p e - S e y . Z e i t . f. P h y s i o l . C h e m . 2 1 7 : 1 - 2 2 . 1933. E r k a m a , J . O n t h e e f f e c t o f c o p p e r a n d m a n g a n e s e on t h e i r o n s t a t u s of h i g h e r p l a n t s . T r a c e e l e m e n t s i n p l a n t p h y s i o l o g y . W a lth a m , M a s s . C h r o n i c a B o ta n ic a C o m p a n y publ. H am d allah , A. E. W. V itam in C - G e h a lt E i s e n - bzw . M a g n e s iu m f r e i g e z o g e n e r P f l a n z e n . P r o t o p l a s m a 3 2 : 3 1 - 4 3 . 1939. H a m n e r , K. C . , a n d L . A . M a y n a r d . F a c t o r s i n f l u e n c i n g t h e n u ­ t r i t i v e v a l u e of t o m a t o . USDA M i s c . B u l l . 5 0 2 : 1 - 2 3 . 194 2. H ein ze, P . H., and A. E . M u r n e e k . C o m p a r a t i v e a c c u r a c y and e f f i c i e n c y i n d e t e r m i n a t i o n of c a r b o h y d r a t e s in p l a n t m a ­ t e r i a l s . M o . A g r . E x p . S t a . , R e s . B u i . 3 1 4 , 1940. H e s s , A . F . , a n d L . J . U n g e r . T h e d e s t r u c t i o n of t h e a n t i s c o r b u t i c v i t a m i n in m i l k b y t h e c a t a l y t i c a c t i o n of m i n u t e a m o u n t s of c o p p e r . P r o c . S o c . E x p t . B i o l . M e d . 1 9 : 1 1 9 - 1 2 0 . 1921. H e s t e r , J. B. M a n g an ese and vitam in C. S c ie n c e 93:401. 1941. H o p k i n s , E . F . T h e n e c e s s i t y a n d f u n c t i o n o f m a n g a n e s e in t h e g r o w t h of C h l o r e l l a s p . S c i e n c e 7 2 : 6 0 9 - 6 1 0 . 1930 . H u m m e l l , F . C . , a n d H. H. W i l l a r d . D e t e r m i n a t i o n of i r o n i n b i o l o g ­ i c a l m a t e r i a l s . T h e u s e of O - P h e n a n t h r o l i n e . J o u r . I n d . E n g . 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D e t e r m i n a t i o n of e x c h a n g e a b l e b a s e s i n s o i l s . C h e m . , A n a l . E d . j_3: 4 3 6 " 4 4 1 - I nd. E n g . 194 1 - P o n t i n g , J . D. E x t r a c t i o n of a s c o r b i c a c i d f r o m p l a n t m a t e r i a l s . R e l a t i v e s u i t a b i l i t y of v a r i o u s a c i d s . I n d . E n g . C h e m . , A n a l . E d . j _ 5 : 3 8 9 - 3 9 1 . 1943. R a n d o i n , L . a n d L e G a l l i c , P . S u r l ’e x i s t e n c e d e c o r r e l a t i o n s e n t r e l e s t e n e u r s e n p r o t e i n e s , en f e r e t en v i t a m i n e s d e s t i s s u s v e g e t a u x . B u l l . S o c . C h e m . B i o l . 2 2 : 5 9 3 - 6 0 7 . 1940. 169 R e e d m a n , E . J . , a n d E . W. M c H e n r y . C o m b i n e d a s c o r b i c a c i d i n p l a n t t i s s u e s . B i o c h e m . Jo.ur. 3 2 : 8 5 - 9 3 . 1938. R e e d s , J . F . , a n d R . W. C u n n i n g s . D e t e r m i n a t i o n of c o p p e r in p l a n t m a t e r i a l u s i n g t h e d r o p p i n g m e r c u r y e l e c t r o d e . I nd. E n g . C h e m . , A n . E d . J_3; 12 4 “ 12 7 * 1941. R e i d , M . E . L o c a l i z a t i o n of a s c o r b i c a c i d i n t h e c o w p e a p l a n t a t d i f f e r e n t p e r i o d s of d e v e l o p m e n t . A m e r . J o u r . B o t . 24: 4 4 5 - 4 4 7 . 1937. R e i d , M . E . A s t u d y of p h y s i c a l a n d c h e m i c a l c h a n g e s i n t h e g r o w i n g r e g i o n o f p r i m a r y r o o t s of c o w p e a s e e d l i n g s . A m e r . J o u r . B o t . 2 8 : 4 5 - 5 1 . 1941. (a) R e i d , M . E . R e l a t i o n of v i t a m i n C to c e l l s i z e i n t h e g r o w i n g r e g i o n of t h e p r i m a r y r o o t of c o w p e a s e e d l i n g s . A m e r . J o u r . B o t . 2 8 : 4 1 0 - 4 1 5 . 1941. (b) R e i t e m e i e r , R . F . S e m i m i c r o - a n a l y s i s of s a l i n e s o i l s o l u t i o n s . I n d . E n g . C h e m . A n a l . E d . 1 5 : 3 9 3 - 4 0 2 . 1943. R o y , W. R . , a n d G . M . B a h r t . T h e e f f e c t of z i n c , i r o n , m a n g a n e s e and m a g n e s i u m a p p lie d to f r e n c h e d and b r o n z e d o r a n g e g r o v e s , on t h e v i t a m i n C c o n t e n t of o r a n g e s . P r o c . F l a . S t a t e H o r t . S o c . 5^3:34-38. 1940. R u d r a , M. N. R o l e of m a n g a n e s e in t h e b i o l o g i c a l s y n t h e s i s of a s c o r b i c a c i d . N a t u r e 143:81 1. 1939. S h a w , A. C . , a n d L . C. P a s c o e . F o r m a t i o n a n d d i s t r i b u t i o n of v i t a m i n C in t h e r a d i c l e a n d c o t y l e d o n of t h e b r o a d b e a n , ( V i c i a f a b a ) . N a t u r e 1 6 4 : 6 2 4 . 1949. S i d e r i s , C. P . M a n g a n e s e i n t e r f e r e n c e i n t h e a b s o r p t i o n a n d t r a n s ­ l o c a t i o n of r a d i o a c t i v e i r o n ( F e 5 9) i n A n a n a s C o m o s u s (L) M e r r . P l a n t P h y s i o l . 2 5 : 3 0 7 - 3 2 1 . 1 95 0. S o m e r s , G. F . , a n d W. C. K e l l y . A s c o r b i c a c i d a n d d r y m a t t e r a c c u m u la tio n in T u r n ip and B r o c c o l i le a f d i s c s a f t e r in f i l ­ tr a tio n w ith in o rg a n ic s a lts , o rg a n ic a c id s , and s o m e e n z y m e i n h i b i t o r s . P l a n t P h y s i o l . 2 6 : 9 0 - 1 0 9 . 1951. ■I)'--'? 170 S o m e r s , I. I ., a n d J . W. S h i v e . T h e i r o n - m a n g a n e s e r e l a t i o n in p l a n t m e t a b o l i s m . P l a n t P h y s i o l . 1 7 : 5 8 2 - 6 0 2 . 1942. S p e n c e r , E . L,., I n h i b i t i o n of i n c r e a s e a n d a c t i v i t y of t o b a c c o m osaic v iru s under n itro g en -d eficien t conditions. P la n t P h y s i o l . J i 6 : 2 2 7 - 2 3 9 . 1941. S t o l o f f , L . S. C o l o r i m e t r i c m e t h o d f o r p h o s p h a t e s . M o d i f i c a t i o n of t h e A . O . A . C . m o l y b d e n u m b l u e m e t h o d . I n d . E n g . C h e m . , A n a l . E d . L4:6 3 6 - 6 3 7 . 1942. S t o t z , E . , H a r r e r , C . J . , a n d K i n g , C . G. A s t u d y of “ a s c o r b i c a c i d o x i d a s ’e ” in r e l a t i o n to c o p p e r . J o u r . B i o l . C h e m . U _ 9 :5 1 1 - 5 2 2 . 1937. S t r o m m e , E . R . A v a i l a b i l i t y to p l a n t s of i r o n a n d m a n g a n e s e in g l a s s y f r i t s . ( T h e s i s ) M i c h i g a n S t a t e C o l l e g e . 1951. S z e n t - G y o r g y i , A. O b s e r v a t i o n s on t h e f u n c t i o n of p e r o x i d a s e on t h e c h e m i s t r y of t h e a d r e n a l c o r t e x . D e s c r i p t i o n of a n e w c a rb o h y d ra te d e riv a tiv e . B iochem . Jo u r. 22:1387-1409. 1928. T h o m p s o n , J . B . , R . B. K o c h e r , a n d H. W. F r i t z s c h e . A b r o w n i n g . r e a c t i o n i n v o l v i n g c o p p e r - p r o t e i n s . A r c h . B i o c h e m . 18: 4 1 - 4 9 . 1948, W a t s o n , S. A . , a n d G . R.- N o g g l e . E f f e c t of m i n e r a l d e f i c i e n c i e s u p o n t h e s y n t h e s i s of R i b o f l a v i n a n d a s c o r b i c a c i d b y t h e o a t p l a n t . P l a n t p h y s i o l . 2 2 : 2 2 8 - 2 4 3 . 1947. W i l l i s , L . G. E v i d e n c e of t h e s i g n i f i c a n c e of o x i d a t i o n r e d u c t i o n e q u i l i b r i u m in so il f e r t i l i t y p r o b le m . Soil Sci. Soc. A m . P r o c . 2 : 2 9 1 - 2 9 7 . 1936. W y n d , F . L,. R e l a t i o n s h i p s b e t w e e n d r y w e i g h t , p r o t e i n , a n d v i t a ­ m i n s in l e a v e s of c e r e a l s . F o o d R e s e a r c h 1 1 : 1 9 5 - 2 0 2 . 1946. W y n d , F . L . A v a i l a b i l i t y to w h e a t p l a n t s of i r o n in v e r y i n s o l u b l e g l a s s f r i t s . L l o y d i a J_4:1- 3 3 . 1951. W y n d , F . L . , a n d G . R . N o g g l e . T h e e f f e c t s of f e r t i l i z e r t r e a t m e n t s on t h e v i t a m i n C c o n t e n t of o a t s h a r b e s t e d a t t h e j o i n t i n g s ta g e (u n p u b lish ed ) (1950a). 171 W y n d , F . L . , a n d G . R . N o g g l e . C h e m i c a l c o m p o s i t i o n of R y e g r o w n o n d i f f e r e n t s o i l t y p e s in O n t a r i o , C a n a d a . P a r t VII, R e l a t i o n s h i p s b e t w e e n c o n c e n t r a t i o n s of a s c o r b i c a c i d a n d o t h e r c o m p o n e n t s i n t h e p l a n t . L l o y d i a 13 (3): 1 9 1 - 1 9 5 . (1950b). | X II. APPENDIXES. ANALYTICAL PROCEDURES 1 . Ashing o f the samples. REAGENTS H ydrofluoric a c id . Concentrated S u lfu ric a c id . Concentrated N itr ic a c id . O il N. PROCEDURE P lace one-gram o f d ry p la n t m a te ria l in to a platinum crucible* Place in to a m uffle oven and in c re a se g rad u ally th e tem perature u n t i l th e sample s t a r t s smoking. Ash a t 850 o C. f o r two hours. Let c o o l. Add 1 m i l l i l i t e r o f 1 :4 s u lf u r ic acid and f i l l th e c ru c ib le h a lf f u l l w ith h y d ro flu o ric a c id . Evaporate to dryness on hot p la te a t in term ed iate h e a t. I n f r a red lamps may be sused to step up th e evaporation of th e h y d ro flu o ric a c id . D issolve th e ash i n 0*1 n i t r i c a c id . Make up to volume in 200 m i l l i l i t e r s volum etric f la s k w ith n i t r i c acid 0 .1 N. 1 , Determ ination o f iro n Iro n was determined in a liq u o ts o f th e ash so lu tio n by th e procedure o f Hunanell and W illard (1938) REAGENTS A cetic a c id . Approximately 2 N s o lu tio n . D ilu te 114 m i l l i l i t e r s of g la c ia l a c e tic acid to one l i t e r w ith d i s t i l l e d w ater. H ydrochloric a c id . 1:1 d ilu tio n H ydrochloric a c id . 1:100 d ilu tio n Ammonium c i t r a t e . 1 per cent so lu tio n Bromophenol blue in d ic a to r s o lu tio n . 0 .4 p er c e n t. Grind 1 gram of bromophenol blue w ith 3 m i l l i l i t e r s of 0.05 N sodium hydroxide, and d ilu te t o 250 m i l l i l i t e r s with w ater. B uffer s o lu tio n s . 1 . S o lu tio n of pH 3*5. Mix 6 .4 m i l l i l i t e r s o f 2 N sodium a c e ta te w ith 93*6 m i l l i l i t e r s of 2 N a c e tic ac id , and d ilu te to 1 l i t e r . 2 . S olution of pH 4*5* Mix 43 m i l l i l i t e r s o f 2 N sodiun a c e ta te w ith 57 m i l l i l i t e r s o f 2 N a c e tic a c id , and d ilu te to 1 l i t e r . Hydroquinone s o lu tio n . D issolve 1 gram of hydroquinone in 100 m i l l i l i t e r s of a b u ffe r so lu tio n of pH 4*5* S to re in a r e f r ig e r a to r . D iscard as soon as c o lo r develops. O -phenanthroline s o lu tio n . D issolve 1 gram of o-phenanthroline monohydrate i n d i s t i l l e d w ater. Warm i f necessary to e f f e c t so lu tio n , and d ilu te to 200 m i l l i l i t e r s . Sodium a c e ta te . 2 M. D issolve 272 grams o f sodium a c e ta te tr ih y d r a te in d i s t i l l e d w ater and d ilu te to one l i t e r . I ro n . Standard so lu tio n D issolve 1.000 gram of e l e c tr o ly tic ir o n in 50 m i l l i l i t e r s of a te n p er cent s u lf u r ic acid so lu tio n . Warm i f necessary to h asten re a c tio n . Cool, and d ilu te to one l i t e r w ith d i s t i l l e d w ater. One m i l l i l i t e r contains one m illigram of iro n . PROCEDURE. 1 .P ip e t 15 m i l l i l i t e r s o f th e ash s o lu tio n , rep resen tin g 0.075 grams o f dry p la n t m a te ria l, in to a 25 m i l l i l i t e r s volum etric f la s k , and a s im ila r a liq u o t in to a 25 m i l l i l i t e r s beaker. The a liq u o t should co n tain from 0.01 to 0.10 m illigram s of ir o n . 2 . Add 5 drops of bromophenol b lu e in d ic a to r to th e a liq u o t i n th e b eaker, and t i t r a t e th e so lu tio n w ith 2 N sodirna a c e ta te u n t i l th e co lo r matches th a t of an equal volume of th e b u ffe r whose pH i s 3 .5 and which a lso contains th e same amount of in d ic a to r. 3 . Add 1 m i l l i l i t e r of th e hydroquinone so lu tio n , and 2 m i l l i l i t e r s o f th e ortho-phenanthroline reagent to th e a liq u o t i n th e volum etric f la s k , 4. Add th e same volume o f 2 N sodium a c e ta te th a t was found necessary to a d ju st th e a liq u o t i n the beaker to a pH of 3*5. I f tu r b id ity develops a t pH 3 ,5 , add 1 m i l l i l i t e r of th e ammonium c i t r a t e s o lu tio n before adding th e sodium t a c e ta te . D ilu te to 25 m i l l i l i t e r s , mix and l e t stand f o r 1 hour to assu re complete co lo r development. 5, Determine th e o p tic a l d e n s ity i n th e Coleman spectrophotom eter a t a wave le n g th o f 510 and w ith a PC-4 f i l t e r , using a w ater blank to balance th e in s tro n e n t. CALCULATIONS: Prepare a c a lib r a tio n curve w ith standard so lu tio n s containing from 0,01 to 0 ,1 m illigram s of iro n . p a rts p er m illio n of iro n per cent o f iro n - = mg. found x 1000 g , sample mg. found x g. sample 0.1 2. Determ ination of calcium Calcium was determined in aliq u o ts of the ash so lu tio n by th e semimicro procedure of R. F. Reitem eier (1943) REAGENTS Amonium o x a la te . 4 p er cent so lu tio n Sodium hydroxide. 30 p er cent so lu tio n . Hydrochloric a c id . 1:15 d ilu tio n P erchloric a c id . Approximately 4 N so lu tio n . D ilu te 340 m i l l i l i t e r s of 70 p er cent p erch lo ric acid to 1 l i t e r . Sodium o x alate. Standard 0.01 N so lu tio n . D issolve 0.670 grams of sodium oxalate in w ater and d ilu te to 1 l i t e r . Ammonium h ex a n itra te m ixture. Approximately 0.01 N standard so lu tio n in p erch lo ric acid . Dissolve 5*76 grams of h ig h est q u a lity ammonium h e x n itra te ce ra te in 250 m i l l i l i t e r s of 4 N p erch lo ric acid and d ilu te to 1 l i t e r . The reagent i s then standardized as follow s: P ipet 5 o r 10 m i l l i l i t e r s of fre sh standard 0.01 N sodium oxalate in to a beaker containing 5 m i l l i l i t e r s of 4 N p erch lo ric ac id . Add 0 .2 m i l l i l i t e r s of n itr o - f e r r o in in d ic a to r and t i t r a t e w ith th e ce ra te so lu tio n to a c o lo rle ss end p o in t. Determine a blank t i t r a t i o n co rrectio n on a sim ila r sample lacking th e oxalate so lu tio n . The norm ality o f th e cerate so lu tio n i s obtained as follow s: N - m is, x 0.01 C- D where mis ■ m is. of ox alate used C - m is. of cerate used to t i t r a t e th e oxalate D = m is. of cerate to t i t r a t e th e blank. Do not attem pt to adjust th e norm ality of th e cerate to ex actly 0.01 N. Store in a dark b o ttl e , and restan d ard ize every 2 o r 3 days. N ltro-F erroin in d ic a to r. E ith e r nitro-orthophenanthroline ferro u s s u lfa te , or n itr o - orthophenanthroline ferro u s p erchlorate may be used. When th e sulphate i s used, add 0 .1 m i l l i l i t e r to the unknown so lu tio n and 0 .2 m i l l i l i t e r s to th e stan d ard izatio n so lu tio n . I f th e p erch lo rate i s used add 0.5 m i l l i l i t e r s . Methyl red in d ic a to r. Dissolve 0.1 grams in 100 m i l l i l i t e r s of 95 p er cent alcohol. PROCEDURE. 1 . P ipet 5 m i l l i l i t e r s of th e ash so lu tio n , representing 0.025 grams of the dry p lan t m a te ria l, in to a co n ical, 1 5 -n d .llilite rs heavy duty cen trifu g e tu b e. 2. Add 1 drop of methyl red in d ic a to r so lu tio n , and then add 30 p er cent sodium hydroxide drop by drop w ith constant s tir r in g u n til a yellow co lo r appears. The pH should be 6.3 3 . Add 1:15 hydrochloric acid drop by drop u n t i l th e so lu tio n b arely tu rn s pink. 4 . Add 2 m i l l i l i t e r s of th e ammonium o x alate, mix and l e t stand over n ig h t. The so lu tio n need stand only 2 hours, but when a la rg e number of samples are being determined a t one tim e, i t i s convenient to l e t stand over night a t t h is stag e. 5. C entrifuge a t 3000 r.p .m . f o r 10 minutes. C arefu lly decant the supernatant liq u id in to a 2 5 - m illilite r s volum etric fla sk and preserve f o r th e determ ination of magnesium. 6. Rinse down the sid es o f th e tube w ith 5 m i l l i l i t e r s of 1:50 ammonium hydroxide blown from a p ip e tte . Break up and s t i r th e p re c ip ita te w ith a th in g lass rod and again cen trifu g e a t 3000 r.p .m . f o r 10 minutes. Decant th e supernatant liq u id in to th e 2 5 - m illilite r s volum etric fla s k containing th e so lu tio n preserved fo r th e magnesium determi­ n atio n . ■7. Drain th e tubes fo r 10 minutes by in v e rtin g them on f i l t e r paper. 8 . Blow 3 m i l l i l i t e r s of th e 4 N p erch lo ric acid in to th e tu b e. When th e p r e c ip ita te has d isso lv ed , tr a n s f e r th e so lu tio n to a 25 m i l l i l i t e r s volum etric f la s k , and rin se the tube with 5 m i l l i l i t e r s o f w ater. 9. Add 0.1 m i l l i l i t e r of th e ferro u s n itr o - fe rr o in (o r 0*5 m i l l i l i t e r of th e p erch lo rate n itro -fe rro in ) in d ic a to r, and t i t r a t e with approximately 0.01 N standard cerate to a co lo rle ss end point by means of a micro b u re tte . The blank t i t r a t i o n , determined in th e same manner i s u su ally about 0.03 m i l l i l i t e r s . CALCULATIONS : per cent calcium ■ (mis, ce ra te) (N of c e ra te) (2.004) 4. Determination of magnesium Magnesium was determined in a liq u o ts o f th e ash so lu tio n by th e method o f R eitem eier. (1943). REAGENTS Ammonium ch lo rid e. 30 p er cent so lu tio n . Dissolve 30 grams of ammonium chloride in w ater and d ilu te to 100 m i l l i l i t e r s F i l t e r before use. Ammonium phosphate. 5 p er cent solution* Dissolve 25 grams of ammonium dihydrogen phosphate, NH/JH^PO^, in water and d ilu te to 500 m i l l i l i t e r s . F i l t e r before u se. Ammonium hydroxide. Concentrated so lu tio n . Ammonium hydroxide wash so lu tio n . Mix 20 m i l l i l i t e r s of concentrated ammoniun hydroxide in SO m i l l i l i t e r s o f w ater, 100 m i l l i l i t e r s of 95 per cent e th y l alcohol, and 100 m i l l i l i t e r s of e th e r. Sulphuric ac id . Approximately 1 N. D ilute 27.7 m i l l i l i t e r s of concentrated sulphuric acid (sp e c ific g ra v ity 1.84) to 1 lite r. Ammonium molybdate. Dissolve 5 grams of ammonium molybdate in 80 m i l l i l i t e r s of warm w ater, add 2.8 m i l l i l i t e r s of concentrated sulphuric acid to th e cool so lu tio n , and d ilu te to 100 m i l l i l i t e r s w ith w ater. Prepare fre sh every 7 days. Hydroquinone. 0.5 per cent so lu tio n . D issolve 0.5 grams of hydroquinone in 100 m i l l i l i t e r s of water made s lig h tly acid w ith one drop o£ concentrated sulphuric ac id . S tore in the r e f rig e ra to r . Sodium su ccin ate. 20 per cent solution* D issolve 20 grams of anhydrous succinate in 100 m i l l i l i t e r s of w ater. I f hydrated sodium succinate i s used, d isso lv e 33,6 grams in 100 m i l l i l i t e r s of w ater. Store in th e re frig e ra to r* Ammonium hydroxide. 1:1 d ilu tio n Boric ac id . Approximately 0*8 M so lu tio n . D issolve 50 grams of boric acid in w ater and d ilu te to 1000 m i l l i l i t e r s . Brom thymol blue in d ic a to r. D issolve 0*05 grams o f brom thymol blue in 2*4 m i l l i l i t e r s of 0*01 N sodium hydroxide and d ilu te to 175 m i l l i l i t e r s w ith w ater. Phenolphtalein in d ic a to r. D issolve 1 gram of phenolphtalein i n 100 m i l l i l i t e r s of 60 p er cent e th y l alcohol. Magnesium su lp h ate. Standard so lu tio n . D issolve 0*1014 o f magnesium su lp h ate, MgS0^*7H20, in 1 l i t e r . Each m i l l i l i t e r contains 0*01 m illigram of magnesium. PROCEDURE 1. D ilu te th e supernatant liq u id obtained by cen trifu g in g th e calcium as th e oxalate to 25 m i l l i l i t e r s . 2* P ipet 5 m i l l i l i t e r s of t h is so lu tio n , rep resen tin g 0*005 grams o f dry p lan t m a te ria l, in to a 15 m i l l i l i t e r s co n ical, heavy duty cen trifu g e tu b e . I f a d iffe re n t aliq u o t i s used, d ilu te o r evaporate to 5 m i l l i l i t e r s . 3* Add 1 m i l l i l i t e r of 30 per cent ammonium ch lo rid e and 1 m i l l i l i t e r of 5 per cent ammonium phosphate, and 1 drop of phenolphtalein in d ic a to r. M L 4. Heat to 90° C. in a w ater bath* and add concentrated ammonium hydroxide drop by drop, with constant tw irlin g , u n til a pink co lo r appears. Cool, 5. Add 2 m i l l i l i t e r s of concentrated ammonium hydroxide and s t i r with a th in g la ss rod. Stopper, and l e t stand over n ig h t. 6. Centrifuge a t 1500 r.p.m . fo r 10 m inutes, c a re fu lly decant th e supernatant liq u id , and d rain th e tube fo r 10 minutes by in v e rtin g i t on a f i l t e r paper. Wipe th e mouth of th e tube with a clean c lo th . 7. Wash th e sides of th e tube with 5 m i l l i l i t e r s of the anmonium hydroxide was so lu tio n , and break up th e p re c ip ita te w ith a th in g lass rod, and again cen trifu g e a t 1500 r.p .m . fo r 10 minutes, decant, and drain as before. S. Repeat th e washing, cen trifu g in g , and decanting fo r th e th ir d tim e. 9* Add 2 m i l l i l i t e r s of 1.0 N sulphuric a c id , and d ilu te w ith w ater to about 5 m i l l i l i t e r s and l e t stand 5 minutes. 10. T ransfer the content of th e tube to a 25 m i l l i l i t e r s volumetric fla s k , washing out th e tube w ith 3 o r 4 m i l l i l i t e r s of w ater. 11. Add 5 m i l l i l i t e r s of th e b o ric acid so lu tio n . I f the so lu tio n i s acid , add 5 drops of brom phenol blue in d ic a to r, and n e u tra liz e by adding ammonium hydroxide dropwise. 12. Add 2 m i l l i l i t e r s of th e ammonium molybdate so lu tio n . Mix. 13. Add 2 m i l l i l i t e r s of th e hydroquinone so lu tio n . Mix. 14. Add 2.5 m i l l i l i t e r s of the sodium succinate so lu tio n . Mix; make up to 25 m i l l i l i t e r s w ith w ater, mix, and l e t stand fo r 30 m inutes. The color i s constant between 0*3 and 4 hours. 15. Measure th e o p tic a l d en sity of th e so lu tio n a t a wave length of 775 using a PC-5 Coleman f i l t e r . 16. Run a blank determ ination, using a l l of th e reagents used in determining th e unknown. 17. Make a c a lib ra tio n curve by running standards containing from 0.01 to 0.10 m illilig ram s of magnesium. CALCULATIONS: per cent Mg = mgs, found x 0 .1 g . sample A 5* Determination of potassium Potassium was determined in th e so lu tio n of digested p la n t m a terial by th e colorim etric method of Peech M.(19Al). REAGENTS Ethanol. 70 p er cent so lu tio n . D ilu te 500 mis of 95 per cent ethanol with 180 m i l l i l i t e r s o f w ater. N itroso R - s a lt. 0.5 per cent so lu tio n . Dissolve 0.5 grams of N itroso R -sa lt (disodium s a lt of l-n itro so -2 -h y d ro x y -3 ,6naphthalene-disulfonic acid) in 100 m i l l i l i t e r s o f water* When not exposed to d ire c t su n lig h t, th e reagent i s sta b le f o r sev eral weeks. Sodium c o b a lti n itr lte . 25 per cent so lu tio n . Dissolve 25 grams of reagent q u a lity trisodium c o b a ltin itr lte in w ater, and d ilu te to 100 m i l l i l i t e r s and f i l t e r . Cool th e so lu tio n before use and keep in a re f rig e ra to r . Prepare d a ily th e necessary amount of th is reagent needed. Sodium pyrophosphate. 5 per cent so lu tio n . D issolve 5 grams of powdered sodium pyrophosphate (N aj^O y.lO ^O ) i n 100 m i l l i l i t e r s of w ater. Sodium a c e ta te . Approximately 2.4 K so lu tio n . Sulphuric a c id . Approximately 0 .1 N solution* N itric a c id . Approximately 0*1 N so lu tio n . Potassium ch lo rid e. Standard so lu tio n . Dissolve 0*9533 grama of dry potassium ch lo rid e in 500 m i l l i l i t e r s of w ater. Each m i l l i l i t e r contains 1 m illigram of potassium. PROCEDURE 1. P ipet an aliq u o t of 3 m i l l i l i t e r s of digested ash so lu tio n , representing 0.013 grams of dr,, p lan t m a te ria l, in to a 15 m i l l i l i t e r s co n ical, heavy duty centrifuge tu b e . I f the aliq u o t i s le s s than 3 m i l l i l i t e r s d ilu te to 3 m i l l i l i t e r s w ith 0 .1 N n i t r i c ac id . Add 1 m i l l i l i t e r of th e sodium c o b a ltin itr lte reagent and mix thoroughly by tw irlin g the tube* Let stand in th e re f rig e ra to r f o r 1 hour. 2. Add 4 m i l i l i t e r s of 70 per cent eth an o l, s t i r with a th in g lass rod, wash th e rod w ith ethanol, and cen trifu g e fo r 15 minutes a t 1700 r.p.m . Decant the supernatant liq u id 7and d rain th e tube fo r se v eral minutes on f i l t e r paper. 3 . Add 5 m i l l i l i t e r s of 70 per cent ethanol down th e w alls of the tu b e, break up th e p re c ip ita te w ith a s tir r in g rod, wash th e rod w ith 1 m i l l i l i t e r of alcohol, and again cen trifu g e fo r 10 minutes a t 1700 r.p.m . 4. Decant th e c le a r so lu tio n , allowing th e tube to d rain fo r sev eral m inutes, and repeat fo r th e th ir d tim e, th e washing w ith 5 m i l l i l i t e r s of 70 per cent alcohol. 5. Dissolve th e p re c ip ita te in 5 m i l l i l i t e r s of 2 N sulphuric acid and place th e tube in a water bath a t about 70® C. A fter th e p re c ip ita te in the bottom of the tube has been disso lv ed , add 5 to 7 m i l l i l i t e r s of water and heat fo r another 5 to 10 minutes to dissolve any p e c ip ita te adhering to th e sid es of th e tu b e. Cool. D ilute to 10 m i l l i l i t e r s , stopper, and mix. 6* Introduce a 1 m i l l i l i t e r a liq u o t of th e so lu tio n in to a 25 m i l l i l i t e r s volum etric fla s k . 7. Add 1 m i l l i l i t e r of 5 per cent pyrophosphate so lu tio n , d ilu te to a volume of 20 m i l l i l i t e r s w ith w ater. 8. Add 1 m i l l i l i t e r of 2.5 N sodiun. ac eta te so lu tio n and mix. 9 . Add 2 m i l l i l i t e r s of 0.5 per cent so lu tio n of N itro ao -R -salt, mix, bring to volume of 25 m is. and mix. A fter 15 minutes, determine the o p tic a l d en sity by the Coleman spectrophotometer, using a wave length of 550 vv\J*~ and a PC-4 f i l t e r . A water blank i s used in balancing th e instrum ent. 10. Hake a standard curve by preparing standard so lu tio n s containing from 0 .1 to 1 .0 m illigram s of potassium in th e a liq u o t placed in the cen trifu g e tu b e. CALCULATION: per cent K ■ (mgs, found) (0.1) (g s. sample) A 6. Determination of phosphorus Phosphorus was determined on aliq u o ts of th e ash so lu tio n by the method described by S to lo ff. L.S. (1942) REAGENTS Ammonium m olybdate-sulphuric a c id . Dissolve 5*0 grams of amnonium molybdate in approximately 80 m i l l i l i t e r s of warm (about 50° C.) w ater, add 2*8 m i l l i l i t e r s of concentrated sulphuric acid to th e cool so lu tio n and d ilu te to 100 m i l l i l i t e r s w ith w ater. This so lu tio n should not be used i f a white residue has s e ttle d o u t. Do not prepare la rg e q u a n titie s . Use a fre sh ly prepared so lu tio n every few days. Hydroquinone. 0.5 per cent so lu tio n . Dissolve 0*5 grams of hydroquinone in 100 m i l l i l i t e r s of water made s lig h tly acid w ith a drop of concentrated sulphuric acid . Store in th e r e f rig e ra to r . Sodium su ccin ate. 20 per cent so lu tio n . Dissolve 20 grams of anhydrous sodium succinate in 100 m i l l i l i t e r s of w ater. Store in the re frig e ra to r. Potassium phosphate. Standard so lu tio n . Dissolve 0.4394 g . of potassium dihydrogen phosphate in 1000 m i l l i l i t e r s of w ater. Each m i l l i l i t e r contains 0 .1 m illigram of phosphorus. Ammonium hydroxide. 1:1 d ilu tio n Boric acid . S aturated so lu tio n . Dissolve 50 grams of boric acid in about 800 m i l l i l i t e r s of warm water and d ilu te to 1 lite r. PROCEDURE: 1, P ip et an aliq u o t of 1 m i l l i l i t e r of th e ash so lu tio n , represen­ tin g 0.005 grams of dry p la n t m a te r ia l,,in to a 25 m i l l i l i t e r s volum etric fla s k . The aliq u o t should not contain more than 0.3 m illigram s of phosphorus. The e ffe c tiv e range of phospho­ rus concentrations f o r t h i s procedure i s from 0,002 to 0,300 m illigram s of phosphorus in th e a liq u o t. I f flu o rid e s are p rese n t, add 5 m i l l i l i t e r s o f th e satu rated b o ric acid so lu tio n . I f th e so lu tio n i s ac id , add 5 drops of th e brom-phenol blue in d ic a to r and n e u tra liz e by adding ammonium hydroxide u n t i l th e color tu rn s b lu e. I f b oric acid has been added, 1:10 ammonium hydroxide i s s a tis fa c to ry , but i f th e o rig in a l acid e x tra c t i s used alone, then 1:1 ammonium hydroxide i s more convenient, 2 , Add th e follow ing reagents in th e order l i s t e d , shaking w ell a f te r each ad d itio n . The maximum lapse of tim e between each ad d itio n which w ill not a ffe c t th e co lo r development i s 5 m inutes, but th e blanks must be tre a te d in th e same manner. (a) 2,0 m i l l i l i t e r s of th e ammonium molybdate so lu tio n , (b) 2.0 m i l l i l i t e r s of the hydroquinone so lu tio n . (c) 2,5 m i l l i l i t e r s of th e sodium succinate so lu tio n . 3 , Make up to volume w ith d i s t i l l e d water and allow to stand fo r 30 m inutes. The co lo r i s sta b le between 30 minutes and 4 hours. Determine th e o p tic a l d en sity o f th e color in a Coleman U niversal Spectrophotometer, using a wave length of 775 and a PC-5 f i l t e r . 7* Determination of manganese Manganese was determined in a liq u o ts of th e ash so lu tio n by th e procedure of W illard and Great-house (1917) REAGENTS Sodlun m etaperiodate. Fine powder Phosphoric ac id . 85 per cent S u lfu ric ac id . Concentrated Sodium s u l f i t e . Fine powder Potassium permanganate. Standard so lu tio n containing 0.0250 m illigram s of manganese p er m i l l i l i t e r . Prepare a 0.10 N standard potassium permanganate so lu tio h . Add 22.8 m i l l i l i t e r s of the standard so lu tio n to a 250 m i l l i l i t e r s Erlenmeyer fla s k . Add about 50 m i l l i l i t e r s of d i s t i l l e d water and 1 m i l l i l i t e r of concentrated su lfu ric acid . Heat to b o ilin g and reduce th e permanganate by adding sodium s u lf ite powder. Avoid a la rg e excess of s u l f i t e . B oil o ff th e excess su lfu r dioxide and d ilu te to one l i t e r . Each m i l l i l i t e r o f th is so lu tio n contains 0.0250 m illigrams of manganese. PROCEDURE 1. P ipet 20 m i l l i l i t e r s o f th e ash so lu tio n , representing 0.1000 grams o f dry p la n t m a terial, in to a 25 m i l l i l i t e r s volum etric fla s k . \ 2. Add 1 m i l l i l i t e r of 85 per cent phosphoric acid and about 50 milligram s of sodium p erio d ate. Mix w ell, and place in water bath a t 95° C. Let stand fo r two hours. 3* Cool, make to volume w ith d i s t i l l e d w ater, mix, and measure o p tic a l d en sity in th e Coleman spectro­ photometer using a PC-4 f i l t e r , a wave length of 530 vrJ*' and a w ater blank. 4* Prepare a s e rie s of standard so lu tio n s containing 0.0125 to 0.250 m illigram s of manganese per 25 m i l l i l i t e r volum etric fla s k . Develop color as above. P lot o p tic a l d en sity against concentration of manganese. 8 . D e t e r m i n a t i o n o f c o pper. Copper was d e te r m in e d p o l a r o g r a p h i c a l l y p r o c e d u re o u t l i n e d by Reeds and Cunnings according t o the (194 1). REAGENTS N i t r i c a c i d . Concentrated s o lu t i o n S u l f u i l c - p e r c h l o r i c m i x t u r e . Mix e q u a l volumes o f c o n c e n t r a t e d s u l f u r i c a c i d and 72 p e r c e n t s o l u t i o n o f p e r c h l o r i c a c i d Ammonium h y d r o x i d e . C o n c e n tr a t e d s o l u t i o n Ammonium h y d r o x id e s o l u t i o n , a 1 : 4 0 s o l u t i o n . Sodium c i t r a t e s u p p o r t i n g e l e c t r o l y t e . D i s s o l v e 105.07 grams o f c i t r i c a c i d and 20 grams o f sodium h y d r o x id e i n d i s t i l l e d w a t e r and make up t o a volume o f 2 l i t e r s . Acid f u c h s i n s o l u t i o n . A 0.05 p e r cent s o l u t i o n i n d i s t i l l e d water. N it r o g e n g a s . PROCEDURE 1 . P l a c e 0 , 5 t o 2 grams o f p l a n t m a t e r i a l i n a 30 m i l l i l i t e r s k j e l d a h l f l a s k . Add 10 m i l l i l i t e r s o f c o n c e n t r a t e d n i t r i c a c id , 2 m i l l i l i t e r s of the s u lf u r ic - p e r c h lo r ic s o l u t i o n and a d r o p o f k e r o s e n e t o p r e v e n t f r o t h i n g . 2. Le t s t a n d o v e r n i g h t . 3 . Heat u n t i l t h e s o l u t i o n i s c o l o r l e s s o r a p a l e y e l l o w . 4. Let c o o l . 5. Add 5 m i l l i l i t e r o f w a te r and h e a t t o b o i l i n g . 6. Add a s l i g h t ex ces s o f c o n c e n t r a t e ammonium hydroxid e (5 m i l l i l i t e r s were used) 7 . B o i l f o r a m i n u t e . Bumping o c c u r s r e a d i l y and ea ch sample must be t a k e n s e p a r a t e l y and shaken a l l t h e time w h i l e h e a t i n g t o p r e v e n t any l o s s o f m a t e r i a l from sudden boiling. 8 . F i l t e r t h r o u g h a Whatman No 2 f i l t e r p a p e r i n t o a 100 m i l l i l i t e r s b e a k e r , wash t h e k j e l d a h l f l a s k and f i l t e r pa p e r w i t h t h e 1 :4 0 ammonium h y d r o x id e s o l u t i o n . 9. E v a p o ra te t h e f i l t r a t e t o d r y n e s s a t slow h e a t on a sand b a t h . Be c a r e f u l t ow ar d t h e end o f t h e e v a p o r a t i o n beca us e t h e d r y in g s a l t s s p l a t t e r e a s i l y . 10. Take up t h e r e s i d u e i n 9 m i l l i l i t e r s o f sodium c i t r a t e s o l u t i o n , add 1 m i l l i l i t e r o f 0 , 0 5 p e r ce nt a c i d fuchsin. 11. Remove t h e oxygen by b u b b l i n g n i t r o g e n 20 m i n u t e s . 12. Run t h e c u r v e s w i t h an a p p l i e d p o t e n t i a l from 0 . 2 v o l t s t o - 2 . 8 v o l t s . The h a l f wave p o t e n t i a l f o r copper i n t h e sodium c i t r a t e s u p p o r t i n g e l e c t r o l y t e i s - 0 . 1 5 v o l t s a g a in s t a s a tu r a te d calomel e le c tr o d e . 13. Measure t h e d i f f u s i o n c u r r e n t from t h e polar og ra ra and c a l c u l a t e t h e c o r r e s p o n d i n g amount o f co pper . I i 'W i 9* Determination of nitrogen REAGENTS Hydrochloric acid . Standard 0.02 N solution Boric acid . 2 per cent so lu tio n . Dissolve 10 grams of acid in 500 m i l l i l i t e r s o f hot w ater. Methyl purple in d ic a to r. Cosmercially prepared so lu tio n . (F le ish e r chemical company) Add 2 drops to th e bo ric acid so lu tio n p er 100 m i l l i l i t e r s . Sodium hydroxide. 30 per cent so lu tio n . Dissolve 150 grams of sodium hydroxide p e lle ts in 350 m i l l i l i t e r s of water. R anker's so lu tio n . Dissolve 32 grams of s a lic y lic acid in 1 l i t e r o f concentrated sulphuxlc acid . Sodimn th io su lp h a te . Dissolve 50 grans of sodium th io su lp h ate in 100 m i l l i l i t e r s of w ater. Selenium . Powdered metal Potassium sulphate-Copper sulphate m ixture. P ulverize in a m ortar 3 p a rts of copper sulphate with 1 p a rt of potassium sulphate. PROCEDURE 1. Weigh from 40 to 50 milligrams of th e fin e ly ground dry plant m aterial in to a 50 m i l l i l i t e r s micro K jeldahl d ig estio n fla s k . A long handled micro weighing tube w ill be found very convenient. 2. Add 2 m i l l i l i t e r s of Ranker's so lu tio n , mix. 3. Add 5 drops of th e th io su lp h ate so lu tio n and g en tly warm. 4. A fter sev eral m inutes, add about 5 milligrams o f th e powdered selenium by means o f a micro spatula* B oil u n til the so lu tio n i s c le a r. Usually AO to 60 minutes are s u ffic ie n t fo r th e d ig estio n of dry powdered g rass m a terial. 5. T ransfer the digested so lu tio n to th e micro s t i l l and ca re fu lly wash out th e fla sk w ith 2 rin sin g s o f w ater. 6. Add 10 m i l l i l i t e r s o f 30 per cent sodiun hydroxide and d i s t i l l fo r 10 minutes in to 5 m i l l i l i t e r s of 2 per cent b o ric acid . 7. T itr a te to a purple co lo r w ith standard 0.02 N hydrochloric acid . 8. Run a blank determ ination using a l l the reagents as in the determ ination of the unknown. CALCULATIONS per cent N ■ (mis. HC1 used - m is. HCL blank) x 0.28 gs. sample 10, Determination of total carbohydrates. The t o t a l carbohydrates in th e dry p lan t m a te ria l was determined by prelim inary h y d ro ly sis, and then estim ating th e reducing sugar by th e micro procedure described by Heinze and Mumeek (1940). REAGENTS Hydrochloric acid . Concentrated so lu tio n Lead a c e ta te . N eutral, 25 p er cent so lu tio n Potassium o x alate. 25 per cent so lu tio n Potassium hydroxide. 25 p er cent so lu tio n Methyl red in d ic a to r. 0 .1 per cent so lu tio n in 60 per cent eth y l alcohol. Copper reagent "5011. Dissolve 25 grams of sodium carbonate and 25 grams of sodium potassium t a r t r a t e (Rochelle s a lt) in about 600 m i l l i l i t e r s o f w ater. Then add 75 m i l l i l i t e r s o f a 10 per cent copper sulphate so lu tio n by means of a p ip e tte dipping below th e surface of th e so lu tio n . Add 20 grams of sodiua bicarbonate and 1 gram of potassium iodide and mix. Add 200 m i l l i l i t e r s of a so lu tio n of potassium io d ate containing 3*567 grams o f io d ate per l i t e r . Mix. Bring to volune in a 1 l i t e r volum etric f la s k . Store in a dark b o ttle . The io d ate content may be varied according to the amount of sugar to be determined. I f th e reagents are made up as d ire c te d , le s s than 4.40 m illigram s of glucose p er 10 m i l l i l i t e r s o f so lu tio n may be determined. Potassium lodide-potaaslum o x a la te . Dissolve 2.50 grams of potasslun Iodide and 2,5 grams of potassium oxalate In w ater and d ilu te to 100 m i l l i l i t e r s . This reagent should be stored In a dark b o ttle and prepared fre sh each week. The development of fre e Iodine on standing w ill se rio u sly d istu rb th e determ ination o f sugar. Sodium th io su lp h ate. Standard 0,0200 N so lu tio n . Dissolve 24*82 grams of sodiua th io su lp h ate, Na^^O ^iS^O , In about 800 m i l l i l i t e r s o f boiled w ater. Add 10 m i l l i l i t e r s of 0,1 N sodiua hydroxide. D ilu te to 1 l i t e r . This solution i s about 0 ,1 . Let stand sev eral days before standardizing. To dtandardize the so lu tio n , accu retely weigh from 0,12 to 0,17 grams of potassium Io d ate in to a 250 m i l l i l i t e r s Erlenmeyer fla s k . D issolve th e s a lt in 50 m i l l i l i t e r s of b o iled w ater. T itr a te immediately w ith th e th io su lp h ate to a straw co lo r. Add 3 drops of a sta rc h In d ic a to r, o r enough to give a blue co lo r, and complete th e t i t r a t i o n to th e disappearance o f th e starch -io d in e blue co lo r. C alculate th e norm ality of th e th io su lp h ate. One m illieq u iv alen t of potasslun io d a te i s 0,03567 grams. N of th io su lp h ate ■ gs. potassium io d ate (mis. th io su lp h ate) x 0.03567 Prepare th e standard 0,0200 N th io su lp h ate by d ilu tio n as follow s: Vx - 0.0200 ViNx where Vx * f in a l volume o f the d ilu tio n Vx s volume of the th io su lp h ate to be d ilu te d Nx s norm ality o f the th io su lp h ate to be d ilu te d . Sulphuric a c id . Approximately 1 N so lu tio n . D ilu te 28.4 m i l l i l i t e r s \ o f concentrated sulphuric acid (sp . g r. 1.835) to 1 l i t e r . Starch in d ic a to r. Add 1 gram of the "soluble" sta rc h to 100 m i l l i l i t e r s of a sa tu ra ted sodium chloride so lu tio n . A gitate, and sto re in the re frig e ra to r. PROCEDURE: 1. Weigh 1 gram of dry p la n t m a terial in to a 500 m i l l i l i t e r s re flu x f la s k , add 200 m i l l i l i t e r s of w ater, and 20 m i l l i l i t e r s of 25 per cent hydrochloric acid . 2 . Reflux fo r 2.5 hours. Sw irl th e fla s k occasionally to wash down p a r tic le s adhering to th e g la s s . 3 . Add 5 drops of the methyl red in d ic a to r, and n e u tra liz e w ith 25 per cent sodium hydroxide. The n e u tra liz a tio n must be done c a re fu lly . 4* Add 30 m i l l i l i t e r s of th e 25 per cent n e u tra l led a c e ta te , mix. Test f o r complete p re c ip ita tio n by adding a drop o f the lead a c e ta te . 5. Add 10 m i l l i l i t e r s o f 25 per cent potassium o x alate . Mix. Test f o r complete p re c ip ita tio n of the lead by adding a drop of th e o x alate. 6. F ilte r in to a 500 m i l l i l i t e r s volum etric fla sk through Whatman No. 2 f i l t e r paper. Bring to volume and mix. 7. The magnitude of the aliq u o t depends on the concentration of reducing sugar. O rdinarily 3 m i l l i l i t e r s w ill be sa tisfac to ry * Three m i l l i l i t e r s correspond to 15 m illigram s of the dry p lan t m a te ria l. 8 . Pipet an aliq u o t of 3 m i l l i l i t e r s of th e so lu tio n in to an 8 x 1 inch t e s t tu b e. Add 10 m i l l i l i t e r s of reagent "50". Mix, cover th e tubes w ith a sm all beaker and immerse in a b o ilin g water bath f o r 15 minutes. 9* Cool in running water f o r 3 m inutes, o r u n til th e tem perature i s about 30° C. 10. Add 2 m i l l i l i t e r s of the potassium iodide-potassium oxalate so lu tio n . Mix. 11. Add 10 m i l l i l i t e r s of the 1 N sulphuric acid . A gitate thoroughly. Let stand f o r 5 minutes. 12. T itra te with 0.0200 N standard th io su lp h ate u n t i l th e straw co lo r has almost disappeared. Add 3 drops of the starch in d ic a to r, and complete th e t i t r a t i o n . 13* Prepare a c a lib ra tio n curve by obtaining th e t i t r a t i o n values f o r a s e rie s of standard glucose so lu tio n s containing from 0.1 to 4*4 m illigram s of glucode per a liq u o t. 14* T itra te a blank, su b s titu tin g 3 m i l l i l i t e r s o f water f o r th e p lan t e x tra c t. CALCULATIONS: The number of m i l l i l i t e r s of th io su lp h ate used to t i t r a t e th e blank, minus th e m i l l i l i t e r s o f th e th io su lp h ate used to t i t r a t e th e sample i s p ro p o rtio n al to the sugar p resen t, calculated as glucose. Read th e amount of sugar in the sample from th e c a lib ra tio n curve. p er cent glucose = mgs, found x 0.1 G8. sample 11. Determination of ascorbic acid* The m e th o d u s e d f o r t h e d e t e r m i n a t i o n o f a s c o r b i c a c i d w a s t h e p h o t o m e t r ic p r o c e d u r e d e s c r i b e d b y H o r e l l ( 1 9 4 1 ) and B e s s e y (1 9 3 8 ) A s o l u t i o n o f 0 . 5 p e r c e n t o x a l i c a c id w a s u s e d a s a n e x t r a c t a n t w h ic h a c c o r d i n g t o P o n t in g ( 1 9 4 3 ) m ay b e s u b s t i t u t e d f o r m e t a p h o s p h o r ic a c i d . REAGENTS: O x a l ic a c i d . 0 . 5 p e r c e n t s o l u t i o n i n d i s t i l l e d w a t e r . D ye s o l u t i o n . 3 4 . 4 m i l l i g r a m s o f s o d iu m 2 , 6 - d i c h l o r o b e n z e n o n e in d o p h e n o l p e r l i t e r o f d i s t i l l e d w a t e r . PROCEDURE: 1 . M a c e r a t e a s a m p le o f a b o u t 5 g r a m s o f f r e s h p l a n t m a t e r i a l w ith 1 5 0 m i l l i l i t e r s o f 0 .5 p er c e n t o x a l i c a c id i n a W a r in g B le n d e r f o r 5 m i n u t e s , t h e n m ake u p t o 2 0 0 m illilite r s . 2 . F i l t e r th e m a c e r a te w ith s u c t io n th r o u g h 2 t h ic k n e s s e s o f C e n c o No 1 3 2 5 0 f i l t e r p a p e r . D is c a r d t h e f i r s t 1 0 m i l l i ­ l i t e r s o f th e e x tr a c t. 3 . S e t t h e C o le m a n S p e c t r o p h o t o m e t e r , u s i n g a w a v e l e n g t h o f 520 an d a P C -4 f i l t e r , t o r e a d 1 0 0 p e r c e n t t r a e m i s s i o n o n t h e g a lv a n o m e t e r s c a l e , a g a i n s t a b la n k t u b e c o n t a i n i n g o n l y d i s t i l l e d w a t e r . U. P ip e t a liq u o t s o f 5 m i l l i l i t e r s o f th e e x t r a c t in t o m a tc h e d C o le m a n p h o t o m e t e r t u b e s , q u i c k l y d e l i v e r 5 m i l l i l i t e r s o f t h e d y e i n t o t h e s a m p le and s h a k e v ig o r o u s ly fo r about 5 se c o n d s. 5 . T ake th e t r a n s m is s io n r e a d in g s 15 and 30 se c o n d s a f t e r t h e t im e t h e d y e h a s b e e n a d d e d . T h e s e r e a d i n g s a r e d e s i g n a t e d a s G]_ an d G g . 6 . R ed u ce t h e e x t r a c t w ith a few c r y s t a l s o f a s c o r b ic a c id t o g i v e t h e t r a n s m i s s i o n r e a d i n g , G sr . 7 . A d j u s t t h e g a lv a n o m e t e r r e a d i n g t o g i v e t h e v a l u e o f G sr w it h 5 m i l l i l i t e r s o f 0 . 5 p e r c e n t o x a l i c a c i d p l u s 5 i a i l ; i l i t e r s o f d y e an d a f e w c r y s t a l s o f a s c o r b i c a c id . 8 . M ix 5 m i l l i l i t e r s o f 0 . 5 p e r c e n t o x a l i c a c i d w it h 5 m i l l i l i t e r s o f d ye t o g iv e t h e t r a n s m is s io n r e a d in g s G b i and Gb2 » CALCULATION: From t h e a v e r a g e v a l u e o f G s and G b, t h e am ou n t o f a s c o r b i c a c i d , C, i s o b ta in e d a s f o llo w s : C = K ( l o g G s - l o g Gb) W ith t h e am ou n t o f d y e u s e d and t h e d i l u t i o n o f t h e s a m p le t o 2 0 0 m i l l i l i t e r s , t h e v a l u e o f K w a s fo u n d t o b e 8 b y c a l c u l a t i o n fr o m t h e s t a n d a r d c u r v e . Preparation of the standard curve: P rep a re a f r e s h s o lu t io n o f a s c o r b ic a c id b y d is s o lv in g 25 m illig r a m s in 2 5 0 m i l l i l i t e r s o f 0 .5 p er c e n t o x a l i c a c i d . Add 1 4 a l i q u o t s , e a c h d i f f e r i n g b y 1 m i l l i l i t e r in c r e m e n ts (c o r r e s p o n d in g t o 1 t o 1 4 m ic r o g r a m s o f a s c o r b i c a c i d p e r m i l l i l i t e r ) t o a s e r i e s o f 1 4 1 0 0 m i l l i l i t e r s v o lu m e tr ic f l a s k s a n d m ak e u p t o v o lu m e w i t h 0 , 5 p e r c e n t o x a l i c a c i d . U se a liq u o t s o f 5 m i l l i l i t e r s o f t h e s e sta n d a r d s t o o b ta in th e sta n d a r d c u r v e , f o llo w in g th e s t e p s d e s c r ib e d above. P lo t t h e v a lu e s o f t h e e x p r e s s io n , lo g Gs - l o g G b, on m i l l i m e t e r g r a p h p a p e r a g a i n s t t h e kn ow n a m o u n ts o f a s c o r b ic a c id in th e a liq u o t . ft ■ —- =S\ 1 < B 's' L== r F i g u r e 1 . M e c h a n ic a l a r r a n g e m e n t o f t h e c u l t u r e s . Experiment 1 Table 1. Experimental results obtained from wheat plants grown on various levels of iron in the nutrient solutions; averages expressed per 10 plants. i P arts p er m illio n of iro n in th e n u trie n t so lu tio n Grains of fresh Grams of Grams t o t a l weight dry m atter carbohydrate Grams t o t a l nitro g en Grams t o t a l M illigrams of chlorophyll iro n 0 22.3 3.65 0.57 0.13 0.073 0.307 1 22.3 3.33 0.63 0.14 0.076 0.283 2 25.6 4.20 0.59 0.16 0.094 0.437 3 32.2 5.54 0.77 0.20 0.117 0.543 4 26.5 4.36 0.62 0.15 0.092 0.340 5 27.0 4.46 0.62 0.14 0.091 0.357 6 28.9 5.24 0.74 0.17 0.106 0.419 7 22.5 4.33 0.60 0.14 0.074 0.429 8 23.9 4.33 0.63 0.14 0.088 0.342 9 22.4 5.18 0.73 0.15 0.090 0.404 Experiment 1. Table 2. Experimental results obtained from wheat plants grown on various levels of iron in the nutrient solutions; averages expressed per 10 plaits. P arts p er m illio n of iro n in th e n u trie n t so lu tio n Milligrams of M illigrams of ascorbic acid ascorbic acid a t harvest ju s t a f te r drying M illigrams of ascorbic acid 31 days a f te r drying M illigrams of ascorbic acid 105 days a f te r drying 0 3.41 2.11 1.96 1 2.83 2.27 1.37 0.31 2. 3.33 2.21 2.17 0.27 3 4.24 2.95 2.87 0.42 4 3.11 2.53 2.29 0.33 5 3.62 2.40 2.35 0.34 6 3.62 3.10 2.82 0.45 7 2.53 2.17 2.30 0.34 8 3.23 2.52 2.23 0.36 9 3.00 2.59 2.76 0.52 Experiment 1 Table 3» Experimental results obtained from wheat plants grown on various levels of copper in the nutrient solutions; averages expressed per 10 plants. P arts p er m illio n Grams of fre sh Grams of Grams t o t a l Grams t o t a l of copper in th e m atter dry m atter carbohydrate n itro g en n u trie n t so lu tio n Grams t o t a l M illigrams of chlorophyll iro n , 0.00 19.8 3.38 0.49 0.12 0.070 0.270 0.01 21.3 3.39 0.47 0.12 0.074 0.264 0.02 22.3 3.72 0.53 0.12 0.076 0.305 0.03 22.3 3.62 0.50 0.13 0.074 ' 0.275 0.04 24.0 4.10 0.57 0.13 0.075 0.320 0.05 36.8 4.34 0.60 0.15 0.034 0.356 0.06 30.4 5.07 0.72 0.16 0.092 0.385 0.07 23.9 3.70 0.52 0.14 0.081 0.274 0.03 24.7 4.23 0.59 0.14 0.069 0.334 0.09 18.5 3.05 0.44 0.09 0.053 0.220 Experiment 1 Table 4* Experimental results obtained from wheat plants grown on various levels of copper In the nutrient solutions; averages expressed per 10 plants. P a rts p er m illio n of copper in the n u trie n t so lu tio n M illigrams of ascorbic acid a t h arvest M illigrams o f ascorbic acid ju s t a f te r drying M illigrams of ascorbic acid 31 days a f te r drying M illigrams of ascorbic acid 105 days a f te r drying 0,00 2.77 2.00 1.82 0.27 0.01 2.62 1.91 1.80 0.19 0.02 2.60 2.09 2.05 0.15 0.03 2.42 2.24 2.00 0.15 0.04 3.06 2.74 2.18 0.27 0.05 3.61 2.72 2.34 0.27 0.06 4.66 3.66 2.35 0.32 0.07 3.30 2.56 2.04 0.33 0.08 3.00 2.96 2.22 0.42 0.09 2.51 2.06 1.64 0.44 Experiment 1 Table 5. Experimental results obtained from wheat plants grown on various levels of iron in tbs nutrient solutions; averages expressed per 100 grains of fresh material. P arts per m illio n of iro n i n th e n u trien t so lu tio n M illigrams of iro n Grams of Grams of t o t a l carbohydrate t o t a l nitrogen Milligrams of Grams of chlorophyll dry m atter 0 1.37 2.5 0.59 349 16.3 1 1.27 2.8 0.63 343 17.2 2 1.70 2.3 0.63 368 16.4 3 1.65 2 .4 0.61 364 17.2 4 1.28 2.3 0.58 346 16.5 5 1.32 2.3 0.52 336 16.5 6 1.45 2.6 0.60 367 18.1 7 1.91 2.7 0.63 330 19.5 8 1.43 2.6 0.57 368 18.3 9 1.66 3.0 0.60 368 21.3 Experiment 1 Table 6. Experimental results obtained from wheat plants on various concentrations of iron in the nutrient solutions; averages expressed per 100 grams of fresh material (continued). M illig r a m s o f a s c o r b ic a c id a t h arvest M illig r a m s o f a s c o r b ic a c id ju st a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 31 d a y s a f t e r d r y in g 0 15.24 9.4 8.8 ---- 1 12.70 10.2 8 .4 1.4 2 13.02 8.6 8.5 1.1 3 13.18 9.2 8.9 1.3 4 11.74 9.5 8.7 1.2 5 13.39 8.9 8.7 1.3 6 12.58 10.7 9.8 1.5 7 11.25 9.7 10.3 1.5 8 13.49 9.3 1.5 9 12.34 11.3 2.1 P a r ts p er m illio n o f ir o n in th e n u tr ie n t s o lu tio n 10.5 10.6 M illig r a m s o f a s c o r b ic acud 105 d a y s a f t e r d r y in g Experiment 1 Table 7. Experimental results obtained from wheat plants grown on nutrient solutions supplied with various amounts of copper; averages expressed per 100 grams of fresh material. P arts per m illio n Milligrams of Grams of Grams of Milligrams of of copper in the iro n t o t a l carbohydrate t o t a l n itro g en chlorophyll n u trie n t so lu tio n Grams of dry m atter, 0.00 1.37 2.5 0.6 354 17.1 0,01 1.21 2.2 0.6 339 15.6 0.02 1.37 2.4 0.5 340 16.7 0.03 1.24 2.3 0.6 332 16.3 0.04 1.34 2.4 0.5 311 17.1 0.05 1.32 2.2 0.5 315 16.2 0.06 1.27 2.4 0.5 304 16.7 0.07 1.14 2.2 0.6 338 15.5 0.08 1.35 2.4 0.5 280 17.3 0.09 1.18 2 .4 0.5 285 16.5 Experiment 1 Table 8. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of copper; averages expressed per 100 grams of fresh material. P arts per m illio n o f copper in the n u trien t so lu tio n M illigram s o f M illigram s o f ascorbic a cid ascorbic acid at harvest j u s t a fte r drying M illigram s o f ascorbic a cid 31 days a fte r drying M illigram s o f ascorb ic acid 105 days a fte r drying 0 .0 0 14*03 1 0 .1 9 .2 1 .4 0 .0 1 1 2 .0 1 8 .7 8 .3 0 .9 0 .0 2 1 2 .5 6 9 .4 9 .2 0 .7 0 .0 3 1 0 .8 7 9 .0 0 .7 0 .0 4 1 2 .7 7 1 1 .4 9 .1 1 .1 0 .0 5 1 3 .4 5 1 0 .1 8 .7 1 .0 0 .0 6 1 5 .3 3 1 2 .0 9 .4 1 .1 0 .0 7 1 3 .8 4 1 0 .7 8 .6 1 .4 0 .0 8 1 2 .1 2 1 2 .0 8 .9 1 .7 0 .0 9 1 3 .5 5 1 1 .1 6 .8 1 .6 1 0 .1 Experiment 1 . Table 9. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of iron; averages expressed per 100 grams of dry material. Parts per m illio n o f iro n in th e s o lu tio n . M illigram s ir o n . o f Grams o f t o t a l Grams o f carbohydrate nitrogen t o t a l Grams o f Grams o f fre sh ch lo ro p h y ll m aterial 0 8 .4 1 5 .6 3 .6 4 2 .1 4 612 1 7 .4 1 6 .4 3 .6 8 1 .9 9 582 2 1 0 .4 1 4 .3 3 .3 5 2 .2 4 610 3 9 .6 1 3 .9 3 .5 5 2 .1 2 581 4 7 .8 1 4 .1 3 .5 3 2 .1 0 608 5 8 .0 1 4 .0 3 .1 8 2 .0 4 606 6 8 .0 1 4 .2 3 .3 0 2 .0 3 551 7 9 .8 1 3 .7 3 .2 2 1 .6 9 513 8 7 .3 1 4 .4 3 .1 1 2 .0 1 546 9 7 .3 1 4 .0 2 .3 0 1 -.7 3 470 Experiment 1 . Table 10. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of iron; averages expressed per 100 grams of dry material, (continued) P a r ts p er m illio n o f ir o n in th e n u tr ie n t s o lu t io n M illig r a m s o f M illig r a m s o f a s c o r b ic a c id a s c o r b ic a c id a t h arvest ju st a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 31 days a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 105 days a fte r d r y in g 0 9 3 .3 5 7 .8 5 3 .8 — 1 7 4 .0 5 9 .2 4 8 .8 8 ,1 2 7 9 .4 5 2 .6 5 1 .6 6 .5 3 7 6 .6 5 3 .2 5 1 .3 7 .6 4 7 1 .3 5 3 .0 5 2 .6 7 .5 5 8 1 .1 5 3 .8 5 2 .6 7 .7 6 6 9 .3 5 9 .2 5 3 .3 8 .5 7 5 7 .7 4 9 .6 5 2 .6 7 .3 8 7 3 .7 5 7 .6 5 0 .8 8 .1 9 5 8 .0 5 0 .0 5 3 .2 1 0 .0 Experiment 1. T a b le 1 1 . E x p e r im e n t a l r e s u l t s o b t a i n e d fr o m w h e a t p l a n t s g r o w n o n n u t r i e n t s o lu t io n s v a r y in g i n t h e c o n c e n tr a tio n s o f c o p p e r ; a v e r a g e s e x p r e s s e d p e r 1 0 0 gram s o f d r y m a t e r ia l. P a r ts p er m illio n o f cop p er in th e n u tr ie n t s o lu tio n M illig r a m s ir o n of G ram s o f t o t a l ca r b o h y d r a te G ran s o f t o t a l n itr o g e n G ram s o f c h lo r o p h y ll G ram s o f f r e s h m a te r ia l 0 .0 0 8 .0 1 4 .6 3 .5 4 2 .0 7 585 0 .0 1 7 .8 1 4 .0 3 .5 5 2 .1 7 643 0 .0 2 8 .2 1 4 .2 3 .2 6 2 .0 4 600 0 .0 3 7 .6 1 3 .9 3 .7 1 2 .0 4 615 0 .0 4 7 .8 1 4 .0 3 .1 7 1 .3 2 584 0 .0 5 8 .2 1 3 .9 3 .3 6 1 .9 4 619 0 .0 6 7 .6 1 4 .3 3 .0 5 1 .8 2 600 0 .0 7 7 .4 1 4 .1 3 .3 4 2 .1 3 644 0 .0 8 7 .8 1 3 .9 3 .1 3 1 .6 2 579 0 .0 9 7 .2 1 4 .5 2 .8 7 1 .7 3 608 Experiment 1 T a b le 1 2 . E x p e r im e n t a l r e s u l t s o b t a i n e d fr o m w h e a t p l a n t s g r o w n on n u t r ie n t s o lu t io n s v a r y in g i n t h e c o n c e n tr a io n s o f c o p p e r ; a v e r a g e s e x p r e s s e d p e r 1 0 0 gram s o f d r y m a t e r ia l P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu t io n M illig r a m s o f M i l l i g r a m s o f a s c o r b ic a c id a s c o r b ic a c id at h arvest ju st a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 31 days a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 105 days a fte r d r y in g 0 .0 0 8 2 .0 5 9 .2 5 3 .8 8 .2 0 .0 1 7 7 .2 5 6 .2 5 3 .2 5 .6 0 .0 2 7 5 .3 5 6 .2 5 5 .2 4 .1 0 .0 3 6 6 .9 6 2 .0 5 5 .2 4 .1 0 .0 4 7 4 .6 6 6 .6 5 3 .2 6 .5 0 .0 5 8 3 .3 6 2 .6 5 3 .8 6 .3 0 .0 6 9 2 .0 7 2 .2 5 6 .2 6 .4 0 .0 7 8 9 .2 6 9 .2 5 5 .2 8 .8 0 .0 8 7 0 .1 6 9 .2 5 1 .8 1 0 .0 0 .0 9 8 2 .3 6 7 .6 5 3 .3 9 .4 Experiment 1 . Table 13. Percentages of the original amounts of adcorbic acid found in wheat plants grown on various levels of iron, at each determinations after the drying of the plants. P a r ts p er m illio n o f ir o n in th e n u tr ie n t s o lu tio n P e r c e n ta g e o f P e rcen ta g e o f a s c o r b ic a c id a s c o r b ic a c id a t h arvest ju st a fte r d r y in g P er c e n ta g e o f a s c o r b ic a c id 31 days a fte r d r y in g P e r c e n ta g e o f a s c o r b ic a c id 105 d a y s a f t e r d r jin g 0 100 61.7 57.7 — 1 100 80.3 66.1 11.0 2 100 66.1 65.3 8.4 3 100 69.8 67.5 9.9 4 100 80.9 74.1 10.2 5 100 66.5 65.0 9.7 6 100 85.1 77.9 11.9 7 100 86.2 91.5 13.3 8 100 77.3 68.9 11.1 9 100 85.9 91.6 17.0 Experiment 1 . Table 14. Percentages of the original anounts of ascorbic acid found in wheat plants grown on various levels of copper, at each determinations after the drying of the plants. P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu tio n P e r c e n ta g e o f P e r c e n ta g e o f a s c o r b ic a c id a s c o r b ic a c id a t h arvest ju st a fte r d r y in g P e r c e n ta g e o f a s c o r b ic a c id 31 days a fte r d r y in g P e r c e n ta g e o f a s c o r b ic a c id 105 days a fte r d r y in g 0 .0 0 100 7 2 .0 6 5 .6 1 0 .0 0 .0 1 100 7 2 .4 6 8 .1 7 .2 0 .0 2 100 7 4 .9 7 3 .3 5 .4 0 .0 3 100 9 2 .9 8 2 .7 6 .2 0 .0 4 100 8 9 .3 7 1 .3 8 .6 0 .0 5 100 7 5 .1 6 4 .7 7 .4 0 .0 6 100 7 8 .5 6 1 .3 7 .2 0 .0 7 100 7 7 .3 6 2 .1 1 0 .1 0 .0 8 100 9 9 .0 7 3 .4 1 4 .1 0 .0 9 100 8 1 .9 6 4 .9 1 1 .8 F ig u r e 2 . A v e r a g e f r e s h w e i g h t , e x p r e s s e d i n g r a m s p e r t e n w h e a t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu t io n , e x p r e s s e d i n p a r ts p e r m illio n . F ig u r e 3 . A v e r a g e d r y w e i g h t , e x p r e s s e d i n g ram s p e r t e n w h e a t p la n t s , p lo t t e d a g a in s t t h e c o n c e n tr a tio n o f ir o n in t h e n u tr ie n t s o lu t io n , e x p r e sse d i n p a r ts p e r m illio n . F ig u r e 4 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r te n w h eat p la n t s , p lo t t e d a g a in s t t h e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu tio n , e x p r e sse d in p a r ts p e r m illio n . F ig u r e 5 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d in m illig r a m s p e r t e n w h ea t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n i n th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts per m illio n . F ig u r e 6 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d j u s t a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r t e n w h ea t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n i n th e n u tr ie n t s o lu t io n , ex p ressed in p a r ts per m illio n . F ig u r e 7 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 3 1 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r t e n w h ea t p l a n t s , p lo t t e d a g a in s t t h e c o n c e n tr a tio n o f ir o n i n t h e n u tr ie n t s o lu tio n ex p ressed in p a r ts p er m illio n . JJ£,itkr FB C sm . C y R f \ f v \3 0 IX o 1% 1 8 6 p.p.M e>F o s 2. p XP»0N p M o f Jo 11 11 u Kt C o O .^ 14 - o s I P i a-oK 6. o 0. p.p.NV. & <4 c.F H ^ o tO 2i o 1tt.ot0 F ig u r e 8 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 1 0 5 d a y s a f t e r d r y i n g , e x p r e s s e d in m illig r a m s p er t e n w h ea t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n i n th e n u tr ie n t s o lu tio n , ex p ressed in p a r ts p er m illio n . F ig u r e 9 . A v e r a g e a m o u n ts o f t o t a l c a b o h y d r a t e , e x p r e s s e d i n gram s p e r t e n w h ea t p l a n t s , p lo t t e d a g a in s t th e c o n c e n tr a ­ t io n o f ir o n in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 1 0 . A v e r a g e a m o u n ts o f t o t a l n i t r o g e n , e x p r e s s e d i n g r a m s p e r t e n w h eat p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu t io n , e x p r e sse d i n p a r ts p er m illio n , F ig u r e 1 1 . A v e r a g e a m o u n ts o f c h l o r o p h y l l , e x p r e s s e d i n g ra m s p e r t e n w h ea t p la n t s , p lo t t e d a g a in s t t h e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 1 2 . A v e r a g e a m o u n ts o f n i t r o g e n , e x p r e s s e d i n gram 3 p e r t e n w h e a t p l a h t s , p l o t t e d a g a i n s t t h e am ou n t o f i r o n , e x p r e s s e d i n m illig r a m s p e r t e n w h eat p la n t s . F ig u r e 1 3 . A v e r a g e a m o u n ts o f c h l o r o p h y l l , e x p r e s s e d i n gram 3 p e r t e n w h e a t p l a n t s , p l o t t e d a g a i n s t t h e a m o u n ts o f ir o n , e x p r e s s e d i n m illig r a m s p e r t e n w h ea t p la n t s . «c a, 0 a. 9 u 1 Figure 9. 0. o 6 JS. |j>. r r \ . 1 S.OK) ,Ufe o 'o.* o oJ f 1A 0 N (.0 (U G o O 0.28 TAPt . OF i RoN o."*)^ ^ o .^)8 ©.‘i i o.48 o o f JP .6H p c r io p l a n t s F ig u r e 1 4 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r t e n w h ea t p la n t s grow n on v a r io u s c o n c e n t r a t io n s o f ir o n , p lo t t e d a g a in s t th e p e r c e n ta g e o f th e o r ig in a l am ou n t o f a s c o r b i c a c i d j u s t a f t e r d r y i n g . F ig u r e 1 5 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r t e n w h e a t p l a n t s g ro w n o n v a r i o u s c o n c e n tr a tio n s o f ir o n in th e n u tr ie n t s o lu t io n , p lo t t e d a g a i n s t t h e a m o u n ts o f i r o n , e x p t e s s e d i n m i l l i g r a m s p er te n p la n ts . A SC oR ftic ACifc Ko P£R w 10 P u f t f o T s fiTnA A A JE 'ST p i &. X** 5 GG ? ; o c m -D (O 7> 0 -o r- 2 Ef £ i r o n p e r id p l a n t s F ig u r e 1 6 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 , g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n i n t h e n u tr ie n t s o lu t io n , e x p r e sse d i n p a r ts p e r m illio n . F ig u r e 1 7 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d j u s t a f t e r d r y i n g e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a t e r ia l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f i r o n in t h e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p e r m illio n . F ig u r e 1 8 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r lO O gram s o f f r e s h m a t t e r fr o m w h e a t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in t h e n u tr ie n t s o lu t io n , e x p r e s s e d i n p a r ts p er m illio n . F ig u r e 1 9 . A v e r a g e a m o u n ts o f a s d o r b i c a c i d 3 1 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a tte r fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f ir o n in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 2 0 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 1 0 5 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p er 1 0 0 gram s o f f r e s h m a tte r fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f ir o n i n th e n u tr ie n t s o lu t io n , e x p r e s s e d i n p a r ts p e r m illio n . F ig u r e 2 1 . A v e r a g e a m o u n ts o f c a r b o h y d r a t e , e x p r e s s e d i n g ra m s p e r 1 0 0 g r a m s o f f r e s h m a t t e r fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t th e c o n c e n tr a tio n o f ir o n i n th e n u t r ie n t s o lu t io n , e x p r e se d in p a r ts p er m illio n . uJ (£ t* K t-«i D v> 0 u I0 (S3 <* 0 o *> 4 2 O 6 a 8 io a I Aow X P P H a F m 6 8 T p. o m tc to 5 DA-Ji AFTER D is)lN tr P-P- of flSt 0AA*£ ft^iO 0 o o.S Pig u r e o p.p. tt Op iA o M 2. o . Pt q u r«. Io o A p.p ea. o f 6 8 X A on . o ^£ 1 F ig u r e 2 2 . A v e r a g e a m o u n ts o f t o t a l n i t r o g e n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u t r ie n t s o lu t io n , ex p ressed in p a r ts p er m illio n . I F ig u r e 2 3 . A v e r a g e a m o u n ts o f c h l o r o p h y l l , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p o a n t s , p l o t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u t r ie n t s o lu t io n , ex p ressed in p a r ts p er m illio n . F ig u r e 2 4 . A v e r a g e a m o u n ts o f d r y m a t e r i a l , e x p r e s s e d i n g r a m s p e r 1 0 0 g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in t h e n u t r ie n t s o lu t io n , ex p ressed in p a r ts p er m illio n . F i g u r e 2 5 . A v e r a g e a m o u n ts o f t o t a l n i t r o g e n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu t io n , ex p ressed in p a r ts per m illio n . F ig u r e 2 6 . A v e r a g e a m o u n ts o f d r y m a t t e r , e x p r e s s e d i n g r a m s p e r 1 0 0 g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s g r o w n o n v a r io u s c o n c e n tr a tio n s o f ir o n in th e n u tr ie n t s o lu t io n , p lo t t e d a g a in s t th e am ount o f a s c o r b ic a c id 3 1 d a y s a f t e r d r y in g , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a te r ia l. F ig u r e 2 7 . A v e r a g e a m o u n ts o f d r y m a t t e r , e x p r e s s e d i n g r a m s p e r 1 0 0 g ra m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s g r o w n o n n u t r ie n t s o lu t io n v a r y in in c o n c e n tr a tio n o f ir o n , p lo t t e d a g a i n s t t h e p e r c e n t a g e o f t h e o r i g i n a l am ou n t o f a s c o r b i c a c i d fo u n d i n t h e f r e s h m a t t e r j u s t a f t e r d r y i n g . M J © © o 3l 6 © P I A* o 8 o e X p y N oF 8 1 R. o m f2 tu u d n ju re o 6 SL 2. h u re 8 T>E1.7 R. © o © & 8-5 q.o A Sc.o«.fty s s .5 <0.0 <0.5 R C e KJ T ft f t e R jP M iN fr ftciD ?E. To 75 go 85 9«| PcRCEMTf t Se, o p -The oPsi St Wf t u f t f Aoof J KSCO^ ^K. «t!p TtliT ft FT E <\ 3>ft *yi UC F ig u r e 2 8 . A v e r a g e a m o u n ts o f t o t a l n i t r o g e n , e x p r e s s e d i n g ra m s p e r 1 0 0 g r a m s o f d r y m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu t io n exp ressed in p a r ts p er m illio n . F ig u r e 2 9 . A v e r a g e a m o u n ts o f c h l o r o p h y l l , e x p r e s s e d i n g r a m s p e r 1 0 0 g r a m s o f d r y m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu tio n , ex p ressed in p a r ts per m illio n . F ig u r e 3 0 . A v e r a g e a m o u n ts o f f r e s h m a t e r i a l , e x p r e s s e d i n g r a m s p e r 1 0 0 g r a m s o f d r y m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u t r ie n t s o lu t io n ex p ressed in p a r ts per m illio n . P£f\ cE tsl v I Ul 0 d u< (iJ 3.0 j N iT fV of»£N s. o a. o 0 9.5 0 -J a o ^ 2.0 u0 1.5 a0 2.5 o oc t ° O O © p. © G) G F ,S u r e ■ 2. U 6 p . p. r^>. o F I R .2. 0 « js.p.M. oF I fteN Z r-r Q oF I fc o N * 8 ■ O N Figure 31. Average amounts of iro n , expressed in milligram s per 100 grams of dry m aterial from wheat p la n ts, p lo tted against th e concentration of iro n in the n u trie n t so lu tio n , express d in p a rts per m illio n . Figure 32. Average amounts of ascorbic acid ju s t a f te r drying, expressed in m illigram s per 100 grams of dry m aterial from wheat p la n ts, p lo tte d ag ain st th e concentration of iro n in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 33. Average amounts of ascorbic acid a t h arv est, expressed in milligram s per 100 grams of dry m aterial from wheat p la n ts, p lo tte d against the concentration of iro n in the n u trie n t solution?, expressed in p a rts per m illio n . Figure 34. Average amounts of ascorbic acid 31 days a f te r drying, expressed in milligrams per 100 grams of dry m aterial from wheat p la n ts, p lo tte d against th e concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 35. Average amounts of ascorbic acid lOr days a f te r drying, expressed in m illigram s per 100 grams of dry m aterial from wheat p la n ts, p lo tted against th e concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 36. Average amounts of carbohydrate, expressed in grams per 100 gramms of dry m aterial from wheat p la n ts, p lo tted against the concentration of iro n in the n u trie n t so lu tio n expressed in p a rts per m illio n . h- '7 l» z «£• - t-> 02 to 0 rf H o 60 o So u re. "2>%. to ■? a; O f i IRON/ 3 fN D P I a A * * 15 <£ fZ u j ‘H <£ o o © o O © O o r £ 3 5. IO p.p . O F I R O N I I. ■!■ . ■ I 3, H 3 u r e. 1. NA. o F p .p . 3 6. Figure 37. Average amounts o f ascorb ic a c id , expressed in m illigram s per 100 gram3 o f dry m aterial from wheat p la n ts , p lo tte d again st the amount o f ir o n , expressed in m illigram s per 100 grams o f dry m atter. Figure 38. Average amounts o f ch lo ro p h y ll, expressed in grams per 100 grams o f fr e sh m atter from wheat p la n ts grown on n u trien t s o lu tio n s supplied w ith various concentrations o f ir o n , p lo tte d a gain st the amount o f ir o n , expressed in m illigram s per 100 grams o f fr e sh matter* t•n lu ■p d A X 9o 70 Pi.au r e. (Ay. 1 *0 N 2>7 I O. I 87 ■?£«. 109 w. ?>&.> (M UTTER. •o 00 360 a. 4S0 33o. u re I. <■ M i ai«.*N PER 100 iS . i.« GRRM i. F S.E S H MATTER Figure 39. Average amounts of ascorbic acid 105 days a f te r drying, expressed in m illigram s per 10 wheat p la n ts , p lo tted against th e concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 40. Average amounts of t o t a l carbohydrate, expressed in grams per 10 wheat p la n ts , p lo tte d ag ain st the concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 41. Average amounts of t o t a l n itro g en , expressed in grams per 10 wheat p la n ts , p lo tte d ag ain st the concentration of copper in th e n u trie n t s o ltu tio n , expressed in p a rts per m illio n . Figure 42. Average amounts of chlorophyll, expressed in grams per 10 wheat p la n ts, p lo tte d ag aisn st th e concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 43. Average t o t a l carbohydrate, expressed in grams p er 10 p la n ts, grown on n u trie n t so lu tio n supplied with various concentations of copper, p lo tte d against th e amount of iro n , expressed in m illigram s per 10 p la n ts . Figure 44. Average dry w eight, expressed in grams per 10 p la n ts, grown on n u trie n t so lu tio n s supplied with various con­ ce n tratio n s of copper, p lo tted against the amount of iro n , expressed in m illigram s per 10 p la n ts. cL VJ t— u- UJ 1• :i; o a tp o 0 o o <0 t G o 0 0 cC O1 j 28 O oi oF CoPPeft. 0 0 *f p p> • a. 5 0.5 o _1 a. ib a X 0 0 © G Q 0 Q O O geo o 2 tii _L. *•» 0.0*2. £ o.oH p .p . M. O.00 oF o 0© 0.08 C oPP£« '-:j> ^ 5.0 H cf <* 7 a o oS, o.oH o-(>£> 5.0 J p p. M. oF C o P P E R 0 0R.5 © O \s^ It 2 cC o o $ © o o P3.0 F ibo R£ ^ 3 • 0.9.0 o a ? o.3 o Me-. I Rom o'iS’ 0 . ^ 0 PfR |o P- ^0 tX lb LJ3.S 3 >oC % 0 05 O Pl'iURE ib o.l o 7S o 2. S' o . i o M &. i R o f J o .3 S PER o.<*o lo PLfcNTj Figure 45. Average fre sh weight, expressed in grams per 10 wheat p la n t8, p lo tted ag ain st the concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 46. Average amounts of dry m a te ria l, expressed in grams per 10 wheat p la n ts , p lo tte d against th e concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 47. Average amounts of iro n , expressed in m illigram s per 10 wheat p la n ts , p lo tte d against th e concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 48, Average amounts of ascorbic acid a t h arv est, expressed in m illigram s per 10 wheat p la n ts, p lo tte d against th e concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 49. Average amounts of ascorbic acid ju s t a f te r drying, expressed in milligram s per 10 wheat p la n ts , p lo tte d ag ain st the concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 50. Average amounts of ascorbic acid 31 days a f te r drying, expressed in m i l l i gram s per 10 wheat p la n ts , p lo tted ag ain st the concentrations of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . »- a ■c n tu o. (t; £> '1 ul 3 / • o t o 3 Fi.c^o r C I- o1 oOH oF o 06 oo CO PpC A uV -> ■o o 0 -01. O.o^ p p fA .0 8 P96 « K, 4 0 .0 6 0 08 wD?P £ * 4 Ui Ui <£ •< 3* u G J I* a O VU‘ i/l *r •»* Q q J, A AL/ p.p.fA . A rt ^ fl r t fi oF Co?P>1P- 0 © 0 0 Q o .Oo _i_ ■X. o op o ox o.o** o . o s o f c o m p. r r M 4 k Figure 51. Average amounts of iro n , expressed in m illigram s per 100 grams of fresh m a terial from wheat p la n ts , p lo tted ag ain st the concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 52. Average amounts of ascorbic acid ju s t a f te r drying, expressed in m illigram s per 100 grams of fresh m a terial from wheat p la n ts , p lo tte d ag ain st th e concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 53. Average amounts of ascorbic acid a t h arv est, expressed in m illigram s per 100 grams of fre sh m a terial from wheat p la n ts p lo tte d ag ain st th e concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 54. Average amounts of ascorbic acid 31 days a f te r h arv est, expressed in m illigram s per 100 grams of fre sh m aterial from wheat p la n ts, p lo tte d against th e concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 55. Average amounts of ascorbic acid 105 days a f te r drying, expressed in milligram s per 100 grams o f fre sh m aterial from wheat p la n ts , p lo tte d ag ain st th e concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 56. Amounts of carbohydrate, expressed in m illigram s per 100 grams of fresh m a te ria l from wheat p la n ts , p lo tted ag ain st the concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . ct A t o£ o ui *• M* ?ch T /7 17 I b t vj J .|< l>1 oK 0.0 6 0 Of l c«rr' «. *V OIO *» 2 ^ MAR.UES1 ?ER, CEwl 11 t) tf 1.5 tA c,. IftofJ £ 0 .0 1 O 0*t 0 OF r r o4> O oft 3 to COPPtLR 2 AjtOR.%ic PtciD (O 13 10 12 e ti 8 0 oo 0 0 2 . 00*1 o F o Of f 0.10 o o 2. o oo C o PP C R PT 0 08 o o 4* o.ofe fo o r o.to C o P P f R, D«T5 AFTER, NV o o 6 i o S vi; ® © © © G ACiD 0 *SLO«.e>*c- ti'/ 0 0 £ 2 . 0° • ► tfr ■--£ .5”6 ‘ o .o o . o F eoPpFB. o o 2 o o 4* 0 0 6 o 08 p j).M. o f t o p p e d . o.io Figure 57. Average amounts of t o t a l n itro g en , expressed in m illigram s p er 100 grains of fresh m a te ria l from wheat p la n ts , p lo tted ag ain st th e concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 58. Average amounts of chlorophyll, expressed in milligram s per 100 grams o f fre sh m a terial from wheat p la n ts , p lo tted ag ain st th e concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 59. Average amounts of dry m a te ria l, expressed in grams per 100 grams of fresh m a te ria l from wheat p la n ts , p lo tte d ag ain st th e concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 60. Average amounts of dry m a te ria l, expressed in grams per 100 grams of fre sh m a terial from wheat p la n ts grown on n u trie n t so lu tio n s supplied with various concentrations of copper, p lo tte d ag ain st the amounts of iro n , expressed in m illigram s per 100 grams of fre sh m a te ria l. Figure 61. Average amounts of t o t a l carbohydrate, expressed in m illigrams p er 100 grams o f fresh m aterial from wheat p lan ts grown on n u trie n t so lu tio n s supplied with various concentrations of copper, p lo tte r against the amounts of iro n , expressed in m illigrams per 100 grams of fresh m a te ria l. Figure 62. Average amounts of dry m a terial, expressed in grams per 100 grams of fresh m a terial from wheat p la n ts grown on n u trie n t so lu tio n supplied with various concentrations of copper, p lo tte d agaisnt the percentage of th e o rig in a l amount of ascorbic acid ju s t a f te r drying. 350 310 270 0.00 0 .02 p. 0 0 o 08 >F co p p eR 'i i <1 17.5 16.0- 1 a d d a or l4* i 0 .0 0 0.02- O.O^i p . p . m . o f 0.0 b 0. 00 OOft c o p p e R O.OX o . o v 0.06 0.08 p . p nr* o r coppE R u) \- d P 6 d) 0) 0 i i S' U 0 o « 2 - o* Me, e. P T« ° F 0*»ioR (i»c f\CiC ?£K«_Gr»\ o ur £ O.oo 0.07 . RfTER p p o oM |*\ . o f 6 5^ o.ofe ore p.p.**. o f CoPPtO. o P C o f P E P o H S :o«.a.c ftC.D 105" 8 o o 2. O.oo 0 .0 2 o . o , N■+ <3k 3i» £2 • *m *m* ? s 0 i\ £ 7t 13 0 0 0 0 0 0 0 >)l 0 3 Q JOk, 30. oof l> j. t* ^ oi’K r . Figure 75. Percentages of th e o rig in a l amounts of ascorbic acid, ju s t a f te r drying, 31 and 105 days a f te r drying, in the tis s u e s from wheat p lan ts grown on n u trie n t so lu tio n s supplied w ith various amounts of iro n , p lo tted against th e amount of iro n , expressed in milligram s per 100 grams of dry m atter. Figure 76. Percentages of th e o rig in a l amounts of ascorbic acid ju s t a f te r drying, 31 and 105 days a f te r drying, in th e tis s u e s from wheat p la n ts grown on n u trie n t so lu tio n s supplied w ith various amounts of copper, p lo tted agaisnst the amount of iro n , expressed in milligram s per 100 grams of dry m atter. A 0 © O © 0 0 , A © A 0 O A A © A 0 JwiT & G It S J A , > A M f A. O^'riWU OAli AFTER, I>A-> i fi (* P I orOft.6 7s 7?ao M > - * - . K. /*/ IS i i ■>, , _L_ <0 Experiment 2. Table 15. Experimental results obtained from ■wheat plants grown on nutrient solutions varying in concentrations of iron; averages expressed per 10 plants. P arts per m illio n of iro n in the n u trie n t so lu tio n Fresh weight Grams of in grams dry m atter Milligrams M illigrams of M illigrams of of iro n ascorbic acid ascorbic acid a t harvest ju s t a f te r drying Milligrams of ascorbic acid 7 days a f te r drying M illigrams of ascorbic acid 14 days a f te r drying 0 6.84 0.84 0.076 1,97 1.30 0.95 0.89 e 6.41 0.93 0.142 3.12 1.10 0.31 0.29 4 7.28 0.95 0.114 2.94 1.20 0.69 0.45 6 7.79 1.02 0.184 3.24 1.34 0.67 0.62 8 7.70 1.00 0.208 2.82 1.22 0.82 0.79 10 7.40 0.98 0.201 3.05 1.31 1.10 0.77 12 7.72 1.00 0.243 3.25 1.34 0.80 0.82 14 7.33 0.94 0.216 2.69 1.42 1.08 0.62 16 7.40 0.90 0.189 2.57 1.35 0.36 0.83 18 9.20 0.98 0.255 2.58 1.11 0.95 0.72 Experiment 2 . Table 16. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of copper; averages expressed per 10 plants. P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu t io n F r e s h w e ig h t in graas Gram s o f d ry m a tte r M illig r a m s o f ir o n M illig r a m s o f M illig r a m s o f a s c o r b ic a c id a s c o r b ic a c id a t h arvest ju st a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 7 days a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 14 days a fte r d r y in g O.OO 7 .3 6 0 .9 1 0 .2 1 4 3 .2 3 1.16 0 .7 5 0 .8 2 0.02 6 .7 0 0 .8 9 0 .1 5 0 2 .4 7 1.20 0 .4 4 0 .2 2 0 .0 4 7 .02 0 .9 6 0.168 2.43 1 .3 4 0 .7 3 0 .7 9 0 .0 6 7.13 1 .0 0 0 .1 8 8 2 .8 9 1 .0 9 0 .2 9 0 .1 6 0 .0 8 7.66 0 .9 1 0 .0 9 4 2 .7 6 1.60 1 .1 5 0 .9 3 0.10 6 .9 4 0 .88 0 .1 1 9 2.32 1.38 0 .8 6 0 .6 5 0 .1 2 7 .7 3 0 .9 3 0.1 6 3 2 .9 5 1 .6 4 0 .9 3 0.98 0 .1 4 7 .7 8 1 .0 2 0 .2 0 9 3 .0 9 1 .4 1 0 .9 9 1.C3 0 .1 6 7 .3 5 0 .9 8 0.176 2 .5 5 1 .0 9 0 .1 9 0.20 0 .1 8 8 .1 1 0 .9 6 0 .1 3 7 2 .9 0 1 .5 6 0 .9 5 0 .8 7 Experiment 2 Table 17. Experimental results from wheat plants grown on nutrient solutions varying in concentrations of iron; averages expressed per 100 grams of fresh material. P arts per m illio n of iro n in the n u trie n t so lu tio n Milligrams iro n of Milligrams of Milligrams of M illigrams of Milligrams of Grams of ascorbic acid ascorbic acid ascorbic acid ascorbic acid dry m atter a t harvest ju s t a f te r 7 days a f te r 14 days a f te r drying drying drying 0 1.09 28.6 18.9 13.3 12.9 12.2 2 2.22 48.7 17.2 4.8 4.5 14.5 4 1.57 40.6 16.5 9.6 6.2 13.1 6 2.36 41.7 17.2 8.6 7.3 13.1 8 2.70 36.7 15.3 10.7 10.3 13.0 10 2.70 41.1 17.6 14.3 10.4 13.2 12 3.16 42.2 17.4 10.3 10.6 13.0 14 2.94 36.6 19.4 14.7 8.4 12.8 16 2.56 34.9 18.2 11.7 11.3 12.2 18 3.33 33.6 14.6 12.4 9.4 12.8 Experiment 2. Table 18. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of copper; averages expres3ed per 100 grains of fresh material. P arts per m illio n of copper in th e n u trie n t so lu tio n Milligrams of M illigrams of M illigrams of iro n ascorbic acid ascorbic acid a t harvest ju s t a f te r drying M illigrams of ascorbic acid 7 days a f te r drying Milligrams of Grams of ascorbic acid dry m atter 14 days a f te r drying 0.00 2.39 43.7 15.7 10.1 11.1 12.3 0.02 2.23 36.3 17.9 6.6 3.3 13.3 0.04 2.38 34.4 13.9 10.4 11.2 13.6 0.06 2.63 40.4 15.2 4.1 2.2 14.0 0.08 1.23 36.1 20.9 15.0 12.2 11.9 0.10 1.70 33.3 19.7 12.4 9.3 12.6 0.12 2.12 38.4 21.3 12.1 12.8 12.1 0.14 2.69 39.7 18.1 12.7 13.2 13.1 0.16 2.39 34.6 14.9 2.6 2.3 13-3 0.18 1.69 35.8 19.1 11.7 10.6 11.8 Experiment 2 Table 19. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of iron; averages expressed per 100 grams of dry plant material. P A r ts p e r m i l l i o n o f ir o n in th e n u tr ie n t s o lu t io n M illig r a m s o f M illig r a m s o f M illig r a m s o f ir o n a s c o r b ic a c id a s c o r b ic a c id a t h arvest ju st a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 7 days a fte r d r y in g M illig r a m s o f a s c o r b ic a c id 14 da*s a fte r d r y in g G rans o f f r e s h m a t te r 0 9 .0 235 1 5 5 .1 1 1 3 .0 1 0 5 .6 820 2 1 5 .3 336 1 1 8 .6 3 3 .4 3 0 .7 690 4 1 2 .0 310 1 2 6 .1 7 2 .9 4 7 .2 763 6 1 8 .0 318 1 3 1 .6 6 5 .3 6 0 .9 763 8 2 0 .9 282 1 2 1 .3 8 2 .2 7 8 .9 769 10 2 0 .5 311 1 3 3 .5 1 1 2 .4 7 8 .9 758 12 2 4 .3 325 1 3 3 .9 7 9 .6 8 1 .7 769 14 2 3 .0 286 1 5 1 .3 1 1 5 .1 6 6 .0 781 16 2 1 .0 286 1 4 9 .5 9 5 .6 9 2 .5 820 18 2 6 .0 263 1 1 2 .9 9 6 .7 7 3 .6 781 Experiment 2 Table 20. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of copper; averages expressed per 100 grams of dry material. Parts per m illio n o f copper in th e nu trient s o lu tio n M illigram s o f , M illigram s o f M illigrams o f iro n ascorbic acid ascorb ic acid at harvest ju s t a ft e r drying M illigram s o f ascorb ic acid 7 days a fte r drying M illigram s o f ascorb ic acid 14 days a fte r drying Grams o f fre3h m atter 0 .0 0 2 3 .5 355 1 2 7 .3 8 2 .4 9 0 .1 813 0 .0 2 1 6 .3 277 1 3 4 .8 4 9 .9 2 4 .3 752 0 .0 4 1 7 .5 253 1 3 9 .2 7 6 .3 8 2 .5 735 0 .0 6 1 8 .3 289 1 0 8 .6 2 9 .3 1 5 .8 714 0 .0 8 1 0 .3 303 1 7 5 .7 1 2 6 .1 1 0 2 .7 840 0 .1 0 1 3 .5 264 1 5 7 .0 9 8 .2 7 3 .9 794 0 .1 2 1 7 .5 317 1 7 6 .1 9 9 .8 1 0 5 .9 826 0 .1 4 2 0 .5 303 1 3 8 .2 9 7 .1 1 0 0 .9 763 0 .1 6 1 8 .0 260 1 1 1 .7 1 9 .6 2 0 .8 752 0 .1 8 1 4 .3 303 1 6 2 .1 9 9 .1 9 0 .2 847 Experiment 2 . Table 21. Percentages of the original amounts of ascorbic acid found in wheat plants grown on nutrient solutions varying in concentrations of iron, at each determination after the drying of the plants. P a r ts p e r m illio n o f ir o n in th e n u tr ie n t s o lu t io n P e r c e n ta g es o f a s c o r b ic a c id a t h arvest P e r c e n ta g es o f a s c o r b ic a c id ju st a fte r d r y in g P e r c e n ta g es o f a s c o r b ic a c id 7 days a fte r d r y in g P e r c e n ta g es o f a s c o r b ic a c id 14 days a fte r d r y in g . 0 100 6 6 .1 4 8 .1 4 5 .0 2 100 3 5 .2 9 .9 9 .1 4 100 4 0 .6 2 3 .5 1 5 .2 6 100 4 1 .3 2 0 .5 1 9 .2 8 100 4 3 .1 2 9 .1 2 8 .0 10 100 4 2 .9 3 6 .1 2 5 .4 100 4 1 .2 2 4 .5 2 5 .1 14 100 5 2 .9 4 0 .2 2 3 .1 16 100 5 2 .2 3 3 .4 3 2 .2 18 100 4 2 .9 3 6 .3 2 8 .0 12 Experiment 2 . Table 22. Percentages of the original amounts of ascorbic acid found in wheat plants grown on nutrient solutions varying in concentrations of copper, at each determination after the drying of the plants. P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu t io n P e r c e n ta g eso f a s c o r b ic a c id a t h arvest P e r c e n ta g es o f a s c o r b ic a c id ju st a fte r d r y in g P e r c e n ta g es o f a s c o r b ic a c id 7 days a fte r d r y in g P e r c e n ta g es o f a s c o r b ic a c id 14 days a fte r d r y in g . 0 .0 0 100 3 5 .8 2 3 .2 2 5 .4 0 .0 2 100 A S .6 1 8 .0 9 .0 0 .0 4 100 5 5 .0 3 0 .2 3 2 .6 0 .0 6 100 3 7 .5 1 0 .1 5 .5 0 .0 8 100 5 8 .0 4 1 .6 3 3 .9 0 .1 0 100 5 9 .5 3 7 .2 2 8 .0 0 .1 2 100 5 5 .6 3 1 .5 3 3 .4 0 .1 4 100 4 5 .6 3 2 .1 3 3 .3 0 .1 6 100 4 3 .0 7 .5 8 .0 0 .1 8 100 5 3 .5 3 2 .7 2 9 .8 F ig u r e 7 7 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 p l a n t s , p l o t t e d a g a i n s t t h e am o u n t o f i r o n i n t h e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p e r m illio n . F ig u r e 7 8 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m illig r a m s p e r 1 0 w h ea t p la n t s , p lo t t e d a g a in s t t h e am ount o f ir o n i n t h e n u t r ie n t s o l u t i o n , e x p r e s s e d i n p a r ts per m illio n . F ig u r e 7 9 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d j u s t a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r 10 w h eat p la n t s , p lo t t e d a g a i n s t t h e am ou n t o f i r o n i n t h e n u t r i e n t s o l u t i o n , ex p ressed in p a r t3 p er m illio n . F ig u r e 8 0 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 7 d a y s a f t e r d r y i n g , e x p r e sse d i n m illig r a m s p e r 10 w h eat p la n t s , p lo t t e d a g a i n s t t h e am ou n t o f i r o n i n t h e n u t r i e n t s o l u t i o n , ex p ressed in p a r ts per m illio n . F ig u r e 8 1 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 1 4 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r 10 w h ea t p la n t s , p lo t t e d a g a in s t t h e am ount o f ir o n i n t h e n u t r ie n t s o l u t i o n , ex p ressed in p a r ts per m illio n • F ig u r e 8 2 . A v e r a g e a m ou n t o f f r e s h w e i g h t , e x p r e s s e d i n m illig x * a m s p e r 1 0 w h e a t p l a n t s , p l o t t e d a g a i n s t t h e a m ou n t o f ir o n in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p e r m illio n . I- 0 A 11 *1 OoS 77. H 6 p.p. f*\. 13. 16 OF I (\o fj ia oP I Row in (U 13. ©F o I t> 8 2 fto8 UJ a? © ro 0 0 0 o 0 li. 0 F.u wr e 0 o p p.M. £>F 'J fi.C 'tJ >i 0 p . PT, 12. 8 O^ it 82. Figure 83. Average amounts of dry weight, expressed in gram3 per 10 wheat p la n ts , p lo tte d ag ain st th e amount of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 84. Average amounts of ascorbic acid a t h arv est, expressed in m illigram s per 10 wheat p la n ts, p lo tte d ag ain st the amount of iro n , expressed in m illigram s per 10 wheat p lan ts Figure 8f>. Average percentages of the o rig in a l amount of ascorbic acid ju s t a f te r drying in wheat p la n ts grown on n u trie n t so lu tio n supplied w ith various amounts of iro n , p lo tted against the amount of iro n , expressed in milligram s per 10 wheat p la n ts. Figure 86. Average percentages of the o rig in a l amount of ascorbic acid 7 days a f te r drying, in wheat p la n ts grown on n u trie n t so lu tio n s supplied with various amounts of iro n , p lo tte d against th e amount of iro n , expressed in m illigram s per 10 wheat p la n ts. Figure 8 7 . Average percentage of th e o rig in a l amount of ascorbic acid 14 days a f te r drying, in wheat p la n ts grown on n u trie n t so lu tio n s supplied with various amounts of iro n , p lo tte d against the amount of iro n , expressed in m illigram s per 10 wheat p la n ts. Figure 88, Average amounts of dry weight, expressed in grams per 10 wheat p lan ts grown on n u trie n t so lu tio n s supplied with various amounts of iro n , p lo tte d against the amount of iro n , expressed in milligram s per 10 wheat p la n ts. i/> uJ O Io 8 o 07 O.lfO Oi* er tl I A «M p e ­ «c Is © *» /, * © 0- 'Pi © C r- 0 ,Vi. P - © © © IS 00 © oi.*? Mb-. r f t o w f>ef* o1-'A io . o.l plan a .j oo X 0 0 0 >.60 o- K 0 0.9 b G X 'Tl © © 0 © 0 0 d w sV 0 tb io.^o 9 0? © J o.W 0 P © -i^u m. © > Oo7 o ■I( Mti. T k o \S <\n o .'i *( 0 || PI'AV >9$ .> -5Ar^i ■>¥&»« F ig u r e 8 9 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e am ou n t o f i r o n i n t h e n u t r i e n t s o lu tio n , ex p ressed in p a r ts p er m illio n . F ig u r e 9 0 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g ra m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e am ou n t o f i r o n in th e n u tr ie n t s o lu tio n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 9 1 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d j u s t a f t e r d r y i n g , e x p r e s s e d in m illig r a m s p e r 1 0 0 gram s o f f r e s h m a t e r ia l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e a m o u n t o f i r o n in t h e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 9 2 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 7 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a t e r ia l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e am ou n t o f i r o n in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p e r m illio n . F ig u r e 9 3 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 1 4 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a t e r ia l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e a m ou n t o f i r o n in t h e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 9 4 , A v e r a g e a m o u n ts o f d r y m a t e r i a l , e x p r e s s e d i n g ra m s p e r 1 0 0 g ra m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e am ou n t o f i r o n i n t h e n u t r i e n t s o lu tio n , ex p ressed in p a r ts per m illio n . *ui n <* a I i.o H d as > £ J <2 Bo i.i.ro.|V\. :x«.olO 2. \o it n it iS k p 8 8 ‘3- J3 . I A , 12. . p . m. I ^ i O N i P. o t\J Ifc 8 . fv\, r r>N p.p.ra iR o M F ig u r e 9 5 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g ra m s o f f r e s h m a t e r i a l fr o m w h ea t p la n t s grow n on n u t r ie n t s o lu t io n s u p p lie d w ith v a r i o u s a m o u n ts o f i r o n , p l o t t e d a g a i n s t t h e am ou n t o f i r o n , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a te r ia l. F i g u r e 9 6 . A v e r a g e am ou n t o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g ra m s o f f r e s h m a t e r i a l fr o m w h e a t p la n t s grow n on n u t r ie n t s o l u t i o n s u p p lie d w ith v a r io u s a m o u n ts o f i r o n i n t h e n u t r i e n t s o l u t i o n , p l o t t e d a g a i n s t t h e a m ou n t o f d r y m a t t e r , e x p r e s s e d i n g r a m s p e r 1 0 0 g r a m s o f fr e sh m a tte r . F ig u r e 9 7 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d j u s t a f t e r d r y i n g , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g ra m s o f f r e s h m a t e r i a l fr o m w h e a t p la n t s grow n on n u t r ie n t s o l u t i o n s s u p p lie d w ith v a r io u s a m o u n ts o f i r o n , p l o t t e d a g a i n s t t h e a m o u n t o f i r o n , e x p r e s s d i n m i l l i g r a m s p e r 1 0 0 g ra m s o f f r e s h m a t e r i a l . F ig u r e 9 8 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 7 d a y s a f t e r d r y i n g , e x p r e s s e d in m illig r a m s p e r 1 0 0 gram s o f f r e s h m a t e r ia l fr o m w h e a t p l a n t s g r o w n on n u t r i e n t s o l u t i o n s s u p p l i e d w it h v a r i o u s a m o u n ts o f i r o n , p l o t t e d a g a i n s t t h e a m o u n t o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g ra m s o f fr e s h m a te r ia l. F ig u r e 9 9 * A v e r a g e a m o u n ts o f a s c o i ’b i c a c i d 1 4 d a y s a f t e r d r y in g e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a t e r ia l fr o m w h e a t p l a n t s g ro w n o n n u t r i e n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o u n ts o f i r o n , p l o t t e d a g a i n s t t h e am ount o f i r o n , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f fr e s h m a te r ia l HP.A.O ('KORRh. t-Vnp ft"T Ui 30 I0..O 1.0 MG-. T>G.yi»ic, p«*“ X«o© ■ M fcr. n lo I. S KJJ-. X« f~Ve * h M4 TTq y CW Ih i V R F T £ R M&. ASCo«.*\C a>»»- 15, 1 •1 .O 1 M£- X A orJ p« r~ | o a o. , fVe s h 3. 0 3 .0 AYS pet- (0 0 ^ . ^ FVy ^ ttw p a t' loo g . l.o i.ff mC-. IR oM rvy&tl-ct'* -2.o p cv UJ Iu L. oc « ill <£ ft fc— u <£ 0 IH y o vJ cC »«„* ‘O £ <£ vb 0 . £ I.® M tr I4.0M I.S P£A loo tr, pre i h rn*ttetr I.o 4a & f J. 1R |. ? MR. X ft »t\J 2 tf JI.O © £.•» |j a f Io ° ^ • f t rrj l i ^ A.IU'I' 0 d «« a o o © ■£- — O Io Q 0 f^C* . af3ofJ iT? I o S’. 57o © 57r © 3.0 ■r,t' *• {V Cii ^ © 7$0 7oo PV«\ u r't’ © o 8 [».{».w . o f 6 i f rof J © p . p . m . 8 IX OF IR.O(\/ 16 Figure 112. Average amounts of ascorbic acid at h arv est, expressed in m illigram s per 100 grams of dry m aterial from wheat p la n ts grown on various n u trien t so lu tio n supplied with d iffe re n t amounts of iro n , p lo tted against the amount of iro n , expressed in m illigram s per 100 grams of dry m aterial. Figure 113. Average amounts of ascorbic acid 7 days a f te r drying, expressed in m illigrams per 100 grams of dry m aterial from wheat p la n ts grown on n u trie n t so lu tio n s supplied with various amounts of iro n , p lo tted ag ain st the amount of iro n in th e p lan t m a terial, expressed in m illigrams per 100 grams of dry m a terial. v4 . 2 - a c«e u c. 'J , '3 o •j M&. © Nf t c »' (co a , dry ^ ‘6 M& . IR » M dw y l? £o 9.5" V 4»o OO'O 05c 0 01' 0 C O1t 51 ^ P F i g u r e 1 2 0 . A v e r a g e am ou n t o f d r y m a t t e r , e x p r e s s e d i n g r a m s p e r 1 0 w h e a t p l a n t s , p l o t t e d a g a i n s t t h e a m ou n t o f co p p er in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F i g u r e 1 2 1 . A v e r a g e a m o u n ts o f d r y w e i g h t , e x p r e s s e d i n g r a m s p e r 1 0 w h e a t p l a n t s g ro w n o n n u t r i e n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o fln ts o f c o p p e r , p l o t t e d a g a i n s t t h e am ou n t o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 p l a n t s . F ig u r e 1 2 2 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 w h ea t p la n t s grow n on n u t r ie n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o u n ts o f c o p p e r , p l o t t e d a g a i n s t t h e a m ou n t o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 p la n ts . F ig u r e 1 2 3 . A v e r a g e a m o u n ts o f f r e s h w e i g h t , e x p r e s s e d i n g r a m s p e r 1 0 w h e a t p l a n t s g ro w n o n n u t r i e n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o u n ts o f c o p p e r , p l o t t e d a g a i n s t t h e a m ou n t o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 p l a n t s . F ig u r e 1 2 A . A v e r a g e a m o u n ts o f a s c o r b i c a c i d j u s t a f t e r d r y i n g , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 w h e a t p l a n t s g ro w n o n n u t r i e n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o u n ts o f c o p p e r , p l o t t e d a g a i n s t t h e am ou n t o f i r o n , e x p r e s s e d in m illig r a m s p e r 10 w h eat p la n t s . F ig u r e 1 2 5 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 7 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r 1 0 p la n t s grow n on n u t r ie n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o u n ts o f c o p p e r , p l o t t e d a g a i n s t t h e a m ou n t o f i r o n , e x p r e s s e d i n m i l l i g r a m s p er 10 p la n ts . t*\Cr. A S C o r t t i c ftciD h* — J 'o S T in A f T g R Mtar. i s r j i N t fST >-o \j> sa. j 0- H ftR vltsV o' 3 e HI ^ »» 0 z q©‘ •n c> 9 00 n tr. A ita r B it ^ ^ Kcij> 7 "PAVi <> t/\ o -L/ Or ArTE R. Jp r y iw& p -© r ~ Faech o^e^h* -w ^ «y r ^ \ j » - r - r c r- F ig u r e 1 2 6 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e a m ou n t o f c o p p e r i n t h e n u t r i e n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n * F ig u r e 1 2 7 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g r a m s o f f r e s h m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e am ou n t o f c o p p e r i n th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 1 2 8 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d j u s t a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a t e r ia l fr o m w h e a t p l a n t s , p l o t t e d a g a i s n s t t h e am ount o f c o p p e r in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n F ig u r e 1 2 9 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 7 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p er 1 0 0 gram s o f f r e s h m a te r ia l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e am o u n t o f c o p p e r in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts per m illio n . F ig u r e 1 3 0 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d 1 4 d a y s a f t e r d r y i n g , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f f r e s h m a t e r ia l fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e am o u n t o f c o p p e r in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 1 3 1 . A v e r a g e a m o u n ts o f 1 0 0 g ra m s o f f r e s h a g a i n s t t h e am ou n t ex p ressed in p a r ts d r y m a t t e r , e x p r e s s e d i n gram s p e r m a t e r i a l fr o m w h e a t p l a n t s , p l o t t e d o f cop p er in th e n u tr ie n t s o lu t io n , p er m illio n . (/> OJ v ib . 0.0 0 0.04 o .o s p . p . u o. £ s a i ‘1 "J. 0.14 0.0*1 C o P P E f l , t/iia. >- r vj - r . w jft.£ 11.S . 0 .0 H r >p . K\ . fi 0.0 8 o . 13L p.p.tv\ coPP«*- m -j p < r ihr 8 <£ O 3 llJ. © © p . p . « f t., G p . ir iifc. E. >.oo 0.00 o. I £. © IX Fi o .c K o. I X co p p e r. ^ . o,o3. . Xo . 3>«.y oM Ao ,« r »e»oaj.T>«*y «v/\*TTeft. v*>?> Figure 135. Average amounts of iro n , expressed in m illigrams p er 100 grains of dry m aterial from wheat p la n ts, p lo tted against the amount of copper in th e n u trien t so lu tio n , expressed in part3 per m illio n . Figure 136. Average amounts of ascorbic acid a t h arv est, expressed in m illigram s per 100 grams of dry m aterial from wheat p la n ts , p lo tte d against the amount of copper in th e n u trie n t so lu tio n , expressed in p a rts pe r m illio n . Figure 137. Average amounts of ascorbic acid ju s t a f te r drying, expressed in m illigram s per 100 grams of dry m aterial from wheat p la n ts , p lo tte d against the amount of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 138. Average amounts of ascorbic acid 7 days a f te r drying, expressed in m illigram s per 100 grams of dry m aterial from wheat p la n ts , p lo tte d against the amount of copper in th e n u trie n t so lu tio n , expressed in p arts per m illio n . Figure 139. Average amounts of ascorbic acid 14 days a fte r.d ry in g , expressed in m illigram s per 100 grams of dry m aterial from wheat p la n ts, p lo tte d against th e amount of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 140. Average amounts of fresh m a terial, expressed in grams per 100 grams of dry m atter from wheat p la n ts, p lo tted against the amount of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . vc/> •3 d *<<0 G © £ o O ILo © G © © G © 0 lo° \"ll o.o^f o.i3 . 0.08 p.p.K V . o.o v 0 oi<» o.o o o.ia o. i fe p p e r oo. 0.08 o.4 d CAo t o P P f R © © Ft'c^oR.(i p .p . fl’q oR 6 o .o o O.oo Co P f E R p . p . ro. © o.oo o.oU p .p .N \. F»’^ u «. e 0.08 0.13. t & P p G R . © i^o. o .l6 . Experiment 3 Table 23. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of iron; averages expressed per 10 plants. P a rts per m illio n Milligrams of M illigrams of M illigrams of of iro n in the ascorbic acid calcium magnesium n u trie n t so lu tio n a t harvest Grans of fre sh weight 0 1 7.5 14 47 2 8 .1 3 3 0 .1 12 64 4 5 .4 6 26.2 20 76 4 6 .7 9 2 3 .0 12 61 5 1 .3 12 2 8.3 11 59 4 5 .4 15 2 9 .0 13 71 4 8 .1 18 3 1 .9 15 73 5 3 .0 21 3 0 .3 16 63 50 .8 24 3 1 .4 16 64 54.9 27 23.2 18 64 4 4 .5 T a b le 24. E x p e rim e n ta l r e s u l t s o b ta in e d from -wheat p l a n t s grown on n u t r i e n t s o lu ti o n s v a ry in g i n c o n c e n tr a tio n s o f i r o n ; a v e ra g e s e x p re ss e d p e r 10 p l a n ts .( c o n ti n u e d ) P a r ts p e r m i l l i o n o f i r o n i n th e n u t r i e n t s o lu t i o n Grains o f M illig ra m s o f d r y w eig h t i r o n of M illig ra m s o f p h o sphorus M illig ra m s p o ta ss iu m of M illig ra m s of n itr o g e n 0 2 .8 0 .4 1 45 184 126 3 5 .9 0 .7 4 76 405 253 6 7 .8 0 .9 8 105 514 321 9 6 .4 0 .3 3 81 429 268 12 5 .8 0 .8 1 75 392 248 15 6 .3 0 .3 8 81 430 272 18 7 .4 1 .0 4 86 460 287 21 7 .1 0 .9 6 89 451 276 24 7 .1 1 .0 3 92 454 273 27 6 .4 0 .8 0 77 364 234 Experiment 3 Table 25. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of copper; averages expressed per 10 plants. P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu t io n M illig r a m s o f M illig r a m s o f a s c o r b i c a c i d m a g n e s iim a t h arvest M illig r a m s o f c a lc iu m G ram s o f M illig r a m s f r e s h w e ig h t c o p p e r 0 .0 2 2 .0 61 19 3 9 .7 0 .0 7 2 0 .3 1 8 .6 54 10 3 4 .1 0 .1 1 4 0 .6 2 0 .4 54 12 4 1 .6 0 .1 4 3 0 .9 1 5 .9 47 11 3 6 .4 0 .1 2 1 1 .2 1 7 .1 41 11 3 1 .6 0 .1 1 4 1 .5 1 4 .0 41 15 3 0 .8 O .C 95 1 .8 1 1 .5 31 11 1 8 .7 O .C 86 2 .1 8 .2 21 11 1 3 .3 0 .0 6 9 2 .4 6 .4 19 10 9 .9 0 .0 5 5 2 .7 4 .9 16 9 7 .7 0 .0 4 7 of Experiment 3 Table 26. Experimental results obtained from wheat plants grown on nutrient solution varying in concentrations of copper; averages expressed per 10 plants, (continued) P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu t io n G rains o f M illig r a m s d r y w e ig h t ir o n of M illig r a m s o f M illig r a m s p h osp h oru s p o ta s s iu m of M illig r a m s o f n itr o g e n 0 ,0 4 .7 0 .6 8 75 351 210 0 .3 4 .3 0 .5 8 57 318 189 0 .6 5 .4 0 .6 8 57 409 244 0 .9 3 .9 0 .5 1 a 325 184 1 .2 3 .5 0 .4 2 36 276 169 1 .5 3 .6 0 .3 8 39 273 167 1 .8 2 .3 0 .3 1 28 173 106 2 .1 1 .6 0 .1 9 23 117 73 2 .4 1 .3 0 .1 4 22 92 57 2 .7 1 .1 0 .1 3 20 77 49 Experiment 3 Table 27. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of ironj averages expressed per 100 grains of fresh plant material. P a r ts per m illio n o f ir o n in th e n u tr ie n t s o lu t io n M illig r a m s o f M i l l i g r a m s o f M i l l i g r a m s o f M i l l i g r a m s p h o sp h o r u sp o ta s s iu m n itr o g e n c a lc iu m of M il lig r a m s o f m a g n e siu m 0 160 657 452 50 169 3 169 900 561 26 141 6 227 1107 692 42 165 9 156 832 520 22 119 12 165 857 542 24 130 15 168 893 565 26 148 18 161 863 539 28 136 21 175 889 545 31 123 24 168 832 499 30 117 27 173 819 526 40 145 iJxperiment 3 Table 28. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of iron; averages expressed per 100 grana of fresh plant material (continued). P a r ts per m illio n o f ir o n in th e n u tr ie n t s o lu t io n M illig r a m s o f a s c o r b ic a c id a t h arvest G rains o f M illig r a m s o f d r y m a tte r ir o n 0 6 2 .2 1 0 .0 1 .4 5 3 6 6 ,4 1 3 .1 1 .6 4 6 5 6 .1 1 6 .3 2 .1 0 9 5 4 .1 1 2 .4 1 .6 1 12 6 2 .4 1 2 .7 1 .7 8 15 6 0 .3 1 3 .1 1 .8 3 18 6 0 .2 1 3 .9 1 .9 5 21 5 9 .7 1 4 .0 1 .8 9 24 5 7 .2 1 3 .0 1 .8 9 27 5 2 .1 1 4 .4 1 .8 0 Experiment 3 Table 29. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of copper; averages expressed per 100 grams of fresh material. P a r ts per m illio n o f copper in th e n u tr ie n t s o lu t io n M illig r a m s o f M illig r a m s o f M illig r a m s o f p h o s p h o r u s p o t a s s iu m n itr o g e n M illig r a m s c a l c iu m of M illig r a m s o f m a g n e siu m 0 .0 189 889 532 48 154 0 .3 166 925 550 30 158 0 .6 137 978 583 30 129 0 .9 111 884 500 30 127 1 .2 114 874 536 33 130 1 .5 124 880 538 50 132 1 .8 154 939 578 59 169 2 .1 172 861 540 84 155 2 .4 216 929 578 98 193 2 .7 259 1007 638 111 215 Experiment 3 Table 30. Experimental results obtained from wheat plants grown on nutrient solutions varying in concentrations of copper; averages expressed per 100 grams of fresh plant material, (continued). P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu t io n M illig r a m s o f a s c o r b ic a c id a t h a rv est G ram s o f M illig r a m s o f d ry m a tte r ir o n M illig r a m s o f copper 0 .0 5 5 .5 1 1 .9 1 .7 3 0 .1 8 0 .3 5 5 .1 1 2 .5 1 .6 9 0 .3 3 0 .6 4 9 .0 1 2 .9 1 .6 1 0 .3 4 0 .9 4 3 .7 1 0 .6 1 .3 8 0 .3 3 1 .2 5 4 .2 1 1 .1 1 .3 3 0 .3 6 1 .5 4 5 .6 1 1 .6 1 .2 2 0 .3 1 1 .8 6 1 .5 1 2 .5 1 .6 9 0 .4 7 2 .1 6 1 .5 1 1 .8 1 .4 2 0 .5 1 2 .4 6 4 .4 1 3 .1 1 .3 8 0 .5 6 2 .7 6 3 .8 1 4 .4 1 .7 3 0 .6 1 Experiment 3 Table 31- Experimental results obtained from wheat plants grown on nutrient solutions varying in the concentrations of iron; averages expressed per 100 grains of dry plant material. P a r ts p er m illio n o f ir o n in th e n u tr ie n t s o lu tio n M illig r a m s ir o n o f G ram s o f G ram s o f G ram s o f Gram s o f Grams o f p h o s p h o r u s P o t a s s iu m n i t r o g e n c a lc iu m m a g n e siu m 0 1 4 .5 1 .6 0 6 .5 7 4 .5 2 0 .5 0 1 .6 9 3 1 2 .5 1 .2 9 6 .8 7 4 .2 8 0 .2 0 1 .0 8 6 1 2 .5 1 .3 5 6 .5 9 4 .1 2 0 .2 5 0 .9 8 9 1 3 .0 1 .2 6 6 .7 1 4 .1 9 0 .1 8 0 .9 6 12 1 4 .0 1 .3 0 6 .7 5 4 .2 7 0 .1 9 1 .0 2 15 1 4 .0 1 .2 8 6 .8 2 4 .3 1 0 .2 0 1 .1 3 IB 1 4 .0 1 .1 6 6 .2 1 3 .8 8 0 .2 0 0 .9 8 21 1 3 .5 1 .2 5 6 .3 5 3 .8 9 0 .2 2 0 .8 8 24 1 4 .5 1 .2 9 6 .4 0 3 .8 4 0 .2 3 0 .9 0 27 1 2 .5 1 .2 0 5 .6 9 3 .6 5 0 .2 8 1 .0 1 Experiment 3. Table 32. Experimental results obtained from wheat plants grown on nutrient solutions varying in the concentrations of copper; averages expressed per 100 grams of dry plant material. P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu t io n M illig r a m s ir o n of Grams o f G ram s o f p h o s p h o r u s p o t a s s iu m G ram s o f G ram s o f n it r o g e n c a lc iu m G ram s o f m a g n e siu m M illig r a m s o f copper 0 .0 1 4 .5 1 .5 9 7 .4 7 4 .4 7 0 .4 0 1 .2 9 1 .5 3 0 .3 1 3 .5 1 .3 3 7 .4 0 4 .4 0 0 .2 4 1 .2 6 2 .6 4 0 .6 1 2 .5 1 .0 6 7 .5 8 4 .5 2 0 .2 3 1 .0 0 2 .6 4 0 .9 1 3 .0 1 .0 5 8 .3 4 4 .7 2 0 .2 8 1 .2 0 3 .1 0 1 .2 1 2 .0 1 .0 3 7 .8 7 4 .8 3 0 .3 0 1 .1 7 3 .2 5 1 .5 1 0 .5 1 .0 7 7 .5 9 4 .6 4 0 .4 3 1 .1 4 2 .6 4 1 .8 1 3 .5 1 .2 3 7 .5 1 4 .62 0 .4 7 1 .3 5 3 .7 4 2 .1 1 2 .0 1 .4 6 7 .3 0 4 .5 8 0 .7 1 1 .3 1 4 .3 0 2 .4 1 0 .5 1 .6 5 7 .0 9 4 .4 1 0 .7 5 1 .4 7 4 .2 4 2 .7 1 2 .0 1 .8 0 6 .9 9 4 .4 3 0 .7 7 1 .4 9 4 .2 3 Experiment 3 Table 33- Percentages of the original amounts of ascorbic acid found in wheat plants grown on nutrient solutions varying in the concentrations of iron, at each determination during the drying of the plants at room temperature. P a r ts p er m illio n o f ir o n i n t h e n u tr ie n t s o lu t io n P e r c e n ta g es o f a s c o r b ic a c id a t h arvest P e r c e n ta g es o f a s c o r b ic a c id 14 hours a fte r h a rv est P e r c e n ta g es o f a s c o r b ic a c id 43 h o u r s a f t e r h arvest P e r c e n ta g es o f a s c o r b ic a c id 62 h ou rs a fte r h arvest P e r c e n ta g es o f a s c o r b ic a d d 110 hours a fte r h arv est P e r c e n ta g es o f a s c o r b ic a c id 157 h ou rs a fte r h arvest 0 100 5 3 .5 3 5 .5 2 1 .7 9 .3 5 .6 3 100 4 6 .4 2 6 .9 1 7 .2 8 .3 2 .4 6 100 7 6 .2 4 4 .9 3 1 .1 1 3 .0 6 .6 9 100 6 5 .0 4 1 .4 2 8 .7 1 0 .4 5 .5 12 100 6 5 .5 3 6 .1 2 5 .5 9 .5 4 .2 15 100 7 2 .5 3 6 .0 2 5 .9 7 .8 4 .8 IS 100 7 7 .6 4 1 .7 2 7 .4 1 1 .1 5 .8 21 100 7 2 .5 ------- 2 7 .0 8 .7 5 .0 24 100 7 8 .0 3 4 .8 2 2 .7 9 .3 4 .9 27 100 6 2 .6 3 9 .2 2 2 .1 8 .3 5 .6 Experiment 3 Table 34. Percentages of the original anounts of ascorbic acid found in wheat plants grown on nutrient solutions varying in the concentrations of copper, at each determination during the drying of the plants at room temperature. P a r t3 p e r m illio n o f copper in th e n u tr ie n t s o lu tio n P e r c e n ta g es o f P e r c e n ta g e s o f a s c o r b ic a c id a s c o r b ic a c id a t h arvest 13 h o u rs s ifte r h arvest P e r c e n ta g es o f a s c o r b ic a c id 3 0 h o u fcs a f t e r h arvest P e r c e n ta g es o f a s c o r b ic a c id 52 h o u rs s ifte r h arvest P e r c e n ta g es o f P e r c e n ta g es o f a s c o r b ic a c id a s c o r b ic a c id 100 hours a fte r 147 hours h arvest a fte r h arvest 0 .0 100 5 6 .6 4 1 .3 2 5 .9 1 3 .0 5 .8 0 .3 100 5 9 .0 4 4 .6 2 7 .6 1 4 .5 5 .4 0 .6 100 6 9 .0 4 7 .1 2 8 .4 1 3 .1 6 .3 0 .9 100 7 5 .3 3 8 .9 2 8 .6 1 4 .4 6 .4 1 .2 100 6 0 .2 4 2 .6 2 2 .8 1 0 .2 5 .7 1 .5 100 6 2 .7 4 5 .0 2 8 .1 1 1 .0 6 .6 1 .8 100 7 7 .2 3 2 .8 2 2 .0 --------- — 2 .1 100 8 5 .5 3 6 .1 --------- ------- — 2 .4 100 7 9 .2 3 7 .7 ------- ------- — 2 .7 100 8 9 .3 3 9 .7 ------- ______ ___ Experiment 3 Table 35. Percentages of the original amounts of water found in the wheat plants grown on nutrient solutions varying in the concentrations of iron, at each determination during the drying of the plants at room temperature. P a r ts p er m illio n o f ir o n in th e n u tr ie n t s o lu tio n P e r c e n ta g es o f w a te r a t h arvest P e r c e n ta g es o f w a te r 1 4 h o u rs a fte r h arvest P e r c e n ta g es o f w a te r 43 hou r3 a fte r h arvest P e r c e n ta g es o f w a te r 62 h o u rs a fte r h arvest P e r c e n ta g e s o f P e r c e n ta g es o f w a te r 11 0 h ou rs w a te r 157 h ou rs a fte r h arvest a fte r h arvest 0 100 7 5 .8 4 1 .6 3 3 .7 1 5 .4 9 .3 3 100 7 7 .0 4 6 .1 3 4 .1 1 6 .5 1 0 .0 6 100 7 9 .3 5 2 .4 4 2 .3 2 3 .5 1 3 .1 9 100 7 8 .3 5 1 .4 4 1 .7 2 0 .9 1 1 .9 12 100 8 0 .4 5 3 .5 4 2 .6 2 3 .8 1 6 .4 15 100 7 7 .7 4 9 .8 3 8 .4 1 9 .1 1 1 .5 18 100 7 9 .2 5 1 .2 3 9 .6 1 9 .7 1 1 .8 21 100 7 7 .6 5 1 .4 3 9 .0 1 9 .0 1 1 .7 24 100 7 7 .0 4 9 .0 3 7 .9 1 8 .0 1 0 .4 27 100 7 3 .0 4 3 .5 3 3 .8 1 4 .8 8 .9 Experiment 3 • Table 36. Percentages of the original amounts of water found in the wheat plants grown on nutrient solutions varying in concentrations of copper, at each determination during the drying of the plants at room temperature. P a r ts p er m illio n o f copper in th e n u tr ie n t s o lu t io n P e r c e n ta g es o f w a te r a t h arvest P e r c e n ta g es o f w a te r 13 h o u rs a fte r h arvest P e r c e n ta g es o f w a t e r 30 h o u r s a fte r h arvest P e r c e n ta g es o f w a ter 5 2 h o u rs a fte r h arvest P e r c e n ta g e s o f P e r c e n ta g es o f w a te r 100 h o u rs w a te r 1 47 h o u rs a fte r h a rv est a fte r h a rv est 0 .0 100 7 8 .4 5 2 .2 3 9 .5 2 1 .9 1 3 .4 0 .3 100 7 8 .6 5 1 .3 3 9 .2 2 1 .2 1 2 .6 0 .6 100 7 8 .7 4 7 .2 3 5 .6 1 8 .4 1 1 .5 0 .9 100 7 9 .6 4 8 .1 3 5 .5 1 8 .3 1 0 ,1 1 .2 100 7 9 .1 4 5 .4 3 3 .0 1 6 .9 1 0 .6 1 .5 100 7 7 .4 4 2 .8 3 0 .5 1 5 .0 8 .7 1 .8 100 7 3 .7 3 2 .9 2 1 .2 8 .7 5 .3 2 .1 100 7 3 .5 3 6 .8 2 5 .2 1 0 .0 6 .4 2 .4 100 7 0 .5 3 5 .1 2 1 .9 7 .4 2 .7 100 6 8 .7 3 1 .8 1 9 .9 5 .7 2 .8 1 .9 F ig u r e 1 4 3 * A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d in m illig r a m s p e r 1 0 w h eat p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n i n th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 1 4 4 . A v e r a g e d r y w e i g h t , e x p r e s s e d i n gram s p e r 1 0 w h e a t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in t h e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p e r m illio n . F ig u r e 1 4 5 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 w h ea t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in t h e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts per m illio n . F ig u r e 1 4 6 . A v e r a g e a m o u n ts o f p h o s p h o r u s , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 w h eat p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu t io n , e x p r e sse d i n p a r ts per m illio n . F ig u r e 1 4 7 . A v e r a g e a m o u n ts o f p o t a s s i u m , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 w h ea t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f ir o n in th e n u tr ie n t s o lu t io n , ex p ressed in p a r ts p er m illio n . F ig u r e 1 4 8 . A v e r a g e a m o u n ts o f t o t a l n i t r o g e n , e x p r e s s e d i n m i l l i g r a m s p er 1 0 w h eat p la n t s , p lo t t e d a g a in s t t h e c o n c e n tr a tio n o f ir o n i n th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . z1 < * P i Q . VI V G. m H . IX. . fl N\ . ot p M X P ,e > * J t p tx. is 0 P T«. on! 1^8. Figure 143. Average amounts of ascorbic acid a t harvest, expressed in m illigram s per 10 wheat p la n ts, p lo tte d against the concentration of iro n in the n u trie n t so lu tio n , expre33ed in p a rts per m illio n . Figure 149. Average amounts of calcium, expressed in milligrams per 10 wheat p la n ts, p lo tted against the concentration of iro n in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 150. Average amounts of magnesium, expressed in m illigrams per 10 wheat p la n ts, p lo tte d ag ain st the concentration of iro n in the n u trie n t so lu tio n , expressed in p arts per m illio n . Figure 151. Average fresh weight, expressed in grams per 10 wheat p la n ts, p lo tte d against the concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Mix. o A ivoH tiit o o M-'iB r tR . I? v» ^>\ A. vl T S «ft- O *> T, 00 a 6. o -fueikt o ujeickW'V ft*3 pe«- 4° u> ‘ “* o 4 J0 M4- . ?i A*JTs O V TJ 00 C.».k i w f A »i ? p« r >'’ U Figure 152. Average amounts of ascorbic acid , expressed in milligram s per hundred grams of fresh m atter from wheat p lan ts, p lo tte d against th e concentration of iro n in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 153. Average amounts of phosphorus, expressed in milligrams per 100 grams of fresh m atter from wheat p la n ts , p lo tted ag ain st the concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 154. Average amounts of potassium, expressed in milligrams per 100 grams of fre sh m atter from wheat p la n ts, p lo tted against th e concentration of iro n in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 155. Average amounts of t o t a l n itrogen, expressed in milligrams per 100 gram3 of fresh m atter from wheat p la n ts, p lo tted against the concentration of iro n in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 156. Average amounts of calcium, expressed in milligrams per 100 grams of fre sh m atter from wheat p lan ts p lo tted against the concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 157. Average amounts of magnesium, expressed in milligrams per 100 grams of fre3h m atter from, wheat p la n ts , p lo tted against th e concentration of iro n in the n u trie n t so lu tio n , expressed in p arts per mi l l ion. l/> o c r-? ( A . o f t 8 p.p.i'A.,?- o f IK on < Ik* P i a u .^.»A . o f l H o O •c 0 vl» Q 0 0 0 w 0 l.V 0 ii.8 i j, j». ‘V o fc IK '* o di 10 II IT I* ■*« Figure 152. Average amounts of ascorbic acid a t h arv est, expressed in mi l l igrams per 100 grams of fresh m atter from wheat p la n ts p lo tte d against the concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 160. Average amounts of ascorbic acid, 14 hours a f te r h arv est, expressed in m illigram s per 100 grams of fresh m atter from wheat p la n ts , p lo tted ag ain st the concentration of iro n in the n u trie n t so lu tio n , expressed in p arts per m illion* Figure 161. Average amounts of ascorbic acid, 43 hours a f te r h arv est, expressed in m illigram s per 100 grams of fresh m atter from wheat p la n ts, p lo tte d against the concentration of iro n in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 162. Average amounts of ascorbic acid, 62 hours a f te r h arv est, expressed in milligram s per 100 grams of fre sh m atter from wheat p la n ts, p lo tte d against the concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 163. Average amounts of ascorbic acid , 110 hours a f te r h arv est, expressed in m illigrams per hundred grams of fresh m atter from wheat p la n ts, p lo tted ag ain st the concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illion* Figure 164. Average amounts of ascorbic acid , 157 hours a f te r harvest, expressed in m illigrams per 100 gram3 of fresh m atter from wheat p la n ts , p lo tted against th e concentraion of iro n in the n u trie n t so lu tio n , expressed in p a rts per m illio n . *T M ^ “^ € . P^ \i»o^3-. P £ Cs. C. »"^ M*». i w A v »i il 18 ,p.(v* of 1A»cJ A ao p.p.fA. e>P a 9.0** A C ii) p .p. K iftw J !>0 I* i ftiC io P iP-ftNl pp Figure 165# Average amounts of ascorbic acid a t h arv est, expressed in m illigram s per 100 grams of fresh m atter from wheat p la n ts , p lo tted ag ain st the amounts of iro n , expressed in m illigram s per 100 grams of fresh m atter. Figure 166A Average amounts of ascorbic acid, 14 hours a f te r h arvest, expressed in milligrams per 100 grams of fresh m atter from wheat p lan ts grown on n u trie n t so lu tio n supplied with various concentrations of iro n , p lo tted against the concentration of iro n , expressed in m illigram s per 100 grams of fre sh m atter. Figure 167. Average amounts of ascorbic acid a t h arv est, expressed in milligram s per 100 grams of fresh m atter from wheat plants grown on n u trie n t so lu tio n s supplied wirh various concentrations of copper, p lo tte d against th e amounts of copper, expressed in m illigram s per 100 grams of fresh matter* Figure 168. Average amounts of ascorbic a c id , 13 hours a f te r harvest, expressed in m illigram s per 100 grams of fresh m atter from wheat p la n ts grown on n u trie n t so lu tio n supplied with various concentrations of copper, p lo tte d against the amounts of copper, expressed in milligram s per 100 grams of fresh m atter. Figure 169. Average amounts of ascorbic acid a t h arv est, expressed in milligram s per 100 grams of fresh m atter from wheat p la n ts grown on n u trie n t so lu tio n supplied with various concentrations of copper, p lo tted against the amounts of iro n , expressed in milligrams per 100 grams of fresh matter* vj k o u 1 A. Ot J >1 MG-. J.O I f t oM o a © © 61 © •y5“ £f v7 0 o/i So ■) 0 © ,.7 o M ; cf j <£ ' / VI cC U- 00 0© »'a L itse i”J 8 . i .- l Mi TR.sk; Figure 180. Percentages of th e o rig in a l amounts of w ater, 14 hour3 a f te r h arv est, in the tis s u e s from wheat p lan ts grown on n u trie n t so lu tio n s supplied with various concentra­ tio n s of iro n , p lo tte d against the amounts of iro n , expressed in m illigrams per 100 grams of fresh m atter. Figure 181. Percentages of the o rig in a l amounts of w ater, 43 hours a f te r h arv est, in the tis s u e s from wheat p lan ts grown on n u trie n t so lu tio n supplied with various amounts of iro n , p lo tted against the amounts of iro n , expressed in m illigram s per 100 grams of fresh m atter. Figure 182. Percentages of th e o rig in a l amounts of w ater, 62 hours a f te r h arv est, in the tis s u e s from wheat p la n ts grown nn n u trie n t so lu tio n supplied with various amounts of iro n , p lo tted against the amounts of iro n , expressed in milligram s per 100 grams of fresh m atter. Figure 183. Percentages of th e o rig in a l amounts of w ater, 110 hours a f te r h arv est, in the tis s u e s from wheat p la n ts grown on n u trie n t supplied with various amounts of iro n , p lo tte d against the amounts of iro n , expressed in m illi­ grams per 100 grams of fresh m atter. » o55 ©Q (TO toft ID.OW Z A o tV (* ol r <£ 3 U> H* <£ U- io 00 <2jO\r «. i.fc 1.8 J 0 .o w J.O a.2. M 2> F ig u r e 1 8 4 * A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 10 w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n tr a tio n o f co p p er in th e n u t r ie n t s o lu t io n , e x p r e s se d in p a r ts p er m illio n . F ig u r e 1 8 5 . A v e r a g e d r y w e i g h t , e x p r e s s e d i n g r a m s p e r 1 0 w h e a t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f co p p er in th e n u tr ie n t s o lu t io n , ex p ressed in p a r ts p er m illio n . F ig u r e 1 8 6 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 10 w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f cop p er in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts per m illio n . F ig u r e 1 8 7 . A v e r a g e a m o u n ts o f p h o s p h o r u s , e x p r e s s e d i n m i l l i g r a m s p e r 10 w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f c o p a r in t h e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p e r m illio n . F ig u r e 1 8 8 . A v e r a g e a m o u n ts o f p o t a s s i u m , e x p r e s s e d i n m i l l i g r a m s p e r 10 w h e a t p l a n t s g r o w n o n n u t r i e n t s o l u t i o n s u p p l i e d w i t h v a r i o u s a m o u n ts o f c o p p e r , p l o t t e d a g a i n s t t h e c o n c e n tr a tio n o f cop p er in th e n u tr ie n t s o lu t io n , ex p ressed in p a r ts p er m illio n F ig u r e 1 8 9 . A v e r a g e a m o u n ts o f t o t a l n i t r o g e n , e x p r e s s e d i n m i l l i g r a m s p e r 10 w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f cop p er i n t h e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts per m illio n . rtJa • lo o l.l p .p .H . d.$ 9 Jo © 26.6 a cC M 1.8 coPPC.K © 0 © © 0..*V c o ? P E R ??■ © Figure 164. Average amounts of ascorbic acid a t h arv est, expressed in m illigrams per 10 wheat p la n ts, p lo tted against the concentration of copper in the n u trien t so lu tio n , expressed in p a r t3 per m illio n . Figure 190. Average amounts of calcium, expressed in milligrams per 10 wheat p la n ts, p lo tted against the concentration of copper in the n u trie n t so lu tio n , expressed in p arts per m illio n . Figure 191. Average amounts of magnesium, expressed in milligrams per 10 wheat p la n ts, p lo tted against the concentration of copper in the n u trie n t so lu tio n , expressed in p arts per m illio n . Figurel92. Average amounts of copper, expressed in milligram s per 10 wheat p lan ts , p lo tte d against the concentration of copper in the n u trie n t so lu tio n , expressed in p arts per m illior Figure 193. Average fresh weight, expressed in grams per 10 wheat p la n ts, p lo tted against the concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illion. Figure 214. Average amounts of copper, expressed in milligrams per 100 grams of oven dry tis s u e from wheat p la n ts, plo tted against the concentration of copper in th e n u trien t so lu tio n , expressed in p arts per m illio n . if i D > J <1 £ O.b p 0.0 p.M o.fc c o P P £ R 80 6o o.U 0. 0 0.0 ^.||9.pv c o frE ^ I 1 \ «.6 p . p . o.6 1-7. p . p . f A . 1-8 C o C P E P . Figure 194. Average amounts of ascorbic acid a t h arv est, expressed in m illigram s per 100 grains of fre sh m atter from wheat p la n ts , p lo tte d ag ain st th e concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 195. Average amounts of phosphorus, espressed in m illigrams per 100 grams of fresh m atter from wheat p la n ts, p lo tted ag ain st th e concentration of copper in the n u trie n t so lu tio n , expressed in p a rts per m illio n . Figure 196. Average amounts of potassium, expressed in milligrams per 100 grams of fre sh m atter from wheat p la n ts, p lo tted against the concentration of copper in the n u trien t solu­ tio n , expressed in p a rts per m illio n . Figure 197. Average amounts of t o t a l n itro g en , expressd in milligrams per 100 grams of fresh m atter, p lo tted ag ain st the concentration of copper in the n u trie n t so lu tio n , expressed in p arts per m illio n . Figure 198 Average amounts of calcium, expressed in m illigram s per 100 grams of fresh m atter from wheat p la n ts, p lo tted against the concentration of copper in th e n u trie n t solu­ tio n , expressed in p a rts per m illio n . Figure 199 Average amounts of magnesium, expressed im m illigram s per 100 grams of fresh m atter from wheat p la n ts, p lo tted against the concentration of copper in th e n u trie n t so lu tio n , expressed in p a rts per m illio n . 'W 'O 'J »3ddo-3 w- 0 0 O'O 0*0 ‘8M 3>ioV.ti j otl j ?

• €> 1^4. o.o 0.0 P. P. R 1.2. P.T fH. cefPE fr C ofP E R o.l O.R o.o o.& 1.2. I* P. P. K i. t o f P f c p . o.o Fi^oA .e 5 ^ a a . *I * r? im 1.2. I.v p . p .K , C.oPP£fl» c.oPPe « f l f l J F ig u r e 1 9 4 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d a t h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 100 g r a m s o f f r e s h m a t t e r fr o m w h ea t p la n t s , p lo t t e d a g a in s t th e c o n c e n tr a tio n o f co p p er in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 2 0 3 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d , 1 3 h o u r s a f t e r h arv est, expressed in m illigram s per 100 grams of fresh m atter from wheat p la n ts , p lo tted ag ain st the concentration of copper in th e n u trie n t so lu tio n , expressed in part3 per m illion* F ig u r e 2 0 4 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d , 3 0 h o u r s a f t e r h a r v e s t e x p r e s s e d i n m i l l i g r a m s p e r 100 g r a m s o f f r e s h m a t t e r fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f cop p er in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 2 0 5 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d , 52 h o u r s a f t e r h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 100 g ra m s o f f r e s h m a t t e r fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f cop p er in th e n u tr ie n t s o lu t io n , ex p ressd d in p a r ts p er m illio n . F ig u r e 2 0 6 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d , 1 0 0 h o u r s a f t e r h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 100 g r a m s o f f r e s h m a t t e r fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f co p p er in th e n u tr ie n t s o lu t io n ., e x p r e sse d in p a r ts per m illio n . F i g u r e 2 0 7 . A v e r a g e a m o u n ts o f a s c o r b i c a c i d , 1 4 7 h o u r s a f t e r h a r v e s t , e x p r e s s e d i n m i l l i g r a m s p e r 100 g r a m s o f f r e s h m a t t e r fr o m w h e a t p l a n t s , p l o t t e d a g a i n s t t h e c o n c e n t r a t i o n o f copppr in th e n u tr ie n t s o lu t io n , e x p r e sse d in p a r ts p er m illio n . i,0 So 0.0 •p . p . m . copper ft.% 1.1 l.« p. p m . c o p p e r 0 © Fi'<^oRE Ho H o.o 0.6 I.a P P M . 1.8 i o f,. 0.0 IM 0 .6 0.0. P. P. NV. C .oP P G R O 6 l-Z p.p. n . l.g copper Figure 21k. This fig u re may be found a f te r fig u re 193. Figure 215. Percentages of the o rig in a l amounts of w ater, 13 hours a f te r h arv est, in th e tis s u e s from wheat p lan ts grown on n u tire n t so lu tio n supplied w ith various amounts of copper, p lo tte d against th e amounts of copper, expressed in m illigrams per 100 grams of fresh matter* Figure 216. Percentages of the o rig in a l amounts of w ater, 30 hours a f te r h arv est, in the tis s u e s from wheat p la n ts grown on n u trie n t solutions supplied with various amounts of copper, p lo tte d against the amounts of copper, expressed in m illigram s per 100 grains of fresh matter* Figure 217. Percentages of the o rig in a l amounts of w ater, 13 hours a f te r h arv est, in th e tis s u e s from wheat p la n t3 grown on n u trie n t so lutions supplied with various amounts of copper, p lo tted against the amounts of iro n , expressed in m illigrams per 100 grams of fresh m atter. Figure 218. Percentages of the o rig in a l amounts of w ater, 30 hours a f te r h arv est, in th e tis s u e s from whe.it p lan ts grown on n u trie n t so lutions supplied with various amounts of copper, p lo tted against the amounts of iro n , expressed in milligram s per 100 grams of fresh m atter. K 62. 110. and 157 hours a f te r h arv est, in the tis s u e s from wheat p la n ts, p lo tted against th e concentration of iro n in the n u trie n t so lu tio n , expressed in p a rts per million* Figure 229. Percentages of th e o rig in a l amounts of ascorbic acid , 14, 43 , 62, HO, and 157 hours a f te r h arv est, in the tis.. ues from wheat p la n ts, p lo tte d against the concentra­ tio n of iro n in the n u trien t so lu tio n , expressed in p a rts per m illio n . O—0 —0 - 0 - 0 —Q - O —(•)—0 im Ko«»* 9 o < n J \ ) ^0 r= vp X r o in o w 'o ^ a. io 50 » x > ? v '| a ? - i > j o «f s IQ. ,4 a o y .0 I N M M ' i y ) r « \ ®> > > o © 3 ■C N*-l IMB 'S'dS i. 0 c« ot u V © © o U 0 Q © © O 0 P .P . M © 10 f— 4 0 0 o» ‘M ^ - o ' O S V i T o niJ «" < X ' ' » \ A Experiment 4. Table 37. Experimental re s u lts obtained from wheat p la n ts grown on n u trie n t so lutions varying in concentrations of iro n ; averages expressed per 10 p la n ts.(to p s ) P arts per m illion of iro n in the n u trie n t solution Grams of Milligrams of fresh weight iro n Milligrams manganese of M illigrams of Grams of ascorbic acid dry weight 0.0 36.1 0.845 0.811 28.99 4.8 0.5 39.9 0.936 0.796 26.09 5.2 1.0 44.4 1.017 0.918 26.11 6.2 5.0 41.0 0.912 0.784 28.99 5.3 10.0 42.0 0.952 0.812 26.38 5.6 Experiment h. Table 38. Experimental re s u lts obtained from wheat p la n ts grown on n u trie n t so lu tio n s varying in concentrations o f iro n ; averages expressed per 100 grams of fre sh m aterial from the to p s and the ro o ts of the D iants. PLANTS ROOTS PLANTS TOPS P arts per m illion of iro n in th e n u trien t solution Milligrams of Milligrams of Milligrams of iro n manganese ascorbic acid Grams of dry m atter Milligrams of ascorbic acid 0.0 2.46 2.37 80.3 14.0 3.79 0.5 2.52 2.14 65.4 14.0 2.11 1.0 2.53 2.28 56.8 15.4 4.80 5.0 2.55 2.19 70.7 14.8 6.20 10.0 2.48 2.12 62.8 14.6 3.95 Experiment 4. Table 39. Experimental re s u lts obtained from wheat p la n ts grown on n u trie n t solutions varying in concentrations of iro n ; averages expressed per 100 grams of dry m aterial from th e 'to p s and th e ro o ts of the p la n ts. PLANTS ROOTS PLANTS TOPS P arts per m illio n Milligrams of ascorbic acid of iro n in the n u trien t so lu tio n M illigrans of Milligrams of manganese iro n M illigrams of iro n Milligrams of manganese 0.0 573.6 17.6 16.9 14.3 2.5 0.5 467.1 18.0 15.3 45.0 10.0 1.0 381.8 16.4 14.8 43.5 4.5 5.0 477.7 17.2 14.8 65.0 4.8 10.0 430.1 17.0 14.5 100.0 5.5 Figure 233. Average amounts of iro n , expressed in milligram s per 10 wheat p la n ts, grown on n u trien t so lutions supplied with various amounts of iro n , expressed in p a rts per m illio n . Figure 234. Average amounts of manganese, expressed in m illigram s per 10 wheat p la n ts, grown on n u trien t so lu tio n s supplied with various amounts of iro n , expressed in p a rts p er m illio n . Figure 235. Avera amounts of ascorbic acid a t h arv est, expressed in milligram s per 10 wheat p la n ts, grown on n u trie n t so lu tio n s supplied with various amounts of iro n , expressed in p a rts per m illio n . Figure 236. Average fresh weight, expressed in grams per 10 wheat p la n ts, grown on n u trie n t so lu tio n s supplied with various amounts of iro n , expressed in p a rts per m illio n . I Pl -8 Id 2 0.8 2 «C 0.6 2 <£ 2 0.H 0.1 (o O.-a t p p . IA 6 10 I AOfJ * ri&wRe Z 3& . ^ .0 <£ tX * < » d 3 no \J £ <0 a ul cd JS 3 bs U. .? ' . S' o F iKotO Io 0 \o cvi- 0 m tb A 5C t o.S I S' p.p. ^ . oF I ft on* Io o.r rr » m. of A1 ft. 0 Fl'GrUftC 2^ S' . F i Or U ft. E 10 p.p. N V oF I RoftJ 1 ^ 5 . o .s p.p.M. S' IftoN to F ig u r e 2 4 6 . A v e r a g e a m o u n ts o f i r o n , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g r a m s o f d r y m a t e r i a l fr o m w h e a t r o o t s g r o w n on n u t r i e n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o u n ts o f i r o n , ex p ressed in p a r ts p er m illio n . F ig u r e 2 4 7 . A v e r a g e a m o u n ts o f m a n g a n e s e , e x p r e s s e d i n m i l l i g r a m s p e r 1 0 0 g r a m s o f d r y m a t e r i a l fr o m w h e a t r o o t s , g ro w n o n n u t r i e n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o u n ts o f ir o n , e x p r e sse d in p a r ts p er m illio n . F ig u r e 2 3 7 . A v e r a g e d r y w e i g h t , e x p r e s s e d i n gram s p e r 1 0 w h e a t t o p s , grow n on n u t r ie n t s o l u t i o n s s u p p lie d w ith v a r io u s a m o u n ts o f i r o n , e x p r e s s e d i n p a r t s p e r m i l l i b n . t 10 m d (>o •/» 2o to o . S (V\. of F vG-u AE oF XA.OW Experiment 5. Table 40. Experimental re s u lts obtained from tomato p la n ts grown on n u trien t so lu tio n s varying in concentrations of iro n ; averages expressed per 100 grams of fresh m aterial from the basal and the term inal leaves of th e p la n ts. P arts per Grams of dry m atter m illio n of iro n in th e ■■ ■ —- ■ n u trie n t Basal Terminal so lu tio n leaves leaves M illigrams of ascorbic acid - 1■ - 1 ■" Basal Terminal leaves leaves Milligrams of iro n Milligrams of manganese ■ ■■■ - - - ----- ■ -----Basal Terminal Basal Texminal leaves leaves leaves leaves 0.0 14.0 18.0 55.2 61.6 84.1 32.9 23.13 12.0 0.5 13.0 17.0 50.1 70.3 90.6 30.0 16.37 9.6 1.0 13.3 20.0 57.7 77.1 79.8 32.7 20.75 10.7 5.0 12.4 15.3 45.8 78.9 75.8 42.1 20.82 13.7 10.0 12.7 15.3 52.8 89.7 76.0 35.0 18.38 16.6 Experiment 5 Table 41. Experimental r e s u lts obtained from tomato p la n ts grown on n u trie n t so lutions varying in concentrations of iro n ; averages expressed per 100 grams of dry m a terial from the basal and the term inal leaves of th e p la n ts. P arts per m illio n of iro n in th e n u trie n t so lu tio n Milligrams of ascorbic acid M illigrams of iro n Basal leaves Basal leaves Terminal leaves Terminal leaves Milligrams of manganese Basal leaves Terminal leaves 0.0 394 342 11.8 6.0 3.24 2.16 0.5 385 414 11.8 5.1 2.13 1.63 1.0 434 386 10.6 6.5 2.76 2.13 5.0 369 516 9.4 6.4 2.58 2.10 10.0 416 586 9.6 5.4 2.33 2.53 Figure 248. Average amounts of iro n , expressed in milligram s per hundred grams of fresh matter from the b asal and the term in al leaves of tomato p lan ts grown on n u trie n t so lu tio n s supplied with various amounts of iro n , expressed in p a rts per m illio n . Figure 249. Average amounts of manganese, expressed in milligrams per hundred grams of fresh m atter from th e b asal and th e term in al leaves of tomato p lan ts grown on n u trien t so lu tio n s supplied with various amounts of iro n , expressed in p a rts per m illio n . Fi 6UJU IS 8 B c T ft r o a ■3. T t ?■? tA , o 5 o.S X M. 3 5. C. l EAOES T 10.0 LEJWES T =. TERMINAL LE AVE S &i q 3 r r T TERM 'iNM - if t o M £>F Fi&uRE 8 T 3 1.0 0.<3 =. v-tftvlti 3 T 1.0 a feftSRU f lA o U S.o T 10.o Figure 250. Average amounts of dry m atter, expressed in grams per 100 grams of fre sh m atter from the basal and the term in al leaves of tomato p la its grown on n u trie n t so lutions supplied w ith various amounts of iro n , expressed in p a rts per m illio n . Figure 251. Average amounts of ascorbic ac id , expressed in milligrams perlOO grams of fre sh m atter from th e basal and th e t e r ­ minal leaves of tomato p la n ts grown on n u trie n t so lu tio n supplied with various amounts of iro n , expressed in p a rts per m illio n . 2 50 fi&jftE fcrftfcSM. UtRVitS T t "TtRlA'vMfvL U&VIES S T o 3 r 6.5 3 T 3 r i. o p . p t*.. o f B: R o t S T o.£ LEAVE s TEftMlNA . LEAVES. 2. 5 ( . i PT o / kA S fU T i Fi'G uR t io 5-0 I fk ow M. I 3 T of i R T 5 .0 o n B T too Figure 252. Average amounts of iro n , expressed in m illigram s per 100 grams of dry m atter from th e b asal and the term in al leaves of tomato p lan ts grown on n u trien t so lu tio n s supplied with various amounts of iro n , expressed in p a rts per m illio n . Figure 253. Average amounts of manganese, expressed in m illigrams per 100 grams of dry m atter from the basal and the term inal leaves of tomato p lan ts grown on n u trie n t so lu tio n s supplied with various amounts of iro n , expressed in p a rts per m illio n . P t‘ o o <\ 6 1 5 a, go 6o Ho lo & T 3 O 3 T *.sr B T 1.0 p .p . M. Pi &u K E T 5.0 of 3 T ( 0. 0 2. 5 2>. It U 3 T o 3 3 T o T 3 s p .p .M . oT T 5.0 (.0 IA.ON B T 10. 0 Figure 254. Average amounts of ascorbic acid, expressed in milligrams per 100 grams of dry m atter from the b asal and the term inal leaves of tomato p lan ts grown on n u trie n t so lu tio n s supplied w ith various amounts of iro n , expressed in p a rts per m illio n . Figure 255. Average amounts of ascorbic acid a t h arv est, expressed in m illigram s per 100 grams of dry m atter from the b asal and th e term inal leaves of tomato p lan ts grown on n u trie n t so lu tio n supplied with various amounts of iro n , p lo tte d ag ain st th e amounts of iro n , expressed in m illigram s per 100 grams of dry m atter. Figure 256. Average amounts of ascorbic acid a t h arv est, expressed in m illigram s per 100 grams of dry m atter from the b asal and the term in al leaves of tomato p lan ts grown on n u trie n t so lu tio n s supplied with various amounts of iro n , p lo tte d ag ain st the amounts of manganese, expressed in m illigram s per 100 grams of dry matter* %S H . F»'^o £ 3 T 0 3 T B T o -5 p -r M i° B T 3 _ _ 5o o r I ft. o M T I o. o o t ftftsfti LEftoes <& iTEfthiNM L6Ad£S F.'& •u(\ £ 2-5 6 Pieju KE 4 2. 5 5 . A © © o © A © A © O © © A A 3o iiiJ m- <&ofl.C u. 0 o I.s W(r. of 1 A.ON l5L NUr. Fi cyy a. e a n . l*t 18 ?£ftC£M Tftit <6» of K\ATT£<^ MfcuSANEsE Experiment 6 Table 42. Average amounts of iro n in milligram s per 100 grams of fresh m atter from tomato p la n ts grown on n u trie n t so lutions containing no copper and 0.04 p a rts per m illion of copper and various concentrations of iro n a t in te rv a ls during the growth of the p la n ts. P arts per m illion of copper in the n u trie n t solutio n P arts p er million of iro n in the n u trie n t so lu tio n November 25, 1950 58 days old 0.00 0.0 0 .0 0 0 0.5 0.855 0.00 1.0 0.00 December 15, 1950 78 days old Terminal leaves Basal leaves January 3, 1950 97 days old Terminal leaves Basal leaves 1 .0 0 0 1.234 1.048 1.347 1.092 0.926 0.917 0.685 1.163 1.032 1.092 1.300 5.0 0.992 1.172 1.155 1.227 1.169 0.00 10.0 2.322 2.002 1.254 1.391 1.196 0.04 0.0 0.924 1.272 1.291 0.985 1.079 0.04 0.5 0.537 1.380 1.010 1.075 1.096 0.04 1.0 0.813 1.452 0.974 1.051 1.197 0.04 5.0 0.655 1.328 0.747 1.607 1.487 0.04 10.0 1.663 1.253 0.762 0.855 1.284 Experiment 6. T ab le 4 3 . Average am ounts o f m anganese i n m illig ra m s p e r 100 grams o f f r e s h m a tte r from tom ato p l a n t s grown on n u t r i e n t s o lu t i o n s c o n ta in in g no co p p er and 0 .0 4 p a r t s p e r m i l l i o n o f copper and v a r io u s c o n c e n tr a tio n s o f i r o n a t i n t e r v a l d u rin g th e grow th o f th e p l a n t s . P a r ts p e r m i l l i o n o f c o p p er i n th e n u t r i e n t s o lu tio n P a r t s p e r m i llio n o f iro n in th e n u t r i e n t s o lu ti o n December 1 5 , 1950 78 day s o ld T erm in al le a v e s B a s a l le a v e s J a n u a ry 3 , 1950 97 day s o ld T erm in al le a v e s B a sa l le a v e s 0 .0 0 0 .0 ------- - 1 .0 0 0 0 .5 5 2 1 .3 6 1 0 .0 0 0 .5 0 .7 7 0 1 .0 2 7 0 .6 4 7 1.288 0 .0 0 1 .0 0 .4 5 4 0 .9 6 9 0 .4 5 9 1 .1 4 6 0 .0 0 5 .0 0 .3 6 1 0 .713 0 .3 6 1 1 .0 4 9 0 .0 0 1 0 .0 0.4 7 7 0 .8 4 5 0 .3 3 6 0.9 2 9 0 .0 4 0 .0 1 .2 8 4 1 .1 5 7 0 .7 6 0 1 .7 9 8 0 .0 4 0 .5 0 .7 9 9 1.043 0.5 9 7 1.0 7 0 0 .0 4 1 .0 0 .6 8 7 0.7 4 3 0.6 7 8 1.4 1 0 0 .0 4 5 .0 0.6 8 3 0 .4 7 6 0 .4 7 0 0.8 7 8 0 .0 4 1 0 .0 0 .7 3 0 0 .7 6 2 0.3 9 2 0 .8 2 7 Experiment 6 T a b le 44. Average amounts o f a s c o r b ic a c id i n m illig ra m s p e r 100 grams o f f r e s h m a tte r from to m ato p l a n t s grown on n u t r i e n t s o lu t i o n s c o n ta in in g no copper and 0 .0 4 p a r t s p e r m i l l i o n o f co p p er and v a r io u s c o n c e n tr a tio n s o f i r o n a t i n t e r v a l s d u r in g th e growth o f th e p l a n t s . P a r ts p e r m illio n o f copper i n t h e n u trie n t s o lu t i o n P a r ts p e r m i llio n o f i r o n i n th e n u trie n t s o lu tio n J a n u a ry 3 , 1950 97 days o ld December 1 5 , 1950 78 days o ld November 2 5 , 1950 53 days o ld T e rm in al le a v e s B a sa l le a v e s T e rm in al le a v e s B a s a l le a v e s 0 .0 0 0 .0 3 0 .1 3 6 .7 8 .8 2 5 .1 1 4 .1 0 .0 0 0 .5 2 9 .9 3 0 .2 7 .6 2 1 .2 1 1 .7 0000 1 .0 2 2 .8 3 2 .7 9 .9 2 5 .3 1 0.3 0 .0 0 5 .0 2 6 .5 2 7 .0 8 .8 2 7 .7 1 0 .9 0 .0 0 1 0 .0 2 6 .7 3 0 .9 9 .1 2 9 .9 1 1 .7 0 .0 4 0 .0 2 9 .9 2 7 .6 9 .5 2 4 .2 1 3 .0 0 .0 4 0 .5 3 2 .0 2 8 .6 8 .9 3 1 .0 1 0 .0 0 .0 4 1 .0 2 7 .4 2 7 .0 7 .7 2 4 .6 9 .7 0 .0 4 5 .0 2 4 .0 3 5 .4 1 0 .3 3 2 .7 1 1 .0 0 .0 4 1 0 .0 2 7 .8 3 7 .2 2 1 .3 1 1 .0 1 2 .6 Experiment 6. T a b le 4 5 . Average d r y w e ig h ts , i n grams p e r 100 grams o f f r e s h m a tte r from tom ato p l a n ts grown on n u t r i e n t s o lu tio n s c o n ta in in g no copper and 0 .0 4 p a r t s p e r m ill io n of copper and v a r io u s c o n c e n tr a tio n s o f i r o n a t i n t e r v a l s d u rin g th e grow th o f th e p la n ts . P a r ts p e r m illio n o f cop p er i n th e n u trie n t s o lu t i o n P a rts per December 1 5 , 1950 November 2 5 , 1950 78 days o ld m i llio n o f 58 days o ld iro n in th e T erm in al le a v e s B a sa l le a v e s n u trie n t s o lu t i o n J a n u a ry 3 , 1950 97 days o ld T erm in al le a v e s B a sa l le a v e s 0 .0 0 0 .0 5 .9 5 6 .76 6 .68 8 .0 2 8 .2 2 0 .0 0 0 .5 5.89 7 .33 6 .6 4 7 .7 2 7 .4 1 0 .0 0 1 .0 6.08 7 .31 6 .6 4 7 .8 0 7 .7 0 0 .0 0 5 .0 6 .1 5 7 .0 2 6 .80 7 .8 5 7 .48 0 .0 0 1 0 .0 6 .3 0 7 .06 6.66 7 .8 9 7 .65 0 .0 4 0 .0 7.18 7.70 6 .9 4 8 .2 1 7 .99 0 .0 4 0 .5 7 .0 9 7 .30 6 .8 5 7.96 7.16 0 .0 4 1 .0 7.08 7.37 6 .6 1 7 .7 4 7.72 0 .0 4 5 .0 6 .9 1 7 .6 6 6 .9 7 7 .8 4 8 .0 7 0 .0 4 1 0 .0 6 .8 5 7.68 6.79 8 .0 4 7 .8 8 Experiment 6. T a b le 46 . Average am ounts o f i r o n e x p re ss e d i n m illig ra m s p e r 100 grams o f d ry m a tte r from tom ato p l a n t s grown on n u t r i e n t s o lu t io n s s u p p lie d w ith no c o p p er and 0 .0 4 p a r t s p e r m i l l i o n o f cop p er and v a r io u s c o n c e n tr a tio n s o f i r o n , a t i n t e r v a l s d u rin g th e grow th o f th e p l a n t s . P a r ts p e r m illio n o f co p p er i n th e n u trie n t s o lu t i o n P a rt3 p e r November 2 5 , 1950 Decembre 1 5 , 1950 m illio n o f 58 days o ld 78 d a y s o ld ir o n i n th e n u trie n t T erm in al le a v e s B a s a l le a v e s s o lu t i o n J a n u a ry 3 , 1950 97 d ay s o ld T erm in al le a v e s B a sa l le a v e s 14.97 --------- 12.75 15.38 1 4 .5 2 16 .4 4 18.3 7 12.38 12.00 1 .0 11.26 15 .5 4 1 5 .9 1 16.8 8 14.0 0 0 .0 0 5 .0 16.13 16.98 16.69 15.63 15.63 0 .0 0 1 0 .0 36.89 18.83 28 .3 6 15.6 3 17.63 0 .0 4 0 .0 12.8 7 18.60 16.5 2 13.50 12.0 0 0 .0 4 0 .5 7.57 14.74 1 8 .9 0 1 5 .2 5 13.50 0 .0 4 1 .0 11.48 14.72 19.70 1 5 .5 0 13.53 0 .0 4 5 .0 9 .4 8 10.71 1 7 .3 4 18.43 20.50 0 .0 4 1 0 .0 24.28 1 1 .2 2 1 6 .3 2 16 .3 0 10.63 0 .0 0 0 .0 0 .0 0 0 .5 0 .0 0 Experiment 6 T a b le 47. Average amounts o f m anganese e x p re ss e d i n m illig ra m s p e r 100 grams o f d ry m a tte r from tom ato p l a i t s grown on n u t r i e n t s o lu ti o n s su p p le d w ith no copper and 0 .0 4 p a r t s p e r m i llio n o f copper and v a r io u s c o n c e n tr a tio n s o f i r o n , a t i n t e r v a l s d u rin g th e grow th o f t h e p l a n t s . P a rts p e r m illio n o f copper i n th e n u t r i e n t s o lu t i o n P a r ts p e r m illio n o f iro n in th e n u t r i e n t s o lu tio n December 1 5 , 1950 78 days o ld T e rm in al le a v e s B a s a l le a v e s J a n u a ry 3 , 1950 97 days o ld T e rm in al le a v e s B a sa l le a v e s 0 .0 0 0 .0 1 4 .9 7 ------- — 16.5 6 6.38 0 .0 0 0 .5 1 5.4 6 1 0 .5 0 17.38 8 .3 8 0 .0 0 1 .0 1 4 .6 0 6 .2 1 14.88 5.88 0 .0 0 5 .0 10.49 12.26 1 4.0 2 4 .6 0 0 .0 0 1 0 .0 1 2.6 8 6 .7 5 1 2 .1 4 4 .2 6 0 .0 4 0 .0 1 6 .6 7 1 7 .9 0 22.50 9 .26 0 .0 4 0 .5 1 5.23 1 0 .9 4 14.88 7 .5 0 0 .0 4 1 .0 1 1 .2 4 9 .3 2 18.26 8 .7 6 0 .0 4 5 .0 8 .9 6 10.88 6 .0 0 0 .0 4 1 0 .0 11.2 2 10.50 4 .8 8 8 .9 2 9 .5 0 Experiment 6 T a b le 4 8 . A verage amounts o f a s c o r b ic a c id e x p re ss e d i n m illig ra m s p e r 100 gram s o f d ry m a tte r from tom ato p l a n t s grown on n u t r i e n t s o lu ti o n s s u p p lie d w ith no co p p e r and 0 ,0 4 p a r t s p e r m i l l i o n o f copper and v a r io u s c o n c e n tr a tio n s o f i r o n , a t i n t e r v a l s d u rin g t h e grow th o f t h e p l a n t s . P a rts p e r P a rts p e r m illio n o f m illio n o f cop p er i n t h e i r o n i n th e n u trie n t n u trie n t s o lu t i o n s o lu t i o n November 2 5 , 1950 58 day s o ld December 1 5 , 1950 78 days o ld T e rm in al le a v e s B asal le a v e s J a n u a ry 3 , 1950 97 days o ld T erm in al le a v e s B a s a l le a v e s 0 .0 0 0 .0 506 541 132 313 172 0 .0 0 0 .5 508 412 114 275 153 0 .0 0 1 .0 375 447 149 324 134 0 .0 0 5 .0 431 385 129 353 146 0 .0 0 1 0 .0 424 438 137 479 153 0 .0 4 0 .0 416 358 137 295 163 0 .0 4 0 .5 451 392 130 389 139 0 .0 4 1 .0 387 366 117 318 126 0 .0 4 5 .0 347 462 148 417 136 0 .0 4 1 0 .0 406 484 186 265 140 F ig u re 260 t o f i g u r e 265 in c lu d e d . Average amounts o f d ry m a tte r , e x p re ss e d i n grams p e r hundred grams o f f r e s h m a tte r from to m ato p l a n t s grown on n u t r i e n t s o lu tio n s s u p p lie d w ith v a rio u s am ounts o f ir o n and w ith o u t o r w ith 0 .0 4 p a r t s p e r m illi o n o f c o p p e r. The v a lu e s f o r th e b a s a l and th e te r m in a l le a v e s a r e re p re s e n te d by b a r g ra p h s f o r each h a r v e s t: 58, 78, and 97 days a f t e r s e e d in g . The shaded p o r tio n s c o rresp o n d t o th e in crem en t o f one c o p p er s e r i e s o v e r th e o th e r . Mo « - C O PP6 ft. O . o i 5 * 7>AYS olD F i &oft.€ 5.60 5" 6 (a d » e p . p M o C . o 5 I A . o F I »o 2 fa n I 0 .5 Bs. fcASAL 1 S «o 1 0. o W oF FT 0.0*1 7 8 7 ft y i P P E f t . R £ M CoppEA Ol D - a t 2, f» i o R e a. OF 3> #»• Y ( i o 2 61 i R o t J 78 I>AYS 6U3 r i C o p p E f t t> A j ) F« dO frE i 0.5 ^ J B T B 0 T O B T S B » p . p . . o F S T a BT C 1 7 o D P A P C Y S BT 0-5 BT JT S' '° « p.p.(A. X f t o i J O.Oft M o HvENT t o l o A4-E p a T f p. p. M oF IftoiJ C o f p £ ft. R O Fi G0 R 6 L R7 D BAYS p. t j U S old a r e U>g V BT B> T 0 o» BT.fcT » S’ >.^v. oF IftftU p . p . B t 10 a r 0 B T B 0.5 T B l p . p . p a T S . x R o r J B T IO igure 266 to figure 271 included. Average amounts of iro n , expressed in m illigram s per 100 grams of fresh m atter from tomato p lan ts grown on n u trien t solutions supplied with various amounts of iron and without or with 0.04 p a rts per m illion of copper. The values fo r the basal and the term inal leaves are represented bybar graphs for each h arv est: 58, 78 and 97 days a fte r seeding. The shaded portions correspond to the increment of one copper se rie s over the other. I i Wo 2 ? S' 8 3> R y s •It r Fi fe j A £ P • i COPPER. 0 • OM OL3> 58 I 5 .6 6 . F t .s i |p.p f> (v\. o f S' X« orJ •o /■ A vs a (.3) o . o 4* p>.p.M c o P P £ R 7 8 4 . Ay s» o i n "it BAfcAL, T i TEAM.(V<\. .1 0 ^ 9 F i (iro ft, £ H b 3 Fi C u f t t I ts 't 1 .5 Ui f t* I 1 S 0 OS ft o "BT BT OS » p.p PA . of 6 1 6 TAoM ft 7 id Wo c OPP 6 fi °n 3>«YS ©L-» 'S " t.5 FifruftE B T O ft T T o. o \'5 > C\ o O R E p.p N\ . C o P P c.R s 2-7 2. I *-i > fA M T t * io 1 OS ( £ ftMJ No fi p p. ft* F9 of lo lAoW Io 6 o p p . M. of I A0N 0. o "? P c A o -T fcr & f i &f t U -r ft y s l OO T * Tt RWi WdJ F 1 CjuR F 2.7«t P E R So 2o r «• ft 1 I O N >6 o o.f 3 T 1.0 of 5 &T B T ST 1.0 o.S I® Xf t o M M i t L i C r R A M S of p . p ca. i V 10o C-OfPf^ **1 pfly s o . o y j^ys.M C o f P e R A7 ^ Ay s ot-3> oc3> F| ‘ Cr o ft E Fi'&o A £ 9.7 6 9. 7 7 . So r l 10 a r o ( S T pr{« IS O. 0 S T> ft > S J tU F i fi* O ft.6 7 « p.y, (V^ ■» ft Y 5 C.O?f £ft O'-B 17 8 n e> 3 o T - BT o.S \ P p w 5 ‘i a>o 3 T lo B T o B T o.S F i (*<-> R t B T v p .p .w 5" «° x«3.«W J,7 2 ft7 s o uX> . o l Z> T> ft Y 5 B 7 o.oft p. p «w . C o p p t C . coffee. mo ^7 "BT 18 0 I I I I BT o e>T ftT i o .^ ppM BT r IRON B T BT i° o S*T 0 .5 l p.p n\ BT BT 5- lo I Rom F ig u r e 2 8 2 t o f i g u r e 2 8 5 i n c l u d e d . A v e r a g e a m o u n ts o f m a n g a n e s e , e x p r e s s e d i n m illig r a m s p e r 1 0 0 gram s o f o v e n d r y t i s s u e fr o m t o m a t o p l a n t s g r o w n o n n u t r i e n t s o l u t i o n s u p p l i e d w i t h v a r i o u s a m o u n ts o f i r o n a n d w i t h o u t o r w ith 0 .0 4 p a r ts p e r m illio n o f c o p p e r . The v a lu e s f o r th e b a s a l and th e te r m in a l le a v e s a r e r e p r e s e n te d b y b a r g r a p h s f o r t h e tw o l a s t h a r v e s t s : 78 a n d 9 7 d a y s a f t e r s e e d i n g . T he s h a d e d p o r t i o n s c o r r e s p o n d t o t h e in c r e m e n t s o f o n e c o p p e r t r e a t m e n t o v e r t h e o t h e r . Mo \> p.RA. ^offeft. 3.oH C. o'? P £ A. 7% ■pfi' ii o O 1% J P l Y S OUJ) 2. 8' It I» A> M r u e d it 10a ACAs ft t a o B> T 0.5 *3 T .1 S B T BT BT o o.r « lo p.p.A\. IfcoN ^6 P £ ft. p p. (*\ . "BT Nt No 5 *° c or*PeA ■> A > s o i J> it fiOrJfte. p l £» 0 A. £ 2. 18 5 oC MhM eft "PAYS B1 l«.ON o.ot C. ® P P €. ft. A1 BT Miuu A WS 2° It a I BT o B T o.S BT ( p.^.NV. B t B T lo B T o 5 T B T 3 t o tf l £ p. p. f* ■ 1 Ao N B T (ft F ig u r e 2 8 6 t o f i g u r e 2 9 1 i n c l u d e d . A v e r a g e a m o u n ts o f a s c o r b i c a c id e x p r e s s e d i n m illig r a m s p e r h u n d red gram s o f f r e s h m a t t e r fr o m t o m a t o p l a n t s g ro w n o n n u t r i e n t s o l u t i o n s s u p p l i e d w i t h v a r i o u s a m o u n ts o f i r o n a n d w i t h o u t o r w it h 0 .0 4 p a r t s p e r m i l l i o n o f c o p p e r . The v a lu e s f o r t h e b a s a l and t h e t e r m in a l le a v e s a r e r e p r e s e n te d b y b a r g r a p h s f o r e a c h h a r v e s t : 5 8 , 7 8 an d 9 7 d a y s a f t e r s e e d in g . The sh a d ed p o r tio n s co rresp o n d t o th e in c r e m e n t o f o n e c o p p e r s e r i e s o v e r t h e o t h e r . No SS o.on p .p .m Cof f CP £ S D Ays <* i-3> C09PC. B. 2>fty:» OlO ‘L 3 £>. M I* T| £ A F O PR.ES** OS ir Af l Ms O.Of p .p. M . wol ’ PEO 7 ? "Pftys o u j . coPPE^ 33 BY S O LT r I vj- 0 S. X &^ < F i (TO p . 6 SL 8 <\ PEP loo <<0 20.ON M. f3 P No 78 IQ I to As*.o«.Ci«c. i X> lo e> t o of Uo a r B r b t o.s 1 r p |» M I^oW frAAMS 6 u f*x £ 0 10 J> aS o a T B t B T 1 o . S 0 . ^ 7 F ' u 0 p . p . J > l b D f P - Y t Io C O **?£ a . O L *> . b G fl 1 0>ON . M r S p . p . aa M ini F i ol 6 a t C. o PP 6 A ^7 J f l l f S <<0 J] r i L i n i . I Iq 5 T O BT oS ST t fcT "BT S 10 oc XFwOOl BT 0 BT o . S G> T I p.p. m . oF G> T S' T 3 ‘ ® x ^oM Figure 292 to fig u re 297 included. Average amounts of ascorbic acid, expressed in m illigram s per hundred grams of oven dry tis s u e from tomato p lan ts grown on n u trien t solutions supplied with various amounts of iron and without or with 0.04 p a rts per m illion of copper. The values fo r the b asal and the term inal leaves are represented by bar graphs fo r each h arvest: 58, 78 and 97 days a f te r seeding. The shaded portions correspond to th e increment of one copper se rie s over the other. Si o copper f 8 C fty S O. Os o lT p . p. Rys o l j F « (r UR £ i q ' i , 5R. > MRTTt p . Cr o (V.E 0 Io p.p.M. ^ o ftMS IOO wft ? £ P* o.S c. o p pe 7 8 ^ ftVS I E o o Io i N p.p. I*. o.oM p. p .p . nv. U o N CoPPEft, 78 I ftVS o L D o-*> F i froP> E 6oo Q.^5 . Hoc Av. >I> loo M III I i d RMi ft s -oft. ftK n 6r i ] e, T o too j j ftr 0. 9 I j j b t *5 j j ex ue t o "* P . p . ftA i e o w I e t i p p . RA Mo co ?Pt* o .o S ^7 1 fty s o L"D, ° n Fi&uft e r ex o.S n S’ bt i° I KflN M.. C . o 7 P e R p .p 3> e v s ou d %e\ i Frcruee ^ b Ae x it>0 QX o a U. T a S T ri e>T oS i 5 p . p na . X Ko (V i l e x lo I o. I t oo.s s T * n II e»T r p . p . PA. a x u e x io 3 R. a N m Figure 298. Average amounts of ascorbic acid a t h arv est, expressed in milligram s per hundred grams of fresh m atter from tomato p la n ts, 58 days old grown on n u trien t solutions supplied with various amounts of iron and without or with 0,04 p a rts per m illio n of copper, p lo tte d against the amounts of iro n , expressed in m illigrams per hundred grams of the corresponding fresh plant m aterial. Figure 299. Same as fig u re 298, fo r th e 78 days old p la n ts. Figure 301. Same as fig u re 298, fo r th e 97 days old p la n ts. Figure 300. Average amounts of ascorbic acid a t h arv est, expressed in milligrams p er hundred grams of fresh m atter from tomato p la n ts, 78 days old grown on various le v els of iro n in the n u trien t so lu tio n without and with 0,04 p a rts per m illio n of copper, p lo tte d against the amounts of manganese, expressed in milligram s per 100 grams of th e corresponding fresh p lan t m a terial. Figure 302. Same as fig u re 300, fo r the 97 days old p la n ts. ^ A S' 8 3o • x> * Y 5 Pi OuP. 6 OA 0. o H © No .W) C«^pC®- 5>t?> i t s t t H ’S P sC fV 'CL a.S 8 A A o O 2fe XJ* "7 8 T> fty> old 1 8 F ioofct yqq. J> f t y c , p i GrU ^ OLD ^ oO <<0 A % A %0 A lo A lo . o. S % 2S 21 o .s o .(o A © o •8 l •© t -2. I. u 0 A 3® ^7 /> 2 3> ft V5 OLD 'lo 'l p i Croat *\1 U fty s © old FYfroftt “i o l 5* A %0 A i« 0 A O a O AA© ©5 M Ct, oF I R 0(0 .A. -X. 0.7 O.S r*&. oF Qfc© l.i © ( *i N\^tJ&PiM9Se A t.S Figure 303. Average amounts of ascorbic acid at h arv est, expressed in milligram s per hundred grams of fresh m atter from tomato p la n ts, 53 days old, grown on n u trie n t so lutions supplied with various amounts of iron and without or with 0.04 p a rts per m illio n of copper, p lo tted against the amounts of dry m atter, expressed in grams per hundred grams of the corresponding p lan t m aterial. Figure 304. Same as fig u re 303* fo r the 78 days old p la n ts. Figure 305. Same as fig u re 303* fo r th e 97 days old p la n ts. ©0 © wo cofpe* s t R i t b t e f P E B , s£ft ' E 5 & A A © © J 8 d fty s P i C*uftE A 3 o3 © © © © 0 A 7 8 I> ftys o t t FiGrui^e 3o*t ✓ i 2 <* j J Fifro^e ioS >p £ o TE(sCfcNT-Aj,£ 7. ° o f 5R.y MATTER. Experiment 7 Table 49. Experimental results obtained from the various leaves and parts of stem of a tobacco plant grown on soil; values expressed per 100 grams of fresh material; the total fresh weight of each is expressed in grams. Distance from Grams of th e th e le a f to t o t a l fre sh the s o il in of each le a f centim eters Grams of dry m atter Grams of water M illigrams of M illigrams of ascorbic acid iro n a t harvest Milligrams of manganese 9 10.5 6.49 93.51 14.8 2.603 --------- 14 12.6 7.09 92.91 18.1 1.330 0.151 IS 16.1 7.89 92.11 18.5 1.422 0.108 23 18.1 8.10 91.90 24.2 0.822 0.301 29 15.2 8.47 91.53 24.3 0.713 0.212 34 13.3 9.11 90.89 28.1 0.817 0.215 39 10.5 10.30 89.70 27.1 1.480 0.300 43 (2 leaves) 14.8 10.95 89.05 25.7 1.403 0 .4 H 51 (6 leaves) 12.3 10.20 89.80 28.6 1.596 0.235 62 (14 leaves) 8.6 10.91 89.09 22.9 1.617 0.412 Stem (upper 19 cm.) 6.85 93.15 16.0 0.238 0.000 Stem (bottom 43 an.) 9.17 90.83 9.6 0.083 0.028 Experiment 7. Prelim inary experiment Table 50. Experimental re s u lts obtained from th e various leaves and p a rts of stem of a tobacco p lan t grown on s o il; values expressed per 100 grams of dry plant m a terial. Distance from the le a f to the s o il in centim eters Milligrams of iro n Milligrams of manganese Grams of fre sh m atter 9 40.10 ------- 1540.8 14 18.76 2.13 1410.4 18 18.02 1.37 1267.4 23 10.15 3.72 1234.6 29 8.42 2.50 1180.6 34 8.97 2.36 1097.7 39 14.37 2.91 970.9 A3 12.82 3.75 913.2 51 15.65 2.30 980.4 62 14.83 3.78 916.6 3.4S 0.00 1459.9 Stem (bottom 43 cm. ) 0.91 0.31 1090.5 Stem (upper 19 cm.) Experiment 7 Table 51. Experimental r e s u lt3 obtained from term in al, median and b a sa l leaves of tobacco p lan ts grown on various cu ltu res d iffe rin g in iro n a v a ila b ility and supplied with no copper and with 0,08 p a rts per m illio n of copper. Averages expressed per 100 grams of fresh p lan t m a te ria l. Culture kind of leaves Milligrams of iro n Series with 0,08 ppm. of copper Milligrams of manganese S eries with no copper S eries with 0,08 ppm. of copper S eries with no copper Milligrams of dry m atter S eries with 0.08 ppm. of copper S eries with no copper Iro n -F rit Terminal 3.24 4.30 0.97 1.13 11.16 10.2 Iro n -F rit Median 4 .SO 5.46 0.94 0.96 ?.42 8.4 Iro n -F rit Basal 3.72 3.87 0.80 0.68 6.90 7.1 Gravel4 ppm. iron Terminal 4.04 4.78 1.09 0.97 10.50 10.0 Median 4.55 7.32 0.81 0.88 8.15 8.1 Basal 3.70 5.47 0.66 0.97 6.73 6.5 NO-iron Terminal 1.64 2.53 0.96 0.84 8.19 9.8 NO-iron Median 2.82 4.42 1.37 0.88 No-iron Basal 3.00 4.58 1.83 0.91 7.04 6.52 8.1 5.9 Experiment 7 . Table 52. Amounts of ascorbic acid a t harvest and a t various times during drying, in the term in al, median and basal leaves of tobacco p lan ts grown on various cu ltu res d iffe rin g in iro n a v a ila b ility and not supplied w ith copper. Averages expressed in m illigrams p er 100 grans of fre sh m atter. Culture Kind of leaves 24 hours a f te r harvest 48 hours a f te r harvest 72 hours a f te r harvest 120 hours a f te r harvest 144 hours a f te r harvest Iro n -F rit Terminal 80.4 45.4 34.9 19.8 10.8 6.1 Iro n -F rit Median 57.1 47.6 13.0 12.8 6.9 5.1 Iro n -F rit Basal 25.4 17.9 12.3 5.8 1.3 1.6 Gravel4 ppm. iro n Terminal 83.4 43.0 27.7 23.5 7.8 5.4 Median 25,7 27.8 25.6 18.1 5.4 3.3 Basal 21.0 21.9 11.1 4.1 No-iron Terminal 70.2 52.4 27.7 21.6 8.9 — NO-iron Median 37.4 33.0 17.0 14.7 7.0 7.1 No-iron Basal 13.8 16.3 6.5 5.9 1.3 1.4 At harvest 1.6 0.7 Table 53. Amounts of ascorbic acid a t harvest and a t various time during drying, in the term in al, median and basal leaves o f tobacco p la n ts grown on various cultures d iffe rin g in iro n a v a ila b ility and supplied with 0.08 p a rts per m illion of copper. Averages expressed in m illigrams per 100 grams of fre sh m atter. Culture Kind of leaves At harvest 24 hours a f te r harvest 48 hours a f te r harvest 96 hours a f te r harvest 120 hours a f te r harvest 192 hours a f te r harvest Iro n -F rit Terminal 72.0 52.6 47.3 14.6 14.1 2.9 Iro n -F rit Median 48.4 34.2 47.1 14.8 13.9 3.3 Iro n -F rit Basal 23.0 21.3 19.8 6.9 4.8 0.9 Gravel4 ppm. iro n Terminal 59.7 57.5 38.2 16.4 11.3 5.0 Median 34.5 31.2 29.2 13.1 12.1 1.7 Basal 10.8 17.4 10.7 5.8 2.4 0.2 No-iron Terminal 47.5 26.6 25.7 6.0 5.7 0.0 No-iron Median 34.9 20.2 14.6 4.3 4.6 ----- NO-iron Basal 17.9 14.6 9.7 ------- 0.1 ----- Experiment 7 Table 54. Experimental re s u lts obtained from term in al, median and basal leaves of tobacco p la n ts grown on various cu ltu res d iffe rin g in iro n a v a ila b ility and supplied w ith no copper and with 0.08 p a rts per m illio n of copper. ilverages expressed p er 100 grains of dry m atter. Culture Kind of M ill! grams of iro n S eries with 0.08 ppm. of copper S eries with So copper M illigrams of manganese S eries with 0.08 ppm. of copper S eries with no copper Iro n -frit Terminal 29.0 42.0 8.8 11.0 I r o n - f r it Median 51.0 61.1 10.0 10.8 I r o n - f r it Basal 50.1 54.6 10.8 9.6 Gravel4 ppm. iro n Terminal 38.5 47.8 10.4 9.7 Median 55.8 90.1 9.9 10.8 Basal 64.5 84.8 11.5 24.4 V No-iron Terminal 20.0 25.8 11.7 No-iron Median 40.0 54.5 19.5 No-iron Basal 46.0 77.5 28.0 8.6 10.8 15.3 Experiment 7 Table 55. Percentages of the original amounts of ascorbic acid found in the terminal, median and basal leaves of tobacco plants grown on various cultures differing in iron availability and not supplied with copper. Culture Kind of leaves At harvest 24 hours a f te r harvest 48 hours a fte r harvest 72 hours a f te r harvest 120 hours a f te r harvest 144 hours a f te r harvest I r o n - f r it Terminal 100 56.5 43.4 24.6 13.4 7.6 I r o n - f r it Median 100 83,3 22.8 22.4 12.1 8.9 I r o n - f r it Basal 100 70.5 48.4 22.8 5.1 6.3 Gravel4 ppm. iro n Terminal 100 51.6 33.2 28.2 9.4 6.5 Median 100 108.2 99.6 70.4 21.0 12.8 Basal 100 104.3 52.9 19.5 7.6 3.3 NO-iron Terminal 100 74.6 39.5 30.8 12.7 — No-iron Median 100 88.2 45.5 39.3 18.7 19.0 No-iron Basal 100 118.1 47.1 42.8 9.4 10.1 Experiment 7 Table 56 . Percentages of the original amounts of ascorbic acid found in the terminal, median, and basal leaves of tobacco plants grown on various cultures differing in iron availability and supplied with 0.08 parts per million of copper. Culture Kind of leaves At harvest I r o n - f r it Terminal 100 73.0 65.7 20.3 19.6 4.0 I r o n - f r it Median 100 70.7 97.3 30.6 23.7 6.8 I r o n - f r it Basal 100 92,6 86.1 30.0 20.9 3.9 Gravel4 ppm. iro n Terminal 100 96.3 64.0 27.5 13.9 8.4 Median 100 90.4 84.6 38.0 35.1 4.9 Basal 100 161.1 99.0 53.7 22.2 1.9 No-iron Terminal 100 56.0 54.1 12.6 12.0 0.0 No-iron Median 100 57.9 41.8 12.3 13.2 --------- No-iron Basal 100 81.6 54,2 — 24 hours a fte r harvest 48 hours a f te r harvest 96 hours a f te r harvest — 120 hours a f te r harvest 0.5 192 hours a f te r harvest Experiment 7. T a b le 57. D if fe r e n c e s betw een th e p e rc e n ta g e s o f th e o r i g i n a l am ounts o f a s c o r b ic a c id found i n t h e te r m in a l, m edian and b a s a l le a v e s o f to b acco p l a n t s grown on v a rio u s c u l t u r e s d i f f e r i n g i n i r o n a v a i l a b i l i t y and s u p p lie d w ith no c o p p e r, and betw een th o s e s u p p lie d w ith 0 ,0 8 p a r t s p e r m i l l i o n o f c o p p e r, a t v a rio u s tim e o f d r y in g . At h a r v e s t Kind o f le a v e s Iro n -frit T erm in al 0 - 1 6 .5 - 2 2 .3 - 6 .2 Iro n -frit Median 0 + 1 2 .6 ~ 7 4 .5 - 1 6 .4 Iro n -frit B asal 0 - 2 2 .1 ' 3 7 .7 - 1 5 .8 B ra v e l 4 ppm. ir o n T erm in al 0 - 4 4 .7 - 3 0 .8 - 9 .5 Median 0 f 1 7 .8 + 1 5 .0 - 1 4 .1 B a sa l 0 - 56.8 - - 1 4 .6 NO-iron T e rm in a l 0 -r 1 8 .6 ~ 1 4 .6 N o -iro n Median 0 -V 3 0 .3 3 .7 •4- 5 .5 N o -iro n B asal 0 + 3 6 .5 7 .1 -t- 8 .9 24 h o u rs a f t e r h arv est 48 h o u rs a f t e r h arv est 120 h o u rs a f t e r h a rv e st C u ltu re - 4 6 .1 -+- 0 .7 Figure 306. Average amounts of iro n , expressed in milligrams per hundred grams of fresh matter from the various leaves of a tobacco p lan ts grown on s o il. Figure 307. Average amounts of manganese, expressed in milligrams per hundred grams of fresh m atter from the various leaves of a tobacco plant grown on s o il. Figure 308. Average amounts of ascorbic acid, expressed in milligrams per hundred grams of fresh m atter from the various leaves of a tobacco p lan t grown on s o il. Fi &j ft. E Jo 2>0 6 , 2 a M vb £ <1 In 18 as “J^ I i s T A M c . 6 P a d i a T iIE Fi &j a. e 0S is uEfwes Si To - n u t Si 62. soit iw CM. i o7 UJ ■^oi Uj 2 < >5 Q 2 < z 31 .b z IS 18 >i5TAWC.t lb pRoM i s I S THt 3 S S i L6AJt«; To 5-1 -TV*e . soil. F i &u a e . 2>oS m 3 ij,T A U C E iff a* FRoKV in T u t 3S l.£ 0 .v jes a“ Si T iT H S => S O I <- fil Iw CM. Figure 309. Average amounts of ascorbic acid, expressed in m illi­ grams per hundred grams of fresh m atter from the various leaves of a tobacco plant grown on s o il, p lo tted against the amounts of manganese, expressed in milligram s per hundred grams of the corresponding fre sh p lan t m a terial. Figure 310. Average amounts of ascorbic acid, expressed in m illigrams per hundred grams of fresh m atter from the various leaves of a tobacco p lan t grown on s o il, p lo tted ag ain st th e amounts of iro n , expressed in milligrams per hundred grams of th e corresponding plant m a terial. Figure 311. Average amounts of ascorbic acid, expressed in milligrams per hundred grams of fresh m atter from the various leaves of a tobacco plant grown on s o il, p lo tted against the amounts of dry m atter, expressed in grams per hundred grams of th e corresponding fresh plant m aterial. M Cr o PibwO^> ^ o o ©© ACi D *3 O —r- * O 2 i- (TJ O © N3-3-tf3 J 3 ‘SVl 3° ASCoRBic 4 a? MG- m o 00 j- G© D 0 o O© cP 0 © CP Z. 0 0 » u A v . i.T> Figure 312. A verse amounts of manganese, expressed in milligrams per 100 grams of fresh m atter from the various leaves of a tobacco p lan t grown on s o il, p lo tted against the distance from th e leaves to the s o il, expressed in centim eters. Figure 313. Average amounts of ascorbic acid , expressed in milligrams per 100 grams of fresh m atter from the various leaves of a tobacco plant grown on s o il, p lo tted against the distan ce from th e leaves to the s o il, expressed in centim eters. © © c o © © 0 o © f i & o a. e 3 -L. IO le J # 3 >»s t a m c e L,o the S'O u&aoes to the 6O sox. ih CAA . o o © o o © o Fi O J k E IO ■J)i S T Ao A w c £ ko 30 T ‘i l ' i W £ i t f t o E So S T o T « £ 6 0 S o il- 1 *^ CM igure 314 to fig u re 319 included. Average amounts of dry m atter, expressed in grams per hundred grams of fresh m atter from tobacco plants,grown on various cu ltu res d iffe rin g in iro n a v a ila b ility without and with 0,08 p a rts per m illio n of copper. The values fo r the b asal, median and term inal leaves are represented by bar graphs. The shaded portions correspond to the increments of one copper se rie s over the other. u> 5 (U o UJ . _ Y UJ __ UJ w d o 0 X u< H X X o 0 & ,< * 0 »-• c>) J «r a. 0 2 0 2 V) U1 Jl IU d d (u 0 CO t' mJ 0 • V) — cO _J UJ ? d «!> 2 d uj UJ CJJ C/1 <2 O r .J ul n rd> UJ <£ 0 0 IX P uJ u. a 0 2 (x j < 2 £ ^ O -T *■* tO -lLV W ^>SCL 0 3 - 5 VJ JL N 3 ^ 0* 3 d. J* » H igure 320 to fig u re 325 included. Average amounts of iro n , expressed in m illigram s per 100 grams of fresh m atter from tobacco p la n ts, grown on various c u ltu re s d iffe rin g in iro n a v a ila b ility without and w ith 0.08 p a rts per m illion of copper. The values fo r th e b asal, median, and term inal leaves are represented by bar graphs. The shaded portions correspond to the increments of one copper se rie s over th e other. LoPPEft C O P P t R. F m T & RfcU EL C . U L T O R . E S C U L T o K E F. rl TEdKiNM 3^0. F i& J f t E K c Di RN Wo CoPPE R S E r R l T C u 2> 2. ' . coPPER OUPivlE 1. F i & j F 6 (_Tj R € 5 no s E ^ \ E s . i«>6N Culture r i E. 2 SEft-'ES iJLTuRt 1 3 '2. *i . . fA & ni Wo C o P P E R. i t P,vEs N D wo C-oPPER I R S E ft. , E * oN 3 iI S . F.cuRc 2i.s I I I I Figure 326 to fig u re 331 included. Average amounts of iro n , expressed in m illigram s per 100 grams of oven dry tis s u e from tobacco p lan ts grown on various cu ltu res d iffe rin g in iro n a v a ila b ility without and with 0.08 p a rts per m illio n of copper. The values fo r th e b asal, median, and term inal leaves are represented by bar graphs. The shaded portions correspond to th e increments of one copper se rie s over the other. CoPPeR. pRi T c o p ? e a S£fti£« & pvftoE.L tuLTORE S 80 Ft 1S.B c u l t u r e s ^ E co p p ea No I fi*ON 3 '-7 ■ se a its C-'Ji-Tufi.E.S Fi &j B. e '32-8. nO-e>(0 f* • *>«. e s e a i e s M t O i AN 6RSAU K h Ml of T£R»a ; n »\U tA K tvio fAn-L * No C o P P ta SP8i£S c o p p e a s e ftiEs &aftVEL £. o L T j R E S Fi Jrw 8 E 330- no c o fp e a NO IR oM P iuae s e a v E s 3 ”i i f m t cuLTuae^ F ii d aa. UJ *j •j ti 0 0 U 2 o 0 U- z ■ ^ u. l- cfl r/ O »* U i d o \r jO 3 C"5 U t0 o K0 tO cO d Ui J tii d •** a ui 0o. o o 0. o. 0 d U i vo to uJ d Ui "2 rf ui 10 D h -j 3 <£ o< O ui t cO cO ti a: U i UJ -1 a o a. a. o d O U. < * 2 a. i <£ w t- _ j_ _ i_ H oo tf> U i Ui cO d , 0 1 Vn r d U i J fO a. D a. d U I 0 < * d U_ M . o* Q T Q a O a ki m a m u t.l s \im * 4 y'^yys cr o Figure 33# to 343 included. Average amounts of manganese, expressed in milligrams per 100 grams of oven dry tis s u e from tobacco p lan ts grown on various cu ltu res d iffe rin g in iro n a v a ila b ility without and with 0.08 p a rts per m illio n of copper. The values for the b a sa l, median, and term inal leaves are represented by bar graphs; the shaded portions correspond to the increments of one copper se rie s over the other. CoPPef t FAi t Seizes c. u i. T 0 A E fi&o R6 T£fcMi(OftL No FA i T Fi&u A E n o AG & e c a a \/ S f A.' iES 3 3 R Wo fi&oAE 3^ I. S € A i6i ASM P> &o B.E 3 ^ 0 I n CofPG A G A f t o E C S copPEA No S tR ifJ M Basal W£ O i A . i \ J C ol T a F iC r o A C 338. C- of PE A coppe S t A\Gs C o L T o A G S 3 m . No No i CoPPGA SG A i £ S X A oN Fi &o A £ 2>W3 . I M & f'A e> 6 Figure 344. Average amounts of ascorbic acid, expressed in m illigrams per 100 grams of fre sh m atter from th e term in al, median, and b asal leaves of tobacco p lan ts grown on an iro n containing f r i t supplied with a n u trie n t so lu tio n with 0.08 p a rts per m illion of copper; p lo tted against the time of th e various determ inations during the diying of the plant m a terial, expressed in hours. Figure 345. Same as figure 344, fo r p la n ts grown on gravel supplied with a n u trien t solution containing 4 p arts per m illio n of iro n and 0,08 p a rts per m illion of copper. Figure 346. Same as figure 344, fo r p lan ts grown on gravel supplied with a n u trien t solution deprived of iro n and with 0.08 p a rts per m illio n of copper. SERIES Ho o \oo & P- C.F F A E i « M ATE CoPPEf t . COff Eft. CEK & ft* ft u e l F i &• y ft. 6 3 *» 5 lo o M i l l i &• «. ft AAV of f t S t o d e i c R«_ » X> 6o culture s Co f P Ef t - fo No S E< ME5 X Rofi FifruKE 2)^6. T <<0 \ 3 • i. 0 io * - 6o So I*# i** Ti m ® XM w m* o » So F'.i-ufl.e 3>5o a *»» a lo o lo © ‘ ■ i.S 4.0 •■------------ J.s" • ...... 3-© * ^ ‘ - ■ ------- ■A. ‘i . o 3 5 MG- p F ‘i - S S 5 ■ o S iftoU a 0 a ----- Fi Cr 0 fc E 3 S' 1, 13 col- a A a a A A o o a» 0 0 T ^s I^ S 5*75 3*0 r - a 3’ fAG>. 0 C® C"5 V ^ P r~o fe.S 7,£> V. Figure 352. Average amounts of ascorbic acid a t h arv est, expressed in milligram s per 100 grams of fresh m atter from the term in al, median, and basal leaves of tobacco p la n ts grown on various cu ltu res d iffe rin g in iro n a v a ila b ility , supplied with a n u trien t so lu tio n containing 0.08 p a rts per m illio n of copper, p lo tte d against the amounts of manganese in the corresponding fresh tis s u e , expressed in m illigram s. Figure 353. Same as figure 352, fo r the tobacco p la n ts grown on a so lu tio n deprived of copper. G TtR*A«NAL & MEDv AM G> S a s a »- □ Ft fru R 6 0 S’ 2. . A O © J2_ O of o.4 t.O (.1 N\Cr. t-Z O f !.*> N \ A.VJ & A i.W M E I. f 1C 1.7 I. s S f rfl So o .7 o-3 tACr . . t t . A. o f fA A »0 & f t H t s £ I.'i |. A Figure 354. Average amounts of ascorbic acid a t harvest, expressed in milligrams per hundred grams of fresh m atter from the term in al, median, and basal leaves of tobacco p la n ts grown on various cu ltu res d iffe rin g in iro n a v a ila b ility , supplied without and with 0.08 p arts per m illio n of copper, p lo tted against th e amount of dry m atter, expressed in grams per hundred grams of fresh m atter. 0 0 0 0 0 0 & © © O □ TEfctAvNfu. & AACfciKfJ 0 feftSKL A © © © R&-OB.E •» ? ?£(>vC.£N Tft (VS °» OF 5>f>»y IO <* A T T £ R 3>rt Figure 355 to fig u re 360 included. Amounts of ascorbic acid a t h arv est, 24, 48, and 120 hours a f te r h arv est, expressed in milligram s per 100 grams of fresh m atter from the term in al, median and basal leaves of tobacco p lan ts grown on various cu ltu res d iffe rin g in iro n a v a ila b ility , supplied without and with 0.08 p a rts per m illion of copper. The shaded portions of th e bar graphs correspond to the increments of one copper se rie s over the o th er. tn A iu £ ffl ui A A d (/} .•> u. rVl d ui & d 4 a. UJ 0 > O «c cf O UJ 3 »b Ul UJ Ui ct 0 H J 0 rO o cd d H Ul UJ O VO Ul v\ .U •+i d ui 0c> £ Ui •0 O) In (O A Ui <£ 0 f- • o r _j I n 0 o m Ui CL iu J Ui d T A Ul in aa. <- ui d 3 l*J ** u. Ui o •va Sty on 3 In »o J 0 d U) o d ■> »V 0 d i- 2 U- j= E Figure 361. Percentages of th e o rig in a l amounts of ascorbic acid found in the fresh tis s u e s from the term inal, median and b asal leaves of tobacco p lan ts grown on an iro n containing f r i t , supplied with a n u trien t so lu tio n with 0.08 p arts per m illio n of copper; p lo tted against th e time of th e various determ inations during the drying of the p lan t m aterial, expressed in hours. Figure 362. Same as figure 361, fo r the tobacco plants grown on gravel supplied with a n u trie n t solution containing k p arts per m illio n of iro n and 0.08 p a rts per m illion of copper. Figure 363. Same as figure 361, for th e tobacco p la n ts grown on gravel supplied with a n u trien t so lu tio n deprived of iro n and with 0.08 p arts per m illio n of copper* c o p p c 6 A w PRi T SERiEfi. CoLTuRfcS Pi G o A 6. i6S . - TE R M i U AL - NV E O f A ti Co o Eo is o oS f ts c a ( \e ■ a> a s ft u Wo COPPER & A. A0 € t- 6£P(ES CULTURE 5 T I* t o EU W PiGuP-e SER'iES Figure 367. D ifferences between the percentages of th e o rig in al amounts of ascorbic acid found at each determ ination in the term inal, median and basal leaves of tobacco p la n ts grown on the iron-containing f r i t deprived of copper and the percentages obtained from the plants grown on the same media but supplied with 0,08 p a rts per m illio n of copper, p lo tted against the time of each determ ination, expressed in hours a f te r harvest. Figure 368. Same as fig u re 367, fo r the tobacco p lan ts grown on gravel supplied with U p arts per m illion of iron. Figure 369. Same as figure 367, fo r the p lan ts grown on gravel supplied with a n u trien t solution deprived of iro n . TfcKl'V. fvjfcu Oi M 4 P tT f ' Gr O (L 6 oats 3 6 7 t o L ' T o a C £> Figure 370. Percentages of the o rig in a l amounts of ascorbic acid 24 hours a f te r h arv est, in the b asal, median and term inal leaves of tobacco plants grown on various cu ltu res d iffe rin g in th e ir iron a v a ila b ility and supplied with 0.08 p a rts per m illion of copper in the n u trie n t so lu tio n , p lo tted against the amounts of iro n in the corresponding plant m aterial, expressed in milligrams per 100 grams of fresh m aterial, expressed in milligrams per 100 grams of fresh m aterial. Figure 371. Percentages of th e o rig in a l amounts of ascorbic acid 48 hours a fte r h arv est, in th e b asal, median and th e term inal leaves of tobacco p lan ts grown on various cu ltu res d iffe rin g in th e ir iro n a v a ila b ility and supplied with 0.08 p a rts per m illion of copper in the n u trien t so lu tio n , p lo tted against the amounts of iro n in the corresponding plant m aterial, expressed in milligrams per 100 grams of fresh m a terial. . r- u* d n s 0 ill 0 0 0 P3* O */> U i j v r o t Ic5 4- >■» V ■>'.*» V •"» ® 0 in Figure 372. Percentages of the o rig in a l amounts of ascorbic acid 96 hours a f te r h arv est, in th e b asal, median and term inal leaves of tobacco p la n ts grown on various cultures d iffe rin g in th e ir iro n a v a ila b ility and supplied w ith 0.08 p a rts per m illio n of copper in th e n u trie n t so lu tio n , p lo tte d against th e amounts of iro n in the corresponding plant m a terial, expressed in milligrams per 100 grams of fresh m aterial. Figure 373. Same as figure 372, 120 hours after harvest. ? e AC. PMT A>* £ of T«£ oAx V* t" t\tAOJ i of '.6 f»sc.0< 0 2. *> 5 ■p m P e ^ c t w T R 6-e of T*r£ (o 2 at r. s * m p m ✓ 0© AMajiiT aPA iCaftfcic Am. ‘ 5 i 2. o Viojh o A ^ T c li bAW.c'j” Figure 374. Percentages of tte o rig in a l amounts of ascorbic acid 24 hours a f te r harvest in the b asal, median and term inal leaves of tobacco p lan ts grown on various cultures d iffe rin g in th e i r iro n a v a ila b ility and deprived of copper in th e n u trien t so lu tio n , p lo tted against the amounts of iro n in the corresponding p lan t m aterial, expressed in milligram s per 100 grams of fresh m aterial. Figure 375. Same as fig u re 374* 48 hours a f te r harvest. "Ve*. c. &^~ o•»> 2 t. f \ * c o a. ft i c* •N o Ho O ^ s J& 0 R C l t R Rl\K O fc O 0 o *■ © \S3.’^5S V ^ 0 M 7' o ? &> to >A P. o 0. V| © 0(5>D 7» _ -3 «* (P ^ r r-■ (P j> m pj v H2 ^ tc. P (O J> P 11 X ip ?• * -I-J -4 O I'tAcEWCfl. e o -» 3-Wo, 0 _EL O »/■> *4 <3 e^l <£.'''y 3 l <0'ijo ■><;y .J: >V 5 ! * 8 » o :»3y .Q uO fO i ^ oi . rJ 3-* Wa w 3 3 ' o 3