THE WTAMIN B] AND C CONTENT OF MUNG BEANS AM," mum BEAN SPROUTS Thesi: for the Degree of M. S. WCHCAN STATE CCLLEGE Hui-Jan Yeh 1939 n. .5} T. T .3 r.. . ., . , aka .4... ,J ‘wmw...&g;3.~fitnfl.. .n.r... THE'VITAMIN Bl AND C CONTENT OF HUNG BEANS AND HUNG BEAN SPROUTS B: Huiplan‘Zch A.THESIS Submitted to the Graduate School.of Michigan State Collage of Agriculture and Applicd Science in partial fulfilment of the rcquiremcnta for thc dcgrcc of EASTER OF SCIENCE Department of Nutrition Division of Home Economics 1939 THESIS ACKNOHLEDGMENTS The.vriter wishes to express her sincere appreciation to Miss Flora M. Hanning of the Home Economics Division of Michigan State College, under whose supervision this study Ias conducted, for her interest and helpful criticism, and to Dr. Marie Dye, Dean of the Home Economics Division of Hichigan State College for her helpful.suggsstions. Acknowledgment is also given to the La Choy Food Product Company, Detroit, Michigan for generously donating the bean sprouts used in this study. TABLE OF CONTENTS Page Introduction 3 Literature Review 6 Determination of Moisture and the Proportion of the Cotyledonal and Non-cotyledonal Parts Experimental Procedure 10 Results and Discussion 11 Vitamin B1 Experimental Procedure 13 Results and Discussion 20 Vitamin G Experimental Procedure 26 Results and Discussion 30 Summary 35 Bibliography 36 Appendix 58 THE VITAMIN B1 AND C CONTENT OF HUNG BEANS IND HUNG BEAN SPROUTS _;ntroggction Green mung beans (Phaseolus 522333), sometimes called green gram beans, are very commonly used in China either in the form of dry beans or sprouts. but more often as sprouts. Mung bean sprouts take the place of vegetables in the diet. They are available all through the year and are quite cheap. In some parts of the country the sprouts are sold.vith the cotyledonal part out off, probably for better palat- ability and appearance. Mung bean sprouts are undoubtedly an important item in the Chinese dietary. There are many kinds of mung beans. Those commonly used in China are more or less round in shape, green in color. and about 2.5 mm. by 2.5 mm. by 3 mm. in size. This kind of beans was also used in th1I~IXP periment. The method often used in cooking the dry beans is boiling in a large quantity of water for hours until very tender. The details of the procedure of sprouting the beans differs slightly in different sections of the country; but the general principles are almost the same. They are: l. lashed clean. 2. Soaked for about 5 to 10 hours. 3. Sprouted in a perforated bottom container. 4. Sprinkled.vith eater many times a day to insure dampness. 5. Kept in a perfectly dark and warm condition. The sprouts are usually 7 to 9 cm. in length from root tip to the expanded cotyledons, of which about 5 cm; is white, fleshy, and thick. The cotyledons are pale yellow in color and have expanded. Often the plumuls-has emerged from between the cotyledons. The plumule is usually yellow in color and § to 1 cm. in length. Though some investigations have been made of the value of mung beans and their sprouts as sources of vitamins, the information is far from‘complete. This work was undertaken in the hope of increasing our knowledge of this important Chinese food. The object was to investigate the three points mentioned below: I. The vitamin B and 0 content of mung bean sprouts. 1 2. The difference of the content of these vitamins between dry and sprouted beans. 3. The distribution of these two vitamins in cotyledonal and non-cotyledonal parts. Literature Review Vitamin B1 Most of the work on the vitamin value of mung beans and mung bean sprouts was done in India.and China and published in those countries. Miller (11) reported that Embrey (5) had found that dry mung means, when fed as 452 of the weight of the diet, contained enough vitamin B1 for the growth.of mice. The bean sprouts also were fed in dried form, and at a 251 level, contained enough vitamin B1 for the growth of mice. About l0 mice were used in each group. Embrey suggested as a result of her experiments that vitamin Bl increased when the beans were sprouted. Santos (12) also stated that the cooked sprouted beans showed greater vitamin Bl potency than cooked beans. He used recovery of weight of animal as a measure of the vitamin B of the food tested. The age of the rats was not stated, but most of them weighed considerably over one hundred grams when placed on the experimental diet. Both the beans and the sprouts were cooked, then dried and fed as a.vitamin B1 supplement. Seven rats were fed on the beans, 4 receiving 1 gm., and 3 receiving i gm. daily supplements; and 6 rats on the sprouts, 3 re~ ceiving 1 gm., and 3 receiving é gm. supplements daily. Miller (ll) reported the vitamin value of mung bean sprouts, supplied by the local market in Honolulu. Sherman's method was fol- lowed in her research for the comparison of vitamin B1 value between raw and cooked sprouts (steamed for 5 minutes). Nine rats were put on 3 gram raw sprouts level and l on 4 gm. level. For cooked sprouts 10 rats were put on 2.7 gm. level and 2 on 3.6 gm. level. The rats were fed daily except Sundays for a period of 8 weeks. She concluded that the amount of raw sprouts for maintaining net weight in standard rats for a period of 8 weeks was between 2.5 gm. and 3.0 gm., and between 2.2 and 2.7 gm. for the cooked sprouts (equivalent to 2.5 and 3.0 gm. of raw). According to Sherman's unit basis, raw bean sprouts con- tained 150 to 180 units per pound and the cooked sprouts 150 to 170 units per pound of which correction had been made for the difference in water content. Jansen (9) stated that about 30% of katjang idjoe (mung beans) was enough to cure polyneuritis in pigeons. Donath (4) found that katjang idjoe were quite rich in vitamin Bl. van Veen (14} reported one International unit of vitamin 31 per gm. of dry black mung beans (Phaseolus radiatus). Spruyt (13) found that black mung beans had the same amount of vitamin B1 as green.mung beans. The work of Wilson (16} showed that the green mung beans con- tained 150 to 160 units of vitamin Bl per 100 gm. of dry beans. The unit of vitamin 81 was defined as the quantity which, when given to a rat each day would produce an increase of weight of 10 gm. per week for at least 3 weeks. They claimed that this unit was equal to an Inter- national unit and to 4 micrograms of the Jansen and Donath‘s crystals. The rats were put on.vitamin 81 free diet when they weighed about 30 gm. At least 6 rats were used for each test. Sue-l5... . I‘D-T Vitamin C Donath (4) found that dry katjang idjoe (mung beans) contained no vitamin C. Ghosh (7) found by titration method that dry beans contained 4.0 mg. of vitamin 0 per 100 gm. and sprouted beans con- tained 31.0 mg. per 100 gm. Ahmad (7) found that the vitamin C content of dry mung beans was 3.0 mg. per 100 gm. and of sprouted beans was 23.0 mg. per 100 gm. Guha (8) found that ascorbic acid in Phaseolus Muggg_ was increased 7 to 8 times by germination. Miller (ll) reported that 2.5 to 3.0 gm. of raw sprouts seemed to protect guinea pigs from scurvy, while 2.? gm. of cooked sprouts (equivalent to 3.0 gm. raw) did not. However, according to her ex- perimental data, she thought that the amount needed to protect against scurvy probably lay between 3 to 4 gm. Nine guinea.pigs distributed on 4 different levels of raw sprouts, and 8 guinea.pigs on 4 different levels of cooked sprouts were used in this experiment. The results obtained indicated that there was considerable destruction of vitamin 0 even in the 5 minutes of cooking. She concluded that, on the basis of Sherman's unit, raw mung bean sprouts contained 180 units of vitamin 0 per pound; and in the cooked state, the bean sprouts probably had ap» proximately 150 units per pound or possibly a little less. She claimed that in the raw state mung bean sprouts have a vitamin c content equal to that of lemon, orange, and tomato Juice; and.when cooked 5 minutes, a slightly lower value. flats (15) reported that 3 gm. of sprouts of each of the varieties of beans tested was enough to protect guinea pigs from scurvy. His result checked very closely with Killer's (ll) findings. Obi (3) found that mung bean sprouts contained 0.022.mg. per am. determined by the Harris method, 0.15 mg. per gm. by the iodometric method, and [+(I)] by biological assays. I. II. 10 Eeiszsinaiige.efflusisiu£2.ead.ihs.Ezensziisn.21.ins. Esixlsdenalrend.£22:22ixlssseel.Basis Experimental Procedure Moisture: Since the method and condition for sprouting the beans is slightly different in different places, the moisture content is expected to be different. It would be of interest if the vitamin value could be compared on the dry basis so these determinations were made. About 20 gm. of fresh or frozen bean sprouts, and about 10 gm. of dry beans were used in each determination. Two or four de- terminations were made on each sample. They were weighed on an analytical balance to one tenth of a mg. The samples were first dried at a temperature below60°0. in.en oven which had an electric motor to circulate air. Then, the samples were transferred into another ordinary drying oven at 65°C., and dried to constant weight. The moisture content of fresh sprouts from Detroit was determined at 60°C., but it was found there were not enough changes in weight to cause e.decrease of 0.11 when put into the oven at 65°C. Proportion of the two parts -~ cotyledonal and non-cotyledonal: In each determination 100 gm. of bean sprouts were used. The sprouts were cut at the point right next to the cotyledonal pert. After all of them were cut, the two parts were weighed again. The weighings were made on a torsion balance. All of the processes were completed as quickly as possible and with minimum I. II. handling so as to prevent loss of moisture. Results and Discussion Hoisture: Moisture content is given in Table I. According to the moisture content of sprouts from Lansing, 100 gm. of dry beans as purchased would give 1125 gm. of bean sprouts. According to the moisture content of bean sprouts from Detroit, 100 gm. of dry beans as purchased would give 1506 gm. of sprouts, assuming that the addition of water would account for the total change in 'Cight. Table I. Moisture Content Material .219: Moistgzg Dry beans 6.6 Powdered dry beans 6.5 Fresh bean sprouts from Lansing 91.? Fresh bean sprouts from Detroit 93.8 Frozen bean sprouts from Detroit 93.4 Proportion of two parts -- cotyledonal and non-cotyledonal parts: There were 19 determinations of 5 different samples made. The percentage of cotyledonal part ranged from 10.6 to 12.82 with an average of 11.41 by weight. The percentage of non-cotyledonal part ranged from 86.4 to 89.01 with an average of 87.82. The sum of the percentage of these two parts was 99.21. The weight lost due to evaporation was 0.81. Assuming that the rate of evaporation I .. ,! she'lrf I 12 was the same on these parts, the percentage of the cotyledonal and nonpcotyledcnal parts would be 11.5 and 88.52; This was termed as corrected proportion of the two parts. The results are given in Table 2e Table 2. Proportion of cotyledonal part and non-cotyledonal part of the bean sprouts. No. of Cotyledonal Nonpcotyledonal determinations part part 1 10.81 87.51 3 11.0 89.0 2 12.8 86.4 4 12.0 87.0 9 10.6 88.8 iverage 11.42 87.82 Corrected average 11.51 88.51 13 Vitamin B1 Experimental Procedure I. Preparation of supplements: I. 2. 3. The bean sprouts. The bean sprouts for the vitamin B1 assay were kindly supplied by the La Choy Food Products Company in Detroit. During the period of transportation the sprouts were kept in dry ice. They looked perfectly fresh upon arrival. Im- mediately after the bean sprouts arrived, a small portion of them.was cut in the same manner as was done in the determina» tion of the proportion of the cotyledonal and non, cotyledonal parts of the sprouts. After this was completed (about 3-4 hours, during which they were kept cold) both the whole sprouts and the non-cotyledonal parts were put into wax paper cups of one pint capacity, placed in the freezing room in the Horticulture Building, and kept frozen. The dry beans. The dry beans were purchased from Lansing. These beans and the beans used by the La Choy Food Products Company for sprouting were shipped from.Manchuria. The beans were washed with water, dried in air, and ground to fine powder. The standard solution. The standard vitamin B1 solution was prepared by dis- solving exactly 10 mg. of thiamin chloride hydrochloride in 14 100 0.0. of 20: alcohol solution. This solution was called stock solution. Two 0.0. of the stock solution was diluted to 250 0.0. with distilled water. Five 0.0. of the final so» lution equaled 4 gamma of thiamin chloride hydrochloride. II. Preparation of basal ration: Chase and Sherman's (2) vitamin B1 deficient diet was used. The composition of the ration is as follows: 1. 2. Casein free from vitamin Bl 181 Salt mixture (McCallum.and Steenbock) 4 Butter fat 8 God liver oil 2 Autoclaved yeast 15 Starch 53 Purification of casein. The casein.was purified by extracting with 601 cold alcohol (by weight). With 1.5 liter of 60: alcohol, 300 gm. of casein was treated and the whole was shaken for § hour and then allowed to stand for 5% hours. The mixture was fil- tered with suction and thoroughly washed with 750 c.c. of 60% alcohol. The casein was shaken with another lnliter portion of 60% alcohol for one half hour. After standing 19 hours, it was filtered, washed with 750 0.0. of 601 alcohol, and finally with one half liter of 95% alcohol and then dried in air. Autoclaving yeast. Dried powdered baker's yeast was mixed with 0.1 NaOH to make a smooth paste in a proportion of 100 gm. yeast to 15 125 0.0. NaOH. The mixture was then heated in a pressure cooker at 15 pounds pressure for 6 hours. The resulting yeast was neutralized with standard HCl. It was dried in an oven at 60° C. and then ground. 3. Salt mixture. Salts were weighed on a torsion balance. They were ground separately in a mortar, mixed and ground. The final mixture.was passed through a 40 mesh seive. Composition of salt mixture #2 --- U.S.P.X. (Modification of McCollum and Steenboch #40) Sodium chloride (NaCl) 0.173 Anhydrous magnesium sulfate (MgSO4) 0.266 Sodium phosphate (NaH‘zPO4 0 820) 0.347 Potassium phosphate (K2HPO4) 0.954 Calcium acid phosphate CaH4 (P04)2 . H20 0.540 Ferric citrate (léHéO) 0.118 Calcium lactate 1.300 4. Butter fat. ~~Commsrcial butter was melted, decanted, and filtered. 5. Cod liver oil. --Patches' cod liver oil was used. 6. Corn starch. --Argo's corn starch was used. III. Selection and care of animals. Rats:were used as experimental animals. They were of known heredity. when the rats were about 4 weeks old and weighed about 47-55 gm., they were put on vitamin Bl deficient diet for about IV. 16 2 weeks. At the end of the first week harnesses were put on so as to prevent coprophagy. After the first week, rats were weighed every other day, and some times each day at the end of the second week. Growth curves were made. The weight of the rats usually went down after harnesses were put on but it would usually be up again on the fourth day. lhen the weight of a rat went down the second time, that rat was considered to have been depleted in vitamin 31. The average weight of the whole group at the end of the depletion period was about 70 gm. Rats were distributed into different groups according to sex and litter. About 4-5 rats were put on each supplement at the be- ginning of the experiment. Later on, more rats were put on the level where the growth curve was considered to be nearer to the positive control group. Table III shows the distribution. After rats were put on supplement, they were weighed weekly and the food consumption was also recorded. it the time of weigh- ing, harnesses were taken off and were not put back until late in the afternoon. During this period the rats could clean themselves. Feeding supplements. 1. Experimental period. A.4 weeks_growth test period was used. It has been found in this laboratory that the 4 weeks period has approximately the same significance as longer periods. 2. Technique in feeding. All supplements were fed each day including Sunday. Supplements were limited, but basal diet was fed ad. libitum. 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Water and food were available all of the time except for special reasons as mentioned above. Bean sprouts. The approximate amount of sprouts needed daily was taken out each day from the freezing room. The amount left was put into-an evaporating dish, wrapped with wax paper, and kept in the freezing unit of an electric refrigerator. The sprouts were weighed on a small sensitive torsion balance immediately after they were taken out from the freezing room. Care was taken to keep them from thawing. Weighing while they were cold was, of course, not the most accurate way, but, if time were taken to let them come up to room temperature, the bean sprouts would thaw and water would come out from the tissue. Since vitamin B is water soluble, the loss of water this way 1 probably induced higher error than weighing while they were cold. 4. 5. Dry beans. The dry beans were kept in,a glass Jar at room temperature. They were weighed also on the same torsion balance on which the sprouts were weighed. Each gm. of beans was mixed with about 1-2 gm. of basal ration and then fed to the rats. During the first week of the experiment beans were moistened with water; but later it was found that after drying, the beans became a hard mass, and the rats did not like it. Hence, the beans were mixed.with food instead of moistened with water. Thiamin chloride hydrochloride solution. It was pipetted out with a.5 0.0. pipette into a small cup. Vitamin B1 Results and Discussion The records of individual rats are given in Tables 9 to 24 in the appendix, and the summary of data may be found in Table 4. The curve of the growth of rats and the histogram of the food consumption of the rats are shown in Fig. l. The vitamin B1 value, expressed in terms of micrograms of thiamin chloride hydrochloride and International units (equivalent to 3 micro~ grams of thiamin chloride hydrochloride), was calculated on the basis of gain in weight with the positive control group as a standard of comparison. Theqresults from levels of 6 gm. of both the non-cotyledonal part of sprouts and the whole sprouts were used in calculating the vi~ tamin B1 value. The results are tabulated as follows: Table 5. Vitamin 31 Content of Mung Beans and Mung Bean Sprouts Vitamin £1 Value 9; IQQ gm, on raw weight M Microgram of International licrogram of International thiamin unit thiamin unit Food Miner 100 gm. per lQQAgms, per LQQ gm, pg; IQQ gm, Dry beans 618 206 662 221 Whole bean sprouts 85 28 1288 429 Hen-cotyledonal part of bean sprouts 66 22 -.— -.. 21 n.bl uoHH ooaa pom m.m row mooH moo >¢ «.ma m.es «.ms o.o~ e.em m.o¢ m.mv «.me o.mn a.mn o.ov v.ee e.ov o.mn m.an o.mn o.mn m.mm o.nn o.mn n.0e o.ee o.me «.mn m.mn o.an n.nn n.on o.on n.on m.mn m.mn m.mn o.en m.en m.en m.mn m.mn m.mn «.mn >4 .3 . . 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Some of the rats died. \‘~. 1 unit equal to 30 gm. \\ 22 The results showed that dry beans contained 618 micrograms of thiamin per 100 gm. raw weight, and 662 micrograms of thiamin per 100 gm. dry weight; and sprouts contained 85 micrograms of thiamin per 100 gm. raw weight and 1288 micrograms of thiamin per 100 gm. dry weight. Apparently the vitamin 81 increased about 2 times as much as that of the original dry beans after sprouting. The growth rate per week in the dry beans group was considerably higher than the positive control group. If the supplement level were decreased about one fourth of the amount fed in this experiment, the vitamin Bl°value might be a little higher than the present figure; but this would not effect the result very much as shown by the fact that the value calculated from the 8 gm. level sprouts gave almost the same value as from the 6 gm. level of sprouts.. When the results from the 8 gm. level sprouts, in which the growth rate of the rats was even higher than that of the rats supplemented with 1 gm. of dry beans, was used in calculating, the vitamin Bl content of bean sprouts was 81 micrograms of thiamin chloride per 100 gm. ras'weight. The vitamin Bl value was only 4.microgrsms thiamin chloride less than that calculated from the results of the 6 gm. level of sprouts. When the vitamin B1 value was calculated from the results of the 10 gm. level of bean sprouts, the vitamin B1 content per 100 gm. raw weight dropped to 69 micrograms of thiamin chloride. This might in» dicate that the amount of vitamin Bl in.8 gm. of bean sprouts was almost enough fer the maximum growth of rats. When the vitamin B1 in- take was increased beyond what the body needed, it light be simply stored in the body, probably excreted, or it might be utilized less efficiently by the body. When the amount of sprouts in the supplement 24 was increased from 6 to 8 gm., the gain in weight increased accordingly; but when it was increased from the 8 to the 10 gm. level, the gain in weight of the rats was considerably less per unit weight of the sup- plement. Therefore, the vitamin Bl value calculated from the 10 gm. level of sprouts would not represent the true value of the sprouts. Furthermore, the number of animals used in the 8 gm. and the 10 gm. levels was less than that in the 6 gm. level; the average value from the 8 and 10 gm. levels would not be so significant as that from the 6 gm. level. The vitamin Bl value of non-cotyledonal part of the mung bean sprouts was decidedly lower than that of the whole sprouts. There were only 66 micrograms of thiamin chloride in 100 gm. of raw non- cotyledonal part of sprouts, while there were 85 micrograms in 100 gm. of raw whole bean sprouts. In other words, the vitamin 81 was not evenly distributed in the whole sprouts but more concentrated in the cotyledonal part. When the vitamin value was calculated from the results of the 10 gm. level, the vitamin 81 content of the non-cotyledonal part of the sprouts was 58 micrograms of thiamin chloride per 100 gm. raw weight. This value was also lower than that calculated from the results of 6 gm. level. There were only 4 rats on each of these levels, which is rather a.small group of rats in view of the individual variations of the rate. This test has found that dry mung beans are a good source of vitamin 31 (206 International unit per 100 gm. raw weight); whole bean sprouts are a fairly good source (28 International unit per 100 gm. raw weight): and the non-cotyledonal part of bean sprouts is a fair source of vitamin B1 (22 International unit per 100 gm. raw weight). After sprouting, the vitamin Bl content of the beans increased to approximately two times the original value. 25 26 V tam n C Experimental Procedure The beans used for this part of the experiment were purchased in two Chinese restaurants in Lansing. In both of the restaurants, the beans were generally sprouted for seven days. The length of the sprouts was about 7-9 cm., which was almost the same as those from the La Choy Food Products Company, used in the vitamin B test. The l sprouts were stored in a tightly covered glass Jar and kept in an ice box before analysis. They were analyzed as they were, without being washed. The vitamin C content was determined by the microchemical titration method. The details of Tillman's method modified by Bessey and King (1) were followed with few modifications. Bessey and King (I) feund that the dye titration method checked with the biological assays. We have assumed that to be true of mung bean sprouts, and in this study, the reducing value of the materials tested is reported as vitamin C, I. Standardization of 2-6 dichlorophenolindophenol solution. Meander and Guerrant's method (10) was used. They found that the curve obtained for the titrations against sodium thiosulfate was comparable with that in which ascorbic acid was used. In successive portions of hot water,0.l gm. of dye was dissolved and filtered into a.200 c.c; volumetric flask. When all the dye was dissolved, the filter was washed with a small amount of hot water until the washings were nearly colorless. II. 2? Fifteen c.c. of the dye solution was pipetted into a.100 c.c. Erlenmeyer flask; 0.5 to 1.0 gm. potassium iodide and 0.5 to 1.0 c.c. of diluted sulfuric acid (1 to 4) were added. After shaking to facilitate the oxidation of the potassium iodide, the liberated iodine was titrated with 0.01 sodium thiosulfate, using 1 c.c. of 12 soluble starch as an indicator. The sodium thiosulfate was standardized with standard potassium dichromate solution. The potassium dichromate was of good quality, dried, and had been kept in a dessicator. Pales' procedure (6) of standardization was followed. One c.c. of 0.01 N sodium thiosulfate was equivalent to 0.88 mg. of ascorbic acid. Extraction and titration. About 5 gm. of dry beans or bean sprouts were weighed and ground with 2-3 gm. of acid washed white sand and a small amount of 21 meta-phosphoric acid until a paste was formed. The mixture was washed with some more 21’metapphosphoric acid and transferred to a.50 c.c. centrifuge tube. The mixture was centrifuged then for about 5 minutes. The clear solution was filtered through a. quick filter (Whatman No. 41) into a 100 c.c. Erlenmeyer flask. Another 10 c.c. or 15 c.c. of metapphosphoric acid were used to wash the mortar and stirred into the solids, which were again centrifuged. The washing and rinsing were repeated twice more. The total volume of about 50 c.c. was titrated with standardized dye. A persistence of a pink color for about 15 seconds was taken as the end point of the titrations. The.titration was carried out within two minutes. The time between weighing and titration was kept as short as possible -- usually within two hours. III. 28 During this period, direct sunlight was kept out of the laboratory as much as possible. Distilled water was used instead of redistilled water as recommended by Bessey and King(l). The use of distilled water in the manipulations was checked with redistilled water twice. Lemon Juice.was used as a source of vitamin C. It.was found that even when the distilled water was concentrated 5 times, the difference of the results between distilled and redistilled water was less than 11, which is within the experimental error of the method. Therefore, distilled water was used all through the experiment. The method of standardization was checked many times. The value of dye solution standardized by Heander's (10) method was practically the same as the value standardized against pure ascorbic acid. A difference of 1-28 was within the experimental error. During the period of acquiring technique, at least four determinations were run on each sample. Afterwards only duplicates or triplicates were run. Only two determinations were carried on at one time so as to lessen the atmospheric oxidation. The vitamin c content of dry beans, of whole sprouts, and of the non-coty- ledonal part of bean sprouts was determined. The cotyledonal part was calculated by difference. ‘ Recovery tests: 1. Test for atmospheric oxidation. _ 0.5 mg. portiom of pure ascorbic acid solution dis- solved in 21 metaphoric acid were pipetted out into 100 c.c. Erlenmeyer flasks and were let stand without covering for 2. 3. 29 the same amount of time as the grinding process would require. Then, these solutions were diluted to about 50 c.c. with more 21 metaphcsphoric acid and let stand for the same amount of time as the actual determination would require. The results indicated that the destruction of vitamin c by atmospheric oxidation was less than 21 in cool weather, and the percentage of destruction was higher in hot summer weather. It went up to about 101 destruction on a very hot day. Test for completeness of extraction: 0.5 gm portions of pure ascorbic acid solution were ground, washed, and filtered in the same manner as the actual de- termination. The results were compared.with the same amount of ascorbic acid which was exposed to air for the same amount of time. The results showed that the extraction was complete. Oxidation in presence of‘oxidaset 0.5 gm. portions of ascorbic acid solution were added to weighed samples of sprouts and the amount of vitamin 6 found in excess of the average amount of the vitamin c of the same sample was taken as the amount of recovery. In this case, it was very difficult to get a very uniform sampling of sprouts, but the average of the recovery might give some idea of the action of’oxidase under the particular situation. Recovery tests were done on two samples of sprouts and one of dry beans. The average percentage of recovery of two determinations on one I of the samples of sprouts was lOlZ, and the average of three determinations on the other sample was 99.7Z'(97.0Z, 98.91, 103.2%). The average percentage of recovery of two deter- minations on dry beans was 962 (94.02, 98.01). Results and Discussion The results of the vitamin c determination are given in Table 6. The average vitamin C content of dry beans was 6.? mg. per 100 gm. raw weight, and 7.2 mg. per 100 gm. of dried weight (dried at 65°C.). The aserage vitamin C content of the whole bean sprouts was 9.4 mg. per 100 gm. fresh weight and 113.3 mg. per 100 gm. dried weight. The average vitamin c content of the non-cotyledonal part was also 9.4 mg. per 100 gm. fresh weight. Apparently the vitamin c distributed uni- formly in these two parts, and the proportion would be the same as the proportion of fresh weight. In (17) reported that the moisture content of whole bean sprouts was 91.71 and of the non-cotyledonal parts was 93.2%. If the vitamin c content of the non-cotyledonal parts was calculated on this basis, it would become 138.2 mg. per 100 gm., indicating a slightly higher con- tent of vitamin C in the non-cotyledonal part on the dried weight basis. One hundred gm. of raw dry beans contained 6.7 mg. of vitanin C, and the amount of sprouts (1125 gm.) from 100 gm. of beans would have 105.8 mg. of vitamin C. The vitamin 0 value of mung beans was in- creased 15.8 times when the beans were sprouted. A summary of the data is given in Table 7. The reported value of viatmin O in green mung bean sprouts ranged from 2.2 mg. to 31.0 mg. (7) per 100 gm. of fresh sprouts. Miller (11} used biological assays and reported more than two times as much vitamin C as reported in this study. The methods of sprouting, the time of sprouting. the kind of beans, and the freshness of the sprouts may account for these differences. 31 Table 7. Summary of Vitamin 0 Content of Dung Beans and lung Bean Sprouts. Percentage Eitamig 9 content mg, pgr 199 gm in: moisture Eggsh has}: Dried basis Bean sprouts 9l.7 9.4 113.3 Nonpcotyledonal parts of sprouts 93.2 9.4 138.2’ Dry beans 6.6 6.7 7.2 ' Moisture content was not determined in this laboratory. 32 Table 6. Vitamin 0 Content of Mung Beans and Their Sprouts NO. Of San» deterb Vitamin 0 value in mg/g ple. min- of fresh weight of each 3g. Fggg atigns titration Av, Remarks 1 Dry beans 2 0.062 0.066 0.064 2 Z metaphos— 2 ' 4 0.062 0.065 0.064 0.067 0.066 phoric acid was used. 3 9 2 0.068 0.070 0.069 52 trichloro- 4 ' 4 0.063 0.070 0.070 0.072 0.069 acetic acid was used. AV. . 0.067 5 Bean sprout 2 0.099 0.104 0.102 The sprouts were fresh. 6 " 3 0.096 0.100 0.103 0.100 Determinations were made 1 to 3 111.8. “tCr 7 ' 1 0.084 0.084 delivery. 8 ' 1 0.093 0.093 9. ' 2 0.090 0.091 0.091 10 " 2 0.110 0.111 0.111 11. * 3 0.067 0.082 0.081 0.077 Av. ‘ 0.094 Fresh deter- minations were 12¢ . 2 0.089 0.092 0009]. m0 “tar stored over night. 13 ’ 3 0.086 0.088 0.098 0.091 Non-cotyled- 14 onal.part of 3 0.075 0.085 0.085 0.082 bean sprout. They were fresh. 15 9 2 0.088 0.105 0.097 Determinations were made 1 to 16 ' 2 0.099 0.100 0.100 4 hrs. after delivery. 17 “ 2 0.095 0.099 0.097 AV. . 0.094. Fresh, stored 18 . 2 0.090 0.092 00091 over night. 33 Khan the bean sprouts did not look very fresh, slightly dry, or light brownish in color at the tip of the sprouts, the vitamin 0 con- tent was usually about one half of the fresh sprouts. The average vitamin c contents of two samples (21 and 23, not fresh) were 5.3 mg. per 100 gm. and 5.9 mg. Per 100 gm. of the sprouts. Sample 20, though stored over one night, locked fresher than sample 21. The vitamin C content of sample 20 was accordingly higher. The vitamin 0 value of samples 19 and 22 varied so much in different determinations, that the results were not averaged. Uhen the bean sprouts looked quite brownish all over or in part, and slightly soft, the vitamin 0 value according to the titration method, dropped to almost none. This effect of the condition of the samples is tabulated on Table 8. If, however, the fresh sample was stored over night in a re- frigerator and appeared in good condition, it gave results comparable to those determined with the shorter time before testing, as shown in samples 12, 13, and 18 of Table 7. This test has found that dry mung beans are a poor source of vitamin C (6.7 mg. per 100 gm. raw weight); but the vitamin C content increases 15.8 times after sprouting. Both whole bean sprouts and the non-cotyledonal part of the sprouts are fairly good sources of vitamin c (9.4 mg. per 100 gm. fresh weight). .omdv sou eoom sees no: was a“ census peeaseemoc scam «soamm osmosu scam no: van mmo.o owo.o mmo.o «mo.o n a nu .mpapcdsc omada ca Hop so“ cu passe capo unease .eHo «use a 093.0 eeo.o m . mm .hnc eoaooa .hpapcdsa aaeau cannon so“ ca use”: capo cocoa» .eHo made a nmo.o mmo.o aeo.o N . Hm .nop nod cw cams: capo cocoa. .eao «use a mmo.o eao.o amo.o mmo.o Hmo.o mao.o m . om .voa use macaw vases» neamoa .Non new ma cam“: cube escape .nHo whee m NmH.o oncoo mucoo n essence seem mH mmmwaom «MM coupmwadp ecoammc each small code we pswaow seen» no tasaouou oaascm .sm\ms as ends» 0 cassaa> «o .oz .annsau pony mesons” seem no vasecoo o oasopdb om snack III. IV. V. 35 Summary Dry mung beans, mung bean sprouts, and the non-cotyledonal part of the sprouts were analyzed for their vitamin B1 and C content. The average vitamin Bl content (microgram thiamin chloride hydrochloride per 100 gm. raw weight) of (a) dry mung beans was 618 micrograms, (b) bean sprouts was 85 micrograms, (c) non-cotyledonal part was 66 micrograms. The average vitamin C content (miligrams per 100 gm. raw weight) of (a) dry mung beans was 6.7 miligrams, (b) bean sprouts was 9.4 miligrams, (c) non-cotyledonal part of the sprouts was 9.4 miligrams. The vitamin C content of mung beans was increased 15.8 times after the beans were sprouted, and the vitamin B1 content of mung beans was increased about 2 times after the beans were sprouted. The vitamin C of the bean sprouts was evenly distributed in the whole sprout, while vitamin B1 probably was more con- centrated in the cotyledonal part of the sprouts. 2. 3. 4. 5. 5. 7. 11. 12. 13. 36 Bibliography Bessey, 0. A., and C. C. King. The distribution of vitamin C in plant and animal tissues, and its determination. J. Biol. Chem.. 103, 687, (1933). Chase, E. F., and H. C. Sherman. A quantitative study of the determinations of the antineuritic vitamin B. J.A.C.S.. 9E5 3505, (1931). Chi, and B. B. Read. The vitamin C content of Chinese foods and drugs. Chinese J. Physiol. 9, 47, (1935-6). Donath, W. F. Synopsis of the antixerophthalmic antineuritic, and antiscorbutic vitamin percentages, besides the pro- tein, fat, carbohydrate and water contents of different Indian foodstuffs. Mededeel. Dienst Voldsgezondheid N,ed8rl&nd.-Ind10.. lg, :54, (1929). Embwey, H. Investigation of some Chinese foods. China Med. J., 32, 405, (1921). False, H. 4. "Inorganic Quantitative Analysis." p. 354. Fixsen, A. and Roscoe. Tables of the vitamin content of human and animal foods. Nutr. Abs. and Rev., 1, 823, (1937-8) Guha, B. C. and A. R. Ghosh. Vitamin C in Indian foodstuffs. Current Sci., 3, 210, (1934). Jansen, B. C. P. On the need of anti-beri beri vitamins of the animal organism and on the amount of this vitamin indifferent foodstuffs. Dutch East Indies-Mededeelingen van den Burgerlijk Geneeskundige Dienst. 61, (1923). Menader, M. H. and N. B. Guerrant. Standardization of 2, 6-dichlorophenolindophenol. An improved method. Indst. Eng. Chem. Anal. Ed.’ lg, 25’ (1938). Miller, C. D. and D. B. Hair. The vitamin content of mung bean sprouts. J. Home Econ., 29, 263 (1928). Santos, F. 0. Some plant sources of vitamin B and 0. Am. J. Physiol. g2, 310, (1922). Spruyt, J. P. and W. J. Donath. Antineuritic (Bl) vitamin content of 'black' katjang idJoe (Phaseolus radiatus). Geneesk. Tinschr. Ned.-Ind., 1g, 601, (1935 . 14. 15. 16. 17. van Veen, A. C. Vitamin B content of some foods. Gennesk. Tijdschr. Ned-Ind., 1;, 2500, (1935). Bats, R. C. and w. J. Woodhouse. Some sources of vitamin C in India. The antiscorbutic value of various kinds of sprouted mung (Phasiolus Muggo) and their extracted Juices. Indian J. Med. Res., g;, 167, (1934). Wilson, H. E. 0., B. A. Ahmad, 0. Ray, and R. 0. Cuba. The vitamin B content of some common Indian foodstuffs. Indian J. Med. Res., as, 813, (1937). \ £3» —%fi- Wu, H. “Introduction to Nutrition." 5: 3;. ‘3 12131.”; 37 1.111132%}: 39 m.u n.me m.m o.a m.m o.m m.m n.o o.m m.m m.o~ n.m aaam aaxoo; owduo>d n.9m new mm am an m en mm mm mm me Hm nwdm Haves a; me n o .2 m 2 a mm m S n J? :3 m 3 a N.» mm 0 ca H- m- u m ma ca w o .mn nun mm 0.. me n n ma n e n o e «H n .xu ecu .u T! u 9.. an ca nH HH m HH m am a ma a .ma and «.mm «no em an up an ma Hm mu Hm ms nu “some: adapaca IumauaaaIaaMmHwImaMmullaauumn11uwwawanaqqfiaa " .. .¢ _. ..... .... . a .4 macaw oaoct o o no u no «nu no aqua ovum Houpaoo opapamoa no gauche .a oanau m4: n62 ”6 at: “1.3 m.» 3: mg a.» ad.“ ”.3 find 3% ago... ewdnmbd flme awe am 3 3 S. 9 an .3 S we 3 5% H33. a; mm a. a S w m o 3 S 2 .3 3:. fie a 0.3 8H a m a ma cm 3 _. cm .3 3 :3 En W ma mm a ma 3 S .7 m d n ma mm .ma Em m v.2 3H d 3 3 a. ma «H o 3 m 2 .xa a: m. .38 Se 2. 3 an 8 ma. 5 mo 2. 2. 8 232. Gonna do“: case m 3 ,womn ”Macao £50” £38 bag... 3.3% $33 qwfimn mwfimn mam! . .5» mo sumo mom and «zoom was: man .am H np«h_vuucoaenmasm audm no £93050 .OH candy 1.8.1310 11 9 Growth of Rats Supplemented with 6 gm Non~ cotyledonal Part of Mung Bean Sprouts Per Day. Gain of the whole group 11;; number 59579“ 59673“ 5979? 59582” Total Average Initial weight _ 68 62 63 64 257 64.3 a let wk. 8 8 7 6 29 1.3 0H 3'9 2nd wk. 7 4 e 2 21 5.3 :? -§ 3rd wk. 16 4 10 10 40 10.0 C) a: 4th wk. 0 5 3 a 15 4.0 Total gain 31 21 28 26 105 ’ 26.6 Average weekly gain '7.8 5.3 7.0 6.5 26.6 ' 6.7 Table 12. Growth of Rats Supplemented with l0 gm. Non- cotyledonal Part of Mung Bean Sprouts Per Day. 41 Gain of the 0 whole group Rat number 59449119 3941313 5945.11 59393LL 79131 Merge Initial weight 53 74 54 74 280 70 let wk. 3 12 8 12 35 8.8 ‘1 H a, 2nd wk. 7 12 7 10 56 9.0 :3 3rd wk. 8 12 7 _ 14 41 10.5 0 0 3 4th wk. 10 12 8 15 46 11.5 Total gain. 28 48 30 52 158 39.6 Average Eggklx_g§in ;:;.gg __12.Q7 7 5 3 0‘1 39.§ 9.9 42 mum use 3. 9: ma .3. . 0.3 né a; m.» a; ”.0 Sam 3%... omduw>< a.en aen mm we mm on we . an _mm mm an an eaem Haven 1.; S a d 3 m a 2 o m ma a .5; fie u 92 m2 2 S a 2 ma a 3 3 S m :3 B... W. a... d. w 3 m." n S a o m o m .53 gm m n6. 8 o m n m 3 a. a. S 3 e .uw «3 m. «1% new 8 mm 2. 2. we 2. S E. ,2. me 23.; H353 33,“. .593 a; A; Us» ea 58 and hem ausouam doom was: macs: no .aw o spew vopcoaoaaasm muom mo cabana .na candy Table 14. 43 Growth of Rats Supplemented with 8 gm. of Whole Mung Bean Sprouts Per Day. Gain of the whole ggggp Bat 11ng 393988 39442“ 394598 3941311 393791. 761,31 Averagg Initial weight 77 73 61 75 68 354 71 let wk. 16 13 7 19 8 63 12.6 £1 ”é 2nd wk. 18 0 1 10 10 39 7.8 p... 33 3rd wk. 20 12 9 17 9 67 13.4 O 34’ 4th wk. 16 6 4 14 11 51 10.2 Total gain 70 31 21 60 38 220 44.0 Average weekly gain 17.5 7.8 5.3 15.0 9.5 55 11.0 Table 15. Growth of Rats Supplemented with 10 gm. of Whole Mung Bean Sprouts Per Day. Gain oft— whole grp. Rat numbgr 394183 39453L 39453; 39412R 394423 3937‘!5 Tota1 Av, Initial weight 74 65 71 68 59 78 415 69.2 a let wk. 17 8 11 14 11 4 65 10.8 «4 a, 2nd wk. 4 8 10 1 3 18 44 7.3 I». 3‘; 3rd wk. 21 9 18 8 9 18 83 13.8 0 O _ E 4th wk. 17 14 16 18 6 19 90 15.0 Total gain 59 39 55 41 29 59 282 46.9 Average weekly gain 14.8 9.8 13.8 10.3 7.3 g 14.8 70.5 11.7 V m 44 3... «:3 an- 3. .. Sn. 3... 3... me. n6- 58m 366.. 03925 nave... 2.7 a? 8. em- mm- mm. mm- mm- flaw H38. 33 we: 8:. .637 mm... a- e .. 3.. n a m? .aw fie M . 8 3:. on: m. 3.... S... n- e .. m? a .. n .. a. .. a... .64.. Zn 1 n 3... mm- m .. e .. a... 2.. 2.. 3.. ma .3? new m. 0.9. mm- o? 2.. m... m- ma a... H .3. £3 3.6 m5. an em 8. me 2.- ma 2. .232. H325 amlamnnn 4436a. amoaan 59.2 «£99.. £33. £33 £53 kanan ennmae pen a o E» 2: ea 55 even Hoaueeu eafinmez ae fiaonu .3 633. 45 m.en men m.en m.mn n.mn m.an m.oe n.en en m.oe o.me n.mn nonpaaaneeo hHMeew .hd o.meH oeeH anH mea an ema mea ana eeH men wnH awn nonpaaaneoe H38. m.nn «an an an 3 mm 3. an ca 9. 3. am .xw fie e O 8 men an... S me an am 3 en an d. an mm .6? En mm. .01 men men 8 me. an an 3. mm mm d. we on .4? new. m... u 4.8 3... mm. en 04 an an an an ca 8 mm a? and Ila. edllieae H38. amend... «Sen 4352 wee dduuluaonw one no godpa83mcou aflobmn miwrmmm qmrwmn mwbnmn ommvmn magma mwflémn uwvvmn mpdm Monacoo abavfimom no :0fl9a82mcoo doom .rH wands 46 m.en men n.nn n.nn mm mm n.nn m.an n.en n.an n.mn o.ee eeaaaaaeeee haxmew .bd e.eeH eeefl meH mna and mmH and ens and one mea «we eeafiaaaneoe Haney a.nn awn oe mm an mm oe an me me an we .aa nae m u m . 8 9 33 mm... on .3 mm. on 3. 9. en .3 me we .xw En mm. .a.L 11 n.4n men an an am an on en mm mm an «n .k. can m. m.en men en en an an an an em oe am we .uw pea .uammnmawn Hnfiea 7meoaam iwmean mannan mmaean mamean -maean .weaan gunman qwumamwamwammmls.ummammgmam m&dum.fi&ozs asylum cedaQBSumoo hen yum «seem was: age No .aw H span vopcoeoaaasm mde we coapaasncoo uooh O .mH manda Table 19. Food Consumption of Rats Supplemented with 6 gm. of Non-cotyledonal Port of Mung Bean Sprouts Per Day. Consumption o ' _ g: the wh91s gzoup " 59678' 39198‘ 59593 Totg gvergge Rat number 3967* lot wk. 31 31 30 29 121 30.3 8 “'3 2nd wk. 31 29 31 27 118 30.0 poo. gag: 3rd wk. 34 27 32 28 121 30.3 3’: O ‘3 4th wk. 35 32 35 31 133 33.3 Total consumption 131 119 128 115 4934 123.9 Av. weekly consumption 32.8 29.8 32 28.8 123.3 31.0 Table 20. Food Consumption of Rats Supplemented with 10 gm. of Non-cotyledonal Part of flung Bean Sprouts Per Day. Consumption of thg who}; 5:022 Rat number 59448113 394183 394523 59393141. $01.11 Averagg lst wk. 55 :57 32 38 142 55.5 .3 2nd wk. 5:5 43 59 43 158 39.5 D. g; 3rd wk. 41 44 36 47 168 42.0 I c: 8 4th wk. 41 4a 38 49 176 44.0 Total consumption 150 172 145 177 644 161.0 kw. weekly consumption 37.5 43. 36.3 44.3 161 40.3 .Nqihfll 31”": ll?“ 1“ “1 .l.|t. 1- 48 can 33... 9mm pawn 9mm 0.3 nan Con 83 man n.mn 9mm 8332928 has... .5 823 $2 «2.. SH. mm." 03 SH «3 93 SH .2: m3 53695.18 H38. 6.8 man 5. an an 8. 3. G a... S 8 Q .4; 5.. m 83 mm» mm mm on mm no an an m9 3 mm .3: En mm 0.3 on». 3 on mm mm S 3 mm mm on mm .8- 2.... WW 9m... 36 S .3 S «n mm mm mm an 5 mm .9: 33 33503 £39.. madam mwmwmn 593% £35» 33 souamaamcou o no: pom «vacuum zoom mos: macs: no em m swan nouooauanasm mama no comaassmcoo voom .HN manna 49 Table 22. Food Consumption of’Bsts Supplemented with 8 gm. of whole Mung Bean Sprouts Per Day. Consumption - of the WW Rat number 3939331 39442 394529 394111L 393793 1W lat wk. 40 37. 32 42 37 188 37.6 n - 0 gr: 2nd.wk. 50 36_ 27 45 40 198 39.6 D. o {a 3rd wk. 44 40 35 47 37 203 40.6 a - o ” 4th wk. 60 38 31 53 40 222 44.4 Total consumption 194 151 125 187 154 811, 162.2 Av. weekly consumption 48.5 37.8 31.3 46.8 38.5 20.3 40.6 Table 23. Food Consumption of Rats Supplemented with 10 gm. of While Mung Bean Sprouts Per Day. Rat number Consumption — - W 39413“ 394591. 39453 394198 395433 39318-Totg Avegggg 8 1st wk. 40 33 43 34 31 33 214 35.7 €23 2nd wk. 40 37 41 31 34 45 228 38.0 .23 3rd wk. 50 41 so 38 33 42 254 42.3 ° 4th wk. 52 47 49 '41 33 51 273 45.5 Total consumption 182 158 183 144 131 171 969 16.2 kn. weekly consumption 45.5 39.5 45.8 36. 32.8 42.8 242.3 40.5 50 m.mH m.8nH 8.8H 8.8H o.Hm 8.8H 8.8H 8.8M 8.8m 88388288888 sauna; .88 H.88 888 mm 88 mm H8 88 H8 888 88288288888 H8989 I 8.38 8838 a 8.3 mm o 8 NH 8 S .98 58 m 8.38 8.28 a «.ma am an 8 8H NH 8H ma mm .9: 8.8 w 0.8... 03 3 3 2 2 om mm mm .28. 88mm. T! O 8.88 as“ am 88 em em mm 88 an .x- was u lawns-M8488. 9.8 ”2.2 ummvmm; 8:483 a 888 88 8888 839188 awn 3 8828-185 .3 7” m A... w my. e ..w coaumssucou ovum Houucoo coaunMoz mo soupmssmoou moon .cm oHnnB ROOM usr. om -~ Aprm 2:: Se 2’43 " ' , .P-"i-P‘ LU” 'J‘Y LOW AprIS’M Jun 12 45 S6,“ ‘7 11A Apr 5 ’49 . h 9 W 0W \‘7 I;‘ ‘1 J I '. 3‘. v I l l i ' \ l I v r l I . ‘ . l‘ .- .-— -._ '. -AIJ. ‘— .t-.. v ~9-9- bunf— -' ' ._'-!‘_1 :L'__‘l 0 3'!- 'I ‘