I FACTORS AFFECTING N - (PHOSPHONOMETHYL) GLYCINE- (GLYPHOSATE) ACTIVITY IN TURFGRASS AND ALFALFA E MEDICAGD SATIVA (L) J SEEDLING ENVIRONMENTS AND DEGRADATION IN THE SOIL Dissertation for the Degree of Ph. D. MICHIGAN STATE UNIVERSITY ' LOREN JAMES MOSHIER ’ 1977 - . I\\\\\\\\\\\ \IIIIIIIIIIIIIIIIIII I- n v Michigan State University This is to certify that the thesis entitled FACTORS AFFECTING N-(PHOSPHONOMETHYL)GLYCINE (GLYPHOSATE) ACTIVITY IN TURFGRASS AND ALFALFA [MEDICAGO SATIVA (L.)] SEEDLING ENVIRONMENTS AND DEGRADATION IN THE SOIL presented by Loren James Moshier has been accepted towards fulfillment of the requirements for Ph.D. degree in Dept. of Crop and Soil Sciences q .. __..=——- -::; flw/ 6 Major professor Date 'IQDK .3/ /f 7] 0-7 639 fia/o ABSTRACT FACTORS AFFECTING Nfi(PHOSPHONOMETHYL)GLYCINE (GLYPHOSATE) ACTIVITY IN TURFGRASS AND ALFALFA [MEDICAGO SATIVA (L.)] SEEDLING ENVIRONMENTS AND DEGRADATION IN THE SOIL BY Loren James Moshier The effects of several concentrations of B—(phosphono- methyl)glycine (glyphosate) on germination and seedling growth of four Kentucky bluegrass (Poa pratensis L.), two creeping bentgrass (Agrostis palustris Huds.) and two red fescue (Festuca rubra L.) cultivars were evaluated in petri dishes under controlled environmental conditions. None of the glyphosate concentrations had a detrimental effect on germination of any of the turfgrasses examined. A high con- centration of glyphosate (10-4 M) did reduce shoot growth in all but two cultivars. Siduron, a selective preemergence herbicide used in turfgrass establishment, had no effect on either germination or shoot lengths. Glyphosate or siduron applied to the soil and incorporated did not affect turfgrass emergence or growth. When glyphosate was sprayed over the soil with the seed lying on the soil surface, red fescue germination was reduced and shoot growth of all three turf- grass species was inhibited at the high application rate (17.9 kg/ha). Under field conditions, none of the glyphosate treatments reduced turfgrass emergence or growth. Loren James Moshier The extent that glyphosate affected alfalfa (Medicago sativa L.) germination, emergence and seedling growth in several systems was also evaluated. Glyphosate and EPTC, a selective preemergence herbicide used in alfalfa establish- ment, caused no significant reduction in percent germination of either Saranac or Vernal cultivars grown in petri dishes. Glyphosate reduced shoot length of both cultivars. Vernal appeared to be more susceptible to glyphosate than Saranac. Soil applications of glyphosate also had no effect on percent emergence of either cultivar and only reduced plant height when applied to the soil at 17.9 kg/ha. Glyphosate applica- tions to a Kentucky bluegrass sod in a simulated sod seeding alfalfa establishment study in the greenhouse reduced dry weight of both alfalfa cultivars. Application of glyphosate in four management systems showed that a 3-day planting delay of water application after spraying reduced growth inhibition of alfalfa seeded in treated sod. 14 Factors influencing degradation of C-glyphosate to 14CO2 was studied in three soils. Glyphosate degradation occurred primarily by microorganisms and at variable rates. Lhosphate additions stimulated degradation to a limited extent in the Collamer silty clay loam but not in the Norfolk loamy + and A1+++ reduced degradation in sand. Additions of Fe++ the Spinks sandy loam. It is postulated that formation of colloidal Fe and Al precipitates in modified soils with concomitant adsorption of glyphosate was responsible for Loren James Moshier decreased availability of glyphosate to microorganisms. Mn++ additions were found to increase degradation. Spinks soil and organic carbon amendments failed to increase gly- phosate degradation in soils with low degradation rates. FACTORS AFFECTING Nf(PHOSPHONOMETHYL)GLYCINE (GLYPHOSATE) ACTIVITY IN TURFGRASS AND ALFALFA [MEDICAGO SATIVA (L.)] t" _. SEEDLING ENVIRONMENTS AND DEGRADATION IN THE SOIL BY Loren James Moshier A DISSERTATION Submitted to Michigan State University in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Department of Crop and Soil Sciences 1977 ACKNOWLEDGEMENTS The author wishes to express his sincere appreciation to Dr. Donald Penner for his guidance and patience at all times during this study and in the preparation of this dissertation. Gratitude is expressed to Dr. W. F. Meggitt for his cooperation and guidance and to Dr. B. G. Ellis, Dr. A. R. Putnam and Dr. J. M. Tiedje who also served on my guidance committee. Appreciation is extended to Dr. A. D. Kern and Dr. D. B. Sharp, Monsanto Agricultural Products Co., for their contribution of ideas. The assistance of numerous work- study students in the greenhouse and laboratory aspects of this study, and the assistance of my wife, Deanna, in typing the manuscript is gratefully acknowledged. Appreciation is also extended to Monsanto Agricultural Products Co. for their financial assistance and for supplying 14C-glyphosate. ii TABLE OF CONTENTS Page ACKNOWLEDGEMENTS. . . . . . . . . . . . . . . . . . . . ii LIST OF TABLES. . . . . . . . . . . . . . . . . . . . . v LIST OF FIGURES . . . . . . . . . . . . . . . . . . . .viii INTRODUCTION. . . . . . . . . . . . . . . . . . . . . . 1 CHAPTER 1: LITERATURE REVIEW: DISSIPATION OF GLYPHOSATE IN SOIL. . . . . . . . . . . . . 3 Introduction. . . . . .‘. . . . . . . . . . . . . . 3 Chemical and Photodecomposition . . . . . . . . . . 3 Microbial Decomposition . . . . . . . . . . . . . . 4 Influence of Management Practices on He bicid Dissipation . . . . . . . . . . . . . . . . . . 10 Summary .'. . . . . . . . . . . . . . . . . . . . . 15 CHAPTER 2: EFFECTS OF GLYPHOSATE AND SIDURON ON TURFGRASS ESTABLISHMENT . . . . . . . . . . 16 Abstract. . . . . . . . . . . . . . . . . . . . . . 16 Introduction. . . . . . . . . . . . . . . . . . . . 17 Materials and Methods . . . . . . . . . . . . . . . 17 Results and Discussion. . . . . . . . . . . . . . . 20 Literature Cited. . . . . . . . . . . . . . . . . . 30 CHAPTER 3: .USE OF GLYPHOSATE IN SOD SEEDING ALFALFA ESTABLISHMENT . . . . . . . . . . . . . . . 31 Abstract. . . . . . . . . . . . . . . . . . . . . . 31 Introduction. . . . . . . . . . . . . . . . . . . . 32 Materials and Methods . . . . . . . . . . . . . . . 34 Results and Discussion. . . . . . . . . . . . . . . 37 Literature Cited. . . . . . . . . . . . . . . . . . 50 iii CHAPTER 4: FACTORS INFLUENCING MICROBIAL OF 14C-GLYPHOSATE To 14CO2 IN Abstract. . . . . . . . . . . . . . . Introduction. . . . . . . . . . . . . Materials and Methods . . . . . . . . Results and Discussion. . . . . . . . Literature Cited. . . . . . . . . . . CHAPTER 5: SUMMARY AND CONCLUSIONS . . . LIST OF REFERENCES. . .,. . . . . . . . . iv DEGRADATION SOIL. Page 51 51 51 53 56 75 76 78 LIST OF TABLES Page CHAPTER 2 l. TThe effect of glyphosate and siduron treatments on germination of four Kentucky bluegrass varieties grown in petri dishes. . . . . . . . . 21 2. The effect of glyphosate and siduron on percent germination of bentgrass and red fescue grown in petri dishes . . . . . . . . . . 22 3. The effect of glyphosate and siduron treatments on the shoot length of four varieties of Kentucky bluegrass grown in petri dishes . . . . 23 4. The effect of glyphosate and siduron treatments on the shoot length of bentgrass and red fescue grown in petri dishes. . . . . . . . . . . . . . 24 S. The effect of glyphosate and siduron applica- tions on percent germination of three turfgrass species grown in soil in the controlled environment chamber. . . . . . . . . . . . . . . 26 6. The effect of glyphosate and siduron applica- tions on the plant heights of three turfgrass species grown in soil in the controlled environment chamber. . . . . . . . . . . . . . . 27 7. Effects of preplant incorporated and preemer— gence application on stand density of two turfgrass species 8 weeks after planting . . . . 23 8. Effects of preplant incorporated and preemer- gence applications of glyphosate on clipping yield of two turfgrass species 5 weeks after planting . . . . . . . . . . . . . . . . . 29 CHAPTER 3 l. The effect of glyphosate and EPTC treatments on germination of two alfalfa cultivars grown in petri diSheSO O O O O O O O O O O C O O O O O 38 2. The effect of glyphosate and EPTC treatments on the shoot length of two alfalfa cultivars grown in petri dishes. . . . . . . . . . . . . . 39 V Page 3. The effect of glyphosate applications on percent emergence of two alfalfa cultivars grown in soil in the controlled environment chamber. . . . . . . . . . . . . . . . . . . . . 40 4. The effect of glyphosate applications on plant heights of two alfalfa cultivars grown in soil in the controlled environment Chamber. 0 O O O O O O O O O O O O O C O I O O O 41 5. The effect of glyphosate applications on percent survival of-two alfalfa cultivars 4 weeks after planting in Kentucky bluegrass using a single spray-seed operation. . . . . . . 43 6. The effect of glyphosate applications on dry weight yields of two alfalfa cultivars 4 weeks after planting in Kentucky bluegrass sod using a single spray-seed Operation. . . . . . . . . . 44 7. The effect of paraquat applications and four management systems on dry weight yields of 'Vernal' alfalfa 4 weeks after planting in Kentucky bluegrass sod . . . . . . . . . . . . . 45 8. The effect of glyphosate applications and four management systems on dry weight yields of 'Vernal' alfalfa 4 weeks after planting in Kentucky bluegrass sod. . . . . . . . . . . . 47 9. Dry weight of 'Vernal' alfalfa seedlings 4 weeks after initial contact with paraquat and glyphosate treated clippings . . . . . . . . 49 CHAPTER 4 1. Chemical characteristics of soils used in glyphosate degradation study . . . . . . . . . . 54 2. Effect of phosphate addition on degradation of 14C-glyphosate in a Collamer silty clay loam soil. . . . . . . . . . . . . . . . . . . . 62 3. Effect of phosphate additions on degradation of l4C-glyphosate in a Norfolk loamy sand 8011 O O O O O O O O O O O O I O I O O O O O I O 63 4. Effect of ferric chloride additions on degradation of l4C-glyphosate in a Spinks sandy loam soil. . . . . . . . . . . . . . . . . 65 vi 10. Effect of different ferric compounds on degradation of 14C-glyphosate in a Spinks sandy loam soil. . . . . . . . . . . . . . . . Effect of metal additions on degradation soil Effect of pH as adjusted by ammonium C-glyphosate in a Spinks sandy loam 14 carbonate additions on degradation of C- glyphosate in a Norfolk loamy sand soil. . . . Degradation rates of 14C-glyphosate in various soil mixtures with Spinks sandy loam soil. . . Effect of added glucose on degradation of 14C-glyphosate in a Spinks sandy loam soil . . Effect of organic amendments on degradation of 14 C-glyphosate in a Norfolk loamy sand soil vii Page 66 68 69 70 72 73 LIST OF FIGURES Page CHAPTER 4 l. 2. Patterns of 14C-glyphosate degradation in Spinks $011 0 O O I O I O O O O O O O O O O O O O O 58 Patterns of 14C—glyphosate degradation in several soils. . . . . . . . . . . . . . . . . . . 50 viii INTRODUCTION The discovery of the selective phytotoxicity of the phenoxy herbicides (59) ushered in an era of organic herbicide use as a means of controlling weeds. Biodegradability of organic compounds represented a major advantage over use of persistent inorganic compounds which had been used for selec- tive weed control since 1900 (50). The list of chemicals for use in postemergence applications has grown steadily since and today includes both selective and nonselective herbicides (5). Nonselective postemergence herbicides are now used in conventional preplant, minimum tillage, and chemical fallow cropping systems and include both soil and foliar applied chemicals (5). Nonselective foliar applied herbicides used in these systems include dalapon (2,2-dich- loropropionic acid), paraquat.(l,l-dimethyl-4,4-bipyridinium ion) and glyphosate [kaphosphonomethyl)glycine]. Phytotoxic residues from these applications must be removed from or inactivated within the seed environment to prevent inhibition of germination and seedling growth of succeeding crops. Dalapon residues remain mobile in the soil environment (90) and therefore planting of susceptible crops should be delayed for 4 weeks after dalapon applications (11, 61, 82). Paraguat is readily inactivated within the soil by adsorption to soil colloids (2), but plant residues can remain in sufficient amounts to inhibit growth of turfgrass seedlings in contact with the treated foliage (49). Glyphosate is also rapidly inactivated within soils (6, 80) but residues appeared to be present in sufficient quantities in a treated sod environ- ment to affect legume and grass seedling growth in contact with the treated foliage (14). A period of 3 weeks between spraying and seeding appeared necessary to insure complete dissipation of the residues. Studies (31, 71, 79, 90) with 14C-glyphosate applied to various soils showed a wide varia- tion in degradation rates. The possibility of residual characteristics in glyphosate treated sod and soils with low degradation rates is an environmental concern. The objectives of this study were (1) to determine the magnitude of inhibition on turfgrass and alfalfa seedlings by glyphosate residues in several management systems and (2) to determine why 14C-glyphosate degradation to 14C02 proceeded at a relatively slow rate in some soils and not in others. be th me 1e CHAPTER 1 LITERATURE REVIEW DISSIPATION OF GLYPHOSATE IN SOIL Introduction Numerous processes influence the activity and fate of herbicides introduced into the soil environment (66, 88). Transfer and degradation processes immediately act on the herbicide and are intimately associated with each other (88). Transfer processes include absorption and exudation by plants and animals, retention in vegetation and removal by harvest- ing, adsorption by soil particles, movement in the atmosphere as a gas, surface runoff and movement through the soil via capillary flow or in the leachate. Degradation processes include chemical decomposition, photodecomposition and biological decomposition. Chemical and Photodecomposition Photodecomposition of glyphosate has been reported to be negligible (71, 90). Other studies (71, 81, 90) have shown that chemical degradation is also a minor decomposition mechanism in soil. Evolution of 14C as 14CO2 accounted for less than 1 percent of total 14C'applied as glyphosate in a sterilized soil after 28 days (81). Microbial Decomposition Degradation by soil microorganisms is an important biological mechanism by which nonselective postemergence herbicides such as glyphosate are removed from the environment. Microbial degradation rates of herbicides are influenced by the amount of herbicide available for degradation. Adsorp- tion of herbicides to soil clay and organic matter fractions may play a major role in determining the amount of herbicide degraded. Additions of montmorillonite clay and charcoal to a loamy sand decreased the degradation rate of atrazine [2-chloro-4(ethylamino)-6-isopropylamino-gftriazine] and chlorthiamid (2,6-dichlorothiobenzamide) (63). Microbial decomposition of diquat [6,7-dihydrodipyrido (l,2-§:2',1'-g) pyrazinediium ion] was quantitatively reduced by addition of montmorillonite clay to nutrient solution cultures (86). In the same study, additions of kaolinite clay had no significant effect on the decomposition of diquat. Further 14C-diquat retained on the internal studies revealed that surfaces of montmorillonite clay in aqueous soil suspensions was not degradable by microorganisms over a one year period (88). At the end of the experiment, the herbicide was extracted in its original form. However, these same investi- gators observed that diquat and paraquat (1,1'-dimethy1-4,4'- bipyridinium ion) which was ionically adsorbed to soil organic matter was still available for microbial degradation. Soil pH can also affect availability of herbicides to microorganisms. Microbial degradation of metribuzin [4-amino- G-Egrtfbutyl-B-(methylthio)-§§-triazin-5(4H)one] in pH adjusted soils increased as soil pH increased (53). This phenomenon was attributed to increased mobility of the non- protonated form of the compound at higher pH levels. Glyphosate appears not to be available to plants grown in treated soils (6, 80). Studies with l4C-glyphosate (72, 14 80) have shown that C absorbed by both corn (Zea mays L. and soybean (Glycine max (L.) merr.) plants grown up to 8 weeks in treated soils accounted for 1 percent or less of 14C applied. A bioassay study (80) revealed that adsorption rather than microbial degradation was responsible for glyphos- ate inactivation since plants grown in autoclaved soils exhibited no herbicidal effects. Glyphosate does appear to be readily available to soil microorganisms in some soils Since degradation studies have revealed approximately fifty percent evolution of applied 14C-glyphosate as 14 CO2 occurred after approximately 1 month incubation periods (31, 71, 81). However, other soils have shown extremely low degradation rates (31, 71) even though the number of microorganisms found in these soils were compara- ble to the number found in soils with high degradation rates (71). This is strong evidence that glyphosate availability to microorganisms is affected by soil characteristics rather than microbial populations. Soil adsorbent characteristics and pH are soil factors that may affect availability of glyphosate to microorganisms. Adsorption of 14 C-glyphosate was found to be dependent on clay type and hydroxides present in adsorbent:water suspen- sions (81). A soil thin-layer chromatography study (81) revealed that glyphosate mobility was very limited but was affected by soil type and pH. Increased pH levels in soil resulted in increased mobility of glyphosate and was attributed to changes in ionization of the glyphosate molecule. A bio- assay study (80) showed that at high rates of glyphosate application, phytotoxicity increased as pH increased. How- ever, a correlation study (31) on 9 soils showed that correlation coefficients calculated between glyphosate adsorp- tion and percent clay, percent carbon, or pH were not statistically significant. Adsorption studies with glyphosate (31, 81) have shown that glyphosate is bound to soil constituents through the phosphonic acid moiety and competed with inorganic phosphate for available sorption sites. The correlation coefficients between glyphosate adsorption and unoccupied phosphate sorp- tion sites was statistically significant (31). Mobility on soil thin layer chromatography plates increased when soils with added orthophosphate were used (81). It appears that orthophosphate additions would increase availability of glyphosate to microorganisms although degradation studies (79) showed increased availability in only 1 of 2 soils examined. alsc wit} iIIVI ab di th (if. Valency of cations present in the soil system may also affect glyphosate availability. Bioassay studies (81) with wheat revealed that glyphosate phytotoxicity was inversely related to valency Of cations present on clay and organic matter surfaces. It was suggested that the strong adsorption of glyphosate to Al+++ and Fe+++-saturated sur- faces may be due to a mechanism similar to soil formation of colloidal precipitates of Al and Fe phosphates. These forms of P are known to be somewhat available to plants but are able to form unavailable crystalline phosphates (41). Sequence of microbial degradation of compounds such as glyphosate in soil is also of interest. Various forms of aminoalkylphosphonic acids are known to exist in nature (39, 40, 47, 48, 76). Studies (3, 15, 32, 70, 92) on the growthof soil bacteria on alkyl and aminoalkylphosphonic acids revealed that these compounds could be used as sole P sources. Nine of ten bacterial strains were found to be able to utilize P for growth from at least one of eight different aminoalkylphosphonic acids (32). It was suggested that the ability to catabolize the C-P bond is widespread among bacteria. Studies with an isolated enzyme from cell-free extracts of a strain of Bacillus cereus indicated that breakdown of 2-aminoethylphosphonic acid involved a transamination reaction (57). Other investigators (15) have shown that 3-aminopropylphosphonic acid and 2-(3-alanyl- amino)ethy1phosphonic acid can be degraded by the bacterium, e R‘s DO (.0 hi pII ch Pseudomonas aeruginosa, without modification of the amine groups on the two substrates. Phosphonic acids have also been known for some time to exhibit toxicological properties in some microorganisms, higher animals, and plants (26). A number of aminoalkyl- phosphonic acids were found to be toxic to silkworms and chick embryos and to be able to repress tobacco (Nicotiana tobacum L.) mosaic virus and tobacco rootlets (73). Meta- bolism studies revealed that cleavage of the carbon-phosphor- ous bond did not occur in kidney, liver, and plant tissue from these organisms. Another study (28) indicates that a methylphosphonate compound had slight auxin activity in pea (Pisum sativum L.) epicotyl tests. Disodium methylphosphonate was found to markedly depress the dry weight of the tops of rape (Brassica Napus L.) seedlings. Aminomethylphosphonic acid (AMPA) was found to be an extremely active growth retardant in barley (Hordeum vulgare L.) (4). These observa- tions indicate that cleavage of the carbon-phosphorous bond does not occur readily in these plant tissues. Ethephon [(2-chloroethyl) phosphonic acid] activity appears directly related to the release of ethylene after treatment (20, 23, 67, 85, 91). Ethylene evolution with concomittant release of phosphate and chloride ions occurred when ethephon was placed in a reaction flask with sodium hydroxide or with etiolated pea epicotyls (85). Earlier studies (60) have shown that alcohols and phenols are phosphorylated with high yield by reaction with various 2-chloroalkylphosphonic acids at 20-500C. The presence of three or four molar equivalents of cyclohexylamine or triethyl- amine is required and the products formed in addition to phosphorylated compounds are the corresponding alk-l-ene and chloride ions. These observations indicate that decomposition of ethephon in plant tissue may be nonenzymatic. Ethephon additions to soil have also resulted in ethylene release which has been useful in stimulating witchweed (Striga lutea Lour.) germination (24). The decomposition mechanism here could be either enzymatic or nonenzymatic in nature. Investigations (71, 81) have shown that degradation of glyphosate in soils is predominantly due to microbial activity. Aminomethylphosphonic acid (AMPA) was found to be the only significant metabolite when glyphosate was degraded in soil shake flasks (71). This compound then underwent rapid degradation as evidenced by measurement of 14CO2 evolution. This suggests that soil microorganisms eventually cleave the C-P bond but as the secondary step in the degradation sequence of glyphosate. Metabolism studies (72) in corn, cotton (Gossypium hirsutum L.), soybeans, and wheat (Triticum aestivum L.) also revealed that AMPA was a primary metabolite indicating that the C-P bond cleavage was also not a primary degradation step. Incorporation of labeled material into natural products suggested that eventually the C-P bond was cleaved in these plants. 10 Influence of Management Practices on Herbicide Dissipation In recent years, there has been an increased emphasis on use of nonselective and selective postemergence herbicides for weed control in cropping systems. These herbicides in- clude glyphosate, paraquat, dalapon, bentazon in addition to several new compounds currently being developed. Since mo st of these compounds are not intended to have herbicidal activity in the soil, rapid dissipation of these compounds from the soil is ecologically desirable. Procedures which acknowledge herbicide transfer and decomposition processes occurring in soil (88) may be utilized to hasten dissipation of herbicide residues present in soils (12). These procedures include correct cropping SYstems, fallowing and tillage, plowing, irrigation, chemical additions, and microbial additions. The emphasis in this re‘riew will be placed on the latter three procedures. Irrigation can be used to dissipate herbicide residues in the seed environment by increasing biological and non- biC>:l.ogical decomposition of these residues. Early studies (10 , 21, 37, 52) revealed that 2,4-D [(2,4-dichlorophenoxy) a(zetic acid] dissipated at a faster rate in soil when the moisture content was increased. A study (69) with radio- active ring-labeled atrazine showed that microbial popula- 14 tions and CO evolution were greater in soils with 2 higher moisture content. Non-biological dissipation of C-triflural in (3:3:3-trifluoro-2 , 6-dinitro-N,N—dipropyl-p— 11 toluidine) was also found to be greater in soils with higher moisture (62). However, bioassay studies (29, 56) revealed that phytotoxicity of simazine [2-chloro-4,6-bis (ethylamino)- g—triazine] increased as soil moisture level increased. This would indicate that irrigation should be used as a means of dissipation of herbicide residues prior to planting crops. Leaching by irrigation has been shown to be a useful means of herbicide dissipation in certain soils (13, 58, 68, 8 4 ). In a semi-arid region, herbicides apparently were bound and then retained in an unaltered form for long periods until Water was applied (58) . Rewetting the soil by irrigation water appeared to be a practical means of releasing and removing herbicide residues from these soils. A study (14) conducted on sod seeding of alfalfa (Medicago sativa L.) showed that the residual effect of glyphosate was less severe if a 35 day interval was allowed to occur between spraying and seeding. The rainfall received during this period was felt to be a coIl‘lzributing factor in the dissipation of the residues in the Sod environment. Additions of chemicals to soil may also represent a practical means of herbicide dissipation. Chemicals may j‘r1<211.ude adsorbents, surfactants, catalysts, microbial sub- stEli‘ates, and substances to adjust pH, absorb heat, and coIlserve soil moisture (12). Numerous studies (22, 29, 34, 75. 77, 83) have shown that organic matter content is the dominant soil factor affecting phytotoxicity reduction to 12 herbicides. In modified soil studies, addition of organic matter reduced herbicidal activity of simazine (68) , Pro- metryne [2 , 4-bis (isopropylamino) -6- (methylthio) -_s_-triazine] , fluometuron [1,l-dimethyl-3-(a,a,a—trifluoro—I_I_I_-tolyl)] urea, and trifluralin (89). In both studies, modification of the clay content had less of an effect on biological activity than altering organic matter content. Reduced biological activity was attributed to an increase in cation exchange capacity in the modified soils. Other investigators (51, 68 ) have attributed the reduction in phytotoxicity to an increase in microbial activity in soils with higher organic ma tter contents . A study (78) on pyrazon [5-amino-4-chloro-2-phenyl—3 (2351) -pyridazinone] dissipation showed that phytotoxicity was directly related to organic matter content of the soil. The per sistent level of phytotoxicity was attributed to slow release of pyrazon from the organic matter. Activated charcoal has received considerable attention as a means of inactivation of herbicide residues in soil (25). The mechanism responsible for this detoxication is envisioned as simple adsorption of the herbicide so that it is unavailable .to the crop. Investigators found that inactivation depended on the herbicide used, dosage, plant species, quantity of ac-"“::i.vated .C applied (8) and environmental conditions (35) . Exposure to alternate freezing and thawing reduced the ca:Eability of activated C to detoxify atrazine in a 13 Si sand potting medium (35). It therefore appears that long term effects of organic matter and activated C additions on herbicide activity should be considered and examined in greater detail . Other chemical additives may be added to the soil to modify adsorption, mobility, leaching and non-enzymatic degradation of herbicide residues. .Equilibrium adsorption studies (1) revealed that fluometuron adsorption decreased and prometryne increased when CaCl2 was added to the soil solution. Surfactants were found to affect depth of penetra- tion, activity and persistence of diuron [3-(3,4-dichloro- phenyl)-l,l-dimethyl urea] in soil (7). Researchers (16) found that pyridine and hydroxylamine enhanced non-enzymatic detoxification of simazine. Increased rates of chemical degradation of amitrole (3,amino-§_-triazole) were observed after addition of metallic salts (44). In other studies (46) a mixture of liquid N113 and metallic Na was useful in de 1: oxifying herbicide residues . Additions of substances to serve as microbial sub- strates may be useful in herbicide dissipation. Soils a“tended with glucose, a readily available substrate, were f0land to have greater degradation rates of chlorobromuron E 3‘ (4-bromo-3-chlorophenyl) -l-methoxy-l—methyl urea] (74) and fluometuron (9). Studies revealed that additions such as I“anure (10) and leaf mold (52) favored dissipation of 2,4-D in soils. In another study (30), soil amendments with l 14 percent alfalfa was found to increase dissipation of DDT (dichlorodiphenyl trichloroethane). Garbage compost added to soils reduced phytotoxicity of fluometuron and trifluralin (39). A study (64) on organic matter amendments of variable PG content revealed that lower C:N ratios favored increased Iraites of urea herbicide dissipation. The addition of lime or S to adjust soil pH may be yet another means of herbicide residue dissipation. An extensive study (18) on the influence of indogenous soil pH on herbicide phytotoxicity revealed that phytotoxicity was dependent on herbicide structure. Adjustment of soil pH influenced rate of detoxication of five out of eleven herbi- cides studied (17). Optimum levels for detoxication of these herbicides varied from 5.3 to 7.5. A study conducted on a Hillsdale sandy loam with pH previously adjusted revealed an increase in metribuzin phytotoxicity with in- creasing soil pH (54). Leaching and ultimately dissipation of metribuzin residues in the same soil (53) increased with j~1'lereasing soil pH. Phytotoxicity was found to be less at pH 4.5 than at 6.5 after prometryne was applied to synthetic 3011 media (65) . Investigators have shown that increases in soil temper— ature accelerate biological (10, 52, 69, 74) and nonbiological (8'7) degradation of herbicides in soils. In incubated soils, atrazine degraded 2 to 3 times faster with each 10°C increase in temperature between 15° and 35° (69) . Studies on comparison 15 of herbicide persistence within regions in the United States revealed less persistence to occur in southern states (33) . Previous mention has been made regarding the beneficial effect of soil moisture on herbicide degradation in soil. It therefore appears that materials such as black plastic which absorb heat and hold moisture in soil may be a useful means of herbicide dissipation. Inoculation of soils with specific organisms may be a useful method of herbicide dissipation. Several reviews have been written on the detoxication of herbicides by Specific microorganisms (19, 36, 43). Soil enrichment tech- niques for the proliferation of specific organisms effective in degrading herbicides (42) and other organic compounds (27) have been used by microbiologists for some time. Investiga- t(31:3 (45) were able to successfully enhance DDT degradation by inoculating soils with Aerobacter aerogenes. Summary Dissipation of glyphosate appears to be primarily by I“I'Lcrobial degradation. Procedures which increase microbial activity or glyphosate availability would appear to be useful in dissipation of residues that may be present in soils or 30d environments. Other procedures may involve transfer Processes such as leaching by irrigation or procedures to increase chemical decomposition. However, any method used to increase residue detoxiation must be proven to be effective and economical. Additional research in this area is needed. CHAPTER 2 EFFECTS OF GLYPHOSATE AND SIDURON ON TURFGRASS ESTABLISHMENT Abstract The effects of several concentrations of glyphosate [N-(phosphonomethyl)glycine] and siduron [l-(Z-methylcyclo- hexyl)3-phenylurea] on germination and seedling growth of Kentucky bluegrass (Poa pratensis L. 'Nugget' , 'Prato' , ' Fylking' , 'Park'), creeping bentgrass (Agrostis palustris Huds. 'Penncross' , 'Seaside'), and red fescue (Festuca rubra L. I'Pennlawn' , 'Wintergreen') were evaluated in petri dishes under controlled environmental conditions. A high c<>:I:1centration of glyphosate (10-4 M) reduced shoot growth in all but two cultivars. Siduron had no effect. Glyphosate or siduron applied to the soil and incorporated did not af fect turfgrass emergence or growth. When the glyphosate was sprayed over the soil and on the seed lying on the soil Surface, red fescue germination was reduced and shoot growth of Kentucky bluegrass, creeping bentgrass, and red fescue was inhibited at the high rate of glyphosate application (17.9 kg/ha). Under field conditions, none of the glyphosate t:I':eatments reduced turfgrass emergence or growth. 16 17 Introduction Glyphosate, a nonselective postemergence herbicide introduced in 1971, has excellent herbicidal activity on a diverse group of perennial grasses (1, 3). Glyphosate also 11aas been shown to be absorbed and translocated basipetally in numerous annual and perennial herbaceous plants (4). .Zaqpplications of glyphosate at proposed rates did not injure (:urops planted immediately after herbicide treatment (7). Therefore, it appears that glyphosate would be an excellent czlnemical for renovating turfgrass communities. Siduron is used to control germinating annual weed Elarasses in newly seededeentucky bluegrass, red fescue, and some varieties of creeping bentgrass (2) . The purpose of this study was to determine if treat- “flfiaunts of glyphosate and siduron affected percent germination 511145 growth of seedlings of Kentucky bluegrass, creeping bent- grass, red fescue and perennial ryegrass (Lolium perenne L. ' Pennfine') in the laboratory and the field. Materials and Methods The effects of glyphosate and siduron on percent germ- idrlation and shoot lengths of three turfgrass species were Sitrudied by germinating the seeds in petri dishes in a (lcxntrolled environment chamber. Twenty-five seeds each of 'lvugget' Kentucky bluegrass, 'Prato' Kentucky bluegrass, 'Fylking' Kentucky bluegrass, 'Park' Kentucky bluegrass, 18 'Penncross' bentgrass, 'Seaside' bentgrass, 'Pennlawn' red fescue, and 'Wintergreen"red fescue were placed on two layers of Whatman No. 2 filter papers in the petri dishes. Each turfgrass variety was treated with 10 ml per petri 6 5 4 dish of lo’ M, 10- M, and 10' -6 M glyphosate and 10 ml of 10 5 M, 10' M, and lo'4 M siduron. Additions of 10 ml of distilled water to the seeds of each of the eight turfgrass varieties served as the control. The dishes were placed in a growth chamber with a photoperiod of 14 hr, at 25:1 C and a 10-hr dark period of 20:1 C, and moistened with distilled water when necessary. Germination counts and shoot length measurements were made 12 days later for the bentgrass and red fescue varieties and 19 days later for the Kentucky blue- grass varieties. The treatments were replicated four times in a completely randomized design. A study was initiated in the growth chamber to deter- mine if soil applications of glyphosate and siduron reduced percent germination, emergence and plant heights of the same turfgrass species. Styrofoam flats 14.5 cm by 10 cm were filled with a Conover loam soil containing 2.2% organic matter with a pH of 7.0 and randomly split into three equal sections in order to accommodate one species per section. Preemergence applications were simulated by applying glyphos- ate as a spray at 0, 1.1, 2.2, 4.5 and 17.9 kg/ha and siduron at 11.2 and 22.4 kg/ha at 935 LAha volume and 2.11 kg/cm2 pressure after fifty seeds each of 'Park' Kentucky bluegrass, 'Penncross' bentgrass, and 'Pennlawn' red fescue were placed 19 on the soil surface and pressed lightly into the top 0.1 cm layer of soil. Preplant incorporated applications were simulated by applying glyphosate at 1.1, 2,2, 4.5, and 17.9 kg/ha and incorporating in the upper 0.6 cm of soil followed by placing fifty seeds of each turfgrass species 0.3 cm deep in the treated soil. All flats were subirrigated daily during the first week and then surface irrigated the second week. After 14 days, germination and emergence counts and plant height measurements were made for each turfgrass species. The treatments were replicated three times in a completely randomized design. Data presented in Tables 1 to 4 are the means of two experiments with two replications in each and in Tables 5 and 6 are the means of two experiments with three replica- tions each. A field study was initiated July 31, 1975, in Urbana, Illinois, on a Flanagan silt loam soil containing 6% organic matter with a pH of 6.6 to determine the effects of preplant incorporated and preemergence applications of glyphosate on emergence and subsequent growth of 'A-34' Kentucky bluegrass 1 Seeding rates were 1 and and 'Pennfine' perennial ryegrass. 3 kg/are for the 'A-34' and 'Pennfine', respectively. Gly- phosate was applied at 0, 1.1, 2.2, 4.5, 9.0 and 17.9 kg/ha prior to seeding, and lightly raked into the top 2.5 cm of 1This study was done by A. J. Turgeon, Asst. Prof. Dept. of Hort., Univ. of Illinois, Urbana. 20 soil, or immediately after seeding. Plots were 1.52 at2 each in.area. Irrigation was performed as needed to encourage germination. Treatments were replicated three times in a randomized complete block design. After 5 weeks, clipping yields were measured using a 2.5 cm cutting height. After- ward, the plots were moved twice weekly at 3.8 cm. Stand counts were made 8 weeks after planting from two 25 cm2 plugs taken from each replication. Results and Discussion Treatments of glyphosate and siduron caused no significant reduction in percent germination of the eight turfgrass varieties tested (Tables 1 and 2). In the case of 'Wintergreen' red fescue and 'Park' Kentucky bluegrass an increase in percent germination was observed at the low rates of glyphosate compared to the control. Shoot lengths of all four Kentucky bluegrass varieties were severely reduced when seeds were treated with a 10-4 M glyphosate solution (Table 3). Reduction in shoot lengths were also observed for 'Penn- cross' creeping bentgrass and 'Pennlawn' red fescue (Table 4). Thus, under conditions allowing absorption of glyphosate, turfgrass growth could be inhibited by the 10-4 M glyphosate treatment although germination did not appear to be affected. Preemergence application of glyphosate at 1.1, 2,2, 4.5, and 17.9 kg/ha to the soil did not significantly reduce percent germination of Kentucky bluegrass, creeping bentgrass or red fescue with the exception of the 17.9 kg/ha rate on 21 Table 1. The effect of glyphosate and siduron treatments on germination of four Kentucky bluegrass varieties grown in petri dishes. Percent germinationa Treatment 'Nugget' 'Prato' 'Fylking' ‘Park' (%) (%) (%) (%) Control 66 a 64 a 90 a 57 a 10"6 M glyphosate 72 a 66 a 82 a 72 ab 10-5 M glyphosate 68 a 65 a 79 a 81 b 10"4 M glyphosate 52 a 56 a 73 a 66 ab 10'6 M siduron 80 a 75 a 83 a 78 ab 10"5 M siduron 78 a 68 a 76 a 77 ab 10'4 M siduron 64 a 61 a 73 a 65 ab aMeans within columns with common letters are not significantly different at the 5% level by Duncan's multiple range test. Percent values were transformed to the arcsine for statistical analysis. 22 Table 2. The effect of glyphosate and siduron treatments on percent germination of bentgrass and red fescue grown in petri dishes. . . a Percent germination Treatment 'Penncross' 'Seaside' 'Pennlawn"Wintergreen creeping creeping red red bentgrass bentgrass fescue fescue (%) (%) (%) (%) Control 90 a 56 a 82 a 40 a 10"6 M glyphosate 86 a 68 a 72 a 60 b 10'5 M glyphosate 82 a 57 a 77 a 52 ab 10-4 M glyphosate 86 a 66 a 68 a 50 ab 10"6 M siduron 96 a 62 a 74 a 51 ab 10"5 M siduron 89 a 67 a 37 a 54 ab 10" M siduron 75 a 65 a 71 a 49 ab aMeans within columns with common letters are not significantly different at the 5% level by Duncan's multiple range test. Percent values were transformed to the arcsine for statistical analysis. ‘ 23 Table 3. The effect of glyphosate and siduron treatments on the shoot length of four varieties of Kentucky bluegrass grown in petri dishes. Shoot lengthsa Treatment 'Nugget' 'Prato' 'Fylking' 'Park' (mm) (mm) (mm) (mm) Control 12.6 b 11.1 bc 14.7 c 18.9 b 10’6 M glyphosate 15.9 c 9.8 b 14.8 c 19.3 b 10'5 M glyphosate 14.4 be 9.1 ab 10.5 ab 19.7 b 10'4 M glyphosate 8.6 a 7.4 a 7.9 a 15.3 a 10"6 M siduron 15.6 c 12.7 c 14.4 c 19.6 b 10’5 M siduron 15.4 bc 12.5 c' 13.1 bc 22.3 b 10"4 M siduron 15.3 be 10.7 bc 13.6 bc 22.1 b aMeans within columns with common letters are not significantly different at the 5% level by Duncan's multiple range test. 24 Table 4. The effect of glyphosate and siduron treatments on the shoot lengths of bentgrass and red fescue grown in petri dishes. Shoot lengthsa . . . Treatment Penncross' 'Seaside"Pennlawn"W1ntergreen' creep1ng creep1ng red red bentgrass bentgrass fescue fescue (mm) (mm (mm) (mu) Control) 10.9 b 7.3 a 30.0 c 17.5 ab 10'6 M glyphosate 9.6 b 9.7 a 26.7 bc 24.2 b 10'5 M glyphosate 10.2 b 10.1 a 25.3 be 22.5 ab 10"4 M glyphosate 7.9 a 7.0 a 15.5 a 13.7 a 10"6 M siduron 10.8 b 9.1 a 19.5 ab 25.8 b 10"5 M siduron 10.7 b 9.9 a 28.4 c 24.2 b 10'4 M siduron 10.4 b 8.9 a 25.0 bc 25.0 b aMeans within columns with common letters are not significantly different at the 5% level by Duncan's multiple range test. 25 red fescue (Table 5). Following the 17.9 kg/ha preemergence application where the spray in direct contact with the seed lying on the soil surface, the plant height was reduced for all three turfgrass species (Table 6). However, when gly- phosate was applied at this rate preplant and incorporated, no significant reduction in plant height of the three turf- grass species occurred. This is evidence that the glyphosate was rapidly inactivated by the soil as documented by previous reports (5, 6). Siduron did not appear to have a deleterious effect on germination or the shoot heights of the three turf- grass species grown in the controlled environment chamber (Table 6). In a field study, no significant reductions in clipping yield or stand density of 'A-34' Kentucky bluegrass or 'Pennfine' perennial ryegrass resulted from preplant incorporated or preemergence applications of glyphosate (Tables 7 and 8). However, a trend toward higher density and yield was observed with increasing rate of glyphosate. The results of this study indicate that a number of turfgrass species can be safely established following gly- phosate application even under abnormally high herbicide application rates. 26 Table 5. The effect of glyphosate and siduron applications on percent germination of three turfgrass species grown in soil in the controlled environment chamber. . . a Percent germination Treatment Kentucky Creeping Red bluegrass bentgrass fescue (%) (%) (%) Control 63 ab 66 ab 70 b Glyphosate, 1.1 kg/ha PREb 60 ab 68 ab 69 b Glyphosate, 2.2 kg/ha PRE 57 ab 68 ab 69 b Glyphosate, 4.5 kg/ha PRE 57 ab 77 b 69 b Glyphosate, 17.9 kg/ha PRE 50 a 54 a 51 a Siduron, 11.2 kg/ha PRE 63 ab 70 ab 69 b Siduron, 22.4 kg/ha PRE 67 b 72 ab 65 b Glyphosate, 1.1 kg/ha PPIc 60 ab 64 ab 61 ab Glyphosate, 2.2 kg/ha PPI 67 b 66 ab 66 b Glyphosate, 4.5 kg/ha PPI 60 ab 68 ab 69 b Glyphosate, 17.9 kg/ha PPI 58 ab 70 ab 67 b aMeans within columns with common letters are not significantly different at the 5% level by Duncan's multiple range test. Percent values were transformed to the arcsine for statistical analysis. bPRE = preemergence herbicide application. C PPI preplant incorporated herbicide application. 27 Table 6. The effect of glyphosate and siduron applications on the plant heights of three turfgrass species grown in soil in the controlled environment chamber. Plant heightsa Treatment Kentucky Creeping Red bluegrass bentgrass fescue (cm) (cm) (cm) Control 2.7 b 3.1 c 5.7 b Glyphosate, 1.1 kg/ha PREb 2.4 b 3.0 c 5.7 b Glyphosate, 2.2 kg/ha PRE 2.8 b 3.1 c 5.6 b Glyphosate, 4.5 kg/ha PRE 2.7 b 2.6 bc 5.5 b Glyphosate, 17.9 kg/ha PRE 1.5 a 1.1 a 4.1 a Siduron, 11.2 kg/ha PRE 2.6 b 2.8 bc 5.7 b Siduron, 22.4 kg/ha PRE ' 2.4 b 2.6 be 5.7 b Glyphosate, 1.1 kg/ha PPIc 2.5 b 2.8 bc 5.2 b Glyphosate, 2.2 kg/ha PPI 2.5 b 2.7 be 5.8 b Glyphosate, 4.5 kg/ha PPI 2.5 b 3.0 c 5.2 b Glyphosate, 17.9 kg/ha PPI 3.2 c 2.5 b 5.3 b aMeans within columns with common letters are not significant- ly different at the 5% level by Duncan's multiple range test. bPRE preemergence herbicide application. cPPI preplant incorporated herbicide application. 28 Table 7. Effects of preplant incorporated and preemergence applications of glyphosate on stand density of two turfgrass species 8 weeks after planting. Kentucky bluegrass Perennial ryegrass Giigiiiite PPIb PREc A PPIb2 a PREc (No. shoots/25 cm ) 0 25.7 26.5 a 19.8 24.3 1.1 25.3 31.8 a 23.2 27.2 2.2 33.7 29.0 a 27.8 22.2 4.5 17.5 28.8 a 15.5 34.8 9.0 36.5 29.0 a 28.5 23.3 17.9 30.7 20.3 a 27.3 15.2 aMeans within columns with common letters are not significant- ly different at the 5% level by Duncan's multiple range test. bPPI = preplant incorporated herbicide application. C PRE preemergence herbicide application. 29 Table 8. Effects of preplant incorporated and preemergence applications of glyphosate on clipping yield of two turfgrass species 5 weeks after planting. Kentucky bluegrass Perennial ryegrass Glyphosate b c b c (kg/ha) PPI PRE PPI 2 a PRE (g clipping dry wt/56 dm ) 0 1.29 a 1.29 a 28.25 a 27.53 a 1.1 1.62 a 1.39 a 31.75 a 24.67 a 2.2 ' 1.74 a 1.80 a 30.98 a 28.47 a 4.5 2.67 a 1.51 a 28.59 a 28.28 a 9.0 2.05 a 1.23 a 38.13 a 30.60 a 17.9 2.40 a 1.75 a 34.72 a 31.67 a aMeans within columns with common letters are not significant- ly different at the 5% level by Duncan's multiple range test. bPPI = preplant incorporated herbicide application. C PRE - preemergence herbicide application. 30 Literature Cited Baird, D. D., R. P. Upchurch, W. B. Homesley, and J. F. Franz. 1971. Introduction of a new broad spectrum post- emergence herbicide class with utility for herbaceous perennial weed control. Proc. N. Cent. Weed Contr. Conf. 26:64-68. Herbicide Handbook Weed Science Society of America. 1974. 3rd Ed. Weed Science Society of America, Champaign, Illinois. 430 pp. ' Sprankle, P. and W. F. Meggitt. 1972. Effective control of quackgrass with fall and spring applications of glyphosate. Proc. N. Cent. Weed Control Conf. 27:54. Sprankle, P., W. F. Meggitt, and D. Penner. 1975. Adsorp- tion, action and translocation of glyphosate. Weed Sci. 22:235-240. Sprankle, P., W. F. Meggitt, and D. Penner. 1975. Absorp- tion mobility and microbial degradation of glyphosate in the soil. Weed Sci. 22:229-234. Sprankle, P., W. F. Meggitt, and D. Penner. 1975. Rapid inactivation of glyphosate in the soil. Weed Sci. 22: 224-228. Upchurch, R. P. and D. D. Baird. 1972. Herbicidal action of MON 9573 as influenced by light and soil. Proc. West. Weed Sci. Soc. 25:41-45. CHAPTER 3 USE OF GLYPHOSATE IN SOD SEEDING ALFALFA ESTABLISHMENT Abstract The effect of glyphosate [N-(phosphonomethyl)glycine] on alfalfa (Medicago sativa L.) germination, emergence, and seedling growth was studied to evaluate the potential of glyphosate for weed control in alfalfa establishment in for- age renovation. Glyphosate and EPTC’(§-ethyl dipropylthiocar- bamate), a selective preemergence herbicide used in alfalfa establishment, caused no significant reduction in percent germination of either Saranac or Vernal cultivars grown in petri dishes. Glyphosate significantly reduced shoot length of both cultivars. Soil applications of glyphosate also had no effect on percent germination of either cultivar and only reduced plant height when applied to the soil at 17.9 kg/ha. Applications of glyphosate at 2.2, 4.5, and 9.0 kg/ha to a Kentucky bluegrass (Poa pratensis L.) sod in a simulated sod seeding alfalfa establishment study in the greenhouse reduced dry weight of both alfalfa cultivars. Application of gly- phosate in four management systems showed that a 3 day planting delay or water application after spraying reduced growth inhibition of alfalfa seeded in treated sod. These results 31 32 were similar to those obtained with paraquat (l,l'-dimethyl- 4,4'-bipyridinium ion) in these management systems. Kentucky bluegrass clippings treated with paraquat 3 days or less before contact with alfalfa seedlings significantly reduced seedling dry weight. Clippings treated with glyphosate did not significantly reduce alfalfa seedling dry weights. Introduction Alfalfa establishment in a renovation program can improve productivity and nutritional quality of large acreages of forage within the North Central states. This acreage includes both old hay fields and pastures where perennial grasses such as quackgrass (Agropyron repens L.) and Kentucky bluegrass have become the dominant species. Farmers-can renovate these forage stands by utilizing one of three methods of alfalfa establishment. One method is to till the land, then plant alfalfa and a cereal crop as a companion crOp. A second method is to plant alfalfa alone in cultivated soil and use an appropriate preemergence herbicide to control annual weeds. A third method is no-tillage sod seeding. This method involves applications of a nonselective herbicide to suppress prior existing perennial species followed by seeding of alfalfa directly into sod. Glyphosate has potential in forage renovation for remov- ing existing vegetation because of three key characteristics: 1) it is a non-selective herbicide and therefore will control 33 a wide spectrum of perennial species, 2) it is readily trans- located throughout these treated plants, and 3) it is rapidly inactivated within most soils. Recent reports (3, 4) have shown that glyphosate Sprayed at 4.5 kg/ha, did not inhibit germination of Kentucky bluegrass, red fescue (Festuca rubra L.), tall fescue (F. arundinacea Schreb.) and creeping bentgrass (Agrostis palus- tris Huds.) when applied to the soil before seeding or when applied directly over the seeds. Red clover (Trifolium pretense L.) germination was inhibited when glyphosate was applied at 2.0 kg/ha with seeds covered by soil at the time of treatment (5). Germination of annual ryegrass (Lolium multiflorum Lam.) seed, directly treated or covered by soil, was reduced when glyphosate was applied at 3.0 kg/ha and 4.0 kg/ha, respectively (5). In petri dish studies (2), glyphosate applied at 2.2 kg/ha and in direct contact with seeds, did not inhibit germination of prickly sida (Sida spinosa L.), velvet leaf (Abutilon theophrasti Medic.), barnyard grass [Echinochloa crusgalli (L.) Beauv.], and johnsongrass [Sorghum halepeuse (L.) Pers.]. Glyphosate, at 2.2 kg/ha, caused a 1.6 to 3-fold stimulation in red root pigweed (Amaranthus retroflexus L.) germination. Radicle growth inhibitions of the germination was observed with all five weed species. Other petri dish studies (4) showed that shoot lengths of Kentucky bluegrass, creeping bentgrass, and red fescue were reduced when seeds were treated with 10"4 M glyphosate solution. 34 The purpose of this study was to determine whether glyphosate could be safely used for establishing alfalfa by assaying glyphosate effects on percent germination and seed- lings growth under several management systems.) Furthermore, it was of interest to compare the effects of glyphosate with those of EPTC and paraquat. Materials and Methods The effect of glyphosate on percent germination and shoot lengths of two alfalfa cultivars was studied by germ- inating the seeds in petri dishes in a growth chamber within a day temperature of 24°C and night temperature of 14°C in a 16 hr photoperiod regime. Fifty seeds each of Saranac or Vernal alfalfa were placed on two layers on Whatman No. 2 filter papers in the petri dishes containing 10 ml of 0, '6 5 M. and 10"4 M glyphosate or 10 m1 0f 10-6 M' M, and 10"4 M EPTC. EPTC was included in this study 10 M, 10' 10'5 because it is recommended for use in preemergence weed control in alfalfa (6). Germination counts and shoot length measure- ments were made after 7 days. The treatments were replicated three times in a completely randomized design. A study was initiated in the growth chamber to deter- mine if applications of glyphosate to the soil reduced percent emergence and plant heights of the two alfalfa cultivars. Styrofoam flats 14.5 cm by 10 cm were filled with a Spinks sandy loam containing 3.9% organic matter with a pH of 6.1 and 35 split into two equal sections to accommodate one cultivar per section. Preplant incorporated applications were simu- lated by applying glyphosate at 0, 2.2, 4.5, 9.0, and 17.9 kg/ha a.i. at 935 L/ha volume and 2.11 kg/cm2 pressure and incorporating in the upper 0.6 cm of soil followed by placing 50 seeds 0.3 cm deep in the treated soil. Preemergence applications were simulated by applying glyphosate at 0, 2.2, 4.5, 9.0 and 17.9 kg/ha after 50 seeds of each cultivar were placed 0.3 cm deep in soil. All flats were subirrigated daily throughout the study. After 14 days, emergence counts and plant height measurements were made for each alfalfa cultivar. The treatments were replicated three times in a completely randomized design. The effects of glyphosate applications on alfalfa in a simulated sod seeding system were studied in the greenhouse. Sixty seeds of Saranac and Vernal were planted in Kentucky bluegrass sod grown in flats 35 cm by 51.4 cm and split equally to accommodate one cultivar per section. Glyphosate then was applied at 0, 2.2, 4.5, and 9.0 kg/ha at 234 L/ha volume and 1.83 kg/cm2 pressure. Alfalfa was also planted in nontreated soil which then served as a bare soil control. After 4 weeks, the alfalfa was harvested, surviving plants counted and dry weights measured. The treatments were repli- cated three times in a completely randomized design. A second study was initiated to determine the inter- action between glyphosate applications and four management systems on subsequent growth of alfalfa. Fifty seeds of 36 Vernal alfalfa were planted in a Kentucky bluegrass sod grown in flats 15 cm by 30 cm and glyphosate then applied at 0, 2.2, 4.5, and 9.0 kg/ha. Paraquat was also applied at 0.6 and 1.2 kg/ha. Applications were made at the time of seeding or prior to seeding according to the management system in- volved. The first two management systems consisted of single spray-seed operations. The second management system also involved application of 2.5 cm of water 1 day after herbicide treatment. The third and fourth management systems consisted of a 3-day delay in planting after herbicide application. No water was applied for a period of 7 days in the third sys- tem whereas in the fourth system 2.5 cm of water was applied 1 day after the herbicide was applied. Alfalfa was also planted in non-treated soil which then served as a bare soil control. Four weeks after planting the alfalfa was harvested and the dry weight measured. The treatments were replicated three times in a randomized complete block design. Kentucky bluegrass clippings treated with paraquat or glyphosate were placed in contact with alfalfa seedlings to determine whether they affected subsequent growth. Alfalfa was planted in a potting soil mix, thinned to 100 plants per flat and grown for a period of 2 weeks in the greenhouse. Concurrently, Kentucky bluegrass sod in flats was allowed to grow to a height of 15 cm and then was sprayed with paraquat at 0 or 0.6 kg/ha or glyphosate at 2.2 kg/ha 0, 3, 7, and 10 days prior to clipping and placed in contact with the alfalfa 37 seedlings. The flats were subirrigated and also surface irrigated daily with a 10 second fine mist spray. The alfal- fa plants were harvested 4 weeks later and the dry weight measured. The treatments were replicated three times in a completely randomized design. All data presented are the means of two experiments. Results and Discussion Glyphosate and EPTC treatments caused no significant reduction in percent germination of either cultivar examined in petri dish studies (Table 1). Glyphosate treatments at -6 M concentration did not have a detrimental effect on 10 shoot length of Saranac cultivar but did reduce shoot length of the Vernal cultivar (Table 2). Increasing concentrations of glyphosate further inhibited shoot length of both culti-' vars. It therefore appears that under conditions allowing absorption of glyphosate, alfalfa seedling growth can be inhibited. Soil applications of glyphosate had no detrimental effect on percent emergence of either cultivar even when glyphosate was applied eight times the normal use rate of 2.2 kg/ha (Table 3). Preplant incorporated applications of glyphosate did not significantly reduce plant height (Table 4) of either cultivar examined. Preemergence applications of glyphosate also did not significantly reduce plant heights of either cultivar when applied at 2.2, 4.5, and 9.0 kg/ha. 38 Table l. The effect of glyphosate and EPTC treatments on germination of two alfalfa cultivars grown in petri dishes. I 0 a Percent germination Treatment Saranac Vernal (’35) (’6) Control 86 a 96 a 10-6 M Glyphosate 86 a 92 a 10"5 M Glyphosate 90 a 93 a 10‘4 M Glyphosate 85 a 93 a 10'6 M EPTC 85 a 94 a 10'5 M EPTC 84 a 93 a 10'4 M EPTC 86 a 93 a aMeans within columns with common letters are not signifi- cantly different at the 5% level by Duncan's multiple range test. Table 2. The effect of glyphosate and EPTC treatments on the shoot length of two alfalfa cultivars grown 39 in petri dishes. Shoot lengtha Treatment Saranac Vernal (mm/plant) (mm/plant) Control 29.1 de 29.0 d 10'6 M Glyphosate 27.0 cd 25.8 c 10'5 M Glyphosate 17.4 b 17.4 b 10'4 M Glyphosate 12.9 a 12.5 a 10'6 M EPTC 25.9 c 25.9 c 10'5 M EPTC 27.7 cde 29.4 d 10‘4 M EPTC 30.2 e 27.7 cd aMeans within columns with common letters are not signifi- cantly different at the 5% level by Duncan's multiple range test. 40 Table 3. The effect of glyphosate applications on percent emergence of two alfalfa cultivars grown in soil in the controlled environment chamber. a Percent emergence (figfifia) Method Sargnac . vefig?l Control 41 a i 53 ab 2.2 PPIb 44 a 49 ab 4.5 40 a 52 ab 9.0 38 a 47 a 17.9 38 a 44 a 2.2 PREC 46 a 54 b 4.5 43 a 56 ab 9.0 43 a 46 a 17.9 45 a 49 ab aMeans within columns with common letters are not signifi- cantly different at the 5% level by Duncan's multiple range test. b PPI preplant incorporated herbicide application. A C PRE preemergence herbicide application. 41 Table 4. The effect of glyphosate applications on plant heights of two alfalfa cultivars grown in soil in the controlled environment chamber. Plant heighta (figfifia) Method Saranac Vernal (mm/plant) (mm/plant) Control 34 b 35 bc 2.2 prb 33 ab 35 be 4.5 33 ab 33 ab 9.0 33 ab 35 be 17.9 32 ab 33 ab 2.2 PREC 36 bc 34 ab 4.5 34 b 35 be 9.0 38 C 36 c 17.9 31 a 31 a aMeans within columns with common letters are not significant- ly different at the 5% level by Duncan's multiple range test. bPPI preplant incorporated herbicide application. CPRE "preemergence herbicide application. 42 Only when glyphosate was applied at eight times the normal use rate, did a significant reduction in plant height occur. It therefore appears that glyphosate applications onto bare soil would result in injury to alfalfa seedlings only when applied at high rates and allowed to remain on the soil surface. In the simulated sod-seeding study, percent survival of Saranac alfalfa grown in sod sprayed immediately after planting with glyphosate at 2.2 kg/ha was not significantly different from that grown in non-treated soil (Table 5). Percent survival of Vernal alfalfa was significantly reduced when grown in sod treated with glyphosate at 2.2 kg/ha. Increasing rates of glyphosate further reduced percent sur- vival of both cultivars. Percent survival of alfalfa planted in non-treated sod was drastically reduced due to severe competition of Kentucky bluegrass. Glyphosate applied at the normal use rate did not affect dry weight yields of Saranac but did reduce yields of Vernal (Table 6). Increas- ing rates further inhibited growth of both cultivars. It therefore appears that glyphosate residues are present in treated bluegrass sod and are not as readily inactivated as are soil applications of glyphosate. Interactions between paraquat applications and four management systems utilized in a second simulated sod-seeding study were observed (Table 7). Alfalfa growth in paraquat treated sod was reduced if a single spray—seed operation was 43 Table 5. The effect of glyphosate applications on percent survival of two alfalfa cultivars 4 weeks after planting in Kentucky bluegrass using a single spray-seed operation. Percent survivala Planting Glyphosate medium (kg/ha) Saranac Vernal (’35) (‘8) Soil 0 77 e 87 f Sod 0 15 a 12 a 2.2 74 de 71 de 4.5 67 cd 59 c 9.0 44 b 45 b aMeans with common letters are not significantly different at the 5% level by Duncan's multiple range test. 44 Table 6. The effect of glyphosate applications on dry weight yields of two alfalfa cultivars 4 weeks after planting in Kentucky bluegrass sod using a single spray-seed operation. . a Dry weight Planting Glyphosate medium (kg/ha) Saranac Vernal (mg/900 cmz) Soil 0 1697 e 2003 f Sod 0 83 a 62 a 2.2 1700 e 1627 e 4.5 1615 e 1292 e 9.0 835 c 590 b aMeans with common letters are not significantly different at the 5% level by Duncan's multiple range test. 45 Table 7. The effect of paraquat applications and four management systems on dry weight yields of 'Vernal' alfalfa 4 weeks after planting in Kentucky blue- grass sod. Dry weighta Planting Paraguat b medium (kg/ha) Mgt a Mgt b M t c Mgt d (mg/225 cm ) Soil 0 1081 cd 1117 d 1121 d 1077 cd Sod 0 272 a 513 ab 400 a 338 a 0.6 395 a 932 bed 1189 d 1003 cd 1.2 620 abc 945 bed 1136 d 1056 cd aMeans with common letters are not significantly different at the 5% level by Duncan's multiple range test. b Mgt a = planted, sprayed at time of planting. Mgt b = planted, aprayed at time of planting, 2.5 cm water 1 day later. Mgt c = sprayed, planted 3 days later, no water for 7 days. Mgt d = sprayed, 2.5 cm water 1 day later, planted 3 days later. 46 used.(Management System A). The dry weight yield of alfalfa grown in treated sod was not significantly different from the yield of alfalfa grown in non-treated soil if application of water simulating rainfall was made shortly after spraying (Management System B) or a 3-day lapse allowed to occur 4between spraying and seeding (Management Systems C and D). Alfalfa growth in non-treated sod was again severely reduced due to severe competition of Kentucky bluegrass. Similar interactions were observed in the case of glyphosate applications (Table 8). Glyphosate applied at a normal use rate reduced alfalfa growth when applied at the time of planting (Management System A). Yields were not significantly reduced if application of water (Management System B) was made or a 3-day lapse allowed to occur between spraying and seeding (Management Systems C and D). Glyphosate applications at twice the normal use rate reduced growth when an application of water was made, but yields were not sig- nificantly reduced if a 3-day lapse was allowed to occur. Glyphosate applied at four times the normal use rate reduced growth when a 3-day lapse occurred (Management System C). At this rate, both an application of water and a 3-day lapse (Management System D) was necessary to reduce the inhibition in alfalfa growth. The results presented agree with those of Campbell (1) when he surface seeded alfalfa on glyphosate treated sod. However, in his studies an excess of 14 days was required before the danger of inhibiting alfalfa A 47 Table 8.- The effect of glyphosate application and four management systems on dry weight yields of 'Vernal' alfalfa 4 weeks after planting in Kentucky blue- grass sod. Dry weighta Planting Glyphosate medium (kg/ha) Mgt a Mgt b Mgt c Mgt db (mg/225 cmz) Soil . 0 1081 cde‘ 1117 de 1121 de 1077 c-e Sod 0 272 a 513 ab 400 ab 338 ab 2.2 424 ab 644 a-d 1300 e 1157 de 4.5 343 ab 586 abc 869 a-e 1209 e 9.0 318 a 549 ab 564 abc 783 a-e aMeans with common letters are not significantly different the 5% level by Duncan's multiple range test. b Mgt a = planted, sprayed at time of planting. Mgt b = planted, sprayed at time of planting, 2.5 cm water 1 day later. Mgt c = sprayed, planted 3 days later, no water for 7 days. Mgt d = sprayed, 2.5 cm water 1 day later, planted 3 days later. 48 establishment was no longer a problem. This would indicate that placing the seed in the soil increases the margin of safety. Dry weight of alfalfa seedlings 4 weeks after initial contact with paraquat treated Kentucky bluegrass clippings were significantly reduced when paraquat was applied 3 days or less prior to contact (Table 9). Clippings treated with glyphosate and placed in contact with the alfalfa seedlings did not significantly reduce seedling dry weight even when clippings were sprayed immediately prior to contact. Studies by Klingman and Murray (3) showed that clippings treated with glyphosate did not inhibit Kentucky bluegrass, red fescue, and tall fescue germination. Clippings treated with paraquat were found to inhibit germination of the same three species. Results presented here showed that paraquat or gly- phosate residues remaining in the treated sod environment may inhibit alfalfa growth when applied at time of planting. Initial injury may result when emerging seedlings absorb residues that are present in the litter at the soil surface. Further growth inhibition may occur when treated foliage collapses onto emerged seedlings. However, results from the simulated sod seedling studies indicate that a 3-day lapse between spraying and seeding operations will allow success- ful use of glyphosate in alfalfa establishment. 49 Table 9. Dry weight of 'Vernal' alfalfa seedlings 4 weeks after initial contact with paraquat and glyphosate treated clippings. Time prior Dry weighta Application to(§:;:act (g/lOO plants) Control ' - 1.83 b Paraquat 0.6 kg/ha 0 1.35 a 3 1.38 a 7 1.48 ab 10 1.56 ab Glyphosate 2.2 kg/ha 0 1.47 ab 3 1.72 ab 7 1.49 ab 10 1.50 ab aMeans followed by common letters are not significantly different at the 5% level by Duncan's multiple range test. 50 Literature Cited Campbell, M. H. 1974. Effects of glyphosate on the germination and establishment of surface-sown pasture species. Aust. J. Exp. Anim. Husb. 14:557-560. Egley, G. H. and R. D. Williams. 1977. Effects of glyphosate and paraquat upon weed seed germination. Weed Sci. Soc. Amer. Abstr. p. 81. Klingman, D. L. and J. J. Murray. 1976. Germination of seeds of turfgrasses as affected by glyphosate and para— quat. Weed Sci. 24:191-193. Moshier, L., A. J. Turgeon, and D. Penner. 1976. Effects of glyphosate and siduron on turfgrass establish- ment. Weed Sci. 24:445-448. Segura, J., S. W. Bingham, and C. L. Foy. 1977. Susceptibility of some pasture species to glyphosate. weed Sci. Soc. Amer. Abstr. p. 55. , . Weed Science Society of America. 1974. Herbicide Hand- book, 3rd ed., Champaign, Illinois. 430 pp. CHAPTER 4 FACTORS INFLUENCING MICROBIAL DEGRADATION OF 14 14 C-GLYPHOSATE TO CO2 IN SOIL Abstract Glyphosate degradation occurred primarily by micro- organisms and varied among the three soils examined. Phosphate additions stimulated glyphosate degradation to a limited extent in the Collamer silt loam but not in the Norfolk loamy sand. Additions of Fe+++ and Al+++ ions reduced degradation in the Spinks sandy loam. It is postulated that formation of colloi- dal Fe and A1 precipitates in modified soils with concomitant adsorption of glyphosation is responsible for decreased availability of glyphosate to microorganisms. Mn++ additions were found to increase degradation. Spinks soil and carbon substrate amendments failed to increase degradation in both soils with low degradation rates. Introduction Herbicides, applied to the soil, may be subject to microbial, chemical and photodecomposition. Photodecomposi- tion of glyphosate has been reported to be negligible (7, 12). Chemical degradation has also been found to be of minor importance for glyphosate applied to soil (7, 10, 12). After 51 52 28 days, evolution of 14CO2 accounted for l and 17 percent of total 14 14 C applied as C-glyphosate in a sterilized and non-sterilized soil respectively (10) providing evidence that microbial degradation is the predominant mechanism for glyphosate decomposition in soils. Past studies (1, 7, 10, 12) have shown a wide variation l4C-glyphosate in the soils. This in the degradation rate of variation may be due either to the level of microbial activity present in these soils or to availability of the herbicide in the soil environment to microbial degradation. Factors influ- encing microbial activity such as pH, moisture, temperature, organic substrate levels and nutrient levels are known to affect herbicide degradation in soil (3). Attempts have also been made to increase herbicide degradation by soil inocula- tion techniques (2, 4). Few investigations have dealt with the effect of herbicide availability on microbial degradation. On a Hillsdale sandy loam soil with a previously established pH range, microbial degradation of metribuzin increased with increasing pH (5). This increase was attributed to decreased binding of the nonprotonated form of the compound at higher soil pH. The objectives of this study were to determine the influence of various factors on degradation of glyphosate to C02 in three different soil types. 53 Materials and Methods Soils studied included a Spinks sandy loam, a Collamer silt loam from New York State, and a Norfolk loamy sand from Virginia. Chemical characteristics of the soils studied as determined by the soil testing laboratory at Michigan State University, are presented in Table 1. Glyphosate degradation in various soils collected in summer of 1976 was investigated according to procedures adapted from Tiedje and Mason (11). A 25 gram portion of each soil was placed in 125 ml erlen- meyer flasks and received 5 ml of 6.3 x 10-4 M NH4N03. A l-ml portion of solution containing 0.005 uCi of l4C-methyl- labeled glyphosate (spec. act.=1.0 mCi/mmole) was added to each flask. Three days prior to glyphosate application the 25 grams of the Spinks sandy loam soil also received 3 ml of propylene oxide which was allowed to volatilize. This treatment served as the sterilized soil control. A 2-m1 beaker containing 1 M KOH was suspended in each flask to absorb the 14C02. The flasks were sealed and placed at 25 t 2°C under room light and sampled 1, 2, 4, 8, l6, and 32 days later. At sampling time, the contents of the 2-ml beaker were placed in a vial containing 15 ml of a solution consisting of 1:1 (v/v) CAB-O-SIL1 solution: 60 g naphthalene, 4 g PPO, 0.2 g POPOP, 100 ml methanol, 20 ml ethylene glycol made up to a liter with 2:1,4-dioxane. The samples were radioassayed by liquid scintillation spectrometry. 1Product of Research Products Intl. Corp. Elk Grove Village, ILL 60007. 54 Table 1. Chemical characteristics of soils used in glyphosate degradation study. Organic . Phosphate SOil type pH matger (kg/ha) Collamer silt loam 5.6 3.7 68 Norfolk sand 4.4 1.2 72 Spinks sandy loam 6.6 3.8 302 55 The effect of P addition on glyphosate degradation was examined by incorporating CaHPO4 into the low P Collamer and Norfolk soils at concentrations of 0, 200, 400, and 800 ppm and 0, 200, and 400 ppm oven dry soil basis respectively. After thorough mixing, 25 9 samples of each of the amended soils were incubated for l, 2, 4, 8, 16, and 32 days for the degradation study. Studies on cation influence on glyphosate degradation included additions of FeCl -6H 0, Fe O 3 2 4’2 2 2 3' A1C13, CaClz, MnC12°4H20 and NaCl. Ferric chloride was incor- O, FeNH4(SO '12H porated into the Spinks sandy loam soil at a concentration of 0, 200, 400 Fe+++ ppm. Other ferric compounds and metal salts were incorporated to obtain a concentration of 200 ppm of the cation in soil for comparison. Incubation periods for the ferric chloride study were 1, 2, 4, 8, 16, and 32 days and for the Fe+++ compound and cation comparison studies were 1, 2, and 8 days. The effect of soil pH on microbial degradation of gly- phosate was examined by addition of 0, 0.1, 1.0, and 10.0 milliequivalents of ammonium carbonate to the low pH Norfolk soil. Glyphosate then was added to the soil and degradation monitored for incubation periods of l, 2, 8 and 16 days. To ascertain whether the low level of 14C-glyphosate conversion to 14 CO2 in the Collamer and Norfolk soils was related to lack of microbial content, presence of microbial inhibitors, or insufficient substrate to support microbial population, a series of experiments with soil amendments was 56 initiated. Five grams of non-sterilized Spinks sandy loam soil, known to be active in degrading glyphosate, was added to 20 g of sterilized Spinks, non-sterilized Collamer and Norfolk soils. Five m1 of a combined solution of 6.3 x 10-4 M NH4NO3 and 0.1% glucose was added to a Spinks sandy loam soil prior to addition of glyphosate. Five m1 of 0.5% glucose, sarcosine, and glycine was added to a Norfolk soil as a means of comparing different microbial substrates. Alfalfa meal was also incorporated in soil to obtain a concen- tration equal to that of other organic substrates. The Norfolk soil was selected because of the low organic matter content in this soil (Table l). Incubation periods were 1, 2, 8, and 32 days for the Spinks soil inoculum and glucose amendment studies and 1, 2, and 8 days for the substrate comparison study. All data presented are the means of two experiments with three replications per experiment. Results and Discussion Glyphosate degradation occurred readily in the non- sterilized Spinks sandy loam soil and was severely reduced by soil sterilization (Figure 1). This evidence confirms that microbial degradation is the predominant mechanism in soil degradation of glyphosate. Evolution of 14CO2 accounted for 14 3.0 and 9.5 percent of C applied as glyphosate in the Norfolk and Collamer soils, respectively, whereas in the Spinks soil, 14 14 CO2 evolution accounted for 40 percent of applied C after 32 days (Figure 2). 57 Figure 1. Patterns of 14C-glyphosate degradation in Spinks soil. 58 mm ml om mm mm am mm Am>=nz o.m m.v m.v ©m>uwmno om xaomuoz H.o w.o I H.H n.H mocmummmwo «.ma o.HH m.o N.v nwumsflumm m.mH m.oa v.n m.m ©w>ummno om umEmaaoo N.N v.m v.m m.m OOGOHOMMHD 5.0H m.n m.v ~.m omumsflumm mxcflmm m.~a h.ma >.NH o.m nm>ummno om omufiawuwum Among".H mm ©m>Ho>m 03 My AEmv mm m m H 0089 HHOM mxcfldm Huom omocmam Amhmov cowmmomnmmo mafia coaumnsocH mxcflmm spas monsuxflfi HHOM msoflum> ca mummonmwamIO muouxwe HHom .Hflom Emoa wncmm mo mmumu cowumomummo .m manna 71 higher than estimated degradation rates after incubation periods of 1 and 2 days in the Collamer-Spinks soil mixture but then were essentially equal to each other after 8 and 32 days. The Spinks soil may have inoculated the Collamer soil initially resulting in an increase of degradation of available glyphosate but this stimulatory effect was only temporary. Glyphosate residues after longer incubation periods appeared to become inactivated in the soil and had become unavailable to microorganisms. Observed degradation rates were lower in the Norfolk-Spinks soil mixture than estimated degradation rates for all incubation periods. This indicated that the Norfolk soil was able to adsorb glyphosate in a manner which made it unavailable to the active microbial populations in the Spinks soil inoculum. Perhaps the relatively high aluminum levels in the Norfolk played a role in the biologi- cal inactivation of glyphosate in these two soils. iGlucose additions to the Spinks soil failed to increase rates of glyphosate degradation (Table 9). Glucose, a readily available microbial substrate, apparently allowed rapid growth of microorganisms that were incapable of degrading glyphosate. Various organic amendments, in addition to glucose, did not increase glyphosate degradation in the Norfolk loamy sand soil (Table 10). This soil had a relatively low organic matter content thus, conceivably organic amendments would stimulate microbial activity. The presence of sarcosine and glycine but not alfalfa meal in the soil environment inhibited gly- phosate degradation after an 8 day incubation period. This 72 Table 9. Effect of added glucose on degradation of l4C-glyphosate in a Spinks sandy loam soil. Incubation time Soil (days) medium 1 2 8 32 (% l4C evolved as 14C02)b Spinks soil 12.5 b 18.0 c 31.5 e 39.0 f Spinks soil 9.6 a 16.3 c 26.8 d 32.3 e + glucose aConcentration of glucose in soil equal to 200 ppmw (moist soil basis). bMeans followed by common letters are not significantly different at the 5% level by Duncan's multiple range test. 73 Table 10. Effect of organic amendments on degradation of 14C-glyphosate in a Norfolk loamy sand soil. Incubation time Organic (days) Amendment 1 2 8- (% l4C evolved as 14C02) Control 0.6 ab 1.1 de ' 1.8 h Glucose 0.6 ab 1.0 cd 1.9 h Sarcosine 0.5 a 0.9 cd 1.4 g Glycine 0.5 a 0.9 cd 1.3 fg Alfalfa 0.7 b 1.2 ef 1.9 h aAdded as 200 ppmw oven dry soil basis. b Means followed by common letters are not significantly different at the 5% level by Duncan's multiple range test. 74 was unexpected since both of these compounds are similar in structure to glyphosate. In summary, microbial degradation rates of glyphosate varied with soil type. Addition of phosphate stimulated glyphosate degradation in only one of two soils examined. Fe+++ and A1+++ additions to the Spinks soil drastically decreased glyphosate degradation indicating that formation of colloidal Fe and A1 precipitates may play a role in gly- phosate binding. The presence of additional Ca++ and Na+ ions had essentially no effect whereas Mn++ stimulated degradation. Spinks soil and organic carbon amendments failed to increase glyphosate degradation in soils with low degradation rates. 10. 11. 12. 75 Literature Cited Hance, R. J. 1976. Adsorption of glyphosate by soils. Pestic. Sci. 7:363-366. Kaufman, D. D. 1964. Microbial degradation of 2,2-dich- loropropionic acid in five soils. Can. J. Microbiol. 10:843-852. Kaufman, D. D. 1974. Degradation of pesticides by soil microorganisms. p. 133-202. In W. D. Guenzi (ed.) Pesticides in soil and water. Soil Sci. Soc. Amer., Madison, Wisconsin. Kaufman, D. D., J. R. Plimmer, P. C. Kearney, J. Blake and F. S. Guardia. 1968. Chemical versus microbial decomposition of amitrole in soil. Weed Sci. 16:266-272. Ladlie, J. S., W. F. Meggitt and D. Penner. 1976. Effect of soil pH on microbial degradation, adsorption, and mobility of metribuzin. Weed Sci. 24:477-481. Lindsay, W. L. 1972. Inorganic phase equilibria of micronutrients in soils. p. 43-46. In J. J. Mortvelt (ed.) Micronutrients in agriculture. Soil Sci. Soc. Amer., Madison, Wisconsin. Rueppel, M. L., J. T. Marvel and B. B. Brightwell. 1975. The metabolism and dissipation of N-phosphonomethylglycine in soil and water. Amer. Chem. Soc. Abstract No. 26. Sillen, L. G. and A. E. Martell. 1964. 2nd ed. Sta- bility constants of metal-ion complexes. Spec. Pub. No. 17, The Chemical Society, London. Sprankle, P. L. 1974. Factors affecting the herbicidal activity of N—(phosphonomethyl)glycine ( lyphosate) on quackgrass [Agropyron repens (L.) Beauv. and in the soil. p. 87-89. Ph.D. dissertation, Michigan State University, East Lansing, Michigan. Sprankle, P. L., W. F. Meggitt and D. Penner. 1976. Adsorption, mobility and microbial degradation of gly- phosate in soil. Weed Sci. 23:229-234. Tiedje, J. M. and B. B. Mason. 1974. Biodegradation of nitrilo-triacetate (NTA) in soils. Proc. Soil Sci. Soc. Amer. 38:278-283. Weed Science Society of America. 1974. Herbicide Hand- book, 3rd ed., p. 201-204. WSSA, Champaign, Illinois. CHAPTER 5 SUMMARY AND CONCLUSIONS Laboratory, greenhouse and field studies were initiated to determine the extent glyphosate residues affec- ted turfgrass and alfalfa seedlings grown in several management systems and to determine the extent that various soil amendments affected glyphosate degradation. Germination of four Kentucky bluegrass, two creep- ing bentgrass, and two red fescue cultivars was not affected when these turfgrasses were placed in contact with solution of glyphosate in petri dishes. A high concentration of glyphosate (10-4M) did reduce shoot lengths of all three turfgrass species. Germination and shoot length of the three turfgrass cultivars were not affected when exposed to siduron. Soil applications of glyphosate had no effect on seedling growth of the turfgrasses unless glyphosate was applied at very high rates (17.9 kg/ha) and allowed to remain on the soil surface in contact with exposed seeds. Under field conditions, none of the glyphosate treatments reduced turfgrass emergence or growth. Germination of two alfalfa cultivars, Vernal and Saranac, also was not affected when placed in direct contact with glyphOsate solutions in petri dishes. Glyphosate did reduce shoot lengths of Vernal to a greater extent than 76 77 Saranac. Soil applications of glyphosate had no effect on percent emergence of either cultivar. Reduction in plant height occurred only when glyphosate was surface applied at 17.9 kg/ha. Glyphosate applied to a Kentucky bluegrass sod at 2.2, 4.5, and 9.0 kg/ha in a alfalfa establishment study utilizing a simulated sod seeding procedure reduced dry weight of both cultivars. Examination of various management systems revealed that a 3 day planting delay or water appli- cation after spraying reduced growth inhibition of alfalfa in treated sod. Degradation studies showed that glyphosate is degraded primarily by microorganisms and at variable rates in differ- ent soils. Phosphate additions stimulated degradation to a limited extent in the Collamer silt loam but not in the + + and A1++ to the Norfolk loamy sand. Additions of Fe++ Spinks soil drastically inhibited degradation, indicating that formation of colloidal Fe and Al precipitates may play a role in glyphosate binding. The presence of additional Ca++ and Na+ ions had essentially no effect whereas Mn++ stimulated degradation. Adding Spinks soil and various car- bon substrates as amendments failed to increase glyphosate degradation in soils with low degradation rates. LIST OF REFERENCES 10. 11. LI ST OF REFERENCES Abernathy, J. R. and J. M. Davidson. 1971. Effect of calcium chloride on prometryne and fluometuron adsorption in soil. Weed Sci. 19:517-521. Akhavein, A. A. and D. L. Linscott. 1968. The dipyridy- lium herbicides, paraquat and diquat. Residue Rev. 23: 98-145. Alam, A. V. and S. H. Bishop. 1969. Growth of Esherichia coli on some organophosphonic acids. Can. J. Microbiol. 15:1043-1046. 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