Degradation of Benthiocarb in Soils As Affected by Soil Conditions*

J. Pesticide Sci. 2, 7-16 (1977)

Degradation of Benthiocarb in Soils as Affected by Soil Conditions*

Yasuo NAKAMURA, ** Kanji ISHIKAWA** and Shozo KUWATSUKA Laboratory of Soil Science, Faculty of Agriculture, Nagoya University, Chikusa-ku, Nagoya 464, Japan (Received May 22, 1976)

Some factors affecting the degradation of 14C-benthiocarb (S-4-chlorobenzylN, N- diethylthiocarbamate) labelled at the benzene-ring in soils were studied. The degradation rates of 14C-benthiocarb in three different soils under upland, oxidative flooded and reductive flooded conditions were compared. 14C-Benthiocarb wasrapidly degraded under oxidative conditions, but slowly under reductive conditions. Very small differences in the degradation rates were observed among different soils. Under oxidative conditions 14C-carbon dioxide was liberated remarkably with the degradation of 14C-benthiocarb. The degradation was remarkably retarded by sterilizing the soils. The repeated application of benthiocarb, or its incorporation into the soil with simetryne, CNP or propanil had no significant effect on the degradation rate.

Benthiocarb (Saturn(R),S-4-chlorobenzyl N, N-diethylthiolcarbamate), alone or in combing MATERIALS AND METHODS tion with simetryne, prometryne and CNP, is 1. Chemicals extensively used to control weeds mainly in 14C-Benthiocarb labelled at the benzene-ring paddy fields. was used as in the previous study. U The In the previous paper, ' the persistence of specific activity was 3. 21 mCi/mmole and the benthiocarb and its degradation products in radioactive purity was more than 99%. Non- a soil were reported, and its degradation path- radioactive pure chemicals were also described ways were also proposed. The behavior of the in the previous reports. 1'2) degradation products suggested that benthiocarb itself was comparably persistent but the 2. Soil Samples degradation products were rapidly degraded in Anj o soil, a mineral soil with kaolin clay subsequent steps. It was also reported that mineral, was obtained from paddy fields of benthiocarb was degraded in the soil more Paddy Field Experimental Farm, Aichiken rapidly under upland, than under flooded, Agricultural Research Center in Anj o, Aichi conditions. Prefecture; Nagano soil, a mineral soil with In the present paper, the degradation of montmorillonite clay, from Nagano Pref ectural benthiocarb in soils in laboratory was studied Agricultural Experiment Station in Nagano, as affected by soil properties, moisture content Nagano Prefecture; and Tochigi soil, a humic or redoxy conditions, soil sterilization, repeated volcanic ash soil, from a paddy field near application, and combination with other herbi- Utsunomiya, Tochigi Prefecture. The three cides. soil samples were collected from furrow slices * Studies on the Metabolism of Benthiocarb of paddy fields in winter. The soils were crushed and passed through a 2mm sieve, and (Part IV). See reference'. **Visiting researchers from Life Science Re- stored at 5C. The physicochemical properties search Institute, Kumiai Chem., Ind., Co., of these soil samples are shown in Table 1. Ltd., Kikugawa-cho, Ogasa-gun, Shizu- oka-ken, 439, Japan (Present address). 8 口本農薬学会誌第2巻第1号昭和52年2月

Table 1 Properties of soil samples used.

Oven-dry soil basis.

3. Soil Conditioning (Pre-incubation) the glass rod was finally rinsed into the flask (A) Upland conditions : Fifty grams (oven- with a small amount of water. The soil under dried weight basis) of Anjo or Nagano soil oxidative flooded conditions was mixed well by sample, or 25 g of Tochigi soil sample was shaking. The soil under reductive flooded placed in a 100 ml Erlenmeyer flask. Water conditions was mixed with a glass rod, and was added to adjust the soil moisture to 40% then by shaking gently upside down after the of the maximum water holding capacity. The opening was stopped tight. These soil samples mouth of the flask was covered with aluminum treated with 14C-benthiocarb were incubated foil perforated with several small holes. The for 10, 20, 40 and 80 days under the same soil was incubated at 30C in the dark for 2 conditions of pre-incubation as above. weeks. The moisture content was maintained at the initial level by the addition of water 5. Determination of 14C02Liberation once a week to replace the water lost by evapo- For determining 14C02 liberated from the ration. incubated soil, the soil was incubated as follows. (B) Oxidative flooded conditions: Fifty As shown in Fig. 1, a small vessel containing grams (25 g for Tochigi soil) of each soil sample 3 ml (for the upland soil) or 7 ml (for the was placed in a 500 ml Erlenmeyer flask. The oxidative flooded soil) of 12.5 N NaOH aqueous soil was flooded with water up to 0. 5 cm deep. solution was placed in the soil. 14C02liberated The soil layer of each sample was also about from the soil was largely absorbed by the NaOH 0.5 cm deep. The mouth of the flask was solution. For the soil under upland or oxida- covered with aluminum foil. The soil was tive flooded conditions about 51 Jk of C02-free incubated at 30C in the dark for 5 weeks. air was discharged into the flask containing the Water was supplied once a week to maintain soil sample was passed through toluene to the water depth during the experiment. trap volatile organic materials, and then (C) Reductive flooded conditions : Fifty through an alkali scintillator (5 g of PPO and grams (25 g for Tochigi soil) of each sample 0.3g of POPOP in one liter of ethanolamine- was placed in a big test tube (3 cm inner dia- methyl cellosolve-toluene (1:3: 6) solution3') to meter and 20 cm long). The soil was flooded trap 14C02 which had not been absorbed by with water up to 3 cm deep. The soil layer of the NaOH solution in the incubation flask. each sample was about 8 cm deep. The mouth The NaOH solution, toluene and alkali scientil- of the tube was closed with a rubber stopper lator were renewed every time after C02-free covered with paraffin paper. The soil was air was discharged into the flask. The NaOH incubated at 30C in the dark for 5 weeks. solution was diluted to 50 ml with water and 1 ml aliquot of the diluted solution was radio- 4. Application and Incubation of 14C-benthio- assayed with 10 ml of Bray's liquid scintillator carb (PPO 4 g, POPOP 0. 2 g, naphthalene 60 g, '4C-Benthiocarb (25.8 X 105 dpm) dissolved methanol 100 ml, ethyleneglycol 20 ml, and in 0. 5 ml of acetone was applied dropwise on dioxane, to make 1000 ml) using a liquid the soil surface or added to the flooded water scintillation spectrometer. The toluene and at a concentration of 10 ppm on dry soil basis. alkali solutions were also radioassayed with The soil under upland conditions was mixed the alkali scintillator. After incubation, 1 ml well with a glass rod. The soil remaining on of 10 N H2SO4was added to the soil (40 ml of Journal of Pesticide Science 2 (1), February 1977 9 water was also added to the upland soil), air was neutralized with 20 ml of 2 N NaOH, was bubbled through the mixture, and the extracted by shaking with 100 ml of methanol 14002liberated was trapped in the alkali scintil- two times, and centrifuged by the same pro- lator and determined. cedure as described above. The radioactivity For the soil under the reductive flooded of the methanol extracts was measured using conditions, a small vessel containing 3 ml of liquid scintillation spectrometer. The radio- 12.5 N NaOH was placed on the soil in the big activity in the residual soil was determined by test tube described, which was closed tightly combustion. 1) and incubated. An aliquot of the alkali solution was taken out periodically and radio- 7. Radioassay and Thin Layer Chromatography assayed. After the incubation, the soil was The method were described in the previous transferred into a 500 ml Erlenmeyer flask and paper. 1) 14002 in the soil was determined by the same procedure as above described. 8. Degradation in Sterilized Soil Twenty grams (oven-dried weight basis) of Anjo soil sample was placed in a 200 ml flask. Water was added to adjust the soil moisture to 40% of maximum water holding capacity for upland conditions or to maximum water holding capacity plus 10 ml for oxidative flooded conditions. The soils were pre-incubated at 30C in the dark for 2 weeks, then sterilized at 120C for 30 min in an antoclave. After cooling, 14C-benthiocarb (78. 4 X 104 dpm) dissolved in acetone was added to each soil Fig. 1 Apparatus for the determination of sample (10 ppm of the final concentration on liberated 14C02. dry soil basis) under sterile conditions. The mouth of each container was closed with sterile 6. Extraction and Separation of Radioactive cotton. The soil samples were incubated at Compounds 30C in the dark for designated periods. At After 14C02determination, the soil was trans- the same time, non-sterilized soil was incu- fered into a centrifuge tube, rinsing away with bated by the same procedure. acetone and a small amount of water. A total In this case the radioactive substances were volume of 100 ml acetone was added into the extracted with methanol instead of acetone tube, which was then capped tightly. The to avoid the formation of acetone-complex tube was shaken vigorously for 30 min and during the fractionation process. The incubated centrifuged at 3,000 rpm for 10 min. The soil was flooded, and extracted by shaking supernatant solution was transfered by decanta- twice with 150 ml of methanol and centrifuged tion. The soil residue was again extracted with by the same procedure described above. The 100 ml of acetone by the same procedure. The methanol-water solutions (neutral fraction) acetone solutions were combined and neutral- were pooled. The residual soil was extracted ized with aqueous NaOH solution. Acetone twice with a mixture of 20 ml 1.25 N NaOH aq. was evaporated under reduced pressure using and 150 ml methanol following the same a rotary evaporator. The residual water solu- procedures of shaking and centrifugation, and tion was extracted three times with n-hexane. the supernatant solution (basic fraction) were A small portion (0.5 ml) of each solution was collected. The residual soil was extracted with taken and radioassayed. The n-hexane solution a mixture of 20 ml 2.5 N H2SO4 and 150 ml was dried with anhydrous sodium sulfate, methanol by the same procedures, and the concentrated, and analyzed by thin layer supernatant solution (acid fraction) were col- chromatography. lected. The radioactivity of each fraction was The residual soil after acetone-extraction determined, the three fractions were combined 10 日本農薬学会誌第2巻第1号昭和52年2月

and neutralized, and methanol was evaporated under reduced pressure. The residual water RESULTS solution was saturated with sodium chloride 1. Influence of Moisture Condition and Soil and extracted with ether. The other extract was Properties on the Degradation of Benthiocarb dried with anhydrous sodium sulfate, concen- In the previous report, 1) 14C-benthiocarb in trated, and analyzed by thin layer chromato- Anj o soil was degraded much more rapidly graphy. The radioactivity in the residual soil under upland than flooded conditions. In the was determined by the combustion method. '' present study, the degradation of the chemical in 3 different soils under upland, oxidative 9. Repeated Application of Benthiocarb flooded and reductive flooded conditions, were Twenty grams (dry basis) of Anj o soil sample compared. The progress of degradation in placed in a 200 ml flask were pre-incubated Anjo, Nagano and Tochigi soils under these under oxidative flooded conditions (maximum 3 conditions are shown in Fig. 2. water holding capacity plus 10 ml of water). Under each given conditions, the rates of Non-radioactive benthiocarb 10 ppm on dry disappearence of benthiocarb in the soils tested soil basis was added and the sample was were not extremely different. The rates, how- incubated at 30C. After 2 weeks of incubation, ever, were extremely different among different 10 ppm 14C-benthiocarb (78.4 X 104 dpm) was conditions of soil. In all three soils, 14C- added and the incubation was continued. For benthiocarb was degraded most rapidly under comparison, a soil sample, which was not upland conditions, and to a lesser extent under treated with non-radioactive benthiocarb the oxidative flooded conditions. The slowest first time but was treated with 14C-benthiocarb degradation occured under reductive flooded the second time, was incubated together with conditions. The half life period was calculated the above sample. The samples were extracted to be around 20 days under upland conditions, successively with methanol, alkaline methanol about 50 days under oxidative flooded condi- and acidic methanol, as mentioned above. tions, and around 200 days under reductive flooded conditions. 10. Degradation of Benthiocarb Applied with The changes in the radioactivity levels of Other Herbicides the acetone extract, the methanol extract, the Twenty grams (dry basis) of Anj o soil sample soil residue and liberated 14C02 in Anjo soil was pre-incubated under oxidative flooded are shown in Fig. 3. Approximately the same conditions. The pre-incubated soil was treated patterns were observed for Nagano and Tochigi with 200 µg (10 ppm on dry soil basis) of 14C- soils. benthiocarb alone or in combination with Under upland conditions, the radioactivity either of 43,ag of simetryne, 2-methylthio 4,6- in the acetone extracts decreased rapidly, bis (ethylamino)-s-triazine, 171 , ag of CNP, while 14C02was liberated remarkably as time 2. 4. 6-trichloronhenvl 4'-nitro- phenyl ether, or 70 4ug of propa- (A) (B) (C) nil, 3', 4'-dichloropropionanilide. The incubation and subsequent extraction with methanol followed the same procedures described above. The ratios of benthiocarb to each of the other three herbicides are equivalent to the ratios in the herbicide formulations a- vailable commercially.

Fig. 2 Degradation of benthiocarb in three different soils under upland (A), oxidative flooded (B)and re-

ductive flooded (C)conditions.

●Anjo soil, ▲Nagano soil ■Tochigi soil. Journal of PesticideScience2 (1),February 1977 11 elapsed. About 50% of the ap- (A) (B) (c) plied radio activity was liberated as 14CO2within 80 days of incubation under the upland conditions. Similar results were also obtained under oxidative flooded conditions, where one third of the radioactivity was liberated as 14CO2 within the same period. Under reductive flooded conditions, however, the radioactivity was largely Fig. 3 Changes of the radioactivity levels of each fractions recovered in the acetone-extract, and of acetone-extractable benthiocarb under up- and only a small amount of 14CO2 land (A), oxidative flooded (B) and reductive flood- was produced. The radioactivity ed (C) conditions in Anjo soil. levels of the methanol-extract and ○acetone extracts, △MeOH extracts, ▲soil residue, the soil residue were low during ■14CO2, ●benthiocarb. the course of incubation, and were less in the soils under reductive flooded con- largely recovered in the aqueous NaOH solu- ditions than in those under upland and oxi- tion placed in the incubation flask, and only dative flooded conditions. The radioactive a negligible amount was found in the alkali volatile materials trapped in toluene were scintillator trap. The total radioactivity re- produced only in trace amounts under all covered from all the fractions of each soil three conditions. The 14CO2 liberated was sample exceeded 90% of the initial amounts.

Table 2 Timecourses of degradation products in the n-hexane extracts from Anjo, Nagano and Tochigi soils (percent of applied radioactivity).

tr.: trace amounts (less than 0.1 %). 12 日本農薬学会誌第2巻第1号昭和52年2月

The radioactivity in the acetone-extracts, 14C-benthiocarb using this process is shown in after acetone was distilled off, was largely Fig. 4. After the change in the redox state of transfered into the n-hexane extracts, as shown the soil, the rate of degradation increased in Fig. 2 and Table 2. The parent benthiocarb gradually, and after 20 days nearly equalled was the main compound present in the n- to the degradation rate in a soil sample kept hexane extracts at each step of incubation for continuously for 20 days under oxidative all soil samples and moisture conditions (Fig. flooded conditions. 2). As shown in Table 2, the degradation products were present only in small amounts. 2. Degradation in Sterilized Soil Among these products benthiocarb sulf oxide, The time courses of 14C-benthiocarb degrada- desethyl benthiocarb and 4-chlorobenzoic acid tion and 14C02 liberation in sterilized and un- were found in comparatively large amounts, sterilized soils under upland and oxidative especially after 10 to 20 days of incubation. flooded conditions are shown in Fig. 5. In The degradation products and the ratios of their amounts were not extremely different among the soils tested. Benthiocarb sulf oxide was found in larger amounts under oxidative flooded conditions while larger amounts of desethyl benthiocarb were found under reductive flooded conditions. When the soil condition changed in the course of incubation from reductive flooded to oxidative flooded conditions, the degradation of benthiocarb in the flooded soil became more rapid. Forty days after incubating 14C-bentiocarb in Anj o soil under reductive flooded conditions, the soil mixture was transf ered into a 500 ml Erlenmeyer flask to simulate oxidative flooded conditions. The sample was incubated once more. The progress of degradation of

Fig. 5 Time courses of degradation of benthiocarb and liberation of 14C02 in sterilized and unsterilized soils under upland and oxidative flooded condi- tinns -benthiocarb, …14CO2,

●sterilized soil under oxidative flooded con-

ditions, ○ unsterilized soil under oxidative flooded

conditions, ▲sterilized soil under upland conditions,

△unsterilized soil under uPland conditions.

unsterilized soils, the degradation of 14C- benthiocarb and the liberation of 14C02 were observed markedly under both oxidative con- Fig. 4 Comparison between the degradation of benthiocarb under reductive ditions. In sterilized soils, however, the flooded conditions (A) and under degradation of benthiocarb was very slow and reductive, followed by oxidative only trace amounts of 14002 were liberated, flooded conditions (B). Journal of Pesticide Science 2 (1), February 1977 13

Fig. 6 Degradation of 14C-benthiocarb ap- Fig. 7 Degradation of benthiocarb in soil plied 2 weeks after pretreatment with With simetry7ne, CNP and propanil. or without non-radioactive benthio- ●bentiocarb only, ▲with simetryne,

carb in soil. -with CNP, ▼with propanil. ●pre-treatment with non-radioactive benthiocarb, ○without non-radioactive benthiocarb. that simetryne, CNP and propanil have little effect on the degradation rate. Furthermore, 3. Effect of Repeated Treatment of Benthiocarb the degradation products of benthiocarb in- The degradation of 14C-benthiocarb in a soil cubated in combination with other herbicides sample pre-treated with unlabelled benthio- were also the same as those of benthiocarb carb was compared with the rate in another alone. No other additional products were not sample which did not contained unlabelled detected, and the accumulation of any specific benthiocarb. The results obtained are shown product was not observed. in Fig. 6. Only a slight difference was observed between the soils pre-treated with and DISCUSSION without unlabelled benthiocarb, indicating that The degradation of benthiocarb in soils was 14C-benthiocarb in the soil pre-treated with investigated as affected by various soil factors. unlabelled benthiocarb was degraded in an The degradation rates and the degradation almost equal rate compared with the soil con- products in three different soils were compared taining no unlabelled benthiocarb. under three conditions, namely, 1) upland or wet conditions, 2) aerobic flooded conditions 4. Degradation of Benthiocarb in Combination under which soil (about 0. 5 cm deep) flooded with Other Herbicides with water up to about 0.5 cm deep, and 3) Formulations of benthiocarb combined with anaerobic flooded conditions under which soil simetryne, prometryne and CNP are used more (about 8 cm deep) were flooded with water up extensively than the compound alone. A com- to 3 cm deep in a test tube closed with a rubber bination of benthiocarb and propanil is also stopper. The soil under upland conditions is in now under test in fields. Therefore, the degra- general aerobic or oxidative, except for the dation rates of benthiocarb in soil, when inner portion of soil particles which are anaero- applied in combination with these herbicides, bic or reductive. 4' It is well known that the were also compared. Fig. 7 shows the results flooded soil is rapidly transformed into the obtained. The dissipation of benthiocarb was reductive state, but the soil surface remains only in an almost equal rate when it was com- under the oxidative conditions. 5' The color of bined with other herbicides than when it was soils under the aerobic flooded condition tested applied alone. The small difference indicates showed that the soils were kept under oxidative 14 日本農薬学会誌第2巻第1号昭和52年2月

condition during the whole period tested, and to microbial activities and not to non-biological the Eh values ranged between +400 and +500 chemical reactions. The redox state of each mV. Under the anerobic flooded condition soil perhaps mainly affected the development tested, the color of soils changed to dark within of benthiocarb-degrading microorganisms, and the period of pre-incubation. The Eh values affected the chemical degradation of benthio- of Anj o, Nagano and Tochigi soils after pre- carb itself only to a small extent. It was incubation were +200 mV, +70 mV and reported that the kinds of microorganism in- -100 mV, respectively. habiting in upland and flooded soils were Benthiocarb was far rapidly degraded under quite different. 8) Under upland conditions upland and oxidative flooded conditions, than bacteria, molds, actinomycetes, and phyco- under reductive flooded conditions, and it was mycetes were active, while under flooded most rapidly degraded under upland conditions conditions under bacteria were predominant. (Fig. 2). The process of changing the condi- The kinds of bacteria present were also different tions from reductive to oxidative accelerated under both conditions. Benthiocarb is pre- the degradation (Fig. 4). It was presumed from sumably degraded mainly by molds and/or these findings that differences in the redox aerobic bacteria from the fact that it was states, not in moisture contents, of soilslargely rapidly degraded under oxidative conditions. affected the degradation of benthiocarb. The However, because usually only small amounts degradation in the 3 soilslisted in Table 1 was of molds exist in flooded soil even under oxida- compared under the 3 different conditions. The tive conditions, it is tentatively presumed that results are shownin Fig. 2. Although these aerobic bacteria are largely responsible for the soils were quite different in clay mineral, organ- degradation of benthiocarb. It was reported ic matter content and other physico-chemical that N-phenyl carbamate compounds in soils properties, the degradation of benthiocarb and were degraded by different kinds of microbes. the formation of the degradation products in IPC and CIPC9) were degraded by soil bacteria these soils followed similar patterns under each such as Pseudomonas, Flavobacterium, Agro- given condition. These results indicate that bacterium and Arthrobacter on planting media. the degradation of benthiocarb is largely af- Barban10)was degraded by a mold, Penicillium fected by the redox state, not by the physico- sp. On the other hand, little information is chemical properties, of the soil. available on the microbial degradation of thio- In practice, benthiocarb is applied on the carbamate herbicides. Isolation of micro- surface water in paddy fields or sprayed on organisms capable of degrading these herbicides surface soils in upland fields. Because benthio- and elucidation of their degradation pathways carb is adsorbed on soil particles to a large have not yet been carried out. extent, it is presumed that a large part of the In the previous papers1)the major degradation benthiocarb applied is left in the oxidative pathways of benthiocarb in a soil were reported. surface layer of soil and is degraded rapidly Three ways were found to initiate the attack on to CO2, and only the small or trace amount is benthiocarb. The first was by deethylation. leached into the lower reductive layer of soil. Dealkylation of urea herbicides was reported Yamadas) reported that 4% of the applied to be mostly due to microbial activities. 5) benthiocarb remained in the 0-5 cm layer of The second process found was the hydroxy- Konosu soil about 230 days after treatment lation of the benzene ring, which is also thought and it was not detected in the lower layer to be due to microbial activities, as shown in (5-10 cm deep) of the soil. Ishikawa, et al. 7) phenoxy herbicides5) and many other aromatic also reported that in the 0-2 cm layer of comounds.11) The rate and position of hydroxy- Kikugawa soil about 10% of the applied lation of the benzene ring varied according to amount of benthiocarb remained one month the structure of the phenoxy herbicides used after treatment, and trace in the deeper layer. as substrate compounds. Benthiocarb was In the sterilized soil the degradation process hydroxylated on the 2-position of the ring. was extremely retarded (Fig. 5). Accordingly, 2-Hydroxy benthiocarb was also detected in the degradation was presumed to be mainly due plants as ore of the major metabolites.12) The Journal of Pesticide Science 2 (1), February 1977 15 third process was by sulfoxidation of a sulfur whether the chemical was applied once or atom. The extent to which microorganisms twice. The degradation of phenoxy herbi- influenced the sulf oxidation process was not cides21) such as 2, 4-D and MCPA and also determined. The production of benthiocarb carbamate herbicides such as IPC and CIPC22) sulfoxide, however, was possibly due to micro- began after a lag period of about 2 weeks for bial activities, based on the finding that it was the first application, but immediately after the produced in mixed function oxidase systems second application. In the case of benthiocarb of mice livers, 13)moreover, it is not conceivable degradation, such a lag time was not observed that the process occurs so readily by chemical after the first application. Moreover no large reaction in the soil. Sulf oxide of butylate, a differences in the degradation rates were ob- thiocarbamate herbicide, was also reported to served between single and repeated treatments be produced in soil. 14) under the experimental conditions used. How- Furthermore, benthiocarb may be cleaved ever, the initial concentrations of benthiocarb directly in the thio ester linkage by microor- after the second treatment were different be- ganisms in the same way that CIPC was tween once and twice applications. When cleaved in the ester linkage by enzymes ob- 14C-benthiocarb was added into the soil contained from Pseudomonas stkiata. 15) taining the unlabelled benthiocarb after incu- In sterilized soil benthiocarb was degraded bation for 2 weeks, about 70% (about 7 ppm) extremely slowly. This finding suggests that of the unlabelled chemical initially applied microorganisms are largely involved in the remained in the soil. Therefore, the total degradation process from the initial step. concentration of labelled and unlabelled ben- The disappearance of radioactivity from the thiocarb in the pre-treated soil was about 17 soil system, described in the previous paper, )) ppm. In the sample containing only labelled was now confirmed to be release as 14C02. A benthiocarb, the total concentration of the notable amount of 14C02was liberated, showing chemical was 10 ppm. This may accout for that it comprised a large fraction of the de- the finding that 14C-benthiocarb was degraded gradation products. About 50 molar % of the in an almost equal rate in soils pretreated with added radioactivity under upland conditions and without unlabelled benthiocarb. and about 30% under oxidative flooded condi- Benthiocarb is applied in the single formu- tions were released as C02 during the testing lation, but it is largely used in paddy fields in period. On the contrary, other radioactive combination with simetryne and CNP, as well degradation products recovered from the sys- as propanil. The chemical did not affect the tem comprised only a small portion. From degradation of benthiocarb in soil (Fig. 7). these findings it is presumed that the benzene Whether benthiocarb was applied alone or in ring was rapidly oxidized to C02 by micro- combination with simetryne had no effect on organisms shortly after the degradation of its persistence in a paddy field. ") benthiocarb was initiated. The process of cleavage of the benzene ring ACKNOWLEDGEMENT has been studied in detail in many com- This study was largely supported by a research pounds. )s,17) Mechanism of microbial degrada- grant from Kumiai Chemical Ind. Co., Ltd., which tion of benzoic acid and its derivatives have also supplied the radioactive benthiocarb and also been studied. 10) It was reported that 4- many non-radioactive synthetic chemicals. The chlorobenzoic acid, a major intermediate of authors wish to express their thanks to the Com- benthiocarb degradation, was converted to pany for their support. Thanks are also due to Professor K. Kumada in the authors) laboratory chlorinated catechol by microbial activity. 18) for his valuable advice and encouragement during The catechol produced may be leaved in the the course of this study. The authors also ex- same way as those derived from 2,4-D, 17, 19 press their thanks to Dr. A. Nakanishi of Aichi- carbaryl, 20) etc. to produce aliphatic acids, ken Agricultural Research Center, Mr. A. Miko- which are finally oxidized to C02. shiba of Nagano Pref ectural Agricultural Station, As shown in Fig. 6, only a slight difference in and Mr. H. Kato of Utsunomiya University, for degradation of benthiocarb was observed collecting the soil samples and to the members of 16 日本農薬学会誌第2巻第1号昭和52年2月 the authors' laboratory for the analyses of pro- 6) T. Yamada: Zassokenkyu (Weed Research, perties of the soil samples. This study was also Japan) 16, 14 (1973) supported in part by research grants from the 7) K. Ishikawa, Y. Asano, Y. Nakamura & K. Ministry of Education and from the Ministry of Akasaki: ibid 21, 16 (1976) Agriculture and Forestry. 8) S. Ishizawa & K. Toyoda: Bulletin of the National Institute of Agric. Sci. (Japan) B14, 要約 203 (1964) 9) P. C. Kearney & D. D. Kauf man: Science ベンゼン環を14Cで標識したベンチオカーブを用い, 147, 740 (1965) 室内実験により, 土壌中の分解に関与する土壌要因の影 10) S. J. L. Wright & A. Forey: Soil Biol. Bio- 響について研究した. 性質の異なる3種類の土壌を, 畑 chena. 4, 207 (1972) 地状態, 酸化的湛水および還元的湛水状態とし, その中 11) D. D. Kaufman: "Pesticides in Soil & Water, " ed. by W. D. Guenzi, Soil Sci. Soc. でのべンチオカークの分解を比較すると, 土壌間ではあ America (Wisconsin), p. 133, 1974 まり差は認められず, 土壌の酸化還元状態が分解に大き 12) Y. Nakamura, K. Ishikawa & S. Kuwatsuka: く影響した. いずれの土壌でも, 酸化的(好気的)条件 Ann. Meeting of Agric. Chem. Soc. of Japan ではベンチオカーブは速かに分解し, 同時に14CO2が顕 (Abstr. ) p. 443, (1974); Agric. Biol. Chem. in press 著に生成した. 還元的(嫌気的)条件では分解が遅く, 13) J. E. Casida, E. C. Kimmel, H. Ohkawa & 14C2の生成も少なかった. また, 滅菌土壌中ではベンチ R. Ohkawa: Pesticide Biochem. Physiol. オかーブの分解はほとんど認められなかった. 中間生成 5, 1 (1975) 14) J. E. Casida, R. A. Gray & H. Tilles: Science 物相互間の比率は土壌条件の差によっても大差がなく, 184, 573 (1974) 土壌条件が異なっても同一の分解経路で分解するものと 15) P. C. Kearney: J. Agric. Food Chem. 13, 推定された. さらに, ベンチオカーブの反覆施用, シメ 561 (1965) トリン, -CNPおよびプロパニルとの混合施用はベンチ 16) D. B. Harper & ER. Blakley: Can. J. オカーブの分解に大きな影響を及ぼさなかった. Microbiol. 17, 1015 (1971) 17) J. M. Bollag, G. G. Briggs, J. E. Dawson & M. Alexander: J. Agric. Food Chem. 16, REFERENCES 829 (1968) 1) K. Ishikawa, Y. Nakamura & S. Kuwa- 18) A. Ichihara, K. Adachi, K. Hosokawa & tsuka: J. Pesticide Sci. 1, 49 (1976) Y. Takeda: J. Biol. Chem. 237, 2296 2) K. Ishikawa, I. Okuda & S. Kuwatsuka: (1962) Agric. Biol. Chem. 37, 165 (1973) 19) J. M. Tiedje, J. M. Duxbury, M. Alexander 3) K. Kametani, Y. Inoue & K. Maruyama: & J. E. Dawson: ibid. 17, 1021 (1969) Radioisotopes 16, 607 (1967) 20) Shu-Yen Liu & J. M. Bollag: ibid. 19, 487 4) AD. McLaren & G. H. Peterson: "Soil (1971) Biochemistry," ed. by A. D. McLaren & 21) L. J. Audus: "Herbicides and the Soil, " ed. G. H. Peterson, Marcel Dekker, p. 1, 1967 by E. K. Woodf ord & G. R. Sager, Black- 5) F. N. Ponnamperuma: "Advances in A- well Oxford, p. 1, 1964 gronomy," Vol. XXIV, Academic Press, New 22) D. D. Kaufman & P. C. Kearney: Applied York and London, p. 29, 1972 Microbiol. 13, 443 (1965)