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EC number: 619-020-1 | CAS number: 94361-06-5
Biodegradation in soil
Administrative data
Link to relevant study record(s)
- Endpoint:
- biodegradation in soil: simulation testing
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 12 Jun 1991 to 26 Aug 1993
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- other: BBA Part IV 4-1
- Version / remarks:
- December 1986
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Test type:
- laboratory
- Radiolabelling:
- yes
- Oxygen conditions:
- aerobic
- Soil classification:
- not specified
- Year:
- 1 991
- Soil no.:
- #1
- Soil type:
- other: sandy loam with low organic matter content
- % Clay:
- 22.7
- % Silt:
- 26.6
- % Sand:
- 50.7
- % Org. C:
- 1.13
- pH:
- 7
- CEC:
- 16.6 meq/100 g soil d.w.
- % Moisture content:
- 32
- Details on soil characteristics:
- SOIL COLLECTION AND STORAGE
- Geographic location: Flaach 2/90, Im Boden, canton Zürich, Switzerland
- Collecting date: July 17th, 1990
- Organic matter: 1.96%
- Storage conditions and length: After collection, the soil sample was shipped to the test facility and stored there in an open container under field conditions (spring to autumn) or in a glass house (winter). Immediately before use, the samples were transported by car to the testing facility.
- Soil preparation: The soil was passed through a 2 mm sieve and the actual moisture content was determined by drying a sample at about 120°C for 48 hours. On the basis of these results and the maximum water capacity, the soil was mixed with deionized water to reach about 20 % and 37 % of the maximum water capacity (for experiment A and B, respectively). The soil was equilibrated at conditions which were close to the test conditions before the start of the laboratory test. - Soil No.:
- #1
- Duration:
- 140 d
- Soil No.:
- #1
- Initial conc.:
- 0.25 mg/kg soil d.w.
- Based on:
- act. ingr.
- Parameter followed for biodegradation estimation:
- CO2 evolution
- radiochem. meas.
- Soil No.:
- #1
- Temp.:
- 21
- Humidity:
- about 40 % of the maximum water capacity
- Microbial biomass:
- See table in 'Any other information on materials and methods incl. tables'
- Details on experimental conditions:
- Experiment A: Open Test System (to determine the degradation rate)
EXPERIMENTAL DESIGN
- Test vessel: 300 mL Erlenmeyer flasks
- Soil condition: Air dried
- Soil (g/replicate): 100g
- Test solution (mL/replicate): 1.0 mL (followed by an additional quantity of deionized water to reach the final 40 % of its maximum water capacity.). After each addition, the flasks were shaken vigorously and finally stoppered with cotton plugs.
- No. of replication: 20
- Moisture maintenance method: The humidity was controlled regularly by weighing the samples and balanced by adding deionized water to reach about 40 to 50 % of the maximum water capacity.
- Continuous darkness: Yes
- Incubation temperature: 21 ± 4°C
- Sampling intervals: Immediately after application, two soil samples were analyzed. After 2, 4, 8, 12, 16 and 20 weeks, two soil samples each were analyzed.
Experiment B: Close Test System (to determine the volatiles and the material balance)
- Test vessel: 200 mL Erlenmeyer flasks
- Soil condition: Air dried
- Soil (g/replicate): 50 g
- Test solution (mL/replicate): 0.5 mL (followed by an additional quantity of water to reach the final 40 % of its maximum water holding capacity. After each addition, the flasks were shaken vigorously.
- No. of replication: 20
- Continuous darkness: Yes
- Incubation temperature: 20 ± 1°C
- Sampling intervals: Immediately after application, two soil samples were analyzed. After 2, 4, 8, 12, 16 and 20 weeks, two soil samples each were analyzed.
- Details of traps for CO2 and organic volatile: Before disconnecting the trapping systems, the Erlenmeyer flasks were flushed with nitrogen and by occasional shaking for about 15 minutes in order to evaporate the carbon dioxide and volatile compounds from the soil. In some cases, the soil of the reserve flasks were treated with 5 mL concentrated hydrochloric acid. Immediately after addition of the acid, a new trapping system was connected to the flask and the flask was flushed again with nitrogen. Then, the trapping system was removed from the flasks and the soil samples and the traps were analysed separately. The paraffin containing quartz wool was removed from the trap and extracted with 100 mL ethyl acetate.
The soda lime was removed from the trap and placed in apparatus. The adsorbed 14C-carbon dioxide was liberated with 60 mL 18 % hydrochloric acid and collected in the traps containing 15 mL absorber solutions (absorption- and scintillation cocktail). If it was not possible to analyse the soda lime immediately after sampling, the traps were wrapped into aluminium foil and stored in the refrigerator until analysis.
For the biodegradation of soil-bound radiocarbon a slightly different procedure was applied (a Vigreux column containing the absorber solution was used instead of three scintillation vials). The recovery for this procedure was found to be 94 %. The values given are corrected for the recovery.
Experiment D and C: Untreated Samples (to determine the microbial biomass)
- Soil condition: Air dried
- Soil (g/replicate): 50 g
- Test solution: No application solution was added. Appropriate volume of water was added to each flask to reach the final 40 % of its maximum water holding capacity.
- No. of replication: 3
- Incubation condition: The flasks of the experiment C were incubated as those of the experiment A, the flasks of the experiment D as those of the experiment B.
Biodegradation of the Soil-Bound Residues
- Test type: Closed
- Soil condition: 20 g (dry weight) of the pooled soil sample were mixed with 25 g (dry weight) fresh untreated Flaach soil
- Incubation temperature: 20 ± 1°C
- Humidity: 40 % of the maximum water capacity
- Sampling intervals: About one and two months after incubation, two samples were analysed as described for the soil samples of the experiment B (no Mega Bond Elut clean-up before chromatographic analysis). Further eight samples were prepared and incubated. Immediately after preparation, as well as after one and two month, two samples were analysed for the microbial biomass.
- Control group: An aliquot of the extracted soil sample was heat sterilized and incubated for one month as control sample.
- Other: The amounts of 14C-carbon dioxide, the extractable radiocarbon and the metabolic profile were determined in all samples.
- Continuous darkness: Yes - Soil No.:
- #1
- % Non extractable:
- 21.5
- % Recovery:
- 51.6
- Remarks on result:
- other: Open system, on Day 140
- Soil No.:
- #1
- % Non extractable:
- 20.8
- % CO2:
- 1.9
- % Other volatiles:
- < 0.1
- % Recovery:
- 86.8
- Remarks on result:
- other: Close system; on Day 140
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 67.2
- Parameter:
- radiochem. meas.
- Sampling time:
- 140 d
- Remarks on result:
- other: Open system
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 36.5
- Parameter:
- radiochem. meas.
- Sampling time:
- 140 d
- Remarks on result:
- other: Close system
- Key result
- Soil No.:
- #1
- DT50:
- 97 d
- St. dev.:
- 10
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 21 °C
- Remarks on result:
- other: Open system
- Soil No.:
- #1
- DT50:
- 193 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Remarks on result:
- other: Closed system; calculated in a separate study
- Transformation products:
- no
- Details on transformation products:
- - Metabolism of the test substance: The test substance is degraded in the soil environment. Besides the soil-bound residues, the only degradation product detected is 14C-carbon dioxide which is formed upon mineralisation of the phenyl ring.
This finding is in line with the results of previous studies carried out with benzyl-labelled test substance. There, it was established that the first step of the microbial degradation involves the split-off of 14C-carbon dioxide from the parent molecule. No other significant degradation products were observed.
It is therefore concluded that the degradation of the test substance occurs in the following way:
1. elimination of the benzyl carbon atom to evolve carbon dioxide (as the rate determining step),
2. formation of a p-chlorophenyl derivative, most probably p-chlorophenol itself, and
3. degradation and mineralisation of this intermediate which is known to occur very easily by soil microorganisms l)(fast follow-up step).
The amount of the test substance found as soil-bound residues was only about 2 % of the applied radiocarbon. Admixture of fresh soil and re-incubation resulted in the evolution of further 14-carbon dioxide either form the parent a.i. itself or from one of the intermediates. This process is again mediated by microorganisms as demonstrated by the effect of heat sterilisation. - Evaporation of parent compound:
- not specified
- Volatile metabolites:
- yes
- Residues:
- yes
- Details on results:
- An overview of the results is provided in Table 1 – Table 10 in ‘Any other information on results incl. tables’.
- Material Balance: The total recovered radiocarbon content in the experiment A was calculated from the sum of the extractable and non-extractable radioactivity. It amounted to 97.7% of the applied quantity at the beginning of the study and declined continuously to 51.6% after 140 days of incubation. The recovery for the closed experiment B was accounted from the sum of volatiles, 14C-carbon dioxide, extractable and non-extractable radiocarbon, and averaged at about 95.5% of the initially applied amount. The mass balance for the 84 and 140 days sampling dates was slightly below 90% of the applied radioactivity, probably due to incomplete trapping of the volatile components.
- Volatiles: Volatile compounds were only determined in the closed system experiment B. During the course of the study, up to 16.4% of the applied radioactivity could be collected in the soda lime traps. It consisted of 14C-carbon dioxide and was confirmed by liberating it from the traps using hydrochloric acid and collecting it in a specific absorption solution (Carbo-Sorb). In order to check if the 14C-carbon dioxide was collected quantitatively from the trap and the soil, three additional soil samples (B84/R1, B112/R2 and B112/R3) were treated after the first extraction with hydrochloric acid and the air was again flushed into new soda lime traps. The procedure increased the yield of 14C-carbon dioxide by about 15 % in the case of the B84/R1 sample, but only 1 % or less in the case of the B112/R2 and B112/R3 samples. No significant amounts of other organic volatiles were found.
- Extraction Characteristics and Identification of the extractable Radiocarbon: Methanol and methanol - water (8:2) mixtures solubilized between 94% and 99% of the applied radioactivity immediately after the soil treatment. During the incubation period, the quantities of the radiocarbon extractable by the aqueous methanol (extract A) declined and finally reached about 30% of the initially applied radioactivity in the experiment A, and about 64% in the experiment B. After evaporation of the organic portion of the extracting solvent, the aqueous extracts were cleaned-up using a reversed phase C18 column. The eluting solvent consisting of 80% aqueous methanol recovered the test substance almost quantitatively from the sorbent. The so obtained organic extracts (extract B) comprised approximately the same radiocarbon content as the aqueous methanol extract. Only small quantities(< 2.4% of the applied radioactivity) were lost during the clean-up procedure and/or could not be eluted with this solvent mixture. The further analysis of the organic extract B by radio-TLC demonstrated the presence of unchanged test substance as the main component and only traces of metabolites (less than 2.1% of the applied radiocarbon content).
- Characterization of the Remaining Soil Radioactivity: the soil using methanol and methanol – water. In order to identify and/or characterize this remaining soil radiocarbon, the soil samples which have been collected at the 56, 84, 112 and 140 day after treatment and which have already been extracted with methanol and methanol water, were pooled and the radiocarbon content was determined by combustion. Three further extractions were conducted with a mixture of methanol – water (8:2). They released a total radiocarbon content of 7.4% related to the applied radioactivity.
On the basis of the radio-TLC analysis, the amount of the test substance in this extract was calculated to be 5.5% of the applied radioactivity. Only traces of other compounds with polar properties were observed(< 0.2% of the applied radioactivity). The radioactive quantities in the aqueous extracts sampled between day 56 and 140 were increased by an average value of 7.4% related to the applied radioactivity, those of the test substance by an average figure of 5.4%. For the same time period, the quantities of the remaining soil radioactivity were diminished by 7.4%. Finally, the soil-bound residues ranged between 12 and 14% of the applied radiocarbon content.
- Characterization of the Soil-Bound Residues: In an additional experiment, the soil-bound residues in the previously extracted soil samples were treated successively with water, a mixture of acetonitrile - water - acetic acid (14:6:1) and acetonitrile - water - ammonia (14:6:1). These solvent mixtures were reported to release significant amounts of the bound residues of diclofop acid, picloram, simazine and triallate from weathered field soils. This procedure released additional 1% of the applied radioactivity with water, 2.2% using acetonitrile - water – acetic acid and again 1.5% using acetonitrile - water - ammonia. The total quantity of all three extraction steps reached 4.7%, about 2% of it being identified as the parent compound by radio-TLC. At the same time, the remaining soil-bound residues were reduced from about 10% to about 4.5% of the applied radiocarbon content. The recovery was about 90% related to the radiocarbon contained in the pooled soil samples after the additional extractions with methanol – water.
- Biodegradation of the Soil-Bound Residues: Further testing included the bio-availability of the soil-bound residues for microbial degradation. Aliquots of the soil-bound residues, after the additional extractions with methanol - water, were mixed with fresh untreated Flaach soil and incubated in the closed test system. Within four weeks of incubation at 20°C and at 40% of the maximum water holding capacity, about 6% of the soil-bound residues were mineralized and could be trapped as 14C-carbon dioxide. After eight weeks, the amounts of 14CO2 formed were about 8% of the radioactivity used in this test. The test substance was detected at a level of about 11% and 3% after four and eight weeks of incubation, respectively. The main portion of radioactivity was demonstrated to be non-extractable residues.
The recoveries ranged from averaged values of 92% to 108% of radiocarbon contained in the pooled soil samples after the additional extractions. A heat sterilized control sample showed, as expected, almost no mineralization after one months period (only 0.2% of soil-bound radiocarbon applied to the biodegradation experiment).
- Dissipation of the test substance: The total amount of 14C-labelled test substance determined after the first extraction with methanol - water (8:2) and corrected for the results of the additional extractions. The test substance was proven to degrade in Flaach soil without any lag phase. Assuming first order kinetics, the calculated DT50 value was 97 ± 10 days for the experiment A. Such a value is similar to those determined in previous degradation studies of the test substance in Flaach soil showing half-life periods of 73 and 91 days. The degradation of the parent compound in the closed test system (experiment B) was considerably slower probably due to a lower microbial activity or by a substantial decrease of the microbial biomass during the course of the experiment.
- Determination of the Microbial Biomass: The biomass in the treated and untreated soil was determined by measuring the quantities of adenosine-5'-triphosphate (ATP) released from the living microorganisms by a specific detergent. Although this method is not specific for the type and the activity of the soil microbes, it allows a relative estimation of the total activity of the microorganisms in the soil. It could be shown by comparing the ATP content of the treated soil samples to that of the untreated samples that the test substance treatment did not significantly affect the activity of the microorganisms in the experiment A and B. During the course of the experiment B, a substantial increase of the ATP content within the first four weeks was observed followed by a large decrease in both the untreated and treated soil samples towards the end of the study. In the biodegradation study, an increase of the ATP content was noticed probably due to the re-cultivation of the soil which has been previously extracted with methanol and methanol - water. - Conclusions:
- Based on the findings, it was concluded that test substance is degraded in Flaach soil. The DT50 value was calculated to be 97 ± 10 days in open system (first order kinetics). The main metabolite was 14CO2. No other volatiles and only traces of extractable metabolites could be detected. Mineralization by soil microorganisms was the main route for the dissipation of the test substance in the environment under aerobic conditions. The test substance did not produce any polar metabolites containing the intact phenyl ring. Therefore, no risk for contamination of deeper soil layers and especially no harm to the groundwater has to be expected.
- Executive summary:
The aerobic soil metabolism of [14C-phenyl]-labelled test substance was investigated in Flaach soil (sandy loam with low organic matter content) according to the BBA Guidelines Part IV, 4-1. The study was in compliance with GLP criteria.The application rate of the test substance was 252 µg/kg soil (dry weight), equivalent to a field rate of 189 g a.i./ha assuming a homogeneous distribution in the upper 5 cm soil layer and a soil density of 1.5 kg/L (the normal annual application is two times 80 - 100 g a.i./ha).The study was conducted in the dark over a period of 140 days using an open test system (experiment A) in order to determine the degradation rate and a closed test system (experiment B) to investigate the degradation pathway of the test substance and the mass balance.
The recoveries for the open system (experiment A) declined from 97.7% at the beginning of the study to 51.6% of the applied radioactivity at termination, caused by the mineralization of the a. i. to carbon dioxide. The radiocarbon released after successive extraction declined to 37.5% at the end of the study. The recovery rate for the closed test system (experiment B) averaged at 95.5% of applied radioactivity. During the course of the study, up to 16.4% of applied radiocarbon was trapped as radioactive carbon dioxide. No other organic volatiles were found. The test substance extracted from the soil samples decreased with time and represented 63.5 % of the applied amounts at the last sampling date. Only traces of metabolites could be detected (< 0.1 % of the applied radiocarbon content). The soil-bound radiocarbon accounted for 12 - 14% at the end of the experiments. To check the biodegradability of the soil-bound residues, aliquots of the already extracted soil sample were incubated with fresh untreated Flaach soil using the closed test system for two months. About 8 % of soil-bound radiocarbon was mineralized to 14C-carbon dioxide (average recovery 92 - 108%), whereas a sterilized soil sample only produced traces of 14CO2 (0.2% of soil-bound radiocarbon applied to this test).
Based on the findings, it was concluded that test substance is degraded in Flaach soil. The DT50 value was calculated to be 97 ± 10 days in open system (first order kinetics). The main metabolite was 14CO2. No other volatiles and only traces of extractable metabolites could be detected. The DT50 value in closed system was calculated outside of the study and it was determined to be 193 days. Due to the insufficient aeration of the soil in the closed system, this DT50 value was considered to be not relevant for risk assessment of the biodegradation of the substance in soil. Mineralization by soil microorganisms was the main route for the dissipation of the test substance in the environment under aerobic conditions. The test substance did not produce any polar metabolites containing the intact phenyl ring. Therefore, no risk for contamination of deeper soil layers and especially no harm to the groundwater has to be expected.
- Endpoint:
- biodegradation in soil: simulation testing
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 17 Aug 1992 to 13 Oct 1993
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- other: BBA Part IV 4-1
- Version / remarks:
- 1986
- Deviations:
- no
- GLP compliance:
- yes (incl. QA statement)
- Test type:
- laboratory
- Radiolabelling:
- yes
- Oxygen conditions:
- aerobic
- Soil classification:
- not specified
- Year:
- 1 990
- Soil no.:
- #1
- Soil type:
- sandy clay loam
- % Clay:
- 23.2
- % Silt:
- 27.3
- % Sand:
- 49.6
- % Org. C:
- 1.1
- pH:
- 7.2
- CEC:
- 15.3 meq/100 g soil d.w.
- % Moisture content:
- 35.3
- Details on soil characteristics:
- SOIL COLLECTION AND STORAGE
- Geographic location: Flaach 2/92, Im Boden, canton Zürich, Switzerland
- Collecting date: July 1990
- Organic matter: 1.90%
- Soil preparation: The soil was passed through a 2 mm sieve and the actual moisture content was determined by drying a sample at about 120 °C for 48 hours. On the basis of these results and the maximum water capacity, the soil was mixed with deionized water to reach about 35 % of the maximum water capacity. The soil was equilibrated at conditions which were close to the test conditions before the start of the laboratory test. - Soil No.:
- #1
- Duration:
- 140 d
- Soil No.:
- #1
- Initial conc.:
- 0.3 mg/kg soil d.w.
- Based on:
- act. ingr.
- Parameter followed for biodegradation estimation:
- CO2 evolution
- radiochem. meas.
- Soil No.:
- #1
- Temp.:
- 20 ± 1 °C
- Humidity:
- 40% maximum water capacity
- Microbial biomass:
- 153 - 239 μg ATP/kg soil
- Details on experimental conditions:
- EXPERIMENTAL DESIGN
- Test vessel: 300 mL Erlenmeyer flasks
- Soil condition: Air dried
- Soil (g/replicate): 50 g
- Solution (mL/replicate): Adding 0.5 mL application solution by means of a pipette
- Details of traps for CO2 and organic volatile: The flasks were connected to the system for trapping carbon dioxide and volatile compounds. For analysis of the trapping system, the paraffin containing quartz wool from the trap was removed and was extracted with 100 mL ethyl acetate in order to determine the volatile organic metabolites. Duplicate aliquots of the ethyl acetate extract were taken and the content of radioactivity was determined. Radioactive carbon dioxide generated in the soil sample by mineralization of the test substance 14C-labelled test substance was trapped in the soda lime. To determine the 14CO2, it was released quantitatively from the solid absorbent by means of hydrochloric acid and absorbed in liquid absorber cocktail. This solution was analysed for its radiocarbon content by liquid scintillation counting (LSC).
The recovery was determined using an aqueous 14C-sodium carbonate solution containing a known amount of radioactivity. The recovery determined was 94 %. At the beginning of the study a slightly different procedure was applied (instead of a Vigreux column three connected scintillation vials were used).
- Moisture maintenance method: The soil samples were mixed with deionized water to reach 40 % of the maximum water capacity. The moisture content of the soil samples was controlled about every two weeks and the losses were replaced by adding deionized water.
- Temperature: 20 ± 1 °C
- Continuous darkness: Yes
- Others: After each addition, the flasks were shaken well.
TREATMENT SERIES
- Series A: 18 flasks (for the 0, 2, 4, 8, 12, 16 and 20 weeks sampling date in duplicate and 4 flasks as reserve; all flasks treated)
- Series A Biomass: 6 flasks (for the 0, 2, 8 and 20 weeks sampling date for the determination of the microbial biomass and 2 flasks as reserve; all flasks treated)
- Series B: 6 flasks (for the 0, 2, 8 and 20 weeks sampling date for the determination of the microbial biomass as control and 2 flasks as reserve; all flasks untreated)
SAMPLING DETAILS
- Sampling intervals:
Series A: 0 days (within approximately 2 hour), about 2,4,8,12,16 and 20 weeks after application (two replicates each)
Series A Biomass: 0 days, about 2, 8 weeks after application and at the end of the study
Series B: 0 days, about 2,8 weeks after application and at the end of the study
- Samples storage: At the end of the study the remaining samples were stored in a climate cabinet in the testing facility under identical conditions (20 °C ± 1 °C; in the dark). - Soil No.:
- #1
- % Non extractable:
- 16.19
- % CO2:
- 0.33
- % Recovery:
- 96.59
- Remarks on result:
- other: on day 140
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 60.03
- Parameter:
- radiochem. meas.
- Sampling time:
- 140 d
- Key result
- Soil No.:
- #1
- DT50:
- 148 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Remarks on result:
- other: First order kinetic calculation
- Soil No.:
- #1
- DT50:
- 130 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Remarks on result:
- other: Experimental result
- Transformation products:
- no
- Remarks:
- M2 & M3
- Details on transformation products:
- -Degradation Products of the test substance: M2 was detected as the main degradation product of the test substance. The percentage detected was up to 17.36 % of applied radiocarbon (average of two replicates) after 140 days. The concentration of triazole increased continuously during the study. At the same time the test substance was degraded to a metabolite tentatively identified as M3. The maximum concentration of M3 was found at the end of the study (6.7 % of AR). Additional degradation products exhibiting polar properties were ≤ 1.4 % of AR in the methanol/water extraction and ≤ 2.2 % in the water extraction.
- Evaporation of parent compound:
- not specified
- Volatile metabolites:
- yes
- Residues:
- yes
- Details on results:
- An overview of the results is provided in Table 2 – Table 5 in ‘Any other information on results incl. tables’.
- Material Balance: The recoveries were between 96.47% and 99.85% of applied radiocarbon (AR).
- Volatiles: Only small amounts of 14-CO2 were trapped during the course of the study (≤ 0.33% AR). Organic volatiles were not found.
- Extraction Characteristics of Radiocarbon: The soil samples were extracted successively with methanol, methanol/water (8:2) and with pure water. The amount of radiocarbon released by the methanol extraction and methanol/water extractions decreased from 95.7% (0 day) to 62.8% of AR at termination. At the same time the radioactivity content of the water extract increased from 0% to 17.3% of applied radiocarbon. The sum of radiocarbon found in the methanol/water and water extract declined slowly during the course of the study to 80.07% of AR. The percentage of soil bound radioactivity increased to 16.19% of AR after 140 days.
- Methanol/Water Extract: The methanol/water extract (methanol and methanol/water extraction) was subjected to a C18 column clean up. Only the Band R2 extracts contained more than 1.5 % of AR and were analysed by TLC. The B extract was chromatographed using the TLC system V. The two peaks found were identified by co-chromatography with 14C-labelled test substance as the diastereomers A and B of the test substance using the normal phase TLC system V and the reversed phase HPLC system C. The quantitation of the test substance was based on the results of the thin layer chromatograph. The R2 extracts were characterized by the TLC system VI. The major peak was identified by co-chromatography with a standard of 14C-triazole using the TLC System VI and the HPLC system A and C as M2. The results of the TLC were used for quantitation. In the course of the study the two other peaks detected in the chromatogram (at the origin and between Rf= 0.3 -0.4) accounted for a maximum of 0.85 and 1.4 % of applied radiocarbon, respectively, and were not further characterized.
- Water Extract: After concentration of the water extracts the residue was suspended in 10 mL of methanol/water (9:1). After filtration the liquid (water fraction I) contained the major amount of radiocarbon of the water extract. The water fraction I was subjected to chromatographic analysis. The residual solid after suspension with methanol/water (9:1) was dissolved in water and the content of radiocarbon of this aqueous solution (water fraction II) was determined by LSC. The contents of radiocarbon detected in the water fractions II were ≤ 2.21 % of AR (average of two replicates) and were not further characterized. The water fraction I was chromatographed using the TLC system VI, which was already used for the characterization of triazole in the R2 extract (methanol/water extract). M2 and parent compound were identified by co-chromatography with reference standards. The other metabolite found in the extract was tentatively identified by co-chromatograhpy with reference standard as M3.
- Dissipation of the test substance: The parent compound was detected in the methanol/water extract and the water extract as well. The amount of the test substance was determined by radio TLC after clean-up. The results showed that the test substance, calculated as sum of the amount found in the methanol/water extract and in the water extract (each time the sum of the diastereomers A and B), dissipated in the aerobic Flaach soil from the beginning of the study. Assuming a first order kinetic the DT50 was calculated to be 148 ± 22 days. The experimentally determined DT50 value was about 130 days. The DT90 value was not reached within the test period.
- Characterization of the Soil Bound Radioactivity: Three extracted soil samples (12W/1, 16W/1 and 20W/2) were used to characterize the soil bound radiocarbon. These soil samples were additionally extracted twice with water/acetonitrile/acetic acid and once with water/acetonitrile/ammonia (after methanol/water and water extraction). The recovery including the characterization of the soil bound radiocarbon was between 93.76 and 97.45% of AR. The additional extractions reduced the percentage of soil bound radioactivity to ≤ 8.18 % of applied radiocarbon. The pooled extracts were subjected to C18 column clean up. The B extract and the R2 extract of the characterization of the soil bound radioactivity contained the major amount of radiocarbon and were analysed using the TLC system V and VI, respectively. The chromatograms of the B extracts showed two 14C-peaks, which were identified as the diastereomers A and B of the test substance. The sum of both peaks represented 92.63%, 97.65% and 96.92% of radiocarbon applied to the TLC plate corresponding to 2.67%, 2.45% and 2.1% of AR (for the soil samples 12W/1, 16W/1 and 20W/2, respectively). In the chromatograms of the R2 extracts the radioactivity applied was detected at the origin, mainly M2 or M3 were not found.
- Microbial Biomass: The biomass of the treated and untreated soil was determined by measuring the quantities of the adenosine-5'-triphosphate (ATP) released from the living microorganisms by a specific detergent. Although this method is not specific for the type and the activity of the soil microbes, it allows a relative estimation of the total activity of the microorganisms in the soil. It could be shown by comparing the ATP content of the treated soil samples with the untreated samples that the test substance treatment did not significantly affect the activity of the microorganisms. The ATP content during and especially at the end of the incubation period was more or less the same as at the beginning of the experiment. This leads us to the conclusion that the overall activity during the experiment has not been changed significantly. - Conclusions:
- Based on the findings, it was concluded that test substance is degraded in Flaach soil. The DT50 value was calculated to be 148 ± 22 days (first order kinetics), while the experimentally determined DT50 was found to be 130 d. The main route of dissipation of 14C-triazolyl labelled test substance in aerobic soil is the degradation to M2.
- Executive summary:
The aerobic soil metabolism of [14C-Triazolel]-labelled test substance was investigated in Flaach soil (sandy loam with low organic matter content) according to the BBA Guidelines Part IV, 4-1. The study was in compliance with GLP criteria. The application rate of the test substance was 303 µg/kg soil (dry weight), equivalent to a field rate of 227 g a.i./ha assuming a homogeneous distribution in the upper 5 cm soil layer and a soil density of 1.5 kg/L (the normal annual application is two times 80 - 100 g a.i./ha).The study was conducted using a moisture of 40 percent of the maximum water capacity soil at about 20˚C in the dark over a period of 140 days.
The recoveries were between 96.47 and 99.85 percent of applied radiocarbon.Only a small amount of the test substance was mineralized to 14-CO2 (maximum of 0.3% of applied radioactivity) during the study. No organic volatiles were found. The radioactivity released by the extractions decreased with time and represented 80.07% of applied radiocarbon (AR) at termination. The soil bound radioactivity accounted for 16.19% of AR at the end of the study. Metabolite M2 was detected as the main degradation product of the test substance, 17.36 % of applied radiocarbon after 140 days incubation. Furthermore, its concentration increased continuously during the study. Another metabolite, M3 was accounted for a maximum concentration of 6.7% of AR at termination. Additional degradation products exhibiting polar properties were observed amounting to ≤1.4 percent of AR in the methanol/water extraction and to ≤2.2 percent in the water extraction.
Based on the findings, it was concluded that test substance is degraded in Flaach soil. The DT50 value was calculated to be 148 ± 22 days (first order kinetics), while the experimentally determined DT50 was found to be 130 d.
- Endpoint:
- biodegradation in soil: simulation testing
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 27 Nov 1990 to 9 Dec 1991
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- other: EPA 162-1 Aerobic Soil Metabolism Studies
- Version / remarks:
- Subdivision N, October 18 1982
- Deviations:
- yes
- Remarks:
- See Deviations in 'Any other information on materials and methods incl. tables'
- GLP compliance:
- yes (incl. QA statement)
- Test type:
- laboratory
- Radiolabelling:
- yes
- Oxygen conditions:
- aerobic
- Soil classification:
- USDA (US Department of Agriculture)
- Year:
- 1 990
- Soil no.:
- #1
- Soil type:
- loam
- % Clay:
- 22.7
- % Silt:
- 26.6
- % Sand:
- 50.7
- % Org. C:
- 1.13
- pH:
- 7
- CEC:
- 16.6 meq/100 g soil d.w.
- % Moisture content:
- 24.2
- Soil no.:
- #2
- Soil type:
- silt loam
- % Clay:
- 14.6
- % Silt:
- 53.7
- % Sand:
- 31.7
- % Org. C:
- 2.1
- pH:
- 4.3
- CEC:
- 7.13 meq/100 g soil d.w.
- % Moisture content:
- 21.7
- Soil no.:
- #3
- Soil type:
- loamy sand
- % Clay:
- 4.3
- % Silt:
- 12.1
- % Sand:
- 83.6
- % Org. C:
- 0.4
- pH:
- 5.3
- CEC:
- 1.93 meq/100 g soil d.w.
- % Moisture content:
- 6.8
- Details on soil characteristics:
- The information of geographic location and properties of the soil was provided in Table 1 in 'Any other information on materials and methods incl. tables'.
- Storage: After collection, the soil samples are shipped to the test facility and stored there in an open container under field conditions (spring to autumn) or in glass house (winter).
- Soil preparation: The wet soils were air-dried overnight and sieved through a 2 mm sieve. The actual moisture content of these samples was determined by drying at 105 °C for at least 15 hours. - Soil No.:
- #1
- Duration:
- 112 d
- Soil No.:
- #2
- Duration:
- 112 d
- Soil No.:
- #3
- Duration:
- 210 d
- Initial conc.:
- 0.25 ppm
- Based on:
- act. ingr.
- Remarks:
- applicable to all soils
- Parameter followed for biodegradation estimation:
- CO2 evolution
- radiochem. meas.
- Soil No.:
- #1
- Temp.:
- 21.6 ±3.2 °C
- Humidity:
- 75% (at 0.33 bar)
- Microbial biomass:
- 6.5E+05 (as Bacillus subtilis equivalents/g)
- Soil No.:
- #2
- Temp.:
- 21.6 ±3.2 °C
- Humidity:
- 75% (at 0.33 bar)
- Microbial biomass:
- 1.0E+06 (as Bacillus subtilis equivalents/g)
- Soil No.:
- #3
- Temp.:
- 21.6 ±3.2 °C
- Humidity:
- 75% (at 0.33 bar)
- Microbial biomass:
- 6.0E+05 (as Bacillus subtilis equivalents/g)
- Details on experimental conditions:
- EXPERIMENTAL DESIGN
- Preparation of the Application Solution: 5.0 mL of the test substance solution and 2.8 mL of the test substance standard solution were mixed and made up to 250 mL with water.
Experiment A
1360 g of the Flaach soil were weighed and mixed with the required amount of water to reach about 40% of the moisture content at 0.33 bar.)
- Soil condition: Air dried
- Soil (g/replicate): 100
- Test vessel: 300 mL Erlenmeyer flasks (were stoppered and stored at room temperature overnight.)
- Volume of test solution: 1.0 mL of the application solution was added by means of a pipette, followed by an additional quantity of water to each soil sample so as to reach 75 % moisture content at 0.33 bar.
- No. of replication treatments: 12
- Others: After each addition, the flasks were mixed well. Within less than 1 minute after the last addition of water, two soil samples were extracted. The remaining flasks were connected to the apparatus for trapping carbon dioxide and incubated.
- Continuous darkness: Yes (flasks were wrapped with aluminum foil)
- Aeration: An air stream at about 17 mL/min. was passed through the parallel test series (consisting each of one Erlenmeyer flask and four collectors) for about half an hour per working day.
- Temperature: 21.6 ± 3.2°C
- Moisture maintenance method: The moisture content of the soil samples was controlled three times by weighing the flasks during incubation (after 2, 4 and 8 weeks of incubation). Since no loss of humidity was observed, no water had to be added.
Experiment B
About 1425 g of the Louisiana soil were treated in the same way as the Flaach soil samples described in the experiment A.
Experiment C
About 1320 g of the North Carolina soil were treated in the same way as the Flaach soil samples described in the experiment A.
SAMPLING DETAILS
- Sampling intervals: Two soil samples each of the experiment A, B, and C were analysed immediately after treatment. Further samples were taken at 2, 4, 8, 12, and 16 weeks after incubation. In the case of the North Carolina soil, the incubation was continued at a higher moisture content for 30 weeks). Each sampling (soil and absorption traps) was done immediately after having passed the air stream through the trapping apparatus. - Soil No.:
- #1
- % Non extractable:
- 23.9
- % CO2:
- 32.9
- % Recovery:
- 100
- Remarks on result:
- other: on day 112
- Soil No.:
- #2
- % Non extractable:
- 13
- % CO2:
- 26.8
- % Recovery:
- 91.7
- Remarks on result:
- other: on day 112
- Soil No.:
- #3
- % Non extractable:
- 20.4
- % CO2:
- 2.2
- % Recovery:
- 100
- Remarks on result:
- other: on day 210
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 57.7
- Parameter:
- radiochem. meas.
- Sampling time:
- 112 d
- Parent/product:
- parent
- Soil No.:
- #2
- % Degr.:
- 54.1
- Parameter:
- radiochem. meas.
- Sampling time:
- 112 d
- Parent/product:
- parent
- Soil No.:
- #3
- % Degr.:
- 16.9
- Parameter:
- radiochem. meas.
- Sampling time:
- 210 d
- Key result
- Soil No.:
- #1
- DT50:
- 104 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 22 °C
- Key result
- Soil No.:
- #2
- DT50:
- 124 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 22 °C
- Soil No.:
- #3
- DT50:
- > 1 yr
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 22 °C
- Transformation products:
- no
- Details on transformation products:
- - Degradation Products: The major degradation product was identified to be carbon dioxide. The organic extracts were furthermore analysed for the presence of degradation products by radio thin-layer chromatography. In general, the results revealed only small and insignificant amounts of transformation products. The unpolar components were always less than 1.2%, the quantity of polar fractions did not exceed 4.4% of the applied radioactivity. Some very polar products (at the origin of the TLC plate) of less than ca. 6% were also observed. Additional materials were detected in the remaining aqueous phases after extraction with dichloromethane – ethanol mixtures. These components were usually less than 2% related to the applied radioactivity.
- Evaporation of parent compound:
- not specified
- Volatile metabolites:
- yes
- Residues:
- yes
- Details on results:
- An overview of the results is provided in Table 2 and Table 3 in 'Any other information on results incl. tables'.
- Material Balance: The total radioactivity in each sample was calculated from its radiocarbon contents in the trapping solvent, in the aqueous extract and from the unextractable radioactivity determined by combustion. The recoveries were generally between 90 and 105% except in the case of the 14 day samples which had lower values (83 - 98%).
- Volatiles: The volatiles were trapped in absorption solutions especially designed for collection of carbon dioxide. The amounts gradually increased with time in the experiments A and B, where the Flaach and Louisiana soil were tested. They finally reached a value of about 30% related to the applied radiocarbon. Only very limited quantities of volatile materials amounting to less than 3% were collected in the North Carolina soil.
- Dissipation of Radioactivity in the Extracts: The radiocarbon content in the methanol - water extracts and in the organic extracts from the Flaach and Louisiana soils slowly decreased with the ongoing incubation time. At the end of the experiments, the averaged quantities in the organic extracts were about 47% and 49% of the applied radioactivity. Generally, the decrease of the extractable amounts paralleled the increase of the carbon dioxide evolution. However, the extracts from the North Carolina soil were normally between ca. 75 and 100%. Since carbon dioxide production was also low, this type of soil was expected a very slow dissipation.
- Dissipation of the test substance: The organic extract of the soil sample contained the parent compound which was determined by radio thin-layer chromatography. The results showed that the test substance calculated as the sum of the diastereomers A and B, dissipated in the Flaach and Louisiana soil from the beginning of the experiment, whereas the a.i. was fairly stable in the North Carolina soil. Assuming first order kinetics in all cases, a plot of the percentage versus time showed that the half-life periods of the applied test substance were about: 104 days in the Flaach soil, 124 days in the Louisiana soil, > 1 year in the North Carolina soil.
- Soil-Bound Residues: Some radioactivity could not be extracted from the soil by use of methanol - water mixtures and was therefore determined by combustion of the residual soil. The so-called 'soil-bound' residues increased relatively fast to a constant level amounting to a maximum value of 17% in the Flaach and Louisiana soil. At the end of the study, all soils contained about 20 to 24%. - Conclusions:
- Based on the findings, it was concluded that test substance is degraded slowly in Flaach and Louisiana soils. The DT50 values were 102 days and 124 days in Flaach and Louisiana soils, respectively. The major degradation product of 14C-benzyl labelled test substance was CO2.
- Executive summary:
The aerobic soil metabolism of [14C-benzyl]-labelled test substance was investigated in Flaach (Loam), Louisiana (Silt loam) and North Carolina (Loamy sand) soils according to the EPA 162-1 Guideline. The study was in compliance with GLP criteria. The yield concentration of the test substance was 0.25 mg/kg soil (dry weight). The soils (75% moisture at 0.33 bar) were incubated at 21.6±3.2˚C in the dark for up to 16 weeks (Flaach and Louisiana soils) or 30 weeks (North Carolina soil).
The recoveries were between 90 and 105% of applied radiocarbon, except in the case of the 14 day samples which had lower values (83 - 98%). 14CO2 was the predominant degradation product in all incubations, with levels increasing over time in each case. The radiocarbon content in the methanol-water extracts and in the organic extracts from the Flaach and Louisiana soils slowly decreased over the course of the experiment. Generally, the decrease of the extractable amounts paralleled the increase of the carbon dioxide evolution. Only insignificant amounts of other transformation products were observed. However, 'soil-bound' residues were detected to an extent of 24 % (averaged value). Based on the findings, it was concluded that test substance is degraded slowly in Flaach and Louisiana soils. The DT50 values were 102 days, 124 days and > 1 year in Flaach, Louisiana and North Carolina soils, respectively. However, data from North Carolina soil will not be used in risk assessment as the soil is unsuitable for technical reasons. The major degradation product of 14C-benzyl labelled test substance was CO2.
- Endpoint:
- biodegradation in soil: simulation testing
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 21 Jul 2004 to 28 Mar 2006
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 307 (Aerobic and Anaerobic Transformation in Soil)
- Version / remarks:
- 24th April 2002
- Deviations:
- yes
- Remarks:
- The choice of the soils was required by the sponsor and are a deviation of the OECD TG 307.
- GLP compliance:
- yes (incl. QA statement)
- Test type:
- laboratory
- Radiolabelling:
- yes
- Oxygen conditions:
- aerobic
- Soil classification:
- USDA (US Department of Agriculture)
- Year:
- 2 005
- Soil no.:
- #1
- Soil type:
- clay
- % Clay:
- 51
- % Silt:
- 28
- % Sand:
- 21
- % Org. C:
- 9.6
- pH:
- 4.6
- Bulk density (g/cm³):
- 0.8
- Soil no.:
- #2
- Soil type:
- clay
- % Clay:
- 58
- % Silt:
- 24
- % Sand:
- 18
- % Org. C:
- 2.1
- pH:
- 5
- Bulk density (g/cm³):
- 1.1
- Soil no.:
- #3
- Soil type:
- loamy sand
- % Clay:
- 10
- % Silt:
- 2
- % Sand:
- 88
- % Org. C:
- 0.8
- pH:
- 4.7
- Bulk density (g/cm³):
- 1.5
- Soil no.:
- #4
- Soil type:
- clay
- % Clay:
- 50
- % Silt:
- 26
- % Sand:
- 24
- % Org. C:
- 4
- pH:
- 6
- Bulk density (g/cm³):
- 1.4
- Details on soil characteristics:
- SOIL COLLECTION AND STORAGE
- Geographic location: Four Brazilian soil types, the geographic coordinates of the sampling points were:
> GM soil: 22°35'12" S and 47°35'34" W
> RQ soil: 22°45'18" S and 47°53'75" W
> PV soil: 22°40'20" S and 47°37'31" W
> LVdf soil: 21°57'43" S and 47°50'34" W
- Pesticide use history at the collection site: GM soil was sampled from a hygrophyte vegetation area. PV soil was sampled from pastures. LVdf and RQ soils were sampled from forest vegetation. No pesticides were ever applied on the four sites.
- Sampling depth (cm) and date: Soil samples were collected between 0 - 20 cm depth in September/2004 (LVdf, PV and GM soils) and February/2005 (RQ soil)
- Soil preparation: After collection, soils were air-dried and sieved using a 2 mm mesh.
PROPERTIES OF THE SOILS: See Table 1 in 'Any other information on materials and methods incl. tables' - Duration:
- 120 d
- Initial conc.:
- 0.2 mg/kg soil d.w.
- Based on:
- act. ingr.
- Remarks:
- Applicable to all soils
- Parameter followed for biodegradation estimation:
- CO2 evolution
- radiochem. meas.
- Soil No.:
- #1
- Temp.:
- 19 - 21 °C
- Humidity:
- 40 % of the maximum water holding capacity
- Microbial biomass:
- 0.46 mg C/kg
- Soil No.:
- #2
- Temp.:
- 19 - 21 °C
- Humidity:
- 40 % of the maximum water holding capacity
- Microbial biomass:
- 0.13 mg C/kg
- Soil No.:
- #3
- Temp.:
- 19 - 21 °C
- Humidity:
- 40 % of the maximum water holding capacity
- Microbial biomass:
- 0.13 mg C/kg
- Soil No.:
- #4
- Temp.:
- 19 - 21 °C
- Humidity:
- 40 % of the maximum water holding capacity
- Microbial biomass:
- 0.11 mg C/kg
- Details on experimental conditions:
- EXPERIMENTAL DESIGN
- Soil preincubation conditions: Before application, soil samples in the flasks were maintained at 40 % of the maximum water holding capacity during five days, at 20 ± 2 °C in a dark room.
- Soil condition: Air dried
- Soil (g/replicate): 60 g
- No. of replication controls: 2
- No. of replication treatments: 14
- Test apparatus: Eight 250 mL Erlenmeyer flasks
- Others: A vacuum pump was connected at the end of the line which was controlled by timer working 1 h each two hours. The air flow in the apparatus was controlled (10-20 mL/min) by needle valve. Every day, the air flow was measured for each apparatus, using a flow meter. The aerobic samples were stored under ventilation with moistened air (flow-rate: 10 - 20 mUmin) in incubation chambers.
- Continuous darkness: Yes
Test material application
- Application method: The compound was added individually (200 μL), drop wise to the soil surface (each flask) (by means of a syringe) as a solution in acetonitrile. Then, the soil was carefully mixed with a spatula to ensure good mixing and contact of the treatment solution with the soil. In addition, an equivalent volume of test solution was placed in a 10-mL volumetric flask for direct determination of the added radioactivity.
SAMPLING DETAILS
- Sampling intervals: The measurements of 14CO2 evolution and 14C-organic volatiles, as well as the extractions for characterization of the extractable radioactivity were performed at the following intervals: 0, 7, 14, 28, 60, 90 and 120 days for GM, LVdf, RQ and PV soils
MICROBIAL ACTIVITY
To perform this assay, four flasks for each soil were treated and maintained at the same conditions employed in the dissipation study. However, two flasks were treated with the test substance (the same concentration and volume employed in the dissipation study) and two flasks without the compound (control). Measurements were performed at the following intervals: 0 and 120 days after treatment, for all soils.
The activity of the soil microorganisms was evaluated by radio respirometry, using 14C-glucose.
At each sampling time, two grams of GM, LVdf, RQ and PV soil samples were taken from the Erlenmeyer and transferred to a respirometric flask. On the soil surface was added 500μL of 4 μmol/mL 14C-glucose solution, with radioactivity of about 0.06 μCi/ml. The flasks were incubated in the dark, at 20 ± 2 °C, for 1 hour. The 14CO2 evolved from 14C-glucose was trapped in 200 μL of 2 mol/L NaOH solution. The radioactivity was assayed by LSC, and afterwards it was transformed into μg of glucose metabolized*g-1*h-1. - Soil No.:
- #1
- % Total extractable:
- 88.88
- % Non extractable:
- 12.9
- % CO2:
- 1.42
- % Recovery:
- 100
- Remarks on result:
- other: on day 120
- Soil No.:
- #2
- % Total extractable:
- 72.02
- % Non extractable:
- 17.18
- % CO2:
- 1.19
- % Other volatiles:
- 0.01
- % Recovery:
- 90.4
- Remarks on result:
- other: on day 120
- Soil No.:
- #3
- % Total extractable:
- 39.82
- % Non extractable:
- 46.5
- % CO2:
- 3.87
- % Other volatiles:
- 0.1
- % Recovery:
- 90.29
- Remarks on result:
- other: on day 120
- Soil No.:
- #4
- % Total extractable:
- 77.82
- % Non extractable:
- 23.94
- % CO2:
- 0.57
- % Other volatiles:
- 0.01
- % Recovery:
- 100
- Remarks on result:
- other: on day 120
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 59.27
- Parameter:
- radiochem. meas.
- Sampling time:
- 120 d
- Parent/product:
- parent
- Soil No.:
- #2
- % Degr.:
- 66.69
- Parameter:
- radiochem. meas.
- Sampling time:
- 120 d
- Parent/product:
- parent
- Soil No.:
- #3
- % Degr.:
- 78.99
- Parameter:
- radiochem. meas.
- Sampling time:
- 120 d
- Parent/product:
- parent
- Soil No.:
- #4
- % Degr.:
- 39.96
- Parameter:
- radiochem. meas.
- Sampling time:
- 120 d
- Key result
- Soil No.:
- #1
- DT50:
- 77.88 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Key result
- Soil No.:
- #2
- DT50:
- 68.62 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Key result
- Soil No.:
- #3
- DT50:
- 49.87 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Key result
- Soil No.:
- #4
- DT50:
- 161.19 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Transformation products:
- no
- Remarks:
- M2
- Details on transformation products:
- - Extraction and Characterization of the Extracted Radioactivity: The extractable radioactivity decreased from 110.16, 108.60, 100.49 and 105.64% of the applied radioactivity at the beginning of the study to 88.88, 72.02, 39.82 and 77.82% of the applied radioactivity at the end for GM, LVdf, RQ and PV soils, respectively. Extractable radioactivity was characterized by HPLC and showed a transformation of the original test substance to the metabolite M2 and other unknown metabolites. The test substance declined from mean levels of 106.96, 108.59, 100.49 and 105.01% of the applied radioactivity on day o to 40.73, 33.31, 21.01 and 60.04% of the applied radioactivity after 120 days of aerobic incubation in the soils GM, LVdf, RQ and PV, respectively. M2 was the major metabolite observed in this study, increasing during incubation time. This metabolite reached 20.79, 16.35, 16.73 and 7.62 % of the applied radioactivity at 120 days of incubation for GM, LVdf, RQ and PV soil, respectively.
- Evaporation of parent compound:
- not specified
- Volatile metabolites:
- yes
- Residues:
- yes
- Details on results:
- An overview of the results is provided in Table 2 – Table 7 in ‘Any other information on results incl. tables’.
- Balance of Radioactivity: Radioactivity recovery ranged from 110.16 to 94.55% of the total applied in the GM soil, from 108.60 to 90.40% in the LVdf soil, from 100.32 to 103.67% in the RO soil and from 99.52 to 105.64% of the total applied in the PV soil.
- 14CO2 Evolution: At the end of the incubation time 120 days, the cumulative percentages of 14CO2 evolved were 1.42, 1.19, 16.28 and 0.57% of the applied radioactivity from GM, LVdf, RQ and PV soils, respectively.
- 14C-organic volatile: No significant organic volatile were observed for any of the studied soils.
- Unextractable Radioactivity: Non-extractable residues increased throughout the incubation period for all four soils and reached at the end of the study 12.90, 17.18, 44.73 and 23.94% of the applied radioactivity for GM, LVdf, RQ and PV soils, respectively.
- Organic Matter fractionation: At the end of the incubation time 120 days, the non-extractable fraction was submitted to organic matter fractionation. The major part of the radioactivity of the non-extractable was found in the humin fraction which amounted to 8.51, 14.67, 13.34 and 17.35% of the applied radioactivity for GM, LVdf, RQ and PV soils, respectively.
- Half-life: The half-lives for the test substance in the GM, LVdf, RQ and PV soils were respectively 77.88, 68.62, 49.87 and 161.19 days - Conclusions:
- Based on the finding, the test substance can be considered moderately persistent for all soils tested. The DT50 values in the GM, LVdf, RQ and PV soils were 77.88, 68.62, 49.87 and 161.19 days, respectively. M2 was the major metabolite observed in this study. This metabolite reached 20.79, 16.35, 16.73 and 7.62% of the applied radioactivity at 120 days of incubation for GM, LVdf, RQ and PV soil, respectively.
- Executive summary:
The aerobic soil metabolism of [14CTriazole]-labelled test substance was investigated in GM (clay), LVdf (clay), RQ (loamy sand), PV (clay) and Arkansas (silt loam) soils according to the OECT TG 307. The study was in compliance with GLP criteria. The yield concentration of the test substance was 0.2 mg/kg soil (dry weight). Soil moisture was maintained during the incubation time at 40% of the maximum water holding capacity and samples were incubated at 20 ± 2˚C in the dark for up to 120 days. Duplicate samples treated with the test item were taken immediately after treatment (day 0) and after 7, 14, 28, 60, 90 and 120 days after the test substance application. Soil microbial activity was measured at o and 120 days after the application.
The overall recovery comprising the soil extracts, non-extractable fraction and volatile products ranged from 94.55% to 110.16% in the GM, from 90.40% to 108.60% in the LVdf, from 100.32% to 103.67% in the RQ and from 99.52% to 105.64% of total applied radioactivity in the PV soil. At the end of incubation period (120 days), the cumulative percentage of 14CO2 evolved in the GM, LVdf, RO and PV soils were respectively 1.42, 1.19, 16.28 and 0.57% of applied radioactivity. No significant organic volatiles were generated during the experimental phase, in any of the four studied soils. The test substance declined from mean levels of 106.96, 108.59, 100.49 and 105.01% of the applied radioactivity on day 0 to 40. 73, 33.31, 21.01 and 60.04% of the applied radioactivity after 120 days of aerobic incubation in soils GM, LVdf, RQ and PV, respectively.Metabolite M2 was the major metabolite in this study, increasing during incubation time. This metabolite reached the value of 20.79, 16.35, 16.73 and 7.62% of the applied radioactivity at 120 days of incubation for GM, LVdf, RQ and PV soil, respectively.
Based on the finding, the test substance can be considered moderately persistent for all soils tested. The DT50 values in the GM, LVdf, RQ and PV soils were 77.88, 68.62, 49.87 and 161.19 days, respectively. M2 was the major metabolite observed in this study.
- Endpoint:
- biodegradation in soil: simulation testing
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 4 Feb 2005 to 27 Feb 2006
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 307 (Aerobic and Anaerobic Transformation in Soil)
- Version / remarks:
- April 2002
- Deviations:
- not specified
- Qualifier:
- according to guideline
- Guideline:
- other: EPA 162 - 1
- Deviations:
- not specified
- GLP compliance:
- yes (incl. QA statement)
- Test type:
- laboratory
- Radiolabelling:
- yes
- Oxygen conditions:
- aerobic
- Soil classification:
- USDA (US Department of Agriculture)
- Year:
- 2 005
- Soil no.:
- #1
- Soil type:
- silt loam
- % Clay:
- 25
- % Silt:
- 68
- % Sand:
- 7
- % Org. C:
- 1.51
- pH:
- 5.5
- CEC:
- 16.4 meq/100 g soil d.w.
- % Moisture content:
- 23.9
- Details on soil characteristics:
- SOIL COLLECTION AND STORAGE
- Geographic location: UMC Bradford Research Farm, Columbia, Missouri, Boone County, USA.
- Pesticide use history at the collection site: This field site had received only herbicide treatments for the last 5 years.
- Sampling date: 4th March 2005
- Sampling depth: The soil was sampled in the field to a total depth of approximately 6 inches.
- Transportation: The sampled soil was transferred unfrozen and under ambient conditions to the test facility.
- Soil preparation (e.g., 2 mm sieved; air dried etc.): On arrival, the soil was initially coarse sieved through a 5 mm sieve. This was further sieved through a 2 mm mesh. - Soil No.:
- #1
- Duration:
- 150 d
- Soil No.:
- #1
- Duration:
- 210 d
- Soil No.:
- #1
- Duration:
- 300 d
- Soil No.:
- #1
- Initial conc.:
- 0.01 mg/kg soil d.w.
- Based on:
- act. ingr.
- Soil No.:
- #1
- Initial conc.:
- 0.02 mg/kg soil d.w.
- Based on:
- act. ingr.
- Soil No.:
- #1
- Initial conc.:
- 0.055 mg/kg soil d.w.
- Based on:
- act. ingr.
- Soil No.:
- #1
- Initial conc.:
- 0.1 mg/kg soil d.w.
- Based on:
- act. ingr.
- Soil No.:
- #1
- Initial conc.:
- 1 mg/kg soil d.w.
- Based on:
- act. ingr.
- Parameter followed for biodegradation estimation:
- CO2 evolution
- radiochem. meas.
- Soil No.:
- #1
- Temp.:
- 20 ± 1 °C
- Humidity:
- 27.9%
- Microbial biomass:
- 20.92 mg/100 soil on 160 DAT and 17.95 mg/100g soil at termination
- Details on experimental conditions:
- EXPERIMENTAL DESIGN
- Soil preincubation conditions: The soil was pre-incubated under study conditions for 12 days prior to treatment with the application solutions.
- Soil condition: Moisture-adjusted soil samples
- Soil (g/replicate): 100g
- No. of replication treatments: 2
- Test apparatus: Glass centrifuge bottles (volume of vessel approximately 330 mL).
- Details of traps for CO2 and organic volatile: The effluent air from each glass vessel was passed through a tube of ethanediol (to absorb any volatile organics) and a tube of 2M sodium hydroxide (to absorb any 14 CO2 produced).
Test material application
- Preparation of treatment solution: The radiolabelled test substance was dissolved in acetonitrile to give the stock solution. This solution was quantified by LSC and the volume required for the preparation of each treatment solution determined. To prepare each treatment solution, appropriate volumes of the stock solution was added to volumetric flasks and made up to volume with acetonitrile.
- Application method and volume: 100 µL (10 x 10 μL aliquots) of 14C-labelled test substance in acetonitrile were applied to the soil surface of each soil vessel using an electronic repeating dispenser fitted with 1.25 mL combitip. The soil was then mixed thoroughly with a spatula to ensure even distribution of the test item.
- Moisture maintenance method: Throughout the incubation period, the moisture content of each soil pot was checked by weighing at regular intervals. Any moisture losses were replaced on these occasions by the addition of ultra-pure water.
- Continuous darkness: Yes
SAMPLING DETAILS
- Sampling intervals: See Table 1 in 'Any other information on materials and methods incl. tables'
- Sampling method for soil samples: At sampling time, duplicate vessels from each treatment rate were disconnected from the flow-through system, transfen-ed back to the laboratory and extracted immediately.
- Method of collection of CO2 and volatile organic compounds: At each sampling interval, the traps associated with the soil vessels sampled were removed and quantified by LSC. The cumulative total level of radioactivity evolved from each vessel was then calculated. Due to the lack of volatile organics being trapped, the ethanediol traps were not included from 127 DAT. - Soil No.:
- #1
- % Total extractable:
- 44.15
- % Non extractable:
- 52.05
- % Recovery:
- 97.1
- Remarks on result:
- other: on day 150
- Remarks:
- treatment rate 0.01 mg/kg
- Soil No.:
- #1
- % Total extractable:
- 45.7
- % Non extractable:
- 49.3
- % Recovery:
- 95.7
- Remarks on result:
- other: on Day 150
- Remarks:
- treatment rate 0.02 mg/kg
- Soil No.:
- #1
- % Total extractable:
- 50.1
- % Non extractable:
- 47.6
- % Recovery:
- 98.2
- Remarks on result:
- other: on Day 210
- Remarks:
- treatment rate 0.055 mg/kg
- Soil No.:
- #1
- % Total extractable:
- 38.5
- % Non extractable:
- 55.7
- % Recovery:
- 94.85
- Remarks on result:
- other: on Day 300
- Remarks:
- treatment rate 0.1 mg/kg
- Soil No.:
- #1
- % Total extractable:
- 61.2
- % Non extractable:
- 39.55
- % Recovery:
- 100
- Remarks on result:
- other: on Day 300
- Remarks:
- treatment rate 1 mg/kg
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 74.5
- St. dev.:
- 0.99
- Parameter:
- radiochem. meas.
- Sampling time:
- 150 d
- Remarks on result:
- other: at 0.01 mg/kg rate
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 71.2
- St. dev.:
- 3.25
- Parameter:
- radiochem. meas.
- Sampling time:
- 150 d
- Remarks on result:
- other: at 0.002 mg/kg rate
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 63.9
- St. dev.:
- 3.82
- Parameter:
- radiochem. meas.
- Sampling time:
- 210 d
- Remarks on result:
- other: at 0.055 mg/kg rate
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 77.95
- St. dev.:
- 2.47
- Parameter:
- radiochem. meas.
- Sampling time:
- 300 d
- Remarks on result:
- other: at 0.1 mg/kg rate
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 40.35
- St. dev.:
- 1.2
- Parameter:
- radiochem. meas.
- Sampling time:
- 300 d
- Remarks on result:
- other: at 1 mg/kg rate
- Key result
- Soil No.:
- #1
- DT50:
- 78.3 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Remarks on result:
- other: at 0.01 mg/kg rate
- Key result
- Soil No.:
- #1
- DT50:
- 91 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Remarks on result:
- other: at 0.02 mg/kg rate
- Key result
- Soil No.:
- #1
- DT50:
- 167 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Remarks on result:
- other: at 0.055 mg/kg rate
- Key result
- Soil No.:
- #1
- DT50:
- 169 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Remarks on result:
- other: at 0.1 mg/kg rate
- Key result
- Soil No.:
- #1
- DT50:
- 444 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Remarks on result:
- other: at 1 mg/kg rate
- Transformation products:
- not measured
- Evaporation of parent compound:
- not specified
- Volatile metabolites:
- yes
- Residues:
- yes
- Details on results:
- An overview of the results is provided in Table 3 – Table 7 in ‘Any other information on results incl. tables’.
- Microbial Biomass: The microbial biomass for the soils analysed prior to treatment and after the incubation period is shown in Table 3. The quantity of microbial biomass carbon constituted 2.21 % of the organic carbon at the start. This indicated the soils supported a viable microbial population and that these soils were suitable for use in a laboratory degradation study. The quantity of microbial biomass carbon measured at 160 days and 300 days after treatment constituted 1.44% and 1.24% for the soil organic carbon respectively. This indicated the soils supported a viable microbial population throughout the incubation period, although it declined with time.
- Mass Balance: Total mass balances from the samples analysed immediately after treatment (zero time) ranged between 99.7% and 107.9% of the applied dose. Total mass balances for the remaining sampling intervals were ≥ 90% for all treatment rates, with one exception of 86.6% for one replicate sample (98 DAT, 0.1 mg/kg rate). The average total radioactive recovery for all samples was 99.8%.
- Volatile Degradation Products: Insignificant amounts of the applied radioactivity was evolved as volatile products, which were trapped in sodium hydroxide. The maximum amounts of volatile radioactivity evolved from soil pots were 0.9%, 0.7%, 0.6%, 0.7% and 0.3% of the applied radioactivity for treatment rates 0.01, 0.02, 0.055, 0.1 and 1 mg/kg, respectively.
- Radioactive Residues in Soil Extracts: The amount of extractable radioactivity was found to decrease with time for all treatment rates. It ranged from 92.7% to 103.3% on day zero to 39.1% to 56.7% by end of incubation. This decline in extractable radioactivity with time suggests that the remaining radioactivity is bound or incorporated into the soil matrix. Aliquot of each extract was taken and diluted in acetonitrile and water (40 : 60 v/v) for analysis by LC-MS-MS to detem1ine the amount of parent substance remaining. The 0 - 98 DAT samples for the 0.1 mg/kg treatment rate were also analysed by 1 D-TLC and the levels of the test substance and found were compared with the results obtained by LC-MS-MS to corroborate all other LS-MS-MS results.
- LC-MS-MS Quantitation: 14C-labelled test substance standards (0.000045 - 0.09 µg/mL) made up in acetonitrile and water (40:60 v/v) generated a linear response when analysed by LC-MS-MS. Degradation of the test substance in 0.01 and 0.02 mg/kg proceeded quickly, such that the parent compound represented only 25.5% and 28.8% (averaged) of the applied radioactivity after 150 days of incubation. Degradation in the 0.055 and 0.1 mg/kg treatment was slower and incubation terminated after 210 and 300 days respectively. At termination, the average cyproconazole determined in the extracts were 36.1 % (0.055 mg/kg) and 22.1% (0.1 mg/kg) of the applied radioactivity. Degradation in the 1 mg/kg treatment was very slow. After 300 days incubation the test substance represented 59.7% (averaged) of the applied radioactivity.
- Degradation Rate of the test substance at Different Concentrations: The percentage of parent substance as determined by the LC-MS-MS was plotted against incubation time and fitted. However, due to unacceptable variability in the LC-MS-MS data for the 0.1 µg/mL rate, the TLC data (up to 98 DAT) were substituted for the modelling. Results demonstrated that an increase in application rate from 0.01 mg/kg to 1.0 mg/kg resulted in significant decrease in the rate of degradation. - Conclusions:
- Based on the finding, the DT50 values in the test soil at 0.01, 0.02, 0.055, 0.1 and 1 mg/kg soil treatment rate were 78.3, 91.0, 167, 169 and 444 days, respectively. Only a small amount of radiolabelled carbon dioxide were produced during the incubation. The maximum amounts of volatile radioactivity evolved from soil pots were < 1% of the applied radioactivity for all treatment rates.
- Executive summary:
The aerobic soil metabolism of [14CTriazolyl]-labelled test substance was investigated in Missouri soil (Silt loam) according to the OECD TG 307 and EPA 162-1 guideline. The study was in compliance with GLP criteria. The applied rate of the test substance were 0.01, 0.02, 0.055, 0.1 and 1 mg/kg soil (dry weight). Soil moisture was maintained during the incubation time at pF2 and samples were incubated at 20 ± 2˚C in the dark for up to 300 days. In all experiments, the vessels were incubated in a flow-through system, which maintained aerobic and flushed volatile degradation products into trapping systems. Duplicate samples were taken from each treatment rate for analysis at up to 8 intervals during incubation.
The overall mean mass balance (across all rates) was 99.8% of the applied radioactivity, with the means for the individual rate ranging from 98.0 to 101. 7%. The maximum level of 14CO2 evolved in each experiment ranged from 0.3% (1µg/mL rate) to 0.9% (0.01 μg/mL rate) of the applied radioactivity and the level of unextractable radioactivity ranged from 39.1% (1µg/mL rate) to 56.7% (0.1µg/mL rate) of the applied radioactivity, by the end of the incubation. Based on the findings, the DT50 values determined using simple first order (SFO) kinetics were 78 days (0.01 mg/kg rate), 91 days (0.02 mg/kg rate), 167 days (0.055 mg/kg rate), 169 days (0.1 mg/kg rate) and 444 days (1 mg/kg rate).
- Endpoint:
- biodegradation in soil: simulation testing
- Type of information:
- experimental study
- Adequacy of study:
- key study
- Study period:
- 24 Oct 2005 to 2 May 2007
- Reliability:
- 1 (reliable without restriction)
- Rationale for reliability incl. deficiencies:
- guideline study
- Qualifier:
- according to guideline
- Guideline:
- OECD Guideline 307 (Aerobic and Anaerobic Transformation in Soil)
- Version / remarks:
- April 2002
- Qualifier:
- according to guideline
- Guideline:
- other: EPA 162-1: Aerobic Soil Metabolism Studies
- GLP compliance:
- yes (incl. QA statement)
- Test type:
- laboratory
- Radiolabelling:
- yes
- Oxygen conditions:
- aerobic
- Soil classification:
- USDA (US Department of Agriculture)
- Year:
- 2 005
- Soil no.:
- #1
- Soil type:
- sand
- % Clay:
- 3
- % Silt:
- 4
- % Sand:
- 93
- % Org. C:
- 0.35
- pH:
- 6.1
- CEC:
- 2.7 meq/100 g soil d.w.
- % Moisture content:
- 4.5
- Soil no.:
- #2
- Soil type:
- sandy loam
- % Clay:
- 12
- % Silt:
- 16
- % Sand:
- 72
- % Org. C:
- 0.7
- pH:
- 5.7
- CEC:
- 4.1 meq/100 g soil d.w.
- % Moisture content:
- 12.7
- Soil no.:
- #3
- Soil type:
- sandy clay loam
- % Clay:
- 22
- % Silt:
- 16
- % Sand:
- 62
- % Org. C:
- 2.09
- pH:
- 6.5
- CEC:
- 18.3 meq/100 g soil d.w.
- % Moisture content:
- 20
- Soil no.:
- #4
- Soil type:
- silty clay
- % Clay:
- 44
- % Silt:
- 50
- % Sand:
- 6
- % Org. C:
- 2.44
- pH:
- 8
- CEC:
- 35.2 meq/100 g soil d.w.
- % Moisture content:
- 27.9
- Soil no.:
- #5
- Soil type:
- silt loam
- % Clay:
- 13
- % Silt:
- 65
- % Sand:
- 22
- % Org. C:
- 1.51
- pH:
- 4.7
- CEC:
- 7.3 meq/100 g soil d.w.
- % Moisture content:
- 29
- Details on soil characteristics:
- - The information of geographic location, properties and sample dates of the soils is provided in Table 1 in 'Any other information on materials and methods incl. tables'.
- Collection procedures: The soils were sampled in the field to a total depth of approximately 15 cm (6 inches).
- Storage: On arrival at the test facility, the soils were stored at 20 ± 2°C until use.
- Soil preparation: Before use in the study, the soils were sieved (2 mm mesh) and moistened with de-ionised water. An aliquot of each sieved soil was air-dried and submitted for determination of its physico-chemical characteristics. Prior to incubation under study conditions, the sieved soil was adjusted to a moisture content approximately equivalent to pF2 by the addition of de-ionised water. The moisture adjusted soil samples were stored in non-airtight plastic containers at 20 ± 2°C in the dark for at most 14 days prior to dispensing into glass vessels and a further 7-14 days prior to application of the test substance. The microbial biomass in the soil was estimated from the respiratory response following addition of glucose. Initial biomass determinations were made on samples of each soil after adjustment of the soil moisture content. Final biomass measurements were made on untreated soil samples maintained under the same soil aerobic study conditions as the treated samples after approximately 365 days incubation. - Duration:
- 365 d
- Soil No.:
- #1
- Initial conc.:
- 0.018 mg/kg soil d.w.
- Based on:
- act. ingr.
- Soil No.:
- #2
- Initial conc.:
- 0.019 mg/kg soil d.w.
- Based on:
- act. ingr.
- Soil No.:
- #3
- Initial conc.:
- 0.021 mg/kg soil d.w.
- Based on:
- act. ingr.
- Soil No.:
- #4
- Initial conc.:
- 0.022 mg/kg soil d.w.
- Based on:
- act. ingr.
- Soil No.:
- #5
- Initial conc.:
- 0.023 mg/kg soil d.w.
- Based on:
- act. ingr.
- Parameter followed for biodegradation estimation:
- CO2 evolution
- radiochem. meas.
- Soil No.:
- #1
- Temp.:
- 20 ± 2 °C
- Humidity:
- 8.1 g/100g dry soil
- Microbial biomass:
- Initial and final: 3.53 and 2.85 mg/100g Soil
- Soil No.:
- #2
- Temp.:
- 20 ± 2 °C
- Humidity:
- 14.4 g/100g dry soil
- Microbial biomass:
- Initial and final: 15.71 and 8.86 mg/100g Soil
- Soil No.:
- #3
- Temp.:
- 20 ± 2 °C
- Humidity:
- 25.8 g/100g dry soil
- Microbial biomass:
- Initial and final: 32.23 and 22.54 mg/100g Soil
- Soil No.:
- #4
- Temp.:
- 20 ± 2 °C
- Humidity:
- 30.5 g/100g dry soil
- Microbial biomass:
- Initial and final: 54.63 and 39.43 mg/100g Soil
- Soil No.:
- #5
- Temp.:
- 20 ± 2 °C
- Humidity:
- 25.2 g/100g dry soil
- Microbial biomass:
- Initial and final: 11.93 and 6.13 mg/100g Soil
- Details on experimental conditions:
- EXPERIMENTAL DESIGN
- Test vessel: Glass centrifuge bottles (volume of vessel approximately 330 mL).
- Soil condition: Air dried
- Soil (g/replicate): 100
- Soil pre-incubation: The soils were pre-incubated under study conditions for at least 7 days prior to treatment with the application solution.
- Details of traps for CO2 and organic volatile: Volatile products produced were trapped using a flow through system. Air was pushed (using a peristaltic pump) through a water trap, then through the incubation vessels and any volatiles were collected in the outlet 2M NaOH traps. At each sampling interval, the traps associated with the soil vessels sampled were removed and quantified by LSC.
Test material application
- Preparation of treatment solution: The radiolabelled test substance was mixed with acetonitrile to give the stock solution. This solution was quantified by LSC and the volume required for the preparation of the treatment solution determined. To prepare the treatment solution, an appropriate volume of the stock solution was added to a volumetric flask and made up to volume with acetonitrile.
- Volume of test solution used/treatment: 200 μL (10 x 20 μL) of 14C-labelled test substance in acetonitrile (equivalent to ca. 1.8 μg test substance) was applied to each soil pot. This was equivalent to a nominal application rate of 40 g ai/ ha (assuming even incorporation into the top 15 cm (6 inches) of soil).
- Application method: The solution was applied in droplets using an electronic dispenser. The soil was then mixed thoroughly by tapping and rolling the vessel. Triplicate 200 μL (10 x 20 μL) aliquots of the treatment solution were transferred to 10 mL volumetric flasks (which were diluted to volume with acetonitrile) and quantified by LSC. This was performed before, during and after application to check the homogeneity of the treatment solution and to determine the exact application rate.
Experimental conditions
- Moisture maintenance method: Throughout the incubation period, the soil moisture levels were maintained by reweighing the vessels at regular intervals and re-moistening back to their original weights with de-ionised water. The weights before and after re-moistening were recorded and used to determine the fluctuation in moisture content with time.
- Continuous darkness: Yes
SAMPLING DETAILS
- Sampling intervals: Duplicate soil vessels were sampled, from each set of incubation conditions, immediately after treatment (zero time) and 3, 6, 9, 20, 28, 64, 150 and 365 days after treatment (DAT). All vessels were transferred back to the laboratory and extracted immediately. - Soil No.:
- #1
- % Total extractable:
- 40.9
- % Non extractable:
- 62.3
- % CO2:
- 0.1
- % Recovery:
- 100
- Remarks on result:
- other: on day 365
- Soil No.:
- #2
- % Total extractable:
- 48
- % Non extractable:
- 48.5
- % CO2:
- 0.2
- % Recovery:
- 96.6
- Remarks on result:
- other: on day 365
- Soil No.:
- #3
- % Total extractable:
- 42.8
- % Non extractable:
- 55.2
- % CO2:
- 1.1
- % Recovery:
- 99.1
- Remarks on result:
- other: on day 365
- Soil No.:
- #4
- % Total extractable:
- 28.1
- % Non extractable:
- 55.7
- % CO2:
- 0.4
- % Recovery:
- 84.2
- Remarks on result:
- other: on day 365
- Soil No.:
- #5
- % Total extractable:
- 16.9
- % Non extractable:
- 77.2
- % CO2:
- 0.4
- % Recovery:
- 94.5
- Remarks on result:
- other: on day 365
- Parent/product:
- parent
- Soil No.:
- #1
- % Degr.:
- 61.3
- Parameter:
- radiochem. meas.
- Sampling time:
- 365 d
- Parent/product:
- parent
- Soil No.:
- #2
- % Degr.:
- 55.7
- Parameter:
- radiochem. meas.
- Sampling time:
- 365 d
- Parent/product:
- parent
- Soil No.:
- #3
- % Degr.:
- 57.5
- Parameter:
- radiochem. meas.
- Sampling time:
- 365 d
- Parent/product:
- parent
- Soil No.:
- #4
- % Degr.:
- 85.1
- Parameter:
- radiochem. meas.
- Sampling time:
- 365 d
- Parent/product:
- parent
- Soil No.:
- #5
- % Degr.:
- 88.7
- Parameter:
- radiochem. meas.
- Sampling time:
- 365 d
- Key result
- Soil No.:
- #1
- DT50:
- 191 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Key result
- Soil No.:
- #2
- DT50:
- 216 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Key result
- Soil No.:
- #3
- DT50:
- 142 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Key result
- Soil No.:
- #4
- DT50:
- 79 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Key result
- Soil No.:
- #5
- DT50:
- 52 d
- Type:
- (pseudo-)first order (= half-life)
- Temp.:
- 20 °C
- Transformation products:
- no
- Remarks:
- M3
- Details on transformation products:
- The identity of parent was confirmed in each of the soil samples by full scan LC/MS analysis comparison with a certified analytical standard of the test substance (The identity of the parent compound was also confirmed in extracts from Georgia soil with results similar to those reported, however, the results are not included in this report). Analysis of the test substance standard using HPLC conditions 3 gave peaks at ~ 17.3 and ~ 17. 9 minutes (isomers) in the total ion chromatogram (TIC) and product ion chromatogram. The mass spectrum shows a pseudomolecular precursor ion m/z 292. Analysis of each of the soil samples using HPLC conditions 3 gave peaks at ~ 17.3 and ~ 17.9 minutes in the total ion and product ion chromatogram. Comparison of the background extracted full scan and product mass spectra with that of the test substance standard confirmed the presence of the test substance in these samples. The secondary confirmation of the parent quantification by HPLC-MS/MS was satisfactory and supported the data from the primary quantification method. The most significant degraded product was a highly polar component (unretained by reversed phase HPLC) which was observed in all of the soils. This component reached a peak of 11.5% of applied radioactivity after 365 days in Illinois soil, 6.7% after 64 days in Arkansas soil, 3.9% after 9 days in North Dakota soil, 3.2% after 365 days in North Carolina soil and 1.5% of applied radioactivity after 365 days in Georgia soil.
The identity of the component was confirmed as metabolism M3 in the concentrated sample by full scan LC-MS and comparison with a certified radio labelled analytical standard of M3. Analysis of the M3 standard using HPLC conditions 4 gave a peak at ~10.5 minutes, and analysis of concentrated sample gave a peak at ~ 10.5 minutes. The mass spectrum shows a pseudomolecular precursor ion m/z 128 including 14C isotope m/z 130. The product ion spectra of m/z 128 shows a loss of carboxylic acid and presence of triazole. Analysis of the concentrated sample shows a pseudomolecular precursor ion m/z 128 and 14C isotope m/z 130 in the full scan mass spectrum. The product ion spectra of m/z 128 shows a loss of carboxylic acid and presence of triazole. These soils also contained up to 5 other minor degradates, with the combined total not accounting for more than 2.4% of the applied radioactivity. - Evaporation of parent compound:
- not specified
- Volatile metabolites:
- yes
- Residues:
- yes
- Details on results:
- An overview of the results is provided in Table 2 – 10 Table in ‘Any other information on results incl. tables’
- Microbial Biomass: The results indicated that the soils supported a viable microbial population at the start of the study and that there was a decline in the biomass, however, this is common in studies of this duration, and there was still a viable microbial population at the end of the incubation period.
- Mass Balance: The total radioactivity recovered was calculated by summation of the activity in the soil extracts, soil residue on combustion and that trapped as 14CO2 in the 2M sodium hydroxide traps. Total mass balances from the samples analysed immediately after treatment (zero time) ranged between 99.4% and 101.3% of the applied dose. Total mass balances for the remaining sampling intervals were generally > 90% for all incubation soils. The average total radioactive recovery for all conditions was 96.4%. The amount of extractable radioactivity was found to decrease with time under all conditions. It ranged from 96.6%, 96.7%, 94.4%, 97.5% and 99.1% on day 0 to 40.9%, 48.0%, 42.8%, 28.1% and 16.9% by day 365 (for Georgia, North Carolina, North Dakota, Illinois and Arkansas soils, respectively). This decline in extractable radioactivity with time and the low amount of volatile radioactivity suggests that the remaining radioactivity was bound or incorporated into the soil matrix.
- Volatile Degradation Products: Very low levels of radioactivity were evolved as volatile products, which were trapped in sodium hydroxide that is consistent with it being 14CO2. The maximum level for any of the soils did not exceed 1.1% of applied radioactivity.
- Radioactive Residues in Soil Extracts: Degradation of the test substance was fastest in Illinois and Arkansas soils. After 365 days the test substance accounted for 14.9% (of the applied radioactivity) in Illinois and 11.3% in Arkansas. Degradation was slightly slower in Georgia, North Carolina and North Dakota soils, with parent representing 38.7%, 44.3% and 42.5% of the applied radioactivity respectively, after 365 days.
- Investigation into the Nature of the Unextracted Residue: In Arkansas soil, between 27.8 and 33.2% of the applied radioactivity was associated with the humin fraction, between 33.9 and 38.1% of the applied radioactivity was associated with the fulvic fraction, whilst 5.5 to 5.8% was associated with the humic acid fraction. The results indicate that, for the soil which displayed the largest unextracted residue, that this unextracted residue was not associated with a single fraction.
- Degradation DT50 and DT 90 for the test substance: The percentage of applied radioactivity present as parent substance was plotted against incubation time and fitted to simple first order kinetics (forced fit through 100% at zero time). Using the simple first order kinetics model, the best visual fit to the data was obtained by forcing the fit through 100% at zero time. Additionally, this modelling fit was most appropriate since it reflects the high extraction efficiencies (94 - 99%) at zero day. The simple first order DT50 values obtained from this analysis were 191, 216, 142, 79 and 52 days for Georgia, North Carolina, North Dakota, Illinois and Arkansas soils, respectively.
- Storage Stability of the Radioactive Components: The chromatographic profiles of the selected concentrated extracts, which had been stored refrigerated and/or frozen for a minimum of 167 days, were compared with those of the original analysis. HPLC analysis of the extracts showed essentially similar profiles. Quantification of the levels of test substance remaining confirmed that, in these cases, no significant additional degradation was observed after 167 days refrigerated and/or frozen storage. - Conclusions:
- Based on the findings, it was concluded that the degradation was faster in Illinois and Arkansas than in Georgia, North Carolina and North Dakota. The respective half-lives using simple first order kinetics were 79, 52, 191, 216 and 142 days, respectively. Loss of the test substance in soil was via degradation and formation of bound residues. Small amounts of 14CO2 were produced by all soils during the incubation, indicating that the mineralisation of the test substance was very limited. M3 is the only significant metabolite observed in the Illinois (11.5% at 365 DAT) and Arkansas (6.7% at 64DAT) soils. M3 was observed at< 3.9% of applied radioactivity, in the Georgia, North Carolina and North Dakota soils.
- Executive summary:
The aerobic soil metabolism of [14CTriazole]-labelled test substance was investigated in Georgia (sand), North Carolina (sandy loam), North Dakota (sandy clay loam), Illinois (silty clay) and Arkansas (silt loam) soils according to the OECT TG 307 and EPA 162-1 guidelines. The study was in compliance with GLP criteria. The yield concentration of the test substance was 0.018 – 0.023 mg/kg soil (dry weight). The soils were incubated at 20 ± 2˚C in the dark for up to 365 days. Duplicate soil vessels were taken, of each soil, for analysis at 0 (zero time), 3, 6, 9, 20, 28, 64, 150 and 365 days after treatment. Soil extracts were analysed chromatographically and the unextracted residues quantified by combustion of the extracted soil.
Mean mass balances were 102, 97, 97, 93 and 93% of the applied radioactivity, for Georgia, North Carolina, North Dakota, Illinois and Arkansas, respectively. Very small amounts of 14CO2 were produced by all soils during the incubation, indicating that mineralisation of the test substance was limited. M3 is the only significant (> 5% applied) metabolite observed in the Illinois (11.5% at 365 DAT) and Arkansas (6.7% at 64 DAT) soils. M3 was observed at< 3.9% of applied radioactivity, in the Georgia, North Carolina and North Dakota soils. Unextractable residues in Georgia, North Carolina, North Dakota, Illinois and Arkansas soils generally increased throughout the 365 DAT incubation period with maximum levels (all at 365 DAT) of 62.3, 48.5, 55.2, 55.7 and 77.2% of the applied radioactivity, respectively. Based on the findings, it was concluded that the degradation of the test substance was faster in Illinois and Arkansas than in Georgia, North Carolina and North Dakota. The respective half-lives using simple first order kinetics were 79, 52, 191, 216 and 142 days.
Referenceopen allclose all
Table 1. Trapping of 14CO2
Denomination |
Days |
Normal trapping |
Trapping after additional treatment with hydrochloric acid |
||
1000 dpm |
% of AR |
1000 dpm |
% of AR |
||
B84/R1 |
84 |
17.5 |
1. 7 |
3.1 |
0.3 |
B112/R2 |
112 |
127.5 |
12.5 |
0.8 |
0.1 |
Bl12/R3 |
112 |
165.0 |
16.4 |
1.8 |
0.2 |
Table 2. Soil Samples Pooled for Additional Extractions, Characterization and Biodegradation of the Remaining Soil Radioactivity
Denomination |
Days |
Soil weight (g) |
Remaining soil radioactivity *) as percentage of applied radioactivity |
Open test system
A56/1 A56/2 A84/1 A84/2 A112/1 Al12/2 A140/1 A140/2 |
56 56 84 84 112 -112 140 140 |
100 100 100 100 100 100 100 100 |
13.3 13.3 13.0 14.2 18.9 20.4 19.9 23.1 |
Average |
- |
- |
17.0 |
Closed test system
B56/1 B56/2 B84/1 B84/2 B112/1 B112/2 B112/R2 B112/R3 B140/1 B140/2 |
56 56 84 84 112 112 112 112 140 140 |
50 50 50 50 50 50 50 50 50 50 |
16.0 18.5 23.3 22.3 18.3 21.3 19.6 26.9 19.1 22.6 |
Average |
- |
- |
20.8 |
Remaining soil residues |
18.4 |
||
*) not solubilized by the first extractions with methanol and methanol- water (8:2); values determined by combustion
Table 3. Characterization of the Additional Extracts
Extracts |
Total radioactivity |
|
in 1000 dpm |
as percentage of applied radioactivity |
|
Remaining soil radioactivity*) (aliquot of 886.36 g) |
2842.4 |
18.4 |
Agueous extract (Extract A) with methanol-water (8:2) first extract second extract third extract total extracts
Organic extract (Extract B) eluate from C18 column
The test substance(radio-TLC) Other components |
642.7 334.8 170.4 1148.0
871.0
845.9 25.1 |
4.2 2.2 1.1 7.4
5.6
5.5 0.16 |
Rinse R1
Rinse R2 - The test substance (radio-TLC)
Rinse R3 - The test substance (radio-TLC)
Rinse R4
Rinse R5 |
2.1
53.9 <11
44.9 <5
16.3
1.2 |
<0.1
0.3 <0.1
0.3 <0.1
0.1
<0.1 |
*) not solubilized by the first extractions with methanol and methanol- water (8:2); values determined by combustion
Table 4. Degradation of the test substance in Flaach Soil/ All Figures Corrected by the Results of the Additional Extractions (given in% related to applied radioactivity)
Denomination |
Days |
Carbon dioxide |
Aqueous extract (Ext A) *) |
Test substance **) |
Soil-bound Residues *) |
Total recovered ***) |
Open test system (Experiment A) |
93.7 83.6 75.5 66.7 59.2 35.9 32.8 |
3.4 10.6 8.5 5.9 6.2 12.2 14.1 |
97.7 95.6 86.1 77.7 70.6 54.2 51.6 |
|||
A0 A14 A28 A56 A84 A112 A140 |
0 14 28 56 84 112 140 |
n.a. n.a. n.a. n.a. n.a. n.a. n.a. |
94.3 85.0 77.6 71.8 64.4 42.0 37.5 |
|||
Closed test system (Experiment B) |
89.7 83.6 82.5 74.2 68.5 63.0 63.5 |
3.5 9.9 8.6 9.8 14.1 14.1 13.4 |
102.7 99.2 99.2 99.2 87.0 97.2 86.8 |
|||
B0 B14 B28 B56 B84 B112 B140 |
0 14 28 56 84 112 140 |
n.a. 0.5 2.8 6.3 2.1 11.2 1.9 |
99.3 88.8 87.7 83.1 70.8 72.0 71.4 |
|||
*) corrected by 7.4 % (related to applied radioactivity) after additional extractions
**) corrected by 5.5 % (related to applied radioactivity) after additional extractions
***) sum of carbon dioxide, extract A and soil-bound residues
n.a. not analysed
Table 5. Characterization of the Soil-Bound Residues
Extracts |
Total radioactivity |
|
in 1000 dpm |
As percentage of applied radioactivity |
|
Remaining soil radioactivity*) (aliquot of 886.36 g) |
2842.4 |
18.4 |
Extraction of soil-bound residues (aliquot of 100 g) |
17.5 38.7 25.4 81.6 |
1.0 2.2 1.5 4.7 |
Aqueous extract (Extract A) - water - acetonitrile-wate-acetic acid - cetonitrile-water-ammonia - total extracts |
||
Organic extract (Extract B) - eluate from Cl8 column
Test substance (radio-TLC)
Soil-bound residues (combustion) |
35.3
34.6
78.3 |
2.0
2.0
4.5 |
Mass balance (aliquot of 886.36 g) Remaining soil radioactivity*) Additional extractions**)
Extracts from soil-bound residues Soil-bound residues (combustion) Total recovered: 90.3 % ***) |
2842.4
1148.0
723.5
694.0
2565.5 |
18.4
7.4
4.7
4.5
16.6 |
Table 6. Biodegradation of the Soil-Bound Residues in Flaach Soil(All figures given in 1000 dpm)
Denomination |
Days |
Carbon dioxide |
Aqueous extracts (Ext. A) |
Test substance |
Not extractable |
Total recovered |
K0 *) |
0 |
n.a. |
6.98 |
n.a. |
n.a. |
(38.23) |
K1**) |
28 |
0.09 |
9.41 |
n.a. |
n.a. |
n.a. |
1M/1 |
28 |
1.86 |
5.17 |
4.14 |
26.46 |
33.49 |
1M/2 |
28 |
3.05 |
7.17 |
n.a. |
26.46 |
36.68 |
1M |
28 |
2.46 |
6.17 |
4.14 |
26.46 |
35.09 |
2M/1 |
56 |
3.17 |
3.41 |
1.26 |
34.38 |
40.96 |
2M/2 |
56 |
3.14 |
3.61 |
n.a. |
35.06 |
41.80 |
2M |
56 |
3.15 |
3.51 |
1.26 |
34.72 |
41.38 |
*) heat sterilized control sample before incubation: 5 g of treated soil containing 38'234 Kdpm (calculated value)
**) heat sterilized control sample: 20 g of treated and 25 g of fresh untreated soil
Table 7. Biodegradation of the Soil-Bound Residues in Flaach Soil
All figures given in% related to applied radioactivity in the soil samples used for the biodegradation experiment (after additional extractions of the pooled soil samples). All figures in parenthesis given in% related to the applied radioactivity in the soil samples for the experiment A and B.
Denomination |
Days |
carbon dioxide |
Aqueous extracts (Ext. A) |
Test substance |
Not extractable |
Total recovered |
K0 *) |
0 |
n.a. |
18.3 (2.0) |
n.a. |
n.a. |
100.0 (18.4) |
K1 **) |
28 |
0.2 (0.03) |
24.6 (2.7) |
n.a. |
n.a. |
n.a. |
1M/1 1M/2 1M |
28 28 28 |
4.9 (0.5) 8.0 (0.9) 6.4 (0.7) |
13.5 (1.5) 18.8 (2.1) 16.1 (1.8) |
10.8 (1.2) n.a. 10.8 (1.2) |
69.2 69.2 69.2 |
87.6 95.9 91.8 |
2M/1 2M/2 2M |
56 56 56 |
8.3 (0.9) 8.2 (0.9) 8.2 (0.9) |
8.9 (1.0) 9.4 (1.0) 9.2 (1.0) |
3.3 (0.4) n.a. 3.3 (0.4) |
89.9 91.7 90.8 |
107.1 109.3 108.2 |
*) heat sterilized control sample before incubation: 5 g of treated soil containing 38'234 Kdpm (calculated value)
**) heat sterilized control sample: 20 g of treated and 25 g of fresh untreated soil
Table 8. Microbial Biomass Determined as Adenosin-5'-triphospate (ATP) (average of at least two replicates)
Days after Application |
Untreated Samples µg ATP/kg soil |
Treated Samples µg ATP/kg soil |
Open test system (Exoeriment A) |
42.3 36.0 63.8 |
|
0 91 147 |
44.3 41.5 71.6 |
|
Closed test system (Experiment B) |
- 86.7 176.6 15.6 |
|
0 14 28 119 |
83.1 80.6 150.9 14.6 |
|
Biodegradation study |
143.6 195.1 508.3 |
|
0 28 56 |
- - - |
|
Table 9. Degradation of the test substance in Flaach Soil (All Figures Given in% Related to Applied Radioactivity)
All figures represent the total radioactivity before the additional extraction steps with methanol - water (8:2), as well as with water, acetonitrile - water - acetic acid, and/or acetonitrile - water - ammonia
Denomi- nation |
Days |
Aqueous extract (Ext A) |
Organic extract (Ext B) |
Test substance |
Diaste- reomer A |
Diaste- reomer B |
Metabolites |
Rinses (R1-R5) |
Remaining soil radioact. |
Total recovered |
||
unpolar |
polar |
v. polar |
||||||||||
A0/1 A0/2 A0 |
0 0 0 |
94.1 94.5 94.3 |
94.7 94.5 94.6 |
94.3 93.2 93.7 |
44.3 44.6 44.0 |
51.0 48.5 49.7 |
<0.1 0.9 0.5 |
0.2 0.4 0.3 |
0.2 <0.1 0.1 |
1.2 0.6 0.9 |
3.9 2.9 3.4 |
98.0 97.4 97.7 |
A14/1 A14/2 A14 |
14 14 14 |
85.1 84.9 85.0 |
83.7 84.3 84.0 |
83.4 83.7 83.6 |
39.0 39.8 39.4 |
44.4 43.9 44.2 |
0.2 o.7 0.4 |
<0.1 <0.1 <0.1 |
0.1 0.1 0.1 |
1.5 1.6 1.6 |
11.3 9.9 10.6 |
96.4 94.8 95.6 |
A281 A28/2 A28 |
28 28 28 |
77.0 78.2 77.6 |
74.1 78.0 76.0 |
74.0 71.0 77.5 |
36.3 37.6 36.9 |
37.7 39.4 38.6 |
0.2 <0.1 0.1 |
<0.1 1.0 0.5 |
<0.1 <0.1 <0.1 |
2.0 2.1 2.0 |
9.0 8.1 8.5 |
86.0 86.2 86.1 |
A56/1 A56/2 A56 |
56 56 56 |
65.3 63.4 64.4 |
62.2 62.1 62.1 |
61.4 60.9 61.2 |
29.3 28.3 28.8 |
32.2 32.7 32.4 |
0.1 <0.1 0.1 |
0.6 1.3 0.9 |
<0.1 <0.1 <0.1 |
1.8 1.8 1.8 |
13.3 13.3 13.3 |
78.6 76.7 77.7 |
A84/1 A84/2 A84 |
84 84 84 |
51.7 56.3 57.0 |
56.5 54.0 55.2 |
55.2 52.3 53.7 |
23.5 22.7 23.1 |
31.7 29.6 30.6 |
0.1 0.2 0.2 |
1.0 1.2 1.1 |
0.1 0.3 0.2 |
2.2 2.1 2.2 |
13.0 14.2 13.6 |
70.6 70.5 70.6 |
A112/1 A112/2 A112 |
112 112 112 |
36.0 33.1 34.6 |
33.9 30.3 32.1 |
32.4 28.5 30.4 |
14.3 12.2 13.2 |
18.2 16.3 17.2 |
0.3 0.4 0.4 |
1.0 1.1 1.1 |
0.1 0.3 o.2 |
1.7 1.8 1.8 |
18.9 20.4 19.4 |
54.9 53.4 54.2 |
A140/1 A140/2 A140 |
140 140 140 |
30.0 30.2 30.1 |
28.7 28.6 28.7 |
27.6 27.0 27.3 |
12.2 11.5 11.8 |
15.4 15.4 15.4 |
<0.1 0.2 0.1 |
1.0 1.3 1.1 |
0.2 0.2 0.2 |
1.4 1.6 1.5 |
19.9 23.1 21.5 |
49.9 53.2 51.6 |
Table 10. Degradation of the test substance in Flaach Soil (All Figures Given in% Related to Applied Radioactivity). All figures represent the total radioactivity before the additional extraction steps with methanol - water (8:2), as well as with water, acetonitrile - water - acetic acid, and/or acetonitrile - water – ammonia
Denomi- nation |
Days |
Volatiles |
Carbon dioxide |
Aqueous extract (Ext A) |
Organic extract (Ext B) |
Test substance |
Diaste- reomer A |
Diaste- reomer B |
Metabolites |
Rinses (R1-R5) |
Remaining soil radioact. |
Total recovered |
B0/1 B0/2 B0 |
0 0 0 |
n.a. n.a. n.a. |
n.a. n.a. n.a. |
99.9 98.6 99.3 |
88.7 94.0 91.3 |
86.9 92.5 89.7 |
43.7 44.2 43.9 |
43.2 48.3 45.7 |
1.8 1.5 1.6 |
1.6 1.1 1.4 |
3.3 3.7 3.5 |
103.1 102.3 102.7 |
B14/1 B14/2 B14 |
14 14 14 |
<0. <0.1 <0.1 |
0.6 0.4 0.5 |
83.4 94.2 88.8 |
83.4 85.0 84.2 |
82.8 84.5 83.6 |
39.4 44.2 41.8 |
43.4 40.3 41.8 |
0.7 0.5 0.6 |
2.1 2.5 2.3 |
10.0 9.9 9.9 |
93.9 104.5 99.2 |
B28/1 B28/2 B28 |
28 28 28 |
<0.1 <0.1 <0.1 |
2.9 2.8 2.8 |
87.1 88.3 87.7 |
83.6 84.5 84.0 |
82.2 82.9 82.5 |
38.9 40.5 39.7 |
43.3 42.3 42.8 |
1.4 1.6 1.5 |
1.6 2.1 1.8 |
9.1 8.2 8.6 |
99.1 99.3 99.2 |
B56/1 B56/2 B56 |
56 56 56 |
<0.1 <0.1 <0.1 |
6.4 6.2 6.3 |
73.5 77.8 75.7 |
69.5 70.3 69.9 |
67.4 70.0 68.7 |
31.1 34.8 33.0 |
36.3 35.2 35.7 |
2.1 0.3 1.2 |
1.8 1.7 1.7 |
16.0 18.5 17.2 |
96.0 102.5 99.2 |
B84/1 B84/2 B84/R1 B84 |
84 84 84 84 |
<0.1 <0.1 <0.1 <0.1 |
2.2 2.2 2.0*) 2.1 |
59.7 59.1 71.6 63.4 |
58.3 64.8 69.2 64.1 |
57.4 63.7 67.9 63.0 |
26.1 29.4 33.1 29.5 |
31.3 34.3 34.8 33.5 |
0.9 1.2 1.2 1.1 |
1.5 1.1 2.4 1.7 |
23.3 22.3 18.8 21.5 |
85.2 83.5 92.4 87.0 |
B112/1 B112/2 B112/R2 B112/R3 B112 |
112 112 112 112 112 |
<0.1 <0.1 <0.1 <0.1 <0.1 |
2.9 12.8 12.6*) 16.4*) 11.2 |
68.6 67.1 64.4 58.1 64.6 |
57.4 62.3 59.9 52.8 58.1 |
55.7 62.4 59.7 52.1 57.5 |
26.4 29.4 29.0 27.4 28.1 |
29.3 33.0 30.7 24.7 29.4 |
1.7 <0.1 0.2 0.7 0.7 |
1.2 2.1 3.1 3.2 2.4 |
18.3 21.3 19.6 26.8 21.5 |
89.8 101.2 96.6 101.3 97.2 |
B140/1 B140/2 B140 |
140 140 140 |
<0.1 <0.1 <0.1 |
1.4 2.4 1.9 |
66.9 61.2 64.0 |
60.5 56.8 58.7 |
59.6 56.4 58.0 |
27.3 27.9 27.6 |
32.3 28.4 30.4 |
0.9 0.5 o.7 |
0.9 1.6 1.2 |
19.1 22.6 20.8 |
87.4 86.2 86.8 |
Table 2. Recovery and distribution of radioactivity in soil treated with 14C-triazolyl labelled test substance (values as % AR)
Incubation days |
Methanol /water extract Before clean-up treatment |
Elution of concentrated extract+ from silica clean-up column |
Water ext* |
14CO2 |
NER |
Total recovery ** |
||||
Fraction R2* [1st eluate from column] |
Fraction R3* [MeOH/ H2O] |
Extract B* [MeOH/ H2O] 8:2 |
Fraction R4* [MeOH] |
Total radio. eluted |
|
|
|
|
||
0 |
95.71 |
1.47 |
0.82 |
91.84 |
1.53 |
95.66 |
< LOD |
< LOD |
0.76 |
96.47 |
14 |
91.45 |
2.37 |
0.83 |
79.18 |
1.07 |
83.45 |
1.37 |
0.05 |
4.64 |
97.51 |
28 |
85.96 |
4.85 |
1.15 |
77.78 |
0.86 |
84.64 |
3.42 |
0.03 |
7.63 |
97.05 |
56 |
86.6 |
6.47 |
0.97 |
75.96 |
0.53 |
83.93 |
4.25 |
0.16 |
8.38 |
99.39 |
84 |
82.92 |
6.58 |
1.07 |
72.84 |
0.66 |
81.15 |
5.82 |
0.15 |
10.95 |
99.85 |
112 |
73.41 |
11.72 |
0.8 |
57.84 |
0.64 |
71 |
12.26 |
0.1 |
13.1 |
98.86 |
140 |
62.8 |
14.72 |
0.93 |
46.42 |
0.44 |
62.51 |
17.27 |
0.33 |
16.19 |
96.59 |
+ Percentages do not incorporate the results of harsh extractions.
*Average of three replicates
** Total recovery = methanol/water extract + water extract + 14CO2 and unextracted residue.
NER Non Extractable Residue.
Table 3. Distribution of radioactivity in extracts of soil (B, R2, water extracts) treated with 14C-triazolyl labelled test substance (values as % AR).
Incubation days |
Sum of radioactivity in fractions B+R2 +water |
TLC analysis of B +R2 +water |
Total radioactivity of parent and metabolites |
||
Test substance* |
M2 |
M3 |
|||
0 |
93.31 |
89.05 |
0 |
0 |
89.05 |
14 |
82.92 |
80.05 |
0.35 |
0.09 |
80.49 |
28 |
86.05 |
72.31 |
0.81 |
0.56 |
73.68 |
56 |
86.68 |
71.84 |
6.36 |
1.35 |
79.55 |
84 |
85.24 |
67.37 |
6.22 |
1.78 |
75.37 |
112 |
81.82 |
50.55 |
13.15 |
2.56 |
66.26 |
140 |
78.41 |
39.97 |
17.36 |
6.7 |
64.03 |
* Percentages do not incorporate the results of harsh extractions.
Fractions R3 and R4 were not characterised as the radioactivity in these samples did not exceed 1.6% AR.
Table 4. Microbial Biomass Determined as Adenosin-5'-triphospate (ATP)
Sampling Date |
Treated Samples (Series A Biomass) µg ATP/kg soil mean ± StDev |
Untreated Samples (Series B) µg ATP/ kg soil mean ± StDev |
at the beginning of the study |
158 ± 8 |
130 ± 8 |
2 weeks after application |
176 ± 5 |
182 ± 10 |
8 weeks after application |
239 ± 8 |
194 ± 13 |
at the end of the study |
153 ± 1 |
129 ± 30 |
Table 5. Characterization of the Soil Bound Residues (Values in Percent of Applied Radiocarbon)
Sample No. |
Days |
Soil bound 1) |
Extract 2) |
B Extract |
R2 Extract |
R3 Extract |
R4 Extract |
Soil bound 3) |
Rec. 4) |
12W/1 |
84 |
10.48 |
5.5 |
2.88 |
1.34 |
0.42 |
0.04 |
3.87 |
97.4 |
16W/1 |
112 |
13.69 |
6.89 |
2.51 |
2.77 |
0.34 |
0.06 |
6.18 |
96.0 |
20W/2 |
140 |
16.87 |
7.74 |
2.16 |
2.90 |
0.36 |
0.05 |
8.18 |
93.8 |
1) soil bound after methanol, methanol/water and water extraction
2) pooled extracts from water/acetonitrile/acetic acid and water/acetonitrile/ammonia extractions
3) soil bound after water/acetonitrile/acetic acid and water/acetonitrile/ammonia extractions
4) recovery calculated from the methanol/water and water extract, 14CO2, volatiles, extract 2) and soil bound 3)
Table 2. Distribution of radioactivity in soils treated with 14C-benzyl labelled test substance (values as % AR).
Incubation days |
test substance |
Non-polar |
Polar |
Very polar |
Aqueous phase |
14CO2 |
Unextracted |
Total recovery |
Flaach soil |
||||||||
0 |
83.9 |
0.1 |
2.4 |
0.5 |
0.1 |
0.0 |
0.7 |
100.7 |
14 |
54.2 |
0.1 |
0.7 |
4.6 |
0.1 |
7.3 |
15.1 |
86.2 |
28 |
64.4 |
0.2 |
1.2 |
1.0 |
0.2 |
13.5 |
17.1 |
92.1 |
56 |
52.2 |
0.3 |
2.4 |
0.5 |
0.4 |
26.7 |
16.3 |
95.2 |
84 |
46.3 |
0.3 |
2.4 |
0.8 |
0.9 |
32.4 |
17.1 |
101.6 |
112 |
42.3 |
0.6 |
2.8 |
1.6 |
1.0 |
32.9 |
23.9 |
102.7 |
Louisiana soil |
||||||||
0 |
84.3 |
0.3 |
0.8 |
0.6 |
0.1 |
0.0 |
1.0 |
99.6 |
14 |
54.0 |
0.2 |
2.2 |
2.7 |
0.0 |
10.9 |
15.5 |
93.1 |
28 |
55.9 |
0.7 |
1.3 |
0.5 |
0.2 |
25.2 |
12.2 |
93.0 |
56 |
38.3 |
1.2 |
2.0 |
0.5 |
0.9 |
44.2 |
12.8 |
98.0 |
84 |
56.6 |
0.6 |
0.9 |
1.2 |
1.4 |
26.1 |
12.0 |
101.2 |
112 |
45.9 |
0.3 |
1.6 |
1.1 |
1.5 |
26.8 |
13.0 |
91.7 |
North Caroline soil |
||||||||
0 |
91.9 |
0.0 |
0.6 |
1.5 |
0.1 |
0.0 |
0.3 |
99.7 |
14 |
66.7 |
0.4 |
0.8 |
3.1 |
0.0 |
0.5 |
9.2 |
85.0 |
28 |
77.3 |
0.1 |
2.8 |
0.6 |
0.2 |
1.2 |
12 |
89.6 |
56 |
83.1 |
0.5 |
0.6 |
0.9 |
0.8 |
1.2 |
4.2 |
90.4 |
84 |
82.5 |
0.3 |
0.7 |
0.7 |
1.2 |
0.6 |
4.1 |
100.5 |
112 |
82.0 |
0.0 |
0.9 |
1.3 |
0.3 |
2.9 |
14.0 |
100.5 |
140 |
82.2 |
0.1 |
1.4 |
1.7 |
0.9 |
2.3 |
7.8 |
104.1 |
210 |
83.1 |
0.0 |
0.9 |
1.4 |
0.5 |
2.2 |
20.4 |
104.6 |
Table 3. Summary of degradation rates of the test substance in laboratory soils
Soil |
Label position |
Moisture content |
Temp (˚C) |
Initial concentration (mg/kg) |
CO2 (%AR) |
Bound residues (%AR) |
DT50 (days) |
Flaach |
Benzyl |
75% 0.33 bar |
22 |
0.25 |
32.9 |
23.9 |
104 |
Louisiana |
26.8 |
13.0 |
124 |
||||
North Carolina |
2.2 |
20.4 |
> 1 year |
Table 2. Radioactive mass balance: GM soil (soil #1)
Sampling time |
Replicate |
Extract 1 |
Extract 2 |
Extract 3 (Soxhlet) |
Total Extracted |
14CO2 |
14CO2 Organic volatlie |
unextracted |
Total recovery |
Days |
% of applied radioactivity |
||||||||
0 |
R1 |
95.81 |
11.32 |
2.41 |
109.54 |
nd |
nd |
nd |
109.54 |
R2 |
96.57 |
11.43 |
2.78 |
110.79 |
nd |
nd |
nd |
110.79 |
|
Mean |
96.19 |
11.38 |
2.6 |
110.16 |
nd |
nd |
nd |
110.16 |
|
7 |
R1 |
89.85 |
6.89 |
6.02 |
102.75 |
0.21 |
0.04 |
0.56 |
103.57 |
R2 |
94.12 |
4.9 |
4.48 |
103.5 |
0.19 |
0.04 |
1.02 |
104.75 |
|
Mean |
91.98 |
5.89 |
5.25 |
103.13 |
0.20 |
0.04 |
0.79 |
104.16 |
|
14 |
R1 |
87.01 |
7.54 |
4.18 |
98.73 |
0.37 |
0.04 |
1.98 |
101.12 |
R2 |
90.62 |
8.65 |
4.32 |
103.59 |
0.35 |
0.04 |
1.58 |
105.55 |
|
Mean |
88.82 |
8.1 |
4.25 |
101.16 |
0.36 |
0.04 |
1.78 |
103.34 |
|
28 |
R1 |
86.79 |
10.99 |
6.04 |
103.83 |
0.67 |
0.06 |
2.83 |
107.40 |
R2 |
79.73 |
7.09 |
9.79 |
96.62 |
0.7 |
0.05 |
1.05 |
98.42 |
|
Mean |
83.26 |
9.04 |
7.92 |
100.22 |
0.69 |
0.05 |
1.94 |
102.91 |
|
60 |
R1 |
74.12 |
10.4 |
9.54 |
94.06 |
0.95 |
0.03 |
1.31 |
96.35 |
R2 |
70.7 |
10.16 |
9.14 |
90.01 |
0.86 |
0.04 |
1.86 |
92.76 |
|
Mean |
72.41 |
10.28 |
9.34 |
92.03 |
0.9 |
0.03 |
1.59 |
94.56 |
|
90 |
R1 |
76.97 |
9.38 |
13.98 |
100.33 |
1.26 |
0.02 |
3.59 |
105.20 |
R2 |
81.18 |
10.14 |
11.48 |
102.8 |
1.12 |
0.03 |
3.10 |
107.05 |
|
Mean |
79.07 |
9.76 |
12.73 |
101.56 |
1.19 |
0.03 |
3.35 |
106.12 |
|
120 |
R1 |
75.69 |
11.28 |
0.02 |
86.99 |
1.49 |
0.01 |
13.14 |
101.63 |
R2 |
80.42 |
10.35 |
0.00 |
90.77 |
1.36 |
0.00 |
12.65 |
104.78 |
|
Mean |
78.05 |
10.82 |
0.01 |
88.88 |
1.42 |
0.00 |
12.90 |
103.20 |
|
nd: not determined
Table 3. Radioactive mass balance: LVdf soil (soil #2)
Sampling time |
Replicate |
Extract 1 |
Extract 2 |
Extract 3 (Soxhlet) |
Total Extracted |
14CO2 |
14CO2 Organic volatlie |
unextracted |
Total recovery |
Days |
% of applied radioactivity |
||||||||
0 |
R1 |
100.66 |
6.38 |
1.44 |
108.47 |
nd |
nd |
nd |
108.47 |
R2 |
101.15 |
6.15 |
1.42 |
108.72 |
nd |
nd |
nd |
108.72 |
|
Mean |
100.9 |
6.26 |
1.43 |
108.6 |
nd |
nd |
nd |
108.60 |
|
7 |
R1 |
91.68 |
11.45 |
3.08 |
106.22 |
0.19 |
0.04 |
0.81 |
107.26 |
R2 |
91.57 |
5.76 |
3.18 |
100.51 |
0.22 |
0.04 |
0.69 |
101.46 |
|
Mean |
91.63 |
8.61 |
3.13 |
103.37 |
0.21 |
0.04 |
0.75 |
104.36 |
|
14 |
R1 |
85.04 |
7.44 |
3.4 |
95.87 |
0.35 |
0.04 |
1.13 |
97.39 |
R2 |
92.66 |
8.94 |
3.33 |
104.92 |
0.41 |
0.04 |
1.03 |
106.4 |
|
Mean |
88.85 |
8.19 |
3.36 |
100.39 |
0.38 |
0.04 |
1.08 |
101.89 |
|
28 |
R1 |
85.35 |
7.76 |
4.54 |
97.65 |
0.54 |
0.08 |
1.77 |
100.04 |
R2 |
90.29 |
8.16 |
4.44 |
102.9 |
0.62 |
0.07 |
2.13 |
105.71 |
|
Mean |
87.82 |
7.96 |
4.49 |
100.27 |
0.58 |
0.07 |
1.95 |
102.87 |
|
60 |
R1 |
68.31 |
10.56 |
14.05 |
92.92 |
0.68 |
0.02 |
3.45 |
97.07 |
R2 |
80.63 |
11.43 |
12.49 |
104.54 |
0.78 |
0.02 |
3.17 |
108.52 |
|
Mean |
74.47 |
10.99 |
13.27 |
98.73 |
0.73 |
0.02 |
3.31 |
102.79 |
|
90 |
R1 |
65.53 |
10.97 |
13.62 |
90.12 |
0.72 |
0.03 |
10.83 |
101.70 |
R2 |
72.86 |
10.32 |
12.56 |
95.73 |
0.89 |
0.02 |
7.91 |
104.55 |
|
Mean |
69.19 |
10.64 |
13.09 |
92.92 |
0.81 |
0.02 |
9.37 |
103.12 |
|
120 |
R1 |
59.94 |
11.5 |
0.05 |
71.49 |
1.06 |
0.01 |
18.7 |
91.26 |
R2 |
62.99 |
9.53 |
0.03 |
72.55 |
1.31 |
0.02 |
15.67 |
89.55 |
|
Mean |
61.46 |
10.51 |
0.04 |
72.02 |
1.19 |
0.01 |
17.18 |
90.40 |
|
nd: not determined
Table 4. Radioactive mass balance: RQ soil (soil #3)
Sampling time |
Replicate |
Extract 1 |
Extract 2 |
Extract 3 (Soxhlet) |
Total Extracted |
14CO2 |
14CO2 Organic volatlie |
unextracted |
Total recovery |
Days |
% of applied radioactivity |
||||||||
0 |
R1 |
96.2 |
5.3 |
1.22 |
102.71 |
nd |
nd |
nd |
102.71 |
R2 |
91.6 |
5.56 |
1.09 |
98.26 |
nd |
nd |
nd |
98.26 |
|
Mean |
93.9 |
5.43 |
1.16 |
100.49 |
nd |
nd |
nd |
100.49 |
|
7 |
R1 |
90.87 |
5.9 |
1.51 |
98.28 |
1.51 |
0.08 |
3.41 |
103.28 |
R2 |
91.56 |
5.73 |
1.76 |
99.06 |
1.48 |
0.06 |
3.46 |
104.06 |
|
Mean |
91.22 |
5.82 |
1.63 |
98.67 |
1.49 |
0.07 |
3.44 |
103.67 |
|
14 |
R1 |
80.46 |
5.83 |
1.94 |
88.23 |
4.67 |
0.07 |
6.12 |
99.09 |
R2 |
81.68 |
5.6 |
1.75 |
89.04 |
4.64 |
0.07 |
8.53 |
102.27 |
|
Mean |
81.07 |
5.72 |
1.84 |
88.63 |
4.65 |
0.07 |
7.33 |
100.68 |
|
28 |
R1 |
65.26 |
4.69 |
3.32 |
73.27 |
4.69 |
0.20 |
19.49 |
97.65 |
R2 |
64.41 |
5.27 |
2.48 |
72.16 |
5.16 |
0.11 |
19.74 |
97.18 |
|
Mean |
64.84 |
4.98 |
2.9 |
72.72 |
4.92 |
0.16 |
19.61 |
97.41 |
|
60 |
R1 |
50.41 |
3.73 |
1.19 |
55.32 |
7.53 |
0.03 |
31.69 |
94.57 |
R2 |
52.25 |
3.82 |
1.92 |
57.99 |
7.98 |
0.03 |
35.46 |
101.46 |
|
Mean |
51.33 |
3.78 |
1.55 |
56.66 |
7.76 |
0.03 |
33.58 |
98.02 |
|
90 |
R1 |
40.20 |
3.94 |
13.3 |
57.45 |
0.28 |
0.09 |
27.76 |
85.58 |
R2 |
41.28 |
2.90 |
10.16 |
54.33 |
4.97 |
0.06 |
30.87 |
90.24 |
|
Mean |
40.74 |
3.42 |
11.73 |
55.89 |
2.62 |
0.08 |
29.32 |
87.91 |
|
120 |
R1 |
33.65 |
2.85 |
1.04 |
37.53 |
1.49 |
0.13 |
49.15 |
88.31 |
R2 |
37.68 |
2.60 |
1.82 |
42.1 |
6.25 |
0.07 |
43.85 |
92.27 |
|
Mean |
35.67 |
2.72 |
1.43 |
39.82 |
3.87 |
0.1 |
46.5 |
90.29 |
|
nd: not determined
Table 5. Radioactive mass balance: PV soil (soil #4)
Sampling time |
Replicate |
Extract 1 |
Extract 2 |
Extract 3 (Soxhlet) |
Total Extracted |
14CO2 |
14CO2 Organic volatlie |
unextracted |
Total recovery |
Days |
% of applied radioactivity |
||||||||
0 |
R1 |
95.67 |
6.87 |
2.7 |
105.24 |
nd |
nd |
nd |
105.24 |
R2 |
96.47 |
6.72 |
2.85 |
106.04 |
nd |
nd |
nd |
106.04 |
|
Mean |
96.07 |
6.8 |
2.77 |
105.64 |
nd |
nd |
nd |
105.64 |
|
7 |
R1 |
86.44 |
8.32 |
4.84 |
99.60 |
0.84 |
0.04 |
1.61 |
102.09 |
R2 |
89.17 |
8.54 |
5.07 |
102.78 |
0.88 |
0.05 |
1.36 |
105.07 |
|
Mean |
87.81 |
8.43 |
4.95 |
101.19 |
0.86 |
0.04 |
1.49 |
103.58 |
|
14 |
R1 |
86.58 |
10.07 |
6.03 |
102.68 |
0.74 |
0.03 |
1.62 |
105.08 |
R2 |
75.08 |
8.55 |
5.84 |
89.47 |
0.72 |
0.05 |
1.83 |
92.08 |
|
Mean |
80.83 |
9.31 |
5.93 |
96.08 |
0.73 |
0.04 |
1.72 |
98.58 |
|
28 |
R1 |
82.07 |
9.96 |
7.42 |
99.45 |
0.59 |
0.04 |
3.64 |
103.72 |
R2 |
77.63 |
7.52 |
6.54 |
91.7 |
0.53 |
0.05 |
3.05 |
95.33 |
|
Mean |
79.85 |
8.74 |
6.98 |
95.57 |
0.56 |
0.04 |
3.34 |
99.52 |
|
60 |
R1 |
71.38 |
12.97 |
9.47 |
93.82 |
0.95 |
0.02 |
5.65 |
100.44 |
R2 |
72.68 |
13.14 |
8.41 |
94.24 |
0.86 |
0.03 |
6.26 |
101.38 |
|
Mean |
72.03 |
13.06 |
8.94 |
94.03 |
0.9 |
0.02 |
5.96 |
100.91 |
|
90 |
R1 |
69.32 |
11.63 |
6.73 |
87.68 |
0.27 |
0.03 |
12.37 |
100.35 |
R2 |
73.73 |
11.14 |
8.41 |
93.28 |
0.23 |
0.02 |
9.83 |
103.36 |
|
Mean |
71.53 |
11.38 |
7.57 |
90.48 |
0.25 |
0.03 |
11.10 |
101.86 |
|
120 |
R1 |
62.96 |
10.73 |
0.02 |
73.72 |
0.57 |
0.01 |
26.08 |
100.38 |
R2 |
71.29 |
10.62 |
0.02 |
81.93 |
0.58 |
0.01 |
21.81 |
104.32 |
|
Mean |
67.12 |
10.68 |
0.02 |
77.82 |
0.57 |
0.01 |
23.94 |
102.35 |
|
nd: not determined
Table 6. Characterization of radioactivity in the soils
Sampling Time (days) |
test substance |
M2 |
Unknown |
GM soil #1; 106.96 (% of applied) |
|||
0 |
106.96 |
0.18 |
3.02 |
7 |
102.3 |
< LQ (1) |
0.25 |
14 |
100.61 |
< LQ |
0.55 |
28 |
99.5 |
< LQ |
0.72 |
60 |
63.71 |
< LQ |
28.33 |
90 |
45.85 |
22.54 |
33.15 |
120 |
40.73 |
20.79 |
27.36 |
LVdf soil #2; 106.96 (% of applied) |
|||
0 |
108.59 |
< LQ |
< LQ |
7 |
100.75 |
< LQ |
1.52 |
14 |
98.57 |
< LQ |
0.35 |
28 |
97.83 |
< LQ |
0.3 |
60 |
84.48 |
0.65 |
8.3 |
90 |
40.79 |
20.94 |
32.35 |
120 |
33.31 |
16.35 |
26.67 |
RQ soil #3; (% of applied) |
|||
0 |
100.49 |
< LQ |
< LQ |
7 |
97.54 |
1.12 |
< LQ |
14 |
86.58 |
2.06 |
< LQ |
28 |
52.68 |
19.78 |
0.17 |
60 |
32.45 |
23.29 |
0.92 |
90 |
26.28 |
28.77 |
0.84 |
120 |
21.01 |
16.73 |
2.08 |
PV soil #4; 106.96 (% of applied) |
|||
0 |
105.01 |
< LQ |
0.63 |
7 |
99.03 |
< LQ |
2.16 |
14 |
93.58 |
< LQ |
2.50 |
28 |
92.91 |
0.19 |
3.17 |
60 |
88.31 |
2.4 |
3.32 |
90 |
70.86 |
9.14 |
10.49 |
120 |
60.04 |
7.62 |
10.17 |
(1) Limit of quantification: 0.05 µg/mL
(2) Sum of all peaks accounting for < 10 % of the applied radioactivity.
Table 7. Parameters of the exponential first order kinetics model
Soil |
C0(1) (%) |
K (2) (/days) |
R2 |
DT50 (days) |
DT90 (days) |
#1 GM |
112.12 |
-0.0089 |
0.968 |
77.88 |
258.72 |
#2 LVdf |
117.25 |
-0.0101 |
0.9115 |
68.62 |
227.98 |
#3 RQ |
94.595 |
-0.0139 |
0.9409 |
49.87 |
165.65 |
#4 PV |
104.03 |
-0.0043 |
0.9478 |
161.19 |
535.48 |
(1) Maximum concentration in the soil.
(2)
Rate constant for soil dissipation of the molecule.
Table 3. Mass Balance Summary
(a) 0.01 mg/kg rate
DAT (a) |
Replicate |
% Extracted (b) |
% Unextracted (c) |
% Total (d) |
0 |
A |
103.3 |
4.6 |
107.9 |
B |
100.1 |
5.6 |
105.7 |
|
3 |
A |
89.3 |
15.8 |
105.2 |
B |
87.5 |
14.8 |
102.4 |
|
7 |
A |
80.9 |
20.0 |
101.1 |
B |
78.3 |
19.8 |
98.3 |
|
16 |
A |
75.5 |
28.2 |
103.8 |
B |
75.0 |
25.2 |
100.4 |
|
23 |
A |
67.6 |
31.6 |
99.4 |
B |
67.5 |
33.0 |
100.6 |
|
29 |
A |
64.0 |
33.0 |
97.2 |
B |
62.6 |
32.7 |
95.6 |
|
63 |
A |
57.2 |
42.3 |
100.0 |
B |
55.7 |
43.5 |
99.5 |
|
98 |
A |
56.9 |
45.0 |
102.4 |
B |
54.9 |
47.2 |
102 .6 |
|
150 |
A |
43.5 |
55.3 |
99.7 |
B |
44.8 |
48.8 |
94.5 |
(b) 0.02 mg/kg rate
DAT (a) |
Replicate |
% Extracted (b) |
% Unextracted (c) |
% Total (d) |
0 |
A |
100.6 |
2.9 |
103.6 |
B |
99.5 |
2.2 |
101.7 |
|
3 |
A |
92.5 |
8.6 |
101.2 |
B |
89.9 |
8.1 |
98.1 |
|
7 |
A |
83.2 |
14.8 |
98.2 |
B |
81.9 |
17.0 |
99.0 |
|
16 |
A |
75.3 |
23.2 |
98.6 |
B |
75.0 |
21.3 |
96.4 |
|
23 |
A |
68.7 |
27.4 |
96.3 |
B |
71.9 |
25.5 |
97.4 |
|
29 |
A |
71.5 |
26.9 |
98.6 |
B |
68.6 |
28.5 |
97.3 |
|
63 |
A |
57.5 |
41.9 |
99.8 |
B |
59.7 |
41.8 |
101.8 |
|
98 |
A |
65.1 |
35.5 |
100.9 |
B |
56.6 |
42.2 |
99.2 |
|
150 |
A |
43.9 |
50.4 |
95.0 |
B |
47.5 |
48.2 |
96.4 |
(c)0.055 mg/kg Rate
DAT (a) |
Replicate |
% Extracted (b) |
% Unextracted (c) |
% Total (d) |
0 |
A |
95.9 |
4.0 |
99.9 |
B |
96.7 |
3.0 |
99.7 |
|
3 |
A |
92.1 |
7.0 |
99.1 |
B |
92.5 |
7.9 |
100.4 |
|
7 |
A |
85.3 |
12.2 |
97.6 |
B |
90.3 |
10.9 |
101.2 |
|
16 |
A |
82.4 |
18.2 |
100.7 |
B |
83.3 |
16.8 |
100.2 |
|
23 |
A |
78.2 |
22.0 |
100.3 |
B |
81.0 |
18.8 |
99.9 |
|
29 |
A |
76.8 |
23.6 |
100.4 |
B |
77.8 |
22.2 |
100.1 |
|
63 |
A |
62.1 |
33.6 |
96.1 |
B |
68.7 |
29.9 |
98.8 |
|
98 |
A |
62.1 |
33.6 |
96.1 |
B |
65.4 |
32.0 |
97.6 |
|
150 |
A |
59.2 |
37.4 |
97.0 |
B |
61.6 |
38.2 |
100.3 |
|
210 |
A |
47.3 |
49.3 |
97.2 |
B |
52.9 |
45.9 |
99.2 |
(d) 0.1 mg/kg rate
DAT (a) |
Replicate |
% Extracted (b) |
% Unextracted (c) |
% Total (d) |
0 |
A |
97.1 |
3.6 |
100.7 |
B |
97.1 |
3.6 |
100.7 |
|
3 |
A |
92.4 |
6.2 |
98.6 |
B |
91.8 |
7.0 |
98.9 |
|
7 |
A |
88.0 |
10.4 |
98.4 |
B |
88.9 |
9.7 |
98.7 |
|
1 7 |
A |
82.3 |
17.3 |
99.7 |
B |
83.4 |
16.3 |
99.8 |
|
23 |
A |
79.4 |
18.0 |
97.5 |
B |
81.9 |
16.9 |
98.8 |
|
42 |
A |
74.1 |
24.5 |
98.6 |
B |
72.8 |
25.7 |
98.7 |
|
63 |
A |
67.4 |
30.0 |
97.6 |
13 |
76.2 |
30.5 |
106.9 |
|
98 |
A |
56.5 |
29.7 |
86.6 |
B |
62.8 |
34 .1 |
97.1 |
|
150 |
A |
57.9 |
39.2 |
97.7 |
B |
63.5 |
33.7 |
97.6 |
|
210 |
A |
52.3 |
43.3 |
96.0 |
B |
52.3 |
45.2 |
98.0 |
|
300 |
A |
40.0 |
54.7 |
95.4 |
B |
37.0 |
56.7 |
94.3 |
(e) 1mg/kg rate
DAT (a) |
Replicate |
% Extracted (b) |
% Unextracted (c) |
% Total (d) |
0 |
A |
94.9 |
9.4 |
104.4 |
B |
92.7 |
9.7 |
102.3 |
|
3 |
A |
91.7 |
10.4 |
102.1 |
B |
89.2 |
12.1 |
101.3 |
|
7 |
A |
92.8 |
9.6 |
102.4 |
B |
89.1 |
11.7 |
100.8 |
|
23 |
A |
89.6 |
15.1 |
104.8 |
B |
87.9 |
14.0 |
102.0 |
|
42 |
A |
82.4 |
18.6 |
101.1 |
B |
84.2 |
19.0 |
103.3 |
|
63 |
A |
78.5 |
21.7 |
100.3 |
B |
79.9 |
22.0 |
102.0 |
|
98 |
A |
80.9 |
22.4 |
103.4 |
B |
78.4 |
23.2 |
101.8 |
|
150 |
A |
73.6 |
26.0 |
99.8 |
B |
74.8 |
24.8 |
99.7 |
|
210 |
A |
69.2 |
30.6 |
100.0 |
B |
70.4 |
30.6 |
101.1 |
|
300 |
A |
60.7 |
40.0 |
100.9 |
B |
61.7 |
39.1 |
101.1 |
(a) Days after Treatment
(b) Combined ASE extracts quantified by LSC
(c) Extracted soil debris combusted and quantified by LSC
(d) Total recovery by radioactive recovery (include activities trapped in the 2M NaOH traps)
Table 4. Summary of the test substance in Soil
(a) 0.01 mg/kg rate
DAT |
Replicate |
% Test substance (a) |
|
0 |
A |
101.2 |
|
B |
98.7 |
||
3 |
A |
80.5 |
|
B |
78.3 |
||
7 |
A |
60.2 |
|
B |
59.1 |
||
16 |
A |
49.4 |
|
B |
51.4 |
||
23 (b) |
A |
52.0 |
|
B |
50.9 |
||
29 |
A |
48.3 |
|
B |
48.5 |
||
63 |
A |
47.5 |
|
B |
48.3 |
||
98 |
A |
39.9 |
|
B |
35.1 |
||
150 |
A |
24.8 |
|
B |
26.2 |
||
(a) As Determined by LC-MS-MS (as% of applied).
(b) Samples reanalysed.
(b) 0.02 mg/kg rate
DAT |
Replicate |
% Test substance (a) |
|
0 |
A |
106.7 |
|
B |
102 .5 |
||
3 |
A |
87.5 |
|
B |
86.1 |
||
7 |
A |
67.3 |
|
B |
63.7 |
||
16 |
A |
66.5 |
|
B |
65.9 |
||
23 (b) |
A |
61.0 |
|
B |
63.7 |
||
29 |
A |
44.6 |
|
B |
62.3 |
||
63 |
A |
50.3 |
|
B |
51.8 |
||
98 |
A |
55.5 |
|
B |
42.7 |
||
150 |
A |
26.5 |
|
B |
31.1 |
||
(a) As Determined by LC-MS-MS (as% of applied).
(b) Samples reanalysed.
(c) 0.055 mg/kg rate
DAT |
Replicate |
% Test substance (a) |
0 |
A |
93.1 |
B |
94.8 |
|
3 |
A |
85.4 |
B |
86.1 |
|
7 |
A |
76.1 |
B |
79.6 |
|
16 |
A |
68.3 |
B |
71.2 |
|
23 (b) |
A |
69.8 |
B |
73.9 |
|
29 |
A |
65.2 |
B |
66.8 |
|
63 |
A |
57.6 |
B |
61.2 |
|
98 |
A |
61.3 |
B |
61.7 |
|
150 |
A |
42.2 |
B |
45.4 |
|
210 |
A |
33.4 |
B |
38.8 |
(a) As Determined by LC-MS-MS (as% of applied).
(b) Samples reanalysed.
(d) 0.1 mg/kg rate
DAT |
Replicate |
% test substance (a) |
0 |
A |
93.8 |
B |
94.6 |
|
3 |
A |
85.9 |
B |
85.2 |
|
7 |
A |
68.1 |
B |
77.1 |
|
17 |
A |
73.9 |
B |
73.7 |
|
23 |
A |
73.2 |
B |
76.6 |
|
42 |
A |
67.5 |
B |
65.0 |
|
63 |
A |
56.5 |
B |
67.9 |
|
98 |
A |
45.6 |
B |
50.9 |
|
150 |
A |
44.6 |
B |
52.2 |
|
210 |
A |
42.8 |
B |
42.1 |
|
300 |
A |
23.8 |
B |
20.3 |
(a) Determined by TLC (0- 98 DAT) and LC-MS-MS (150-300 DAT) (as% of applied). The half-life calculation was based on the TLC and LC-MS-MS results shown in order to obtain the best kinetic fit (lowest R2).
Table 7. Degradation DT 50 Values for the test substance in Missouri Soil
Treatment Rate mg kg·1 |
Simple First Order (SFO) Model Outputs |
|
DT50 (Days) |
R2 |
|
0.01 |
78.3 |
0.6538 |
0.02 |
91.0 |
0.6889 |
0.055 |
167 |
0.8667 |
0.1 |
169 |
0.8965 |
1 |
444 |
0.9102 |
The additional parameter in the FOMC model was statistically significant at the 5% level:
Treatment Rate mg/kg |
First Order Multi- Compartment (FOMC) Model Outputs |
|
DT50 (Days) |
R2 |
|
0.01 |
22.8 |
0.9467 |
0.02 |
38.9 |
0.8888 |
0.055 |
161 |
0.9122 |
0.1 |
151 |
0.9039 |
1* |
444 |
0.9050 |
* The FOMC model was not found to be significant for the highest rate.
Table 2. Mass Balance Summary
Soil |
Sampling time (days) |
% of Applied Radioactivity in |
|||||||
Extracts |
CO2 |
Unextracted |
Total (1) |
||||||
Georgia |
0 |
96.6 |
NA |
2.8 |
99.4 |
||||
3 |
96.8 |
0.1 |
7.2 |
104.0 |
|||||
6 |
92.0 |
0.1 |
11.1 |
103.1 |
|||||
9 |
90.4 |
0.1 |
11.8 |
1022 |
|||||
20 |
84.2 |
0.3 |
21.0 |
105.4 |
|||||
28 |
80.8 |
0.1 |
25.1 |
105.9 |
|||||
64 |
72.2 |
0.1 |
23 4 |
95.7 |
|||||
150 |
59.9 |
0.1 |
40.6 |
100.6 |
|||||
365 |
40.9 |
0.1 |
62.3 |
103.3 |
|||||
Mean |
102.2 |
||||||||
North Carolina |
0 |
96.7 |
NA |
3.0 |
99.7 |
||||
3 |
92.1 |
0.1 |
6.9 |
99.1 |
|||||
6 |
89.5 |
0.1 |
8.7 |
98.2 |
|||||
9 |
88.2 |
0.1 |
7.4 |
95.6 |
|||||
20 |
83.0 |
0.2 |
15.6 |
98.7 |
|||||
28 |
79.2 |
0.1 |
17.6 |
96.8 |
|||||
64 |
71.0 |
0.1 |
21.8 |
92.9 |
|||||
|
150 |
60.5 |
0.2 |
35.5 |
96.1 |
||||
365 |
48.0 |
0.2 |
48.5 |
96.6 |
|||||
Mean |
97.3 |
||||||||
North Dakota |
0 |
94.4 |
NA |
6.6 |
101.0 |
||||
3 |
81.6 |
0.1 |
12.6 |
94.2 |
|||||
6 |
81.4 |
0.1 |
15.7 |
97.2 |
|||||
9 |
82.6 |
0.0 |
18.4 |
100.9 |
|||||
20 |
75.9 |
0.2 |
23.2 |
99.3 |
|||||
28 |
68.3 |
0.2 |
25.1 |
93.5 |
|||||
64 |
59.2 |
0.3 |
31.2 |
90.7 |
|||||
150 |
56.4 |
0.6 |
38.5 |
95.5 |
|||||
365 |
42.8 |
1.1 |
55.2 |
99.1 |
|||||
Mean |
96.6 |
||||||||
Illinois |
0 |
97.5 |
NA |
3.8 |
101.3 |
||||
3 |
82.7 |
0.1 |
9.0 |
91.7 |
|||||
6 (2) |
- |
- |
- |
- |
|||||
9 |
86.6 |
0.1 |
11.9 |
98.6 |
|||||
20 |
73.8 |
0.1 |
20.4 |
94.3 |
|||||
28 |
72.4 |
0.1 |
20.6 |
93.1 |
|||||
64 |
67.4 |
0.1 |
28.5 |
96.0 |
|||||
150 |
51.8 |
0,2 |
38.3 |
90.3 |
|||||
365 |
28.1 |
0.4 |
55.7 |
84.2 |
|||||
Mean |
93.4 |
||||||||
Arkansas |
0 |
99.1 |
NA |
1.3 |
100.4 |
||||
3 |
79.3 |
0.1 |
12.4 |
91.7 |
|||||
6 |
80.8 |
0.1 |
15.7 |
96.5 |
|||||
9 |
78.4 |
0.1 |
13.2 |
91.7 |
|||||
20 |
77.6 |
0.1 |
17.5 |
95.2 |
|||||
28 |
62.2 |
0.1 |
24.1 |
86.3 |
|||||
64 |
55.6 |
0.1 |
34.7 |
90.4 |
|||||
150 |
43.4 |
0.2 |
42.4 |
85.9 |
|||||
365 |
16.9 |
0.4 |
77.2 |
94.5 |
|||||
Mean |
92.5 |
||||||||
Overall mean |
96.4 |
||||||||
NA: not applicable
(1) Slight discrepancies in totals are due to using unrounded values in the spreadsheet
(2) Due to an error with the application, the 6 DAT samples for Illinois were not applied to.
Table 3. Recovery of 14C-labelled test substance (as % Applied Radioactivity)
Sampling time (days) |
Georgia |
North Carolina |
North Dakota |
Illinois |
Arkansas |
0 |
95.9 |
96.1 |
93.7 |
96.9 |
97.0 |
3 |
95.2 |
90.8 |
77.3 |
79.1 |
77.2 |
6 |
88.2 |
87.3 |
75.6 |
N/A |
76.3 |
9 |
87.3 |
83.1 |
72.9 |
78.7 |
74.4 |
20 |
80.7 |
79.3 |
71.4 |
67.1 |
70.4 |
28 |
78.6 |
76.5 |
60.9 |
65.8 |
53.7 |
64 |
69.3 |
69.1 |
55.8 |
58.9 |
46.4 |
150 |
58.3 |
58.5 |
54.3 |
41.3 |
38.4 |
365 |
38.7 |
44.3 |
42.5 |
14.9 |
11.3 |
NIA - Due to an error with the appllcation, the 6 DAT samples for Illinois were not applied to.
Table 4. Distribution of Extracted Radioactivity - Georgia Soil (Mobile phase conditions 2)(as % of applied radioactivity)
Sampling time (days) |
% test substance |
% M3 |
% Other compponents |
0(A) |
101.6 |
nd |
nd |
0 (B) |
90.1 |
nd |
nd |
Mean |
95.9 |
nd |
nd |
3 (A) |
94.2 |
nd |
nd |
3 (B) |
96.1 |
nd |
nd |
Mean |
95.2 |
nd |
nd |
6 (A) |
86.8 |
nd |
nd |
6 (B) |
88.2 |
nd |
0.3 (1) |
Mean |
86.4 |
nd |
0.2 (1) |
9(A) |
86.4 |
nd |
nd |
9(B) |
88.2 |
nd |
nd |
Mean |
87.3 |
nd |
nd |
20 (A) |
80.1 |
nd |
0.8 (2) |
20 (B) |
81.3 |
nd |
nd |
Mean |
80.7 |
nd |
0.3 (1) |
28 (A) |
77.5 |
nd |
nd |
28 (B) |
79.6 |
nd |
nd |
Mean |
78.6 |
nd |
nd |
64 (A) |
67.9 |
NIA |
NIA |
64 (B) |
70.6 |
nd |
0.7 (1) |
Mean |
69.3 |
nd |
0.7 (1) |
150 (A) |
62.3 |
nd |
nd |
150 (B) |
54.3 |
nd |
nd |
Mean |
58.5 |
nd |
nd |
365 (A) |
38.3 |
1.5 |
nd |
365 (B) |
39 |
1.4 |
nd |
Mean |
38.7 |
1.5 |
nd |
(A)+ (B) replicate samples.
Total % of unidentified discrete components. The value in () denotes the maximum number of components.
N/A Metabolite data not included in the results since the HPLC system had not reached equilibrium.
nd - Not detected
Table 5. Distribution of Extracted Radioactivity - North Carolina Soil (Mobile phase conditions 2)
Sampling time (days) |
% test substance |
% M3 |
% Other compponents |
0(A) |
95.7 |
nd |
nd |
0 (B) |
96.4 |
nd |
nd |
Mean |
96.1 |
nd |
nd |
3 (A) |
91.4 |
nd |
nd |
3 (B) |
90.2 |
nd |
0.4 (1) |
Mean |
90.8 |
nd |
0.2 (1) |
6 (A) |
84.8 |
0.8 |
nd |
6 (B) |
89.7 |
nd |
nd |
Mean |
87.3 |
0.4 |
nd |
9(A) |
81.7 |
1.5 |
0.4 (1) |
9(B) |
84.5 |
1.1 |
nd |
Mean |
83.1 |
1.3 |
0.2 (1) |
20 (A) |
79.7 |
1.2 |
0.2 (1) |
20 (B) |
78.8 |
0.8 |
0.4 (1) |
Mean |
79.3 |
1.0 |
0.3 (1) |
28 (A) |
77.5 |
nd |
nd |
28 (B) |
77.5 |
1.4 |
nd |
Mean |
76.5 |
0.7 |
nd |
64 (A) |
NIA |
NIA |
NIA |
64 (B) |
69.1 |
1.4 |
0.4 (1) |
Mean |
69.1 |
1.4 |
0.4 (1) |
150 (A) |
58.8 |
1.1 |
nd |
150 (B) |
58.1 |
2 |
nd |
Mean |
58.5 |
1.6 |
nd |
365 (A) |
44.8 |
3.8 |
nd |
365 (B) |
43.8 |
2.6 |
nd |
Mean |
44.3 |
3.2 |
nd |
(A)+ (B) replicate samples.
Total % of unidentified discrete components. The value in () denotes the maximum number of components.
N/A Metabolite data not included in the results since the HPLC system had not reached equilibrium.
nd - Not detected
Table 6. Distribution of Extracted Radioactivity - North Dakota Soil (Mobile phase conditions 2) (as % of applied radioactivity)
Sampling time (days) |
% test substance |
% M3 |
% Other compponents |
0(A) |
93.1 |
nd |
nd |
0 (B) |
94.3 |
nd |
nd |
Mean |
93.7 |
nd |
nd |
3 (A) |
78.6 |
0.6 |
1.2 (1) |
3 (B) |
75.9 |
1.7 |
1.8 (2) |
Mean |
77.3 |
1.2 |
1.5 (3) |
6 (A) |
74.3 |
2.0 |
1.3 (1) |
6 (B) |
76.8 |
1.6 |
1.3 (1) |
Mean |
75.6 |
1.8 |
1.3 (1) |
9(A) |
73.0 |
4.1 |
2.2 (2) |
9(B) |
72.8 |
3.6 |
2.1 (1) |
Mean |
72.9 |
3.9 |
2.2 (2) |
20 (A) |
71.4 |
1.4 |
0.6 (1) |
20 (B) |
NIA |
NIA |
NIA |
Mean |
71.4 |
1.4 |
0.6 (1) |
28 (A) |
60.4 |
3.9 |
1.9 (2) |
28 (B) |
61.3 |
2.8 |
1.4 (2) |
Mean |
60.9 |
3.4 |
1.7 (2) |
64 (A) |
55.4 |
0.9 |
nd |
64 (B) |
56.2 |
1.1 |
nd |
Mean |
55.8 |
1.0 |
nd |
150 (A) |
51.9 |
2.4 |
nd |
150 (B) |
56.7 |
1.3 |
nd |
Mean |
55.3 |
1.9 |
nd |
365 (A) |
41.1 |
nd |
nd |
365 (B) |
43.8 |
nd |
nd |
Mean |
42.5 |
nd |
nd |
(A)+ (B) replicate samples.
Total % of unidentified discrete components. The value in () denotes the maximum number of components.
N/A Metabolite data not included in the results since the HPLC system had not reached equilibrium.
nd - Not detected
Table 7. Distribution of Extracted Radioactivity - Illinois Soil (Mobile phase conditions 2) (as % of applied radioactivity)
Sampling time (days) |
% test substance |
% M3 |
% Other compponents |
0(A) |
97.0 |
nd |
nd |
0 (B) |
96.7 |
nd |
nd |
Mean |
96.9 |
nd |
nd |
3 (A) |
79.6 |
1.8 |
0.4 (1) |
3 (B) |
78.6 |
0.6 |
0.7 (1) |
Mean |
79.1 |
1.2 |
0.6 (1) |
6 (A) |
NIA |
NIA |
NIA |
6 (B) |
NIA |
NIA |
NIA |
Mean |
NIA |
NIA |
NIA |
9(A) |
NIA |
NIA |
NIA |
9(B) |
78.7 |
4.1 |
0.8 (2) |
Mean |
78.7 |
4.1 |
0.8 (2) |
20 (A) |
69.0 |
2.2 |
nd |
20 (B) |
65.2 |
3.6 |
0.9 (1) |
Mean |
67.1 |
2.9 |
0.4 (1) |
28 (A) |
65.8 |
4.0 |
1.1 (2) |
28 (B) |
NIA |
NIA |
NIA |
Mean |
65.8 |
4.0 |
1.1 (2) |
64 (A) |
60.8 |
3.6 |
1.8 (2) |
64 (B) |
57.0 |
5.5 |
2.4 (2) |
Mean |
58.9 |
4.6 |
2.1 (2) |
150 (A) |
41.3 |
6.5 |
2.4 (1) |
150 (B) |
33.5 |
16.8 |
1.8 (1) |
Mean |
41.3 |
6.5 |
2.4 (1) |
365 (A) |
15.3 |
11.8 |
nd |
365 (B) |
14.4 |
11.2 |
1.4 (1) |
Mean |
14.9 |
11.5 |
1.4 (1) |
(A)+ (B) replicate samples.
Total % of unidentified discrete components. The value in () denotes the maximum number of components.
N/A Metabolite data not included in the results since the HPLC system had not reached equilibrium.
nd - Not detected
Table 8. Distribution of Extracted Radioactivity - Arkansas Soil (Mobile phase conditions 2) (as % of applied radioactivity)
Sampling time (days) |
% test substance |
% M3 |
% Other compponents |
0(A) |
100.9 |
nd |
nd |
0 (B) |
93.0 |
nd |
nd |
Mean |
97.0 |
nd |
nd |
3 (A) |
75.2 |
nd |
0.7 (2) |
3 (B) |
79.1 |
nd |
0.3 (1) |
Mean |
77.2 |
nd |
0.5 (2) |
6 (A) |
77.8 |
1.2 |
0.3 (1) |
6 (B) |
74.8 |
1.2 |
1.4 (1) |
Mean |
76.3 |
1.2 |
0.9 (1) |
9(A) |
72.8 |
2.1 |
0.4 (1) |
9(B) |
75.9 |
2.7 |
nd |
Mean |
74.4 |
1.1 |
0.2 (1) |
20 (A) |
72.8 |
3.8 |
0.7 (2) |
20 (B) |
67.9 |
4.2 |
nd |
Mean |
70.4 |
4.0 |
0.4 (2) |
28 (A) |
53.4 |
6.3 |
1.7 (1) |
28 (B) |
53.9 |
3.9 |
2.2 (5) |
Mean |
53.7 |
5.1 |
2.0 (5) |
64 (A) |
46.5 |
5.6 |
0.2 (1) |
64 (B) |
46.3 |
7.7 |
0.5 (1) |
Mean |
46.4 |
6.7 |
0.4 (1) |
150 (A) |
45.8 |
2.2 |
nd |
150 (B) |
30.9 |
6.4 |
nd |
Mean |
38.4 |
4.3 |
nd |
365 (A) |
9.9 |
7.4 |
nd |
365 (B) |
12.6 |
3.9 |
nd |
Mean |
11.3 |
5.7 |
nd |
(A)+ (B) replicate samples.
Total % of unidentified discrete components. The value in () denotes the maximum number of components.
N/A Metabolite data not included in the results since the HPLC system had not reached equilibrium.
nd - Not detected
Table 9. Summary of Unextracted Residue Fractionation
Soil (DAT) |
Replicate |
Total % in Post extraction soilds (PES) |
% pf Applied Radioactivity in : |
|||
Humin Fraction (a) |
Fulvia Fraction (b) |
Humin actid fraction (c) |
Loss/Gain |
|||
Arkansas (356DAT) |
A |
74.0 |
27.8 |
38.1 |
5.8 |
-2.3 |
B |
33.2 |
33.9 |
5.5 |
-1.4 |
||
Mean |
30.5 |
36.0 |
5.7 |
|
||
(a). Humm Fraction is that which is not solubilised by NaOH (1.e. PES after 0.1 M and 1 M NaOH extraction step)
(b). Fulvic Fraction is that which remains in solution after acidification of the NaOH extract
(c). Humic Acid Fraction is that which is precipitated out by addition of HCl
Table 10. Degradation DT50 and DT90 Values for Cyproconazole using Simple First Order Model
Soil |
Simple First Order (SFO) Model Outputs |
||
DT50 (Days) |
DT90 (Days) |
R2 |
|
Georgia |
191 |
635 (1) |
0.72 |
North Carolina |
216 |
718 (1) |
0.49 |
North Dakota |
142 |
472 (1) |
-0.80 |
Illinois |
79 |
261 |
0.75 |
Arkansas |
52 |
172 |
0.64 |
(1). Values extrapolated beyond the study period should only be considered indicative
Note - Values are forced fit through 100% at zero time.
Description of key information
All available data was assessed and the aerobic studies following standard test guidelines are included here in a weight of evidence approach. The other studies are anaerobic studies and included as supporting information.
Geometric mean DT50 in soil = 117 d, 20 °C, BBA Part IV 4 -1, EPA 162 - 1 and (or) OECD TG 207, Glänzel 1994, Glänzel & Wission 1994, Wission 1992, Oliver & Edwards 2007, Barizon 2007 and Kuet 2006.
Key value for chemical safety assessment
- Half-life in soil:
- 117 d
- at the temperature of:
- 20 °C
Additional information
Nine studies are available for this endpoint. Six of them followed standard guidelines and complied with GLP (Reliability 1). The effect values from all six studies were used in a weight-of-evidence approach for the CSA. The test conditions and key effect values are summarised in the table below.
Table 1. DT50 values for the substance in soil under aerobic standard test conditions.
USDA/Name/ Origin | OC (%)/pH (water) | T (˚C) / Moisture (%) | DT50 (d) (-Kinetic model in original report) | Author/ year |
Sand / Georgia / US | 0.35 / 6.1 | 20 / pF2 | 191 - SFO | Oliver & Edwards, 2007
|
Sandy loam / North Carolina / US | 0.70 / 5.7 | 20 / pF2 | 216 - SFO | Oliver & Edwards, 2007 |
Sandy clay loam / North Dakota / US | 2.09 / 6.5 | 20 / pF2 | 142 - SFO | Oliver & Edwards, 2007 |
Silty clay / Illinois / US | 2.44 / 8.0 | 20 / pF2 | 79 - SFO | Oliver & Edwards, 2007 |
Silt loam / Arkansas / US | 1.51 / 4.7 | 20 / pF2 | 52 - SFO | Oliver & Edwards, 2007 |
Clay / Gleissolo / BR | 9.6 / 4.6 | 19 - 21 / 40 % MWHC | 77.9 - SFO | Barizon, 2007
|
Clay / Latossolo / BR | 2.1 / 5.0 | 19 - 21 / 40 % MWHC | 68.6 - SFO | Barizon, 2007
|
Loamy sand / Neossolo / BR | 0.8 / 4.7 | 19 - 21 / 40 % MWHC | 49.9 - SFO | Barizon, 2007
|
Clay / Argissolo / BR | 4.0 / 6.0 | 19 - 21 / 40 % MWHC | 161 - SFO | Barizon, 2007
|
Silt loam / Missouri / US | 1.5 / 6.1 | 20 / pF2 | 78.3 - SFO | Kuet, 2006 |
Silt loam / Missouri / US | 1.5 / 6.1 | 20 / pF2 | 91.0 - SFO | Kuet, 2006
|
Silt loam / Missouri / US | 1.5 / 6.1 | 20 / pF2 | 161 - SFO | Kuet, 2006
|
Silt loam / Missouri / US | 1.5 / 6.1 | 20 / pF2 | 169 - SFO | Kuet, 2006
|
Silt loam / Missouri / US | 1.5 / 6.1 | 20 / pF2 | 444 - SFO | Kuet, 2006
|
Sandy clay loam / Flaach / CH | 1.1 / 7.2 | 20 / 40 % MWHC | 148 - SFO | Glänzel, 1994 |
Sandy clay loam / Flaach / CH | 1.1 / 7.2 | 20 / 40 % MWHC | 97 – SFO | Glänzel & Wisson, 1994 |
Sandy clay loam / Flaach / CH | 1.1 / 7.0 | 22 / 75 % 0.33 bar | 104 – SFO | Wisson, 1992 |
Silt loam / Louisiana / US | 2.1 / 4.3 | 22 / 75 % 0.33 bar | 124 - SFO | Wisson, 1992 |
Geomean |
|
| 117 days |
|
Three anaerobic studies are included as supporting information. They followed EPA guidelines 162-2 or 162-3 with acceptable restrictions (Reliability 2). The test conditions and effect values are summarised in the table below.
Table 2. DT50 values for the substance in soil under anaerobic standard test conditions.
USDA/ Name/ Origin | OC (%)/ pH (water) | T. (°C) / Moisture | DT50 (d) - Kinetic model in original report | DT50 (d) - Kinetic model recalculated | Author/ Year |
Sandy loam/ Harnett County/ US | 0.5 / 5.4 Water = pond water | 24 / - | > 1 year | - | Blumhorst 1995 |
Loamy sand/ North Carolina/ US |
| 25 / 75 % of 0.33 bar for 30 days | 303 days – first order | - | Tong 1990 |
Loam/ Flaach/ CH | 2.3 / 7.6 | 22 / 40 % MWHC for 33 days | ~ 130 days – first order | - | Wisson 1990 |
Field studies - additional information
A large number of soil dissipation studies were conducted with the test material. As information on soil dissipation is outside the scope of REACH, the studies are not summarised as an endpoint study entry but briefly discussed here. The test material was tested for different applications in different crops and bare grounds with a rate ranging between 44.8 g/ha (equivalent to 0.04 lb/acre) - 321.2 g/ha (equivalent to 130 g/acre). The DT50 as calculated with SFO model were between 21 - 416 days. The geometric mean DT50 (n = 20) for soil dissipation was calculated to be 132 days (Dharmasri & Newcombe 2010; Gasser 1997; Bass 1994; Bourry 1988; Ali 1992a; Ali 1992b; Ali 1990a; Ali 1990b; Cahill & Bade 1988; Jimenez & Bade 1990; Hertl 1992; Bourry 1991a; Bourry 1991b; Bourry 1991c; Bourry 1991d; Hertl & Vogler 1993; Harvey 2009).
Information on Registered Substances comes from registration dossiers which have been assigned a registration number. The assignment of a registration number does however not guarantee that the information in the dossier is correct or that the dossier is compliant with Regulation (EC) No 1907/2006 (the REACH Regulation). This information has not been reviewed or verified by the Agency or any other authority. The content is subject to change without prior notice.
Reproduction or further distribution of this information may be subject to copyright protection. Use of the information without obtaining the permission from the owner(s) of the respective information might violate the rights of the owner.
