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Environmental fate & pathways

Biodegradation in soil

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Reference
Endpoint:
biodegradation in soil: simulation testing
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2009-03-24 - 2009-12-01
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 307 (Aerobic and Anaerobic Transformation in Soil)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Test type:
laboratory
Specific details on test material used for the study:
Chemical name: N-[(3(5)-Methyl-1H-pyrazol-1-yl)methyl]acetamide
Molecular weight: 153.2 g/mol
Molecular formula: C7H^11N3O
Purity: 99.9%
Radiolabelling:
no
Oxygen conditions:
aerobic
Soil classification:
DIN 19863 (Deutsche Industrie-Norm)
Soil no.:
#1
Soil type:
other: silty sand/sand
% Org. C:
0.74
pH:
5.1
CEC:
4 meq/100 g soil d.w.
Soil no.:
#2
Soil type:
loamy sand
% Org. C:
2.09
pH:
5.5
CEC:
10 meq/100 g soil d.w.
Soil no.:
#3
Soil type:
other: silty sand
% Org. C:
0.97
pH:
6.6
CEC:
9 meq/100 g soil d.w.
Details on soil characteristics:
LUFA 2.1: July 15, 2009 (batch no.: F2.1 2909, SGS INSTITUT FRESENIUS GmbH identification: 010/9378965), depth of soil collection was down to 20 cm), Germany, Rheinland- Pfalz, Dudenhofen, Am Hanhofer Weg links, Nr. 3336/58, location of soil collection of no agricultural use, agricultural plant protection products not used for the sampling year and 4 former years, fertilization performed in 2007: 600 kg/ha CaO + 250 kg/ha MgO.
Soil type: silty sand (DIN) or sand (USDA).

LUFA 2.2: July 15, 2009 (batch no.: F2.2 2909, SGS INSTITUT FRESENIUS GmbH identification: 010/9378966), depth of soil collection was down to 20 cm), Germany, Rheinland- Pfalz, Hanhofen, Großer Striet, Nr. 585, meadow, agricultural plant protection products not used for the sampling year and 4 former years, fertilization performed in 2007: 600 kg/ha CaO + 250 kg/ha MgO.
Soil type: loamy sand (DIN and USDA)

LUFA 2.3: July 16, 2009 (batch no.: F2.3 2909, SGS INSTITUT FRESENIUS GmbH identification: 010/9378967), depth of soil collection was down to 20 cm), Germany, Rheinland- Pfalz, Offenbach, Itn Bildgarten Nr. 507, fallow soil and pumpkin, agricultural plant protection products not used for the sampling year and 4 former years, fertilization performed in 2005 (300 kg/ha): 12 % N, 12 % P205, 17 % K20.
Soil type: silty sand (DIN) or sandy loam (USDA).



Soil No.:
#1
Duration:
29 d
Soil No.:
#2
Duration:
29 d
Soil No.:
#3
Duration:
29 d
Soil No.:
#1
Initial conc.:
0.48 mg/kg soil d.w.
Based on:
test mat.
Soil No.:
#2
Initial conc.:
0.48 mg/kg soil d.w.
Based on:
test mat.
Soil No.:
#3
Initial conc.:
0.48 mg/kg soil d.w.
Based on:
test mat.
Parameter followed for biodegradation estimation:
test mat. analysis
Soil No.:
#1
Temp.:
20°C
Humidity:
45 % MWHC
Microbial biomass:
22 mg C/100 g soil
Soil No.:
#2
Temp.:
20°C
Humidity:
45 %MWHC
Microbial biomass:
34 mg C/100 g soil
Soil No.:
#3
Temp.:
20°C
Humidity:
45 % MWHC
Microbial biomass:
14 mg C/100 g soil
Details on experimental conditions:
Amounts of conditioned soil at 45 % MWHC equivalent to 100 g dry weight were bottled into 1000 mL all glass metabolism flasks (conical shoulder bottles of 10 cm inner diameter) and maintained under a dynamic atmosphere of air, in the dark, at 20 °C ± 2 °C (see experimental set-up, Section: 7.3). Air entering the system was passed through a washing bottle to reduce the water loss of soil. The metabolism flasks were connected via tubing. A flow meter was used to measure on a weekly basis the air at a flow rate of about 15 to 30 mL/min.
The application of the test item was made to the soil surface,
Each test vessel was allocated a number that was used internally during the experimental phase.
Fortification
The soil (equivalent to 100 g dry weight) was treated at 0.48 mg a.s./kg soil (dry weight).
The test item (see section 8) was applied in 192 ^iL (249.8 pg/mL) of a mixture of methanol/water (10/90; v/v) onto the surface of the soil. The volume of methanol added to 100 g of dry soil did not exceed 0.1 vol. % (w/w). The treated soils were thoroughly mixed, by manual shaking, to incorporate the test item into soil. Following fortification the specimens were incubated for the appropriate amount of time under the conditions previously described.
The application solution (one at the beginning of dosing and one at the completion of dosing) was analyzed each in triplicate by LC-MS/MS to determine the measured concentration of N-[(3(5)- Methyl-lH-pyrazol-l-yl)methyl]acetamide (P 70/05). The recovery range accounted for 97 - 98 % of the theoretic value, based on the primary mass transition m/z 154 —> m/z 83.
The homogeneity of the application solution and the stability of the test item were reflected by the calculated standard deviation of N-[(3(5)-Methyl-lH-pyrazol-l-yl)methyl]acetamide (P 70/05) concentrations found (mean value of 97.3 % and RSD of 0.5 %).
For determination of the microbial activity at the start and end of the experiments (J. P. E. Anderson and K. H. Domsch, "A physiological method for the quantitative measurement of microbial biomass in soils'1), no test item was used.
7.5 Incubation
The test vessels were placed in a constant temperature room at 20 °C, with a temperature control of ± 2 °C and incubated in the dark. The temperature was recorded. Specimens were weighed approximately every two weeks to ensure that the soil moisture is being maintained. If the soil moisture e.g. dropped more than 10 % below this value (in weight equivalents), reagent water was added until the desired moisture level was obtained.
Sampling
Soil specimens were assayed at zero-time (initial value), 1,3, 7, 13 and 29 days after fortification. At each sampling interval replicate specimens were taken and assayed for the test item (P 70/05) and 3- Methylpyrazole.
The validity of the analytical method was proven for N-[(3(5)-Methyl~lH-pyrazol-l-yl)methyl]- acetamide (P 70/05) and 3-Methylpyrazole within each specimen sequence, using two freshly prepared adequate fortification values for each soil type.
Method Validation
Prior to the start of the N-[(3(5)-Methyl-lH-pyrazol-l-yl)methyl]acetamide (P 70/05) investigations in soil, the analytical method was established and validated by analysis of N-[(3(5)-Methyl-lH- pyrazol-l-yl)methyl]acetamide (P 70/05) and 3-Methylpyrazole (possible degradate of P 70/05) applied to and extracted from the experimental soils (see Section 9).
Soil blanks (100 g dry weight) conditioned at 45 % MWHC were fortified each with approximately 0.48 mg and 0.01 mg N-[(3(5)-Methyl-lH-pyrazol-l-yl)metliyl]acetamide (P 70/05) per kilogram dry soil equivalent to 100 and 2 % of the target rate.
Soil blanks (100 g dry weight) conditioned at 45 % MWHC were fortified each with approximately 0.26 mg and 0.005 mg 3-Methylpyrazole per kilogram dry soil.
The extraction and analytical method is considered valid since the following criteria for each analyte are fulfilled:
the blank values were not higher than 30 % of the LOQ (LOQ equivalent to the lowest fortification level)
the mean recovery at each fortification level and for each soil matrix is in the range 70-110 %
the relative standard deviation for each fortification level and for each soil matrix is less or equal 20 %
the overall relative standard deviation for each soil matrix and analyte is less or equal 20 %
Specimen Processing
The conditioned soil (45 % MWHC equivalent to 100 g dry soil) was placed in a 250 mL centrifuge bottle (method validation/laboratory concurrent recovery specimens: fortification at this point). 150 mL of the extraction solvent methanol/water/formic acid (50/50/0.1; v/v/v) were added. For specimens derived from the soil degradation study, the soil was quantitatively transferred into the centrifuge vessel using portions of the 150 mL volume of the extraction solvent.
The specimen was shaken for several seconds by hand, subjected to ultrasonic agitation for 10 minutes and finally subjected to a shaking machine (30 min at 270 strokes/min). After centrifugation for 5 minutes at 4000 rpm the supernatant was filtered over glass wool into a 250 mL volumetric flask. The extraction procedure was repeated two times using portions of 50 mL of the extraction solvent. The supernatants on a specimen basis were combined. The volumetric flask was filled up to the mark using the extraction solvent (final volume: 250 mL).
Further dilutions of the specimen extract were performed by the addition of methanol/water/formic acid (10/90/0.1; v/v/v). The final specimen extracts were subjected to LC-MS/MS analysis.
Key result
Soil No.:
#1
DT50:
2.31 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Key result
Soil No.:
#2
DT50:
0.7 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Key result
Soil No.:
#3
DT50:
1.05 d
Type:
(pseudo-)first order (= half-life)
Temp.:
20 °C
Transformation products:
yes
No.:
#1
Evaporation of parent compound:
not specified
Volatile metabolites:
not specified
Residues:
not specified
Conclusions:
The degradation of N-[(3(5)-Methyl-1H-pyrazol-1-yl)methyl]acetamide (P 70/05) was investigated under aerobic conditions at 20 °C. Three German soils of different characteristics were used for the experiment (LUFA 2.1 - silty sand/sand, LUFA 2.2 - loamy sand and LUFA 2.3 - silty sand/sandy loam). The formation of 3-Methylpyrazole, a possible degradate of P 70/05, was monitored.
N-[(3(5)-Methyl-1H-pyrazol-1-yl)methyl]acetamide (P 70/05) degraded fastly in the soil test systems.
The DT-50 values of P 70/05 in the soils LUFA 2.1, LUFA 2.2 and LUFA 2.3 soil were 2.31, 0.70 and 1.05 days.
The DT-90 values of P 70/05 in the soils LUFA 2.1, LUFA 2.2 and LUFA 2.3 soil were 7.68, 2.32 and 3.31 days.
Within the course of the P 70/05 degradation, the original isomers ratio of P 70/05 changed.
The formation of 3-Methylpyrazole reached a maximum of 0.1331 mg/kg on day-3 in soil LUFA 2.3. On day-29 the analysed value for 3-Methylparazole accounted for 0.0158 and 0.0193 mg/kg in soil LUFA 2.3.
Executive summary:

N-[(3(5)-Methyl-lH-pyrazol-l-yl)methyl]acetamide (P 70/05) was tested in aerobic soil on the basis of OECD/OCDE Guideline no.307 for the testing of chemicals, adopted April 24, 2002. The study was performed in the laboratory using analytical grade N-[(3(5)-Methyl-lH-pyrazol-l- yl)methyl]acetamide (purity of 99.9 %). The degradation of N-[(3(5)-Methyl-lH-pyrazol-l- yl)methyl]acetamide (P 70/05) was investigated under aerobic conditions at 20 °C.The formation of 3-Methylpyrazole, a possible degradate ofP 70/05, was monitored.Three German soils of different characteristics were used for the experiment (see Section 12.4).The test concentration of 0.48 mg/kg dry soil was based on a penetration depth of the test item in the soil of 10 cm, a soil bulk density of 1 g/cm3and on a total use of 480 g a.s./ha/year (0.2 % active substance based on application of total mass of 240 kg fertilizer N present as ammonium nitrogen and urea nitrogen).The soil systems were acclimatized under a dynamic atmosphere of air to maintain aerobic conditions. The test period consisted of sampling intervals at zero time, 1, 3,7, 13 and 29 days.The recoveries of N-[(3(5)-Methyl-lH-pyrazol-l-yl)methyl]acetamide (P 70/05) for the initial time specimens of the aerobic soil degradation experiment ranged from 92 to 95 % of the applied test item.For the description of the disappearance time of N-[(3(5)-Methyl-lH-pyrazol-l-yl)methyl]acetamide (P 70/05), a 1storder kinetic was chosen. N-[(3(5)-Methyl-lH-pyrazol-l-yl)methyl]acetamide (P 70/05) degraded fastly in the soil test systems.The DT-50 values of P 70/05 in the soils LUFA 2.1, LUFA 2.2 and LUFA 2.3 soil were 2.31, 0.70 and 1.05 days.The DT-90 values of P 70/05 in the soils LUFA 2.1, LUFA 2.2 and LUFA 2.3 soil were 7.68, 2.32 and 3.31 days.Within the course of the P 70/05 degradation, the original isomers ratio of P 70/05 changed.

The formation of 3-Methylpyrazole reached a maximum of 0.1331 mg/kg on day-3 in soil LUFA 2.3. This value is equivalent to 52 % of the possible value of 3-Methylpyrazole formed by degradation of the applied amount of N-[(3(5)-Methyl-lH-pyrazol-l-yl)methyl]acetamide (P 70/05)to the soil. On day-29 the analysed value for 3-Metliylparazole accounted for 0.0158 and 0.0193 mg/kg in soil LUFA 2.3.

Description of key information

The degradation of N-[(3(5)-Methyl-1H-pyrazol-1-yl)methyl]acetamide (P 70/05) was investigated under aerobic conditions at 20 °C. Three German soils of different characteristics were used for the experiment (LUFA 2.1 - silty sand/sand, LUFA 2.2 - loamy sand and LUFA 2.3 - silty sand/sandy loam). The formation of 3-Methylpyrazole, a possible degradate of P 70/05, was monitored.

N-[(3(5)-Methyl-1H-pyrazol-1-yl)methyl]acetamide (P 70/05) degraded fastly in the soil test systems.

The DT-50 values of P70/05 in the soils LUFA 2.1, LUFA 2.2 and LUFA 2.3 soil were 2.31, 0.70 and 1.05 days.

The DT-90 values of P 70/05 in the soils LUFA 2.1, LUFA 2.2 and LUFA 2.3 soil were 7.68, 2.32 and 3.31 days.

Within the course of the P 70/05 degradation, the original isomers ratio of P 70/05 changed.

Key value for chemical safety assessment

Half-life in soil:
2.31 d
at the temperature of:
20 °C

Additional information