ethyl 5,5-diphenyl-2-isoxazoline-3-carboxylate

Toxicity to soil microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to soil microorganisms

Type of information:

experimental study

Adequacy of study:

key study

Study period:

14 Jan - 12 Feb 1999

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

BBA Part VI, 1-1

Version / remarks:

adopted 1990 (2nd edition)

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

yes

Details on preparation and application of test substrate:

AMENDMENT OF SOIL
- Type of organic substrate: D(+)-Glucose (monohydrate) for biochemistry and microbiology from MERCK, Darmstadt, Germany

APPLICATION OF TEST SUBSTANCE TO SOIL
- Method: test substance dissolved in acetone was mixed with quartz sand, the sand mixture was added to soil

VEHICLE:
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Evaporation of vehicle before use: no data

Test organisms (inoculum):

soil

Total exposure duration:

28 d

Test temperature:

20 ± 2 °C

Moisture:

Max. water holding capacity adjusted to 45%

Details on test conditions:

TEST SYSTEM
- Testing facility: Hoechst Schering AgrEvo GmbH, Department of Environmental Sciences, Ecobiology, Frankfurt, Germany
- Test container (type, material, size): 2L container Nova Crominal, stainless steel, diameter 16 cm, height 10 cm
- Amount of soil: 800 g divided into 3 replicates of 260 g soil each
- No. of replicates per concentration: 3
- No. of replicates per control: 3

SOIL INCUBATION
- Method: series of individual subsamples (one subsample 260 g dw)

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographical reference of sampling site (latitude, longitude): (1) loamy sand: South gate, 60529 Frankfurt, Germany; (2) clay silt: Nassgewann, 65795 Hattersheim, Germany
- History of site
- Vegetation cover: (1) 1996 straw departed, 1997 summer wheat/fallow, 1998 fallow. (2) 1996 summer barley, 1997 bush bean/winter wheat, 1998 winter wheat
- Treatments with pesticides or fertilizers: not treated with crop protection agents for > 2 years
- Depth of sampling: 0 -20 cm
- Soil texture of (1) according to DIN 4220
- % sand: 51.1
- % silt: 37.6
- % clay: 11.3
- Soil texture of (2) according to DIN 4220
- % sand: 10.2
- % silt: 70.7
- % clay: 19.2
- Soil taxonomic classification: Sample (1): loamy sand (Mittel lehmiger Sand), sample (2) clay silt (Stark toniger Schluff)
- Soil classification system: DIN 4220
- pH (in water): (1) 6.0, (2) 7.0
- Initial nitrate concentration for nitrogen transformation test (mg nitrate/kg dry weight): (1) 4.1, (2) 6.2
- Maximum water holding capacity (in % dry weigth): (1) 37.3, (2) 43.9

- Pretreatment of soil: Coarse plant remains were removed from the soil. The soils were air dried until sieving was possible.
- Storage (condition, duration): at ca. 6° C, from 16 Dec 1998 to 12 Jan 1999
- Initial microbial biomass as mg/100 g soil: (1) 17.5 , (2) 31.5

DETAILS OF PREINCUBATION OF SOIL (if any): 48 hours prior to experiment start portions of soil, sufficient to carry out this study, were transferred to an air-conditioned room at 20 ± 2 °C.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): The respiration rates (O2 consumption) of the soils were determined on day 0, 14, and 28. A 50 g (dry weight equivalent) sample was removed at each time point and oxygen consumption recorded.

VEHICLE CONTROL PERFORMED: no data

Nominal and measured concentrations:

125.3 µg/kg soil (equivalent to 94.0 g/ha) and 800 µg/kg (equivalent to 600.0 g/ha)

Reference substance (positive control):

yes

Remarks:

2-Tert.-Butyl-4,6-Dinitrophenol (Dinoterb)

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 0.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Remarks on result:

other: =600 g/ha; no adverse effects on soil respiration observed, < 15%

Details on results:

The test substance when applied at 0.1253 mg/kg soil (94.0 g/ha) or 0.8 mg/kg soil (600.0 g/ha), had a negligible effect on short-term respiration in the loamy sand and clay silt 28 days after treatment. Highest oxygen consumption deviated from the control was -7.7% at the field rate 94.0 g/ha in the loamy sand at day 28. The highest deviation in the clay silt was +5.1 at day 0. All deviations measured were not statistically significant from the control at a significance level of α = 0.05 and can be classified as negligible (Domsch et al., 1983, Residue Reviews 86: 65-105).

Results with reference substance (positive control):

In both soils, the soil microflora was of acceptable sensitivity to the reference substance:
Application of the reference substance dinoterb at a rate of 40 mg/kg to loamy sand resulted in a strong decrease (-30.8%) of the oxygen consumption at day 0. At day 14 and day 28 the oxygen consumption had strongly increased to +32.6% and +23.1% respectively.
Application of the reference substance dinoterb at a rate of 40 mg/kg to clay silt resulted a decrease of oxygen consumption at day 0 (-19.4%), while an increase was observed at day 14 (+13.7%). Measurements at day 28 revealed again a decrease in oxygen consumption (-17.2%).

Validity criteria according to OECD 217 (2000) cannot be evaluated, since control raw on replicate samples data are not reported.

Validity criteria fulfilled:

not specified

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Reference 2

Endpoint:

toxicity to soil microorganisms

Type of information:

experimental study

Adequacy of study:

key study

Study period:

21 Jan - 18 Feb 1999

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

BBA Part VI, 1-1

Version / remarks:

adopted 1990 (2nd edition)

GLP compliance:

yes

Analytical monitoring:

no

Vehicle:

yes

Details on preparation and application of test substrate:

AMENDMENT OF SOIL
- Type of organic substrate: 10 mg N/100 g soil in form of lucerne green meal

APPLICATION OF TEST SUBSTANCE TO SOIL
- Method: test substance dissolved in acetone was mixed with quartz sand, the sand mixture was added to soil

VEHICLE:
- Chemical name of vehicle (organic solvent, emulsifier or dispersant): acetone
- Evaporation of vehicle before use: no data

Test organisms (inoculum):

soil

Total exposure duration:

28 d

Test temperature:

20 ± 2 °C

Moisture:

Max. water holding capacity adjusted to 45%

Details on test conditions:

TEST SYSTEM
- Testing facility: Hoechst Schering AgrEvo GmbH, Department of Environmental Sciences, Ecobiology, Frankfurt, Germany
- Test container (type, material, size): 2L container, stainless steel, diameter 16 cm, height 10 cm
- Amount of soil: 800 g divided into 3 replicates of 260 g soil each
- No. of replicates per concentration: 3
- No. of replicates per control: 3

SOIL INCUBATION
- Method: series of individual subsamples (one subsample 260 g dw)

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographical reference of sampling site (latitude, longitude): (1) loamy sand: South gate, 60529 Frankfurt, Germany; (2) clay silt: Nassgewann, 65795 Hattersheim, Germany
- Vegetation cover: (1) 1996 straw departed, 1997 summer wheat/fallow, 1998 fallow. (2) 1996 summer barley, 1997 bush bean/winter wheat, 1998 winter wheat
- Treatments with pesticides or fertilizers: not treated with crop protection agents for > 2 years
- Depth of sampling: 0 -20 cm
- Soil texture of (1) according to DIN 4220
- % sand: 51.1
- % silt: 37.6
- % clay: 11.3
- Soil texture of (2) according to DIN 4220
- % sand: 10.2
- % silt: 70.7
- % clay: 19.2
- Soil taxonomic classification: Sample (1): loamy sand (Mittel lehmiger Sand), sample (2) clay silt (Stark toniger Schluff)
- Soil classification system: DIN 4220
- pH (in water): (1) 6.0, (2) 7.0
- Initial nitrate concentration for nitrogen transformation test (mg nitrate/kg dry weight): (1) 4.1, (2) 6.2
- Maximum water holding capacity (in % dry weigth): (1) 37.3, (2) 43.9
- Pretreatment of soil: Coarse plant remains were removed from the soil. The soils were air dried until sieving was possible.
- Storage (condition, duration): at ca. 6° C, from 16 Dec 1998 to 19 Jan 1999
- Initial microbial biomass as mg/100 g soil: (1) 17.5 , (2) 31.5

DETAILS OF PREINCUBATION OF SOIL (if any): 48 hours prior to experiment start portions of soil, sufficient to carry out this study, were transferred to an air-conditioned room at 20 ± 2 °C.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): The ammonium and nitrate contents of the soils were determined on day 0, 14, and 28

VEHICLE CONTROL PERFORMED: no data

Nominal and measured concentrations:

125.3 µg/kg soil (equivalent to 94.0 g/ha) and 800 µg/kg soil (equivalent to 600.0 g/ha)

Reference substance (positive control):

yes

Remarks:

Nitrapyrin (2-Chloro-6-Trichlormethylpyridine)

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

>= 0.8 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

nitrate formation rate

Remarks on result:

other: = 600 g/ha; no adverse effects on nitrogen turnover, < 15%

Details on results:

The test substance, when applied at 125.3 µg/kg soil (94.0 g/ha) or 800 µg/kg soil (600.0 g/ha) had a negligible effect on nitrogen turn-over in the loamy sand and clay silt between 0 and 28 days after treatment.

At 125.3 µg/kg soil in the loamy sand deviations of total mineral nitrogen compared to the control were below 2% at each measurement (day 0, 14 and 28). At 800 µg/kg soil the highest deviation of total mineral nitrogen compared to the control was +8.0% at day 14 after treatment. At day 28 this difference to the control was reduced to +6.3%. These differences to the control were statistically significant but can be classified as negligible (Domsch et al., 1983, Residue Reviews 86: 65-105).

In the clay silt the initial deviation (at day 0) of total mineral nitrogen compared to the control was 3.2% in the treatments with 125.3 µg/kg soil and 3.9% in the treatments with 800 µg/kg soil. Nevertheless, these deviations did not differ significantly from the control at the 5% level. At day 14 these deviations had increased to 4.8% at 125.3 µg/kg soil and 7.3% at 800 µg/kg soil and were statistically significant but can be classified as negligible (Domsch et al., 1983, Residue Reviews 86: 65-105). At day 28 total mineral nitrogen concentrations at both treatment levels were lower than in the control (-3.7% and -2.6% respectively) but these deviations were not statistically significant.

Results with reference substance (positive control):

In both soils the nitrification was almost completely inhibited by nitrapyrin. These results with the toxic reference substance indicated that the soil microflora was of acceptable sensitivity to the reference substance.
At 14 and 28 days after application a significant deviation from the control was observed in the loamy sand. The deviations of total mineral nitrogen compared to the control were +32.4% at day 14 and +37.2% at day 28 respectively and consisted almost entirely of ammonium.
At 14 and 28 days after application, a significant deviation from the control was observed in the clay silt, deviation of total mineral nitrogen compared to the control was +17.5% at day 14 and +12.6% at day 28 respectively and consisted almost entirely of ammonium.

Validity criteria according to OECD 216 (2000) are met: nitrate concentration variation between replicate control samples was less than ± 15% (9-10%).

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Description of key information

NOEC (28 d): ≥ 0.8 mg/L (nominal, soil microflora, BBA Part VI, 1-1)

Additional information

No adverse effects (< 15% deviation from control) of the test substance on nitrogen transformation (measured as NO3-N production, OECD 216) and on carbon transformation (measured as oxygen consumption, OECD 217) in soil were observed in two GLP guideline studies (Sowig 1999a and 1999b). Concentrations of 0.1253 mg test item/kg dry soil (equivalent to 94.0 g/ha) and 0.8 mg test item/kg dry soil (equivalent to 600.0 g/ha) were investigated. Test concentrations are related to a soil depth of 10 cm and a soil density of 1.5 g/cm3.