2-ethylhexyl benzoate

Toxicity to soil microorganisms

Link to relevant study record(s)

Reference

Endpoint:

toxicity to soil microorganisms

Type of information:

experimental study

Adequacy of study:

key study

Study period:

24 January 2018 to 21 February 2018

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 216 (Soil Microorganisms: Nitrogen Transformation Test)

Qualifier:

according to guideline

Guideline:

EU Method C.21 (Soil Microorganisms: Nitrogen Transformation Test)

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Remarks:

nitrate analysis

Details on sampling:

Days 0 and 28 three aliquots (50 g dw) of soil

Vehicle:

yes

Remarks:

adsorption onto quartz sand prior to incorporation into the soil bulk

Details on preparation and application of test substrate:

Nominal amounts (0, 5, 16, 50, 160 and 500 mg) were each separately weighed onto quartz sand (5.0 g, particles size 0.1 – 0.5 mm). The blank-test item/sand combination and an amount of powdered Lucerne-green-grass-meal (2.5 g) along with an aliquot (37.0 mL) of water was then separately added to a bulk soil weight of 0.5 kg dw and thoroughly mixed to give the required test concentrations of 0, 10, 32, 100, 320 and 1000 mg/kg dw with a nominal moisture content of 45% of the Water Holding Capacity (WHC).

Test organisms (inoculum):

soil

Total exposure duration:

28 d

Remarks:

in glass jars sealed with polyurethane foam stoppers in a temperature controlled room in darkness.

Test temperature:

19-21 °C

Moisture:

9%, adjusted to 45% MWHC by adding 37 mL deionised water/100 g

MWHC 35.4 g/100 g

Details on test conditions:

TEST SYSTEM
- Test container (type, material, size): glass jar
- Amount of soil: 0.5 kg
- No. of replicates per concentration: 3
- No. of replicates per control: 3

SOIL INCUBATION
- Method: bulk (Three soil aliquots (50 g dw) were removed for Day 0 nitrate analysis from
each test concentration prior to the remaining soil bulk being incubated)

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographical reference of sampling site: Offenbach (Germany)
- History of site:
- Vegetation cover: uncultivated
- Treatments with pesticides or fertilizers: none between 2013 and 2017 (except in 2014 CaO/MgO)
- Accidental contamination: none
- Depth of sampling: 20 cm
- Soil texture: no data
- % sand: 57.2%
- Soil taxonomic classification: sandy loam/silty sand (uB)
- Soil classification system: USDA/German DIN
- pH (in water): 6.1
- Initial nitrate concentration for nitrogen transformation test (mg nitrate/kg dry weight):
- Maximum water holding capacity (in % dry weigth): 35.4%
- Cation exchange capacity (meq/100g): 7.5
- Pretreatment of soil: none
- Storage (condition, duration): ca 28 days
On arrival 14 December 2017, the soil was stored at approximately 4 ºC prior to use. A sub sample of the soil (approximately 500 g) was removed on 20 December 2017 and stored at temperatures of between 20 and 21 °C for 7 days and shielded from light prior to the preliminary biomass being determined on 27 December 2017. Prior to the start of the test, the remaining soil was removed from refrigeration on 10 January 2018 and stored at temperatures of between 15 and 21 ºC for 14 days and shielded from light. After the soil for the study was dosed, the remaining soil was stored at temperatures of between 20 to 21 ºC for 8 further days prior to determination of the definitive biomass on 1 February 2018.
- Initial microbial biomass as % of total organic C: 2.6%

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : nitrate concentration on day 0 and 28

Nominal and measured concentrations:

0, 10, 32, 100, 320 and 1000 mg/kg dw

Reference substance (positive control):

no

Duration:

28 d

Dose descriptor:

EC50

Effect conc.:

41 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

nitrate formation rate

Duration:

28 d

Dose descriptor:

NOEC

Effect conc.:

10 mg/kg soil dw

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

nitrate formation rate

Validity criteria fulfilled:

yes

Remarks:

replicate control nitrification rates was less than 15%

Conclusions:

The effect of the test item on the nitrogen transformation activity of the soil microorganisms gave an EC50 of 41 mg/kg soil dry weight, 95% confidence limits 34 - 50 mg/kg soil dry weight. The No Observed Effect Concentration (NOEC) was 10 mg/kg soil dry weight.

Executive summary:

Soil microorganisms were exposed to the test item at concentrations of 10, 32, 100, 320 and 1000 mg/kg soil dry weight for 28 days at temperatures of between 19 to 22 °C in the dark with the addition of powdered Lucerne-green-grass meal to act as a respiratory substrate. The inhibitory effect of the test item on nitrogen transformation was assessed by the determination of nitrate concentration in the soil samples on Days 0 and 28 and compared to data obtained from the control soil samples.

The effect of the test item on the nitrogen transformation activity of the soil microorganisms gave an EC50

of 41 mg/kg soil dry weight, 95% confidence limits of 34 - 50 mg/kg soil dry weight.  The No Observed Effect Concentration (NOEC) was 10 mg/kg soil dry weight.

Description of key information

Soil microorganisms were exposed to the test item at concentrations of 10, 32, 100, 320 and 1000 mg/kg soil dry weight for 28 days at temperatures of between 19 to 22 °C in the dark with the addition of powdered Lucerne-green-grass meal to act as a respiratory substrate. The inhibitory effect of the test item on nitrogen transformation was assessed by the determination of nitrate concentration in the soil samples on Days 0 and 28 and compared to data obtained from the control soil samples.

The effect of the test item on the nitrogen transformation activity of the soil microorganisms gave an EC50 of 41 mg/kg soil dry weight, 95% confidence limits of 34 - 50 mg/kg soil dry weight.  The No Observed Effect Concentration (NOEC) was 10 mg/kg soil dry weight.

Key value for chemical safety assessment

Short-term EC50 for soil microorganisms:

41 mg/kg soil dw

Long-term EC10 or NOEC for soil microorganisms:

10 mg/kg soil dw