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Toxicity to soil microorganisms

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Reference
Endpoint:
toxicity to soil microorganisms
Type of information:
experimental study
Adequacy of study:
key study
Study period:
definitive exposure from 2015-08-17 to 2015-09-14
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Guideline study in compliance with GLP, but no analysis of test concentrations.
Qualifier:
according to
Guideline:
OECD Guideline 216 (Soil Microorganisms: Nitrogen Transformation Test)
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
no
Vehicle:
no
Details on preparation and application of test substrate:
AMENDMENT OF SOIL
- Type of organic substrate: The soil amounts were amended with powdered lucerne-green-grass-meal (0.5 % of soil dry weight). The C/N ratio was between 12/1 and 16/1. A ratio of 5 g Lucerne per kg of soil (dry weight was used.
- Other:

APPLICATION OF TEST SUBSTANCE TO SOIL
- Method: The soil was divided into six portions of equal weight. The respective test item volume for each test item concentration was given on quartz sand (1 % of the entire soil amount per concentration) and blended thoroughly. Afterwards the test item - quartz sand mixture was mixed carefully into the corresponding portion of the soil with a mixer to ensure a homogeneous distribution of the test item in the soil. Subsequently the soil was distributed to the replicates (three replicates each per concentration and control).
Test organisms (inoculum):
soil
Total exposure duration:
28 d
Test temperature:
18-21 °C
Moisture:
Moisture content as [%] of Maximum Water Holding Capacity:
Day 0: 42.5 - 43.9 %
Day 28: 43.4 - 44.7 %
Details on test conditions:
TEST SYSTEM
- Testing facility: DR.U.NOACK-LABORATORIEN
Käthe-Paulus-Str. 1
31157 Sarstedt
Germany

- Test container (type, material, size): Plastic boxes (volume 1.0 L, food grade) with perforated tops to enable gas exchange. Incubation was performed in bulk and sub-samples were taken for the experimental phase.
- Amount of soil: 400 g (dry weight)
- No. of replicates per concentration: 3
- No. of replicates per control: 3
- No. of replicates per vehicle control: N/A

SOIL INCUBATION
- Method: bulk - Incubation was performed in bulk and sub-samples were taken for the experimental phase.

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographical reference of sampling site (latitude, longitude): Offenbach, “rechts der Landauer Str.“ Nr. 826/7, Rheinland-Pfalz, Germany, R-439683; H-5449554.
- History of site: 2011 - 2014: uncultivated.
- Vegetation cover: Not reported
- Treatments with pesticides or fertilizers: Fertilisation:
2015: none
2014: 3500 kg/ha CaO (05.06.2014 and 24.09.2014)
3500 kg/ha CaO; 1463 kg/ha MgO (15.12.2014)
2013: none
2012: none
2011: none

Pesticides No crop protection products applied during the year of sampling and 4 preceding years.

- Accidental contamination: None reported.
- Other:
- Depth of sampling: approximately 20 cm
- Soil texture (dry weight):
- % sand: 2.0 - 0.63 mm = 2.7 ± 0.8
0.63 - 0.2 mm = 30.3 ± 0.6
0.2 - 0.063 mm = 24.6 ± 1.5
- % silt: 0.063 - 0.02 mm = 19.0 ± 1.8
0.02 - 0.006 mm = 11.5 ± 0.6
0.006 - 0.002 mm = 5.4 ± 0.8
- % clay: < 0.002 mm = 6.6 ± 1.7
- Soil taxonomic classification: silty sand (uS) (LUFA-soil 2.3, Batch-No. F2.31615)
- Soil classification system: German DIN classification
- pH (in water): 5.7 ± 0.6
- Initial nitrate concentration for nitrogen transformation test (mg nitrate/kg dry weight): 11.8 mg NO3-N/kg DW
- Maximum water holding capacity (in % dry weight): 35.4 ± 1.5 g/100 g DW
- Cation exchange capacity (mmol/kg): 7.5 ± 0.9 meq / 100 g
- Pretreatment of soil:
- Storage (condition, duration): The soil was stored for 91 days (2015-04-22 to 2015-07-22) in the dark at 6 ± 2 °C in a climatic room (TE1200, VIESSMANN).
- Initial microbial biomass as % of total organic C: 4.44

DETAILS OF PREINCUBATION OF SOIL (if any): The soil was pre-incubated at room temperature (ca. 20°C) for 26 days (2015-07-22 to 2015-08-17) to adapt the micro-organisms to test conditions before the start of the study.

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): Measurements of nitrate concentrations were carried out after 0, 7, 14 and 28 days. The pH values and water contents were determined on day 0 and 28. The room temperature was measured and recorded continuously with a thermohygrograph.

VEHICLE CONTROL PERFORMED: yes/no - N/A

RANGE-FINDING STUDY
- Test concentrations: 1, 10, 100 and 1000 mg/kg soil DW
- Results used to determine the conditions for the definitive study: 28-d % inhibition at 1 mg/kg soil DW = -6%.
28-d inhibition at 1000 mg/kg soil DW = 66%.
Nominal and measured concentrations:
Nominal: 10, 31.6, 100, 316 and 1000 mg/kg soil dry weight.
Reference substance (positive control):
yes
Remarks:
Cyanoguanidine
Duration:
7 d
Dose descriptor:
EC50
Effect conc.:
353 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
nitrate formation rate
Duration:
14 d
Dose descriptor:
EC50
Effect conc.:
462 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
nitrate formation rate
Key result
Duration:
28 d
Dose descriptor:
EC50
Effect conc.:
620 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
nitrate formation rate
Remarks on result:
other: 95% CL 535-701
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
100 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
nitrate formation rate
Results with reference substance (positive control):
Nitrate-N Formation Rate % compared to Untreated Control at 28-d:
50 mg/kg soil dry weight = 44 %
100 mg/kg soil dry weight = 98 %
Reported statistics and error estimates:
One Way Analysis of Variances (ANOVA) and DUNNETT’S test, were carried out for the determination of statistically significant differences of the nitrate formation rate at day 7, 14 and 28 compared to control replicates. When running a One Way Analysis of Variances a Normality test and an Equal Variance test were done first. P-values for both Normality and Equal Variance test are 0.05. The a-value for ANOVA and DUNNETT’S test (acceptable probability of incorrectly concluding that there is a difference) is a = 0.05.

Table 1 Reduction of Nitrate-N Content in soil containing 3-trimethoxysilylpropyl methacrylate, relative to the untreated control

Nitrate-N Content

Test concentration

Inhibition [%] compared to control

[mg/kg soil dry weight]

0 d

7 d

14 d

28 d

  10

0

2

-3

0

  31.6

2

4

4

0

  100

-2

0

6

1

  316

2

14

13

13

  1000

0

48

40

44

negative values = increase

Table 2 Inhibition of Nitrate-N Formation Rates in soil containing 3-trimethoxysilylpropyl methacrylate, relative to the untreated control

Nitrate-N Formation Rate

Test concentration

Inhibition [%] compared to control

[mg/kg soil dry weight]

7 d

14 d

28 d

  10

5

-8

0

  31.6

9

7

-1

  100

5

18

3

  316

38*

29*

20*

  1000

n.d.*

95*

72*

*) Statistically significant difference compared to control (ANOVA, p = 0.05) negative values = increase

n.d. not determinable, due to negative Nitrate-N Formations rates

Validity criteria fulfilled:
yes
Conclusions:
A 28 day EC50 value of 620 mg/kg soil dw has been determined for the effects of 3-trimethoxysilylpropyl methacrylate on nitrogen transformation rate by soil micro-organisms.

Description of key information

Nitrogen transformation by terrestrial micro-organisms: 28-day EC50 620 mg/kg dwt and EC25 359 mg/kg dwt (nominal) (OECD 216). A NOEC of 100 mg/kg dwt (nominal) was derived for the study. The 28-day results are equivalent to EC50 of 515 mg/l and EC25 of 298 mg/kg dwt, and the study NOEC is equivalent to 83 mg/kg dwt, when expressed in terms of the silanol hydrolysis product.

Key value for chemical safety assessment

Short-term EC50 for soil microorganisms:
515 mg/kg soil dw
Long-term EC10 or NOEC for soil microorganisms:
83 mg/kg soil dw

Additional information

A 7-day EC50 353 mg/kg dwt and EC25 267 mg/kg dwt; 28-day EC50 620 mg/kg dwt and EC25 359 mg/kg dwt (nominal) (OECD 216) have been determined for the effects on nitrogen transformation by a native population of soil microbes exposed to 3-trimethoxysilylpropyl methacrylate (CAS 2530-85-0). A NOEC of 100 mg/kg dwt (nominal) was derived throughout the study (Noack 2016).

In view of the hydrolysis rate of the substane, the test media preparation method and conditions during the test, it is likely that the test organisms were exposed predominantly to the hydrolysis products in the test system. Analytical quantification was not attempted.

In this study the ECx values progessively increase between the 7-day, 14-day and 28-day points. This may reflect increasing tolerance in the test system, progressive degradation of the test substance by hydrolysis in the moist soil substrate, or a combination of the two. The 28-day ECx values are of highest relevance to the silanol hydrolysis product, 3-(trihydroxysilyl)propyl methacrylate, which is the subject of the chemical safety assessment for the environment.

The results may be expressed in terms of concentration of the hydrolysis product, 3-(trihydroxysilyl)propyl methacrylate, by applying a molecular weight correction: (MW of silanol=206.27 / MW of parent=248.35) * [Conc. of parent] = [conc. of silanol]. Using this method, the 28-day results are equivalent to EC50 of 515 mg/l and EC25 of 298 mg/kg dwt, and the study NOEC is equivalent to 83 mg/kg dwt, when expressed in terms of the silanol hydrolysis product.