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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:
from 2015-11-12 to 2015-12-11
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP study conducted to the appropriate OECD Test Guideline
Qualifier:
according to
Guideline:
OECD Guideline 216 (Soil Microorganisms: Nitrogen Transformation Test)
Deviations:
no
GLP compliance:
yes
Specific details on test material used for the study:
Details on properties of test surrogate or analogue material (migrated information):
Not applicable
Analytical monitoring:
yes
Vehicle:
no
Test organisms (inoculum):
soil
Total exposure duration:
28 d
Test temperature:
20 ± 2 °C (nominal)
Moisture:
At experimental starting the soil was adjusted to approximately 45 % of its maximum water holding capacity. All replicates were weighed once per week to check for water losses by evaporation (recommended maximum 5 %, actual 0.2 – 0.8 %). Compensation of demineralised water was done as necessary. Replicates were weighed for this procedure.
Details on test conditions:
TEST SYSTEM
- Testing facility: DR.U.NOACK-LABORATORIEN, Käthe-Paulus-Str.1, D-31157 Sarstedt, Germany
- Test container (type, material, size): Plastic boxes (volume 1.0 L, food grade) with perforated tops to enable gas exchange.
- Amount of soil: 400 g soil dry weight per replicate
- No. of replicates per concentration: Triplicates
- No. of replicates per control: Triplicates

SOIL INCUBATION
- Method: The soil was incubated in the above mentioned boxes. The tops of the boxes were perforated to enable gas exchange.

SOURCE AND PROPERTIES OF SUBSTRATE (if soil)
- Geographical reference of sampling site: Offenbach, “rechts der Landauer Str.“ Nr. 826/7, Rheinland-Pfalz, Germany, Gauß-Krüger-Coordinates: R-439683; H-5449554
- History of site: Cultures: 2011 - 2014: uncultivated, 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
- Treatments with pesticides or fertilizers: No crop protection products applied during the year of sampling and 4 preceding years.
- Depth of sampling: ca. 20 cm
- Soil texture: silty sand (uS) acc. to German DIN classification
- % sand: 2.0 - 0.63 mm: 2.7 ± 0.8 % dry weight, 0.63 - 0.2 mm: 30.3 ± 0.6 % dry weight, 0.2 - 0.063 mm: 24.6 ± 1.5 % dry weight
- % silt: 0.063 - 0.02 mm: 19.0 ± 1.8 % dry weight, 0.02 - 0.006 mm: 11.5 ± 0.6 % dry weight, 0.006 - 0.002 mm: 5.4 ± 0.8 % dry weight
- % clay: 6.6 ± 1.7 % dry weight
- pH (in water): 5.7 ± 0.6
- Initial nitrate concentration for nitrogen transformation test (mg nitrate/kg dry weight): 17.3 mg NO3-N/kg DW
- Maximum water holding capacity (in % dry weigth): 35.4 ± 1.5 g/100 g DW
- Cation exchange capacity (mmol/kg): 7.5 ± 0.9 meq / 100 g
- Pretreatment of soil: Prior to final moistening at test start, the soil was adjusted to about 42 % of its maximum water holding capacity with demineralised water. Drying out of the soil was prevented by moistening with demineralised water as necessary. The soil was checked for a detectable microbial biomass (result expressed in terms of percentage of total organic carbon) and the amount of total inorganic nitrogen. 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. Content of total inorganic nitrogen: 2.83 % (dry weight) Content of total organic carbon: 42.8 % (dry weight) C/N-ratio: 15.1
- Storage (condition, duration): The soil was stored for 14 days (2015-10-07 to 2015-10-21) in the dark at 6 ± 2 °C in a climatic room (TE1200, VIESSMANN).
- Initial microbial biomass as % of total organic C: 2.48

DETAILS OF PREINCUBATION OF SOIL (if any): the soil was pre-incubated at room temperature (ca. 20 °C) for 22 days (2015-10-21 to 2015-11-12) 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.

RANGE-FINDING STUDY
- Test concentrations: 10, 100, 1000 mg/kg soil DW
- Results used to determine the conditions for the definitive study: Based on the results of the preliminary study, the test concentrations for the definitive study were selected: 1000 - 316 - 100 - 31.6 - 10 mg /kg soil dry weight (spacing factor √10)
Reference substance (positive control):
yes
Remarks:
Cyanoguanidine
Duration:
7 d
Dose descriptor:
NOEC
Effect conc.:
>= 1 000 mg/kg soil dw
Basis for effect:
nitrate formation rate
Duration:
7 d
Dose descriptor:
other: LOEC
Effect conc.:
> 1 000 mg/kg soil dw
Basis for effect:
nitrate formation rate
Duration:
14 d
Dose descriptor:
NOEC
Effect conc.:
316 mg/kg soil dw
Basis for effect:
nitrate formation rate
Duration:
14 d
Dose descriptor:
other: LOEC
Effect conc.:
1 000 mg/kg soil dw
Basis for effect:
nitrate formation rate
Key result
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
316 mg/kg soil dw
Basis for effect:
nitrate formation rate
Duration:
28 d
Dose descriptor:
other: LOEC
Effect conc.:
1 000 mg/kg soil dw
Basis for effect:
nitrate formation rate
Duration:
28 d
Dose descriptor:
EC25
Effect conc.:
> 1 000 mg/kg soil dw
Basis for effect:
nitrate formation rate
Key result
Duration:
28 d
Dose descriptor:
EC50
Effect conc.:
> 1 000 mg/kg soil dw
Basis for effect:
nitrate formation rate

 Nitrate-N Content             Inhibition [%] compared to control

 Test concentration (mg/kg soil dry weight)  0 d  7 d  14 d  28 d
 10  0  -9  -3  -1
 31.6  -5  -16  -11  -9
 100  1  -6  -2  -1
 316  1  -4  2  1
 1000  6  6  11  16

negative values = increase

 Nitrate-N Formation Rate Inhibition [%] compared to control            

   Test concentration (mg/kg soil dry weight)  7 d  14 d  28 d
 10  -22  -5  -2
 31.6  -31  -16*  -10
 100  -15  -4  -2
 316  -11  3  1
 1000  5  14*  20*

* Statistically significant difference compared to control (ANOVA, p ≤ 0.05)

negative values = increase

Validity criteria fulfilled:
yes
Conclusions:
The effects of the test item on the metabolic activity of the nitrogen-N (nitrate) formation rate were measured on the day of treatment (day 0) and subsequently after 7, 14 and 28 days.
After 14 and 28 days of exposure to POLYOL PX, a significant inhibition of the microbial nitrate transformation was observed for the test item concentration 1000 mg/kg soil dry weight. The mean inhibition on day 28 compared to the control was 20 %. The EC25 and EC50-values could not be determined since no inhibition effects ≥ 25 or 50 % were detected. Therefore the EC25-value and EC50-value were determined to be > 1000 mg test item/kg soil dry weight.
The corresponding no-observed effect concentration (NOEC) was 316 mg POLYOL PX/kg soil dry weight and the LOEC was determined to be 1000 mg POLYOL PX/kg soil dry weight.
Executive summary:

The effects of the test item on the metabolic activity of the nitrogen-N (nitrate) formation rate were measured on the day of treatment (day 0) and subsequently after 7, 14 and 28 days.

After 14 and 28 days of exposure to POLYOL PX, a significant inhibition of the microbial nitrate transformation was observed for the test item concentration 1000 mg/kg soil dry weight. The mean inhibition on day 28 compared to the control was 20 %. The EC25 and EC50-values could not be determined since no inhibition effects ≥ 25 or 50 % were detected. Therefore the EC25-value and EC50-value were determined to be > 1000 mg test item/kg soil dry weight.

The corresponding no-observed effect concentration (NOEC) was 316 mg POLYOL PX/kg soil dry weight and the LOEC was determined to be 1000 mg POLYOL PX/kg soil dry weight.

Description of key information

The effects of the test item on the metabolic activity of the nitrogen-N (nitrate) formation rate were measured on the day of treatment (day 0) and subsequently after 7, 14 and 28 days.

After 14 and 28 days of exposure to POLYOL PX, a significant inhibition of the microbial nitrate transformation was observed for the test item concentration 1000 mg/kg soil dry weight. The mean inhibition on day 28 compared to the control was 20 %. The EC25 and EC50-values could not be determined since no inhibition effects ≥ 25 or 50 % were detected. Therefore the EC25-value and EC50-value were determined to be >1000 mg test item/kg soil dry weight.

The corresponding no-observed effect concentration (NOEC) was 316 mg POLYOL PX/kg soil dry weight and the LOEC was determined to be 1000 mg POLYOL PX/kg soil dry weight

Key value for chemical safety assessment

Short-term EC50 for soil microorganisms:
1 000 mg/kg soil dw
Long-term EC10 or NOEC for soil microorganisms:
316 mg/kg soil dw

Additional information