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Toxicity to soil macroorganisms except arthropods

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Reference
Endpoint:
toxicity to soil macroorganisms except arthropods: short-term
Type of information:
experimental study
Adequacy of study:
key study
Study period:
October 2018 - January 2019
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 207 (Earthworm, Acute Toxicity Tests)
Version / remarks:
1984
Deviations:
no
Qualifier:
according to
Guideline:
other: Ministry of environmental protection of the people's Republic of China, The Guidelines for the Testing of Chemicals - Effects on Biotic Systems, 207 Earthworm, Acute Toxicity Test
Version / remarks:
2013
Deviations:
no
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
no
Vehicle:
yes
Remarks:
acetone
Details on preparation and application of test substrate:
Since the purity of the test substance is 86.3%, an amount of 2920.02 mg of the test substances was dissolved into 10mL acetone to give the solvent stock solution. The solvent stock solution was inverted several times to ensure adequate mixing and homogeneity.
An aliquot solvent solution was separately added to a certain amount of silica sand and the solvent was allowed to evaporate prior to the incorporation into formulated artificial soil with deionized reverse osmosis water. The treatment groups were prepared with a nominal moisture content of 40-60% by mass of the total water-holding capacity with the deionized water. The test medium preparation process of the blank control group was same as the other groups, but without the solvent and the test substance. The test medium preparation process of the solvent control group was same as the other groups, but without the test substance. The mixer was used to blend all the components.
Test organisms (species):
Eisenia fetida
Animal group:
annelids
Details on test organisms:
TEST ORGANISM
- Common name: redworm
- Source: Jiang Su province Jurong Stock Farm. (ID: EF-MKT-180801)
- Age at test initiation: adult (with clitellum)
- Weight at test initiation: 300 - 600mg

ACCLIMATION
- Acclimation period: 24h
- Acclimation conditions: same as test conditions
- Health during acclimation: no mortality observed
Study type:
laboratory study
Substrate type:
artificial soil
Limit test:
yes
Total exposure duration:
14 d
Test temperature:
20.0 to 20.2 °C
pH:
6.07 to 6.13 at initiation and 6.09 to 6.16 at termination
Moisture:
27.02 to 29.00% at initiation and 26.16 to 29.70% at termination
Details on test conditions:
TEST SYSTEM
- Test container: 1L breathable glass bottle with cover
- Amount of substrate: 750 g wet weight
- No. of organisms per container (treatment): 10
- No. of replicates per treatment group: 4
- No. of replicates per control: 4
- No. of replicates per vehicle control: 4

PROPERTIES OF SUBSTRATE
Sphagnum peat: 10% w/w
Kaolinite clay (Sigma-Aldrich): 20% w/w
Silica sand (screened with 0.2 mm sieve): 70% w/w
The pH of the artificial soil was adjusted to 6.0 ± 0.5. Before the start of the test, measured maximum water holding capacity (WHC) of artificial soil was 62%. Moisture content of the test substrate was adjusted to 40-60% by mass of the total water-holding capacity with the deionized water (moisture content was measured to be 26.16-29.70%).

OTHER TEST CONDITIONS
- Light intensity: 597 - 632 lux

EFFECT PARAMETERS MEASURED
Mortalities were recorded at 7 and 14 days after the start of test. The total gross weights of the earthworms added to each vessel in the controls and the treatment were recorded on Day 0 before addition. The surviving earthworms in each vessel were weighed on Day 14. The temperature and the light intensity in the climate chamber were recorded daily throughout the test. At the beginning and the end of the test, soil moisture content and pH value of one test container were recorded in the blank control group, solvent control group and treatment group randomly.

VEHICLE CONTROL PERFORMED: yes

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 10, 100, 1000 mg/kg dry soil
- Results used to determine the conditions for the definitive study: yes, no observed mortality
Nominal and measured concentrations:
Nominal concentration: 1000 mg/kg dry soil
Reference substance (positive control):
yes
Remarks:
Chlorine acetamide
Key result
Duration:
14 d
Dose descriptor:
LC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Duration:
7 d
Dose descriptor:
LC50
Effect conc.:
> 1 000 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Details on results:
- Mortality at end of exposure period: In the solvent control one of the 40 earthworms died before day 7, the mortality was 2.5 %. In the control and the treatment no mortalities were observed. Thus, the test fullfills the validity criterion that the control mortality should not exceed 10%.
- Total mass of adults at beginning of test: 4963 mg
- Weight reduction 14 days ± standard deviation(%): Control: 9.51 ±3.05; Solvent control: 13.91 ±6.19; Treatment: 18.31 ±7.21
- Morphological abnormalities: no
- Behavioural abnormalities: no
- Other biological observations: no
Results with reference substance (positive control):
The reference substance was tested at five concentrations of 3.75, 7.50, 15, 30 and 60 mg/kg dry artificial soil weight. The mortalities in chloroacetamide treatment groups were shown to be between 0% to 100%. The 14-days LC50 for chloroacetamide was 15.4 mg/kg dry soil weight. Tests from this reference substance conducted in the laboratory showed that the range of LC50 was between 12.1 mg/kg to 27.1 mg/kg, so the 15.4 mg/kg is an acceptable value.

Table 1: Mortality of Eisenia fetida exposed to the test substance and controls after 7 and 14 days of exposure

Parameter

Blank control

Solvent control

Treatment (1000 mg active component of the test substance/kg dry soil weight)

Duration (d)

7

14

7

14

7

14

Number of earthworm

40

40

40

40

40

40

Number of dead eartworms (A replicate)

0

0

0

0

0

0

Number of dead eartworms (B replicate)

0

0

1

1

0

0

Number of dead eartworms (C replicate)

0

0

0

0

0

0

Number of dead eartworms (D replicate)

0

0

0

0

0

0

Total

0

0

1

1

0

0

Mortality (%)

0

0

2.5

2.5

0

0

  

Table 2: Parameters of the artificial soil

Groups

Duration (d)

Soil moisture content% (Expressed as % dry weight)

pH

Blank control

0

29.00

6.13

14

29.70

6.09

Solvent control

0

27.02

6.07

14

26.80

6.11

Treatments( 1000 mg/kg dry soil weight)

0

27.99

6.07

14

26.16

6.16

 

Validity criteria fulfilled:
yes
Conclusions:
The acute toxicity of the test substance to Eisenia fetida was determined in a 14 days artificial soil test according to OECD 207. The 7-Day LC50 and the 14-Day LC50 based on the nominal test concentrations were both higher than 1000 mg/kg dry soil weight.
Executive summary:

The acute toxicity of the test substance to Eisenia fetida was determined in a 14 days artificial soil test according to "Ministry of environmental protection of the people's Republic of China, The Guidelines for the Testing of Chemicals - Effects on Biotic Systems, 2013, 207 Earthworm, Acute Toxicity Test" and "OECD, Guidelines for the Testing of Chemicals, 1984, 207 Earthworm, Acute Toxicity Test”. Based on the results of range-finding test, a treatment group, a solvent control group and a blank control group which had four replicates of 10 worms per replicate were included in the limit test. Consideration of the purity of the test substance is 86.3%, the treatment group was prepared based on the active components. The nominal concentration of active component in the treatment group was 1000 mg/kg dry artificial soil weight. The number of mortalities were determined after 7 and 14 days exposure. The mortality of the earthworms in the blank control group was 0%, in the solvent control group was 2.5%, which were not more than 10 percent. So the test was valid. Under the tested conditions, the 7-Day LC50 and the 14-Day LC50 for the test substance to earthworms (Eisenia fetida) based on the nominal test concentrations were both higher than 1000 mg/kg dry soil weight.

Description of key information

The acute toxicity of the test substance to Eisenia fetida was determined in a 14 days artificial soil test according to OECD 207. The 7-Day LC50 and the 14-Day LC50 based on the nominal test concentrations were both higher than 1000 mg/kg dry soil weight.

Key value for chemical safety assessment

Short-term EC50 or LC50 for soil macroorganisms:
1 000 mg/kg soil dw

Additional information

The acute toxicity of the test substance to Eisenia fetida was determined in a 14 days artificial soil test according to "Ministry of environmental protection of the people's Republic of China, The Guidelines for the Testing of Chemicals - Effects on Biotic Systems, 2013, 207 Earthworm, Acute Toxicity Test" and "OECD, Guidelines for the Testing of Chemicals, 1984, 207 Earthworm, Acute Toxicity Test” (reference 6.3.1 -1). Based on the results of range-finding test, a treatment group, a solvent control group and a blank control group which had four replicates of 10 worms per replicate were included in the limit test. Consideration of the purity of the test substance is 86.3%, the treatment group was prepared based on the active components. The nominal concentration of active component in the treatment group was 1000mg/kg dry artificial soil weight. The number of mortalities were determined after 7 and 14 days exposure. The mortality of the earthworms in the blank control group was 0%, in the solvent control group was 2.5%, which were not more than 10 percent. So the test was valid. Under the tested conditions, the 7-Day LC50 and the 14-Day LC50 for the test substance to earthworms (Eisenia fetida) based on the nominal test concentrations were both higher than 1000 mg/kg dry soil weight.