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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:
From June 2017 to March 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 207 (Earthworm, Acute Toxicity Tests)
Deviations:
no
Qualifier:
according to guideline
Guideline:
other: 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
Deviations:
no
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
- Appearance: Pink liquid
- Test item storage: At room temperature protected from light
- Purity/Composition correction factor: No correction factor required
Analytical monitoring:
no
Vehicle:
yes
Remarks:
dichloromethane
Details on preparation and application of test substrate:
One day before the exposure, a given amount of the test substance was dissolved in 10ml dichloromethane to obtain a solvent solution of the test substance. An aliquot of that solvent solution was separately and rapidly added to about 10g silica sand in a blue cap bottle. The cap was screwed and the bottle shaken to make homogeneous mixing. The cap was then opened and the bottle placed in the fuming cupboard to evaporate the solvent. Silica sand was mixed afterwards into formulated artificial soil with deionised reverse osmosis water to give the blank control, solvent control and test concentrations, respectively, with a nominal moisture content of 40-60% by mass of the total water-holding capacity with the deionized water.
Test organisms (species):
Eisenia fetida
Animal group:
nematods
Details on test organisms:
The study was performed with earthworms (Eisenia fetida). The earthworms were purchased from Jiang Su province Jurong Stock Farm (ID: EF-MKT-170905). All newly acquired earthworms were quarantined and observed for 14 days prior to use in the test. Earthworms were bred on a substrate consisting of 50% by volume peat and 50% by volume cow manure and kept under 20±2°C for 7 days prior to testing. The test organisms were with clitellum and were transferred into artificial soil (without test substance) approximately 24 hours prior to the start of the test. At the start of the test, the used adult earthworms, which were aged of at least two months and originated from the same population, were with clitellum and wet weight of 300 to 600 mg. Before use, the earthworms were gently washed with pure water in a Petri dish, and then quickly dried on absorbent cloth.
Study type:
laboratory study
Substrate type:
artificial soil
Limit test:
no
Total exposure duration:
14 d
Test temperature:
20.0 - 20.3°C
pH:
6.02 - 6.19
Moisture:
24.72 - 30.37% (expressed as % dry soil weight)
Details on test conditions:
TEST SYSTEM
- Test container (material, size): 1L glass vessels with a plastic cover with small ventilation holes
- Amount of substrate: 750 g wet weight
- No. of replicates per treatment group: 4 (10 earthworms per replicate)
- No. of replicates per control: 4 (10 earthworms per replicate)
- No. of replicates per vehicle control: 4 (10 earthworms per replicate)

OTHER TEST CONDITIONS
- Photoperiod: continuous illumination
- Light intensity: 596 to 633 lux

EFFECT PARAMETERS MEASURED
- Survival: for the controls and each test concentration, the number of dead earthworms was assessed on the 7th and the 14th day of exposure by emptying the soil medium onto a stainless plate, sorting worms from the medium, testing their reaction to a mechanical stimulus at the front end, and recording the symptoms and deaths. Worms were considered as dead when they did not respond to a gentle mechanical stimulus to the front end. Due to rapid decomposition under test conditions, missing earthworms were also considered dead. At the completion of this assessment on the 7th day of exposure, the soil medium was replaced in the corresponding test container and the surviving earthworms were placed on the surface for the next 7 days of exposure.
- Weight: the total gross weights of the earthworms added to each vessel in the controls and the treatment groups were recorded on Day 0 before addition. The surviving earthworms in each vessel were weighed on Day 14.

OTHER PARAMETERS
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 groups randomly. The temperature and light intensity in the climate chamber were recorded daily throughout the test.

VEHICLE CONTROL PERFORMED: yes

RANGE FINDING TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: 0 (blank control), 0 (solvent control), 10, 100 and 1000 mg/kg dry soil weight (no replicates)
- Results used to determine the conditions for the definitive study:yes
Nominal and measured concentrations:
Nominal concentrations (definitive test): 0 (blank control), 0 (solvent control), 31.25, 62.5, 125, 250, 500 and 1000 mg/kg dry soil weight
Reference substance (positive control):
yes
Remarks:
chloroacetamide
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
Key result
Duration:
14 d
Dose descriptor:
LC50
Effect conc.:
648 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: 95% CI 542-923 mg/kg soil dw
Details on results:
Mortality:
Mortality in the blank control group on the 7th day and 14th day was 0% and 5%, respectively. Mortality in the solvent control group on the 7th and 14th day was 2.5% and 5%, respectively. The results showed that there were no significant difference between the blank control group and the solvent control group. Mortalities in the 31.25 mg/kg and 62.5 mg/kg dry soil weight group on the 7th and 14th day were both 5% and 10%, respectively. Mortalities in the 125 mg/kg dry soil weight group on the 7th and 14th day were 10% and 12.5%, respectively. Mortalities in the 250 mg/kg dry soil weight group on the 7th and 14th day were 12.5% and 17.5%, respectively.
Mortalities in the 500 mg/kg dry soil weight group on the 7th and 14th day were 7.5% and 22.5%, respectively. Mortalities in the 1000 mg/kg dry soil weight group on the 7th and 14th day were 42.5% and 100%, respectively. The 7 day LC50 value of the test substance to Eisenia fetida was greater than 1000 mg/kg dry soil weight. The 14 day LC50 was 648 mg/kg dry soil weight and the 95% confidence interval was from 542 to 923 mg/kg dry soil weight

Morphological and behavioral abnormalities:
Morphological and behavioral abnormalities observed in the 250 mg/kg, 500 mg/kg and 1000 mg/kg dry soil weight groups after 7d and 14d exposures included body surface color changes, slippage, ulceration, restlessness, genital swelling, slow maneuvers (slow motion and no vitality symptoms) and a lot of mucus secretion.


Results with reference substance (positive control):
The 14-days LC50 for chloroacetamide was 17.9mg/kg dry soil weight. Tests from this reference substance conducted in the laboratory showed that the range of LC50 was between 5.41 to 31.22 mg/kg, so the LC50 of 17.9mg/kg is an acceptable value.
Reported statistics and error estimates:
The LC50 value and 95% confidence limit were calculated using software Toxcalc v5.0.32.
Validity criteria fulfilled:
yes
Conclusions:
Under the conditions of the test, the 7-day LC50 was greater than 1000 mg/kg dry soil weight and the 14-day LC50 was 648 mg/kg dry soil weight (95% CI 542-923 mg/kg dry soil weight).
Executive summary:

The acute toxicity of the test item to Eisenia fetida was investigated in a GLP-compliant study performed in accordance with OECD Guideline No. 207. Under the conditions of the test, the 7-day LC50 was greater than 1000 mg/kg dry soil weight and the 14-day LC50 was 648 mg/kg dry soil weight (95% CI 542-923 mg/kg dry soil weight).

Description of key information

The acute toxicity of the test item to Eisenia fetida was investigated in a GLP-compliant study performed in accordance with OECD Guideline No. 207. Under the conditions of the test, the 7-day LC50 was greater than 1000 mg/kg dry soil weight and the 14-day LC50 was 648 mg/kg dry soil weight (95% CI 542-923 mg/kg dry soil weight).

Key value for chemical safety assessment

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

Additional information

The acute toxicity of the test item to Eisenia fetida was investigated in a GLP-compliant study performed in accordance with standard methods, without deviations. The study is considered as reliable (Klimisch 1) and is selected as a key study for the endpoint.