Registration Dossier

Data platform availability banner - registered substances factsheets

Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.

The new ECHA CHEM database has been released by ECHA, and it now contains all REACH registration data. There are more details on the transition of ECHA's published data to ECHA CHEM here.

Diss Factsheets

Ecotoxicological information

Toxicity to soil macroorganisms except arthropods

Currently viewing:

Administrative data

Link to relevant study record(s)

Reference
Endpoint:
toxicity to soil macroorganisms except arthropods: long-term
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study
Qualifier:
according to guideline
Guideline:
OECD Guideline 222 (Earthworm Reproduction Test (Eisenia fetida/Eisenia andrei))
Deviations:
yes
Remarks:
On one day the measured soil temperature increased up to 22.8°C at maximum for five hours. However, test results have not been impacted.
Qualifier:
according to guideline
Guideline:
ISO 11268-2 (Effects of Pollutants on Earthworms. 2. Determination of Effects on Reproduction)
Deviations:
not specified
GLP compliance:
yes (incl. QA statement)
Vehicle:
yes
Details on preparation and application of test substrate:
Pre-treatment of the Test Item:
The test item was not soluble in water; therefore, a volatile organic solvent was used. The stock solution was prepared in acetone. All concentrations to be tested were based on the content of the test substance (96.6 %).
The test item was dissolved in an amount of acetone sufficient to prepare a stock solution. This stock solution was used to produce the various dosage solutions of the test item. An appropriate amount of the stock and the dosage solutions respectively was used to soak a portion of the quartz sand of the artificial soil by the solvent solution. After evaporation of the solvent by placing under a fume hood, the quartz sand was coated by the test item. The test item coated fraction of the quartz sand was than incorporated into the remainder of the artificial soil by thoroughly mixing it in for approx. 10 minutes. Finally, the contaminated artificial soil was filled into the test vessels. Per test vessel an amount corresponding to 500 g dw was used.
Test organisms (species):
Eisenia fetida
Animal group:
annelids
Details on test organisms:
The earthworms used in this test, Eisenia fetida (Lumbricidae, Earthworms) are kept at ECT Oekotoxikologie GmbH since February 1994. They were originally delivered by Co. Landenberger (D-72355 Schömberg). Only adult worms (with clitellum) with a fresh weight (FW) between 250 and 600 mg were used. The worms were at least two months, but not more than one year old, however the age of individuals did not differ by more than four weeks. Animals which do fulfil these requirements, were taken from a synchronised culture. The worms selected for the test were acclimatised in artificial soil under test conditions at least 24 hours before starting the test.

Feeding:
The adult worms were fed with finely ground cow manure (free of growth promoters, nematicides or similar veterinary pharmaceuticals, provided by Jesus Bruderschaft e.V. Gnadenthal, D-65597 Hünfelden). Before starting the experiment an amount of 5 g food per 500 g soil dry weight was mixed into the artificial soil to be used in the test. After the start of the experiment food was first provided one day after application of the test item and introduction of the adult worms. Thereafter, the adult worms were fed weekly during the first 4 weeks of the test. An amount of 5 g moistened with 10 mL de-ionised water was spread on the soil surface of each test container. If food remains uneaten the ration was reduced on demand. After removing of the adults on day 28 further 5 g of food was mixed with the artificial soil of each test vessel.
Study type:
laboratory study
Substrate type:
artificial soil
Limit test:
no
Total exposure duration:
56 d
Test temperature:
18.3 - 22.8°C
pH:
Start: 6.1 - 6.3
End. 6.6- 6.7
Moisture:
soil moisture = 40 – 60 % of the WHCmax (66.2 %) of the artificial soil;
= 26.5 - 39.7 % of the dry weight of the artificial soil.
Details on test conditions:
Test Units:
Test vessels were Bellaplast containers (Company Polarcup GmbH, Alf/Mosel) consiting of inert (non-toxic) plastic (Polystyrol). The test vessels have a bottom area of 11 x 15.5 cm (= 170.5 cm²) and a height of 6 cm. The test vessels were covered by a transparent lid.
- Test Conditions:
The earthworms were tested in an artificial soil. The composition of this artificial soil was based on OECD Guideline No. 222 (the percentages refer to the dry weight):
- 5 % Sphagnum peat (no visible plant remains; finely ground and air-dried)
- 20 % Kaolin-Clay (kaolinite content > 30 %)
- 74-75 % Quartz sand (fine sand content with particles between 50 and 200 μm higher than 50 %)
- 0.3-1 % Calcium carbonate (CaCO3)
With respect to the properties of the active substance (Log POW ≥ 2) 5 % instead of 10 % peat was used considering the influence on bioavailability (EPPO 2003, OECD 2004). The air dried peat was shredded in a chaff-cutter. Afterwards, all parts of the artificial soil were mixed homogeneously.
- Test system: 10 Eisenia fetida per test vessel;
Test design: 5 treatments plus control and solvent control, 4 replicates per treatment and the deionised water control and 8 for the solvent control;
Test conditions: temperature: 18.3 – 22.8 °C (recommended 20 ± 2 °C),
light cycle: 16 : 8 h with a constant light intensity at the substrate of 414 - 630 lx (recommended 400 – 800 lx)
soil pH-value (day 0): 6.1 – 6.3 (recommended 6 ± 0.5),
soil pH-value (day 56): 6.6 - 6.7
soil moisture(day 0): 50.3 – 53.5 %(rec. 40 – 60% of the WHCmax);
soil moisture (day 56): 52.6 – 58.9 % (rec. deviation ≤ 10 % from day 0)
Test parameters: mortality and biomass after 28 days; reproduction after 56 days
Endpoints: NOEC/LOECBiomass, NOEC/LOECReproduction;
Statistics: Welch-t test (2-sided, p ≤ 0.05), Williams test (1-sided, p ≤ 0.05);
Nominal and measured concentrations:
The test item was tested in five nominal concentrations. 31.6, 56.2, 100, 178 and 316 mg test item/kg soil (dw)
Reference substance (positive control):
yes
Remarks:
Carbendazim (CAS 10605-21-7)
Duration:
56 d
Dose descriptor:
NOEC
Effect conc.:
100 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Duration:
56 d
Dose descriptor:
LOEC
Effect conc.:
178 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
reproduction
Duration:
28 d
Dose descriptor:
NOEC
Effect conc.:
>= 316 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Duration:
28 d
Dose descriptor:
LOEC
Effect conc.:
> 316 mg/kg soil dw
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
growth
Results with reference substance (positive control):
Mortality: 2.5 % mortality was observed at the control and at the concentration of 0.32 mg/kg soil (dw), 5 % at 1.0 mg/kg soil (dw) and at 3.2 mg/kg soil dry weight (dw).
Reproduction: Concerning reproduction the statistical analysis (Williams t-test) showed a significant difference between the control and the concentrations of 1.0 and 3.2 mg/kg soil (dw).
Conclusion: The NOECReproduction was determined as 0.32 mg/kg soil (dw) and accordingly the LOECReproduction was determined as 1.0 mg/kg soil (dw). These observed effects are within the range expected from the guideline (1-5 mg Carbendazim/kg soil (dw)) and hence acceptable sensitivity of the test system is assured.

Effects on Mortality, Biomass and Reproduction of Eisenia fetida.

 Concentration

[mg test item/kg soil (dw)]

 Adult mortality

[%]

 Biomass

[% of initial weight]

 Number of Juvenniles

[% of Solvent Control]

 Control  7.5  143.4  96.8
 Solvent Control  1.3  139.5  100.0
 31.6  0.0  142.1  106.0
 56.2  0.0  134.2  106.4
 100  0.0  138.8  91.9
 178  7.5  143.0  80.8*
 316  0.0  136.7  68.7*

 EC50

[mg test item/kg soil (dw)]

   n.a.  n.a.

 NOEC

[mg test item/kg soil (dw)]

   >=316  100

 LOEC

[mg test item/kg soil (dw)]

   >316  178

n.a. = not applicable;

* Significantly different to control (Williams t-test; 1-sided, p ≤ 0.05).

Since inhibition of reproduction was < 50 % compared to the solvent control in all concentrations of the test item tested (31.3 % inhibition was observed in the highest concentration) no EC50 value was calculated.

Validity criteria fulfilled:
yes
Remarks:
mortality of the adults in the solvent control = 1.3 % ; mean number of juveniles in the solvent control = 462.9; CV for the no. of juv. in the solvent control = 14.1
Conclusions:
The long-term toxicity to earthworm (Eisenia fetida) was tested according OECD Guideline 222 (Earthworm Reproduction Test (Eisenia fetida/Eisenia andrei)) and ISO 11268-2 (Effects of Pollutants on Earthworms. 2. Determination of Effects on Reproduction). During 56 days exposure an NOEC(Reproduction) was determined to be 100 mg test item/kg soil (dw) and the LOEC(Reproduction) as 178 mg test item/kg soil (dw).
Executive summary:

This study was designed to comply with the following method:

- OECD Guideline No. 222 for the Testing of Chemicals "Earthworm Reproduction Test (Eisenia fetida/Eisenia andrei)" adopted April 13, 2004

- ISO Guideline 11268-2 "Soil quality - Effects of pollutants on earthworms (Eisenia fetida) Part 2: Determination of effects on reproduction" adopted July 1998

The purpose of this study was to determine a NOEC/LOEC and/or an EC50 for the effects of the test item on the reproduction (56 days after application), mortality and the biomass development (28 days after application) of the earthworm Eisenia fetida (Lumbricidae) by dermal and alimentary uptake using a standardised artificial soil. The test item was tested in five concentrations and was mixed with the substrate. Four replicates per concentration, eight solvent control replicates and additionally four deionised water control replicates (ten worms per replicate) were tested.

7.5 and 1.3 % mortality was observed at the control and the solvent control, respectively. At all concentrations of the test item 0-7.5 % mortality was determined.

Statistical analysis (Williams test; 1-sided, p ≤ 0.05) showed a significant difference concerning the number of juveniles between the solvent control and the concentrations of 178 and 316 mg test item/kg. All other concentrations tested did not differ statistically significant from the solvent control.

Therefore, the NOECReproductionwas determined to be 100 mg test item/kg soil (dw) and accordingly,

the LOECReproductionas 178 mg test item/kg soil (dw).

Since inhibition of reproduction was < 50 % compared to the solvent control in all concentrations of the test item tested (31.3 % inhibition was observed in the highest concentration) no EC50 value was calculated.

Description of key information

The long-term toxicity to earthworm (Eisenia fetida) was tested according OECD Guideline 222 (Earthworm Reproduction Test (Eisenia fetida/Eisenia andrei)) and ISO 11268-2 (Effects of Pollutants on Earthworms. 2. Determination of Effects on Reproduction). During 56 days exposure an NOEC(Reproduction) was determined to be 100 mg test item/kg soil (dw) and the LOEC(Reproduction) as 178 mg test item/kg soil (dw).

Key value for chemical safety assessment

Long-term EC10, LC10 or NOEC for soil macroorganisms:
100 mg/kg soil dw

Additional information