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Short-term toxicity to aquatic invertebrates

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short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
according to guideline
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
not specified
GLP compliance:
yes (incl. QA statement)
Specific details on test material used for the study:
Chemical name: Tetrahydro-imidazo-(4,5d)-lmidazol-2,5-dion.
Batch number: Zwischenprodukt aus Partie 692.
Purity: 97.6% carbon, 100.3% analysis of nitrogen.
Impurities: 0.024% urea, 0.02% water.
Analytical monitoring:
Details on sampling:
Two samples were taken from the test water and analysed to confirm the concentration of the test substance. All of these fell within the agreed parameters.
Test organisms (species):
Daphnia magna
Details on test organisms:
Test Organism
Strain: Daphnia magna STRAUS (1820)
Source: Institut National de Recherche Chimique Appliquée, France.
Age at study initiation: 2-24hrs startinf with the 3rd breed parent animals.
Age of stock animals: 2-4 weeks.

Method of breeding: Not reported.
Feeding during test: The daphnids were not fed throughout the study.
Food type: Not applicable.
Amount: Not applicable.
Frequency: Not applicable.
Test type:
Water media type:
Synthetic fresh water (M4 medium).
Limit test:
Total exposure duration:
48 h
Remarks on exposure duration:
The test was performed in a closed system; glass tubes sealed with gas unpermeable teflobe caps (nominal volume 20mL).
Post exposure observation period:
Visual recording of swimming ability was recorded after 0, 3, 6, 24 and 48 hours.
2.20 - 3.20 mmol/L.
Test temperature:
18 - 22°C; maximum temperature difference 2°C. Temperature was continously measured during the whole test period in a separate vessel close
to the test vessels.
7.5 - 8.5; measured after 0hr in an additional replicate without daphnids and after 48hours in replicate 1 of each tested concentration.
Dissolved oxygen:
7.8 - 9 mg/L; measured after 0hour in an additional replicate without daphnids and after 48hours in replicate 1 of each tested concentration.
Not specified.
550 - 650 uS/cm
Nominal and measured concentrations:
Nominal concentration of the stock solution was 100mg/L. Stock dilution with M4 medium (synthetic freshwater) were prepared and the following concentrations (mg/L) of the test substance were measured: control, 100, 50, 25, 12.5.
Details on test conditions:
Test volume: ~23mL (vials were filled close to the bottom).
Loading (animals/mL): 0.2
Number of animals per vessel: 5.
Total number of animals per concentration: 20.
Number of replicates per concentration: 4.
Number of replicates per control: 4.
Illumination: Artificial light, type warm white (e.g. OSRAM L58 W31).
Photoperiod: 16hrs light: 8hrs dark.
Reference substance (positive control):
48 h
Dose descriptor:
Effect conc.:
>= 100 mg/L
Key result
48 h
Dose descriptor:
Effect conc.:
> 100 mg/L
48 h
Dose descriptor:
Effect conc.:
> 100 mg/L
Details on results:
The results show that there were no mortalities observed in the daphnids across the tested concentration range. The calculated EC50 was >100.0mg/L.
Reported statistics and error estimates:
The test results allowed no statistical evaluation of the data, since no mortalities were observed.
Validity criteria fulfilled:
The test substance did not show any signs of toxicity to aquatic invertebrates Daphnia magna STRAUS.
Executive summary:

A study was performed to determine the acute effect of the test substance on the swimming ability of the water flea Daphnia magna STRAUS 1820 in a 48-hour static test, according to OECD 202 (1984), EEC Directive 92/32 Annex . V Part C.2 (December 1992) and EPA OPPTS 850.1010 (1996). The test was conducted in a closed system (glass tubes sealed with gas unpermeable teflone caps), and the test substance was tested in the range of concentrations; 12.5, 25, 50 and 100 mg/l. Instead of mortality, the test criterion of acute toxicity was immobilization of the daphnids after 48 hours of exposure, which is expressed as EC (effect concentration). Animals considered to be immobile were those which were unable to swim within 15 seconds, even after gentle agitation of the test vessel.

The results showed that the EC0, EC50 and EC100 after 48 hours were all above 100 mg/L. Therefore the test substance is considered non-toxic to aquatic invertebrates under EC 1272/2008 within these study parameters.

Description of key information

To determine the acute toxicity of the test substance in aquatic invertebrates (Daphnia magna STRAUS) the following the guidelines were used; OECD 202 (April 1984), EEC Directive 92/32 Annex . V Part C.2 (December 1992), EPA OPPTS 850.1010 (1996), the tests were carried out in a laboratory with a GLP Certificate. Twenty (20) daphnids were tested per test concentration (12.5, 25, 50, 1000 mg/L) with 4 test replicates each. A control group of 20 daphnids were also tested. The effects of the test substance on the swimming ability (mobility) of the test species were assessed, the test criterion of acute toxicity is immobilisation of the daphnids after 48 hours of exposure.

Observations were recorded at 0, 3, 6, 24, 48hrs post test substance exposure. At the end of the testing period no mortalities were recorded and the test daphnids showed no signs of immobilisation. The effect concentration value (EC50) was >100mg/L based on analytically confirmed nominal concentration. It was therefore concluded that the test substance is non-toxic to fresh water fish in concentrations lower than 100mg/L.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Effect concentration:
100 mg/L

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