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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:
27 - 29 Mar 2012
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
according to guideline
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
according to guideline
EU Method C.2 (Acute Toxicity for Daphnia)
Version / remarks:
GLP compliance:
yes (incl. QA statement)
Sächsische Staatsministerium für Umwelt und Landwirtschaft, Germany
Analytical monitoring:
Details on sampling:
- Concentrations: 0 and 100 mg/L
- Sampling: at the start and at the end of the test
- Sample storage conditions before analysis: samples were frozen
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: A stock solution was prepared by weighing of 53.0 mg test item and filling with dilution water up to 500 mL (= stock solution A, corresponding to 106.0 mg test item/L). The test solution of the chosen concentration was prepared directly in the test vessels by dilution of the stock solution A with dilution water immediately before the transfer of the Daphnia into the test vessels.
- Eluate: no
- Differential loading: no
- Controls: yes, test medium control without test substance
Test organisms (species):
Daphnia magna
Details on test organisms:
- Source: Landesanstalt für Umweltschutz Baden-Württemberg, Karlsruhe, Germany; holding and breeding in the test facility under standardised laboratory conditions test animals derived from a healthy stock, not first brood progeny
- Age at study initiation (mean and range, SD): < 24 hours
- Culturing conditions: in glass beakers (50 mL) under the same conditions as in the test; test animals were separated from the adults by means of a pipette
- Feeding during test: none
Test type:
Water media type:
Limit test:
Total exposure duration:
48 h
2 mmol/L CaCO3
Test temperature:
19.7 - 21.1 °C
7.63 - 7.84
Dissolved oxygen:
8.05 - 8.96 mg O2/L
Nominal and measured concentrations:
Nominal: 0 and 100 mg/L
Recoveries: 90.2 at 0 h and 102.3% at 72 h
Details on test conditions:
- Test vessel:
- Material, size, headspace, fill volume: glass beaker, test volume 10 mL
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4
- Biomass loading rate: 2 mL test solution per Daphnia

- Source/preparation of dilution water: reconstituted water according to ISO 6341, prepared 7 days in advance
- Conductivity of deionised water: ≤ 10 μS/cm
- Culture medium different from test medium: yes
- Intervals of water quality measurement: temperature recorded continuously, pH and dissolved oxygen at the start and at the end

- Adjustment of pH: no
- Photoperiod: 16 h light : 8 h dark, daily
- Light intensity: approx. 1000 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
number of immobilised Daphnia per vessel (including discoloration, anomalies), 24 and 48 hours after start of exposure
Reference substance (positive control):
potassium dichromate
48 h
Dose descriptor:
Effect conc.:
> 100 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
48 h
Dose descriptor:
Effect conc.:
>= 100 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
Details on results:
- No immobilisation was observed during the study
- Behavioural abnormalities: At any assessment no visible signs of abnormalities of surviving daphnids were observed.
- Mortality of control: none
- Other adverse effects control: none
Results with reference substance (positive control):
The reference item potassium dichromate was tested in a separate study at 0.88, 1.14, 1.48, 1.92 and 2.50 mg/L to control the sensitivity of the test system (the most recent study 06.12. – 08.12.2011)
EC50 (48 h) = 1.22 mg/L (95% CI 1.13 - 1.32 mg/L)
The reference item EC50 values are within the expected range of toxicity for reference toxicity tests performed at the test facility (range of 1.0 – 2.0 mg/L).
Reported statistics and error estimates:
Because a limit test was performed, a calculation and statistical evaluation was not applicable.

The measured concentrations of the susbtance in the test solutions remained at 90% of nominal values at the start of the test and at 102% at the end of the test (after 48 h). Therefore the toxicity results are based on the nominal concentrations.

Validity criteria fulfilled:

Description of key information

EC50 (48 h): > 100 mg/L (Daphnia magna; OECD 202)

Key value for chemical safety assessment

Additional information

One study is available on the short-term toxicity of Reaction mass of 2-(3,4-dimethyl-1H-pyrazol-1-yl)succinic acid and 2-(4,5-dimethyl-1H-pyrazol-1-yl)succinic acid to aquatic invertebrates. The study was conducted as a limit test according to OECD Guideline 202 and GLP (Juckeland, 2012). The test organism Daphnia magna was exposed to the test substance at a nominal concentration of 100 mg/L for 48 hours. No immobilisation was observed during the study, and the EC50 is thus determined to be > 100 mg/L (nominal).