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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
experimental starting date: 2015-05-20; experimental completion date: 2015-05-22
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference Type:
study report
Report Date:

Materials and methods

Test guideline
according to
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
resp. EU Method C.2 (Acute Toxicity for Daphnia)
GLP compliance:
yes (incl. certificate)
Landesamt für Umwelt, Wasserwirtschaft und Gewerbeaufsicht; Kaiser-Friedrich-Straße 7; 55116 Mainz; Germany

Test material

Test material form:
other: viscous liquid
Details on test material:
Stabilized EMIM BF4:
Appearance: lightly yellow clear liquid
Purity: 93.36% (EMIM+, HPLC); 86.68% (BF4-, titration); 4.89% (Stabilizer, HPLC)

Sampling and analysis

Analytical monitoring:
Details on sampling:
limit test at the concentration containing 100mg/L test item

Test solutions

Details on test solutions:
100.9 mg/L test item in dilution water.
Dilution water:
Deionised water with an enrichment of certain minerals (as demanded in the guidelines):

Parameter Concentration in mg/L
CaCl2*2H2O 293.80
MgSO4*7H2O 123.30
NaHCO3 64.80
KCl 5.80
Resulting hardness in mmol/L: 2.502
Resulting hardness in mg CaCO3/L: 250

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
Variety STRAUS
Strain Berlin
Sex female
Age between 0 and 24 hours
Origin Umweltbundesamt Berlin
In-house breeding since 27. September 2007

Animal Husbandry
Daphnia magna is bred in the LAUS GmbH throughout the year. The animals are kept for the use in toxicity tests. They multiply by parthenogenesis, thus being genetically identical. The husbandry is performed similar to the method described in EN ISO 6341, following SOP 115 002 01 („Zucht und Hälterung von Daphnia magna STRAUS“), edition 11, adopted 15. January 2013.

Vessels preserving glasses, nominal volume 2 L
Medium M4-Medium (recipe of ELENDT)
Food unicellular green algae (Desmodesmus subspicatus)
Medium renewal twice a week
Photo period 16/8 hours, using neon tubes
Temperature 20 +/- 2 °C

Study design

Test type:
Water media type:
Limit test:
Total exposure duration:
48 h

Test conditions

0 h: 7.6
48 h: 7.8
Dissolved oxygen:
0h: 8.7 mg/L
48h: 8.9 mg'/L
Nominal and measured concentrations:
Nominal Concentration Measured Concentration % of Nominal Concentration Measured Concentration % of Nominal Concentration
t=0h t=0h t=48 h t=48h

mg/L mg/L % mg/L %

0 n. d. -- n. d. --
100 100 100 110 110
Details on test conditions:
Date of performance 20. – 22. May 2015
Treatment 100 mg/L
Temperature 18.5 – 20.3 °C
Duration 48 hours
Observation times 24 and 48 hours
Feeding none
Lighting diffuse lighting
Test vessels glass beakers, nominal volume 50 mL, tall shape
Replicates 4 vessels, each containing 20 mL test solution and 5 Daphnia
Control 4 vessels, each containing 20 mL dilution water and 5 Daphnia
Reference substance (positive control):
Potassium dichromate K2Cr2O7 (CAS No. 7778-50-9)

Results and discussion

Effect concentrations
48 h
Dose descriptor:
Effect conc.:
> 100 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
Details on results:
In the control and the treatment, none of the Daphnia were immobilised

Nominal Concentration in mg/L Immobility 24 hours Immobility 48 hours
absolute in % abs. in %
Blank control 0 0 0 0 0 0 0 0 0 0
100 0 0 0 0 0 0 0 0 0 0
Results with reference substance (positive control):
Potassium dichromate K2Cr2O7 (CAS No. 7778-50-9) was used as positive control in a current reference study (14070901R201). For the estimation of the EC50s of test item and positive control, the fits showed sufficient statistical correspondence of the data with the dose-response-equation. The 24h-EC50 value was determined as 1.2 mg/L, lying within the demanded range of 0.6 – 1.7 mg/L.

Applicant's summary and conclusion

Validity criteria fulfilled:
20 Daphnia were exposed to the test item and the control for 48 hours in a static test system. The treatment showed no toxicity.
Executive summary:

One valid experiment was performed.

The study was performed as a limit test at the concentration containing 100 mg/L. For the test concentration and the control, 20 Daphnia were exposed to the test item for 48 hours in a static test system. After 24 and 48 hours, the immobilised Daphnia were counted.

The concentration showed no toxicity. None of the animals were immobilised in the control.

Potassium dichromate K2Cr2O7(CAS No. 7778-50-9) was used as positive controlin a current reference study (14070901R201).The 24h-EC50 value was determined as 1.2 mg/L, lying within the demanded range of 0.6 – 1.7 mg/L.


At the beginning and at the end of the test, the content of the test item in the test solutions was determined using HPLC-MS-determination. The recovery after 48 hours was 110 % of the start concentration, the correlation between nominal and measured concentration was good. Therefore, the determination of the biological results was based on the nominal concentration.

The following results were determined for the test itemstabilized EMIM BF
4(species:Daphnia magna).


24h-NOEC ≥ 100 mg/L
48h-NOEC ≥ 100 mg/L
h-EC50 > 100 mg/L
48h-EC50 > 100 mg/L