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Please be aware that this old REACH registration data factsheet is no longer maintained; it remains frozen as of 19th May 2023.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

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:
26 November 2008 and 14 January 2009
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study conducted to GLP in compliance with agreed protocols with no deviations from standard test guidelines and/or minor methodological deficiences which do not affect the quality of the relevant results

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guidelineopen allclose all
according to guideline
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
according to guideline
EU Method C.2 (Acute Toxicity for Daphnia)
GLP compliance:
yes (incl. QA statement)

Test material

Sampling and analysis

Analytical monitoring:
Details on sampling:
The test material concentration in the test samples was determined spectrophotometrically using an external standard. The method was developed by the Department of Analytical Services, Harlan Laboratories Ltd, Shardlow, UK. A volume of test sample was diluted with methanol to give a final theoretical concentration of 5 mg/l. Standard solutions of test material were prepared in methanol at a nominal concentration of 5 mg/l.

- Sample storage conditions before analysis:
Room temperature in the dark
Duplicate samples were taken and stored at approximately -20°C for further analysis if necessary.

Test solutions

Details on test solutions:
For the purpose of the definitive test the test material was dissolved directly in reconstituted water. An amount of test material (100 mg) was dissolved in reconstituted water and the volume adjusted to 1 litre to give the 100 mg/I test concentration. The prepared concentration was inverted several times to ensure adequate mixing and homogeneity. The concentration and stability of the test material in the test preparations was verified by chemical analysis at 0 and 48 hours

Details of the reconstituted water
Stock Solutions
a) CaCI2.2H20 11.76 g/I
b) MgSO4.7H20 4.93 g/I
c) NaHCO3 2.59 g/I
d) KCI 0.23 g/I

An aliquot (25 ml) of each of solutions a-d was added to each litre (final volume) of deionised water with a conductivity of <5 pS cm-l. The reconstituted water had a pH of 7.8 ± 0.2 adjusted (if necessary) with NaOH or HCI and was aerated until the dissolved oxygen concentration was approximately air-saturation value.

The reconstituted water had an approximate theoretical total hardness of 250 mg/I as CaCO3.

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
- Common name: Water Flea
- Strain: Daphnia magna
- Source: in-house cultures
- Age at study initiation (mean and range, SD): Ist instar – less than 24 h
- Method of breeding: Parthenogenises
- Feeding during test Each culture was fed daily with a suspension of algae (Chlorella sp.).

- N/A test organisms derived from in-house cultures

Adult Daphnia were maintained in polypropylene vessels containing approximately 2 litres of reconstituted water in a temperature controlled
room at approximately 20°C. The lighting cycle was controlled to give a 16 hours light and 8 hours darkness cycle with 20 minute dawn and
dusk transition periods.

Study design

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

Test conditions

250 mg/L CaCO3
Test temperature:
20 °C
7.8 +/- 0.2
Dissolved oxygen:
Oxygen concentration of test vessels at 0 hours (fresh media) was observed to have an air saturation value (ASV) in excess of 100%
Nominal and measured concentrations:
Based on the results of the range-finding test a "Limit test" was conducted at a concentration of 100 mg/I to confirm that at the maximum concentration given in the OECD/EEC Test Guidelines, no immobilisation or adverse reactions to exposure were observed

Analysis of the test preparations at 0 and 48 hours showed measured test concentrations to be near nominal and so it was considered justifiable to estimate the EC50 values in terms of the nominal test concentrations only.

Details on test conditions:
- Test vessel:
- Type (delete if not applicable): closed
- Material, size, headspace, fill volume: 250 mL glass jar with 200 mL test preparation, covered
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

- Source/preparation of dilution water: reconstituted water

- Photoperiod: 16 hours light : 8 hours dark, 20 minute dawn/dusk periods
Reference substance (positive control):
potassium dichromate

Results and discussion

Effect concentrationsopen allclose all
24 h
Dose descriptor:
Effect conc.:
> 100 mg/L
Nominal / measured:
Basis for effect:
48 h
Dose descriptor:
Effect conc.:
> 100 mg/L
Nominal / measured:
Basis for effect:
Details on results:
Test media was observed to be dark pink clear throughout duration of tests.
The No Observed Effect Concentration (NOEC) after 24 and 48 hours exposure was 100 mg/L. The NOEC is based upon no significant immobilsation at this concentration.
Acute toxicity of the test material to the freshwater invertebrate Daphnia gave a 48-hour EC50 of greater than 100 mg/L. The NOEC was 100 mg/L
Results with reference substance (positive control):
3 hour EC50 >3.2 mg/L (value not calculated as less than 50% immobilisation occurred at this time point)
24 hour EC50 0.82 mg/L (95% Confidence Limits 0.71-0.94 mg/L)
48 hour EC50 0.71 mg/L (95% Confidence Limits 0.61-0.81 mg/L)

No Observed Effect Concentration after 24 and 48 hours were 0.32 and 0.32 mg/L respectively. The NOEC is based upon zero immobilisation at this concentration.

Any other information on results incl. tables

Cumulative immobilisation data from the exposure ofDaphnia magnato the test material during the definitive test are shown below


There was no immobilisation in 20 daphnids exposed to a test concentration of 100 mg/I for a period of 48 hours. Inspection of the immobilisation data gave the following results:

Time (h)

EC50 (mg/I)

95% Confidence limits




> 100

> 100

The No Observed Effect Concentration after 24 and 48 hours exposure was 100 mg/I. The No Observed Effect Concentration is based upon no significant immobilisation at this concentration.

Applicant's summary and conclusion

Validity criteria fulfilled:
The detection system was found to have acceptable linearity. The analytical procedure had acceptable recoveries of test material in test medium. A method of analysis was validated and proven to be suitable for use.
The acute toxicity of the test material to the freshwater invertebrate Daphnia magna has been investigated and gave a 48-Hour EC50 of greater than 100 mg/l. Correspondingly the No Observed Effect Concentration was 100 mg/l.
Executive summary:

In an Acute Toxicity to Daphnia Magnastudy (Harlan project number: 0959/0231) the test material was found to have an EC50 of greater than 100 mg/l. Correspondingly the No Observed Effect Concentration was 100 mg/l. Testing was performed in accordance with the methods described in the OECD Guidelines for Testing of Chemicals (April 2004) No 202,"Daphnia sp, Acute Immobilisation Test" referenced as Method C.2 of Commission Regulation (EC) No 440/2008.