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Short-term toxicity to aquatic invertebrates

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Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Study period:
14 March 2012 to 09 August 2012
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reference:
Composition 1
Qualifier:
according to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Qualifier:
according to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Qualifier:
according to
Guideline:
ISO 6341 15 (Water quality - Determination of the Inhibition of the Mobility of Daphnia magna Straus (Cladocera, Crustacea))
Qualifier:
according to
Guideline:
other: Guidance document on aquatic toxicity testing of difficult substances and mixtures, OECD series on testing and assessment number 23, December 14, 2000.
GLP compliance:
yes
Test material information:
Composition 1
Analytical monitoring:
yes
Details on sampling:
During the final test singular samples for possible analysis were taken from all test concentrations and the control according to the following schedule:
- Frequency: at t=0 h and t=48 h
- Volume: 3 mL from the approximate centre of the test vessels
- Storage Samples were stored in a freezer until analysis.
At the end of the exposure period, the replicates were pooled at each concentration before sampling. Additionally, reserve samples of 3 mL were taken for possible analysis. If not used, these samples were stored in a freezer for a maximum of three months after delivery of the draft report, pending on the decision of the sponsor for additional analysis.
Details on test solutions:
Preparation of test solutions:
The standard test procedures required generation of test solutions, which should contain completely dissolved test substance concentrations or stable and homogeneous mixtures or dispersions. The testing of concentrations that disturb the test system should be prevented (e.g. film of the test substance on the water surface). The batch of CP Formate tested was a clear colourless liquid with a purity of 94.1% and the substance was not completely soluble in test medium at the higher loading rates prepared. Preparation and weighing were as much as possible performed under dimmed light conditions. Test solutions at loading rates of 1 mg/L and higher were all individually prepared. Preparation included one day of magnetic stirring followed by a settlement period of ~1 hour. This generally resulted in a clear and colourless solution with some floating material at loading rates of 10 mg/L and higher. The Water Accommodated Fraction (WAF) was then collected from the middle of the solutions thus removing the floating material from the end test solutions. The solutions without a floating layer were used as such. All final test solutions were clear and colourless. Note that in addition a ten-fold dilution was prepared from the 1 mg/L solution for testing in the combined limit/range-finding test. All was performed in closed units as to minimise any possible loss due to the expected volatile character of the test substance.
Test organisms (species):
Daphnia magna
Details on test organisms:
- Species: Daphnia magna (Crustacea, Cladocera) (Straus, 1820), at least third generation, obtained by acyclical parthenogenesis under specified breeding conditions.
- Source: In-house laboratory culture with a known history.
- Reason for selection: This system has been selected as an internationally accepted invertebrate species.
- Validity of batch: Daphnids originated from a healthy stock, 2nd to 5th brood, showing no signs of stress such as mortality >20%, presence of males, ephippia or discoloured animals and there was no delay in the production of the first brood.
- Characteristics: For the test selection of young daphnids with an age of < 24 hours, from parental daphnids of more than two weeks old.
BREEDING:
- Start of each batch: With newborn daphnids, i.e. less than 3 days old, by placing about 250 of them into 5 litres of medium in an all-glass culture vessel.
- Maximum age of the cultures: 4 weeks.
- Renewal of the cultures: After 7 days of cultivation half of the medium twice a week.
- Temperature of medium: 18-22°C.
- Feeding: Daily, a suspension of fresh water algae.
- Medium: M7, as prescribed by Dr. Elendt-Schneider (Elendt, B.-P., 1990: Selenium deficiency in Crustacea. An ultrastructural approach to antennal damage in Daphnia magna Straus. Protoplasma 154, 25-33).
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Hardness:
180 mg/l expressed as CaCO3
Test temperature:
19.3-20.4 °C
pH:
7.7-8.0
Dissolved oxygen:
8.3-9.6 mg/L
Salinity:
No details in study report
Nominal and measured concentrations:
- Nominal concentrations: WAFs prepared at loading rates of 4.6, 10, 22, 46 and 100 mg/L.
- Measured concentrations (average t=0-48): 2.9, 6.9, 15, 32 and 34 mg/L.
Details on test conditions:
Test procedure and conditions:
- Test vessels: 100 mL, all-glass, closed with a screw cap and minimal headspace.
- Medium: Adjusted ISO medium.
- Number of daphnids: 20 per concentration.
- Loading: 5 per vessel containing 100 mL of test solution (completely filled).
- Light: No light.
- Feeding: No feeding.
- Aeration: No aeration of the test solutions.
- Introduction of daphnids: Within 40 minutes after preparation of the test solutions.

Measurements and recordings:
Immobility (including mortality): At 24 hours and at 48 hours.
pH and dissolved oxygen: At the beginning and at the end of the test, for all concentrations and the control.
Temperature of medium: Continuously in a temperature control vessel, beginning at the start of the test.
Reference substance (positive control):
yes
Remarks:
Potassium dichromate
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
7.7 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
mobility
Remarks on result:
other: 95% C.L.: 6.7-9.5 mg/L
Details on results:
In the control 10% mortality was observed which was within the accepted guideline criterion of ≤10%. At a concentration of 2.9 mg/L no signs of immobility could be detected . First at 6.9 mg/L a 25% immobility response was noted which increased to 100% at all higher concentrations. The responses recorded were in agreement with what was expected based on the results of the combined limit/range-finding test.
Results with reference substance (positive control):
The 48h-EC50 was 0.28 mg/L with a 95% confidence interval between 0.14 and 0.65 mg/L.
Reported statistics and error estimates:
The EC50-value was calculated at 24 and 48 hours of exposure from the probits of the percentages of affected daphnids and the logarithms of the corresponding test substance concentrations using the maximum likelihood estimation method.
Validity criteria fulfilled:
yes
Conclusions:
The 48-h EC50 value is 7.7 mg/L in Daphnia magna.
Executive summary:

A GLP-compliant OECD 202 guideline study with aquatic invertebrates (Daphnia magna) is available for CP Formate. Daphnids (20/concentration) were exposed to WAFs prepared at loading rates of 4.6, 10, 22, 46 and 100 mg/L under static conditions for 48 hours. Exposure was performed in vessels closed with a screw cap and minimal headspace in order to minimize the loss of the test substance due to evaporation. Analysis of the samples taken at the start of the final test showed measured concentrations of 3.19, 7.65, 16.0, 35.1 and 39.4 mg/L in the WAFs prepared at 4.6, 10, 22, 46 and 100 mg/L. During the exposure period the measured concentrations decreased slightly to respectively 2.58, 6.19, 13.9, 28.6 and 28.6 mg/L, i.e. by 13 to 27% below the initial measured concentrations. The average exposure concentrations (t=0 -48h; geometric mean) were calculated to correspond to 2.9, 6.9, 15, 32 and 34 mg/L. In the control 10% mortality was observed which was within the accepted guideline criterion of ≤10%. At a concentration of 2.9 mg/L no signs of immobility could be detected. First at concentration of 6.9 mg/L, a 25% immobility response was noted which increased to 100% at all higher concentrations. The study met the acceptability criteria prescribed by OECD guideline (202) and was considered valid. Based on these data, the 48 -h EC50 value is determined at 7.7 mg/L (95% C.L. is 6.7 -9.5).

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Justification for type of information:
See 'Attached justification'
Reason / purpose:
read-across source
Related information:
Composition 1
Test material information:
Composition 1
Key result
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
8.5 mg/L
Remarks on result:
other: as determined from read-across to CP Formate

Description of key information

The short-term toxicity to aquatic invertebrates is assessed based on read-across from the close structural analogue CP Formate (CAS# 25225-08-5) and the EC50 value determined at 8.5 mg/L.

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
8.5 mg/L

Additional information

A GLP-compliant OECD 202 guideline study with aquatic invertebrates (Daphnia magna) is available for CP Formate. Daphnids (20/concentration) were exposed to WAFs prepared at loading rates of 4.6, 10, 22, 46 and 100 mg/L under static conditions for 48 hours. Exposure was performed in vessels closed with a screw cap and minimal headspace in order to minimize the loss of the test substance due to evaporation. Analysis of the samples taken at the start of the final test showed measured concentrations of 3.19, 7.65, 16.0, 35.1 and 39.4 mg/L in the WAFs prepared at 4.6, 10, 22, 46 and 100 mg/L. During the exposure period the measured concentrations decreased slightly to respectively 2.58, 6.19, 13.9, 28.6 and 28.6 mg/L, i.e. by 13 to 27% below the initial measured concentrations. The average exposure concentrations (t=0 -48h; geometric mean) were calculated to correspond to 2.9, 6.9, 15, 32 and 34 mg/L. In the control 10% mortality was observed which was within the accepted guideline criterion of ≤10%. At a concentration of 2.9 mg/L no signs of immobility could be detected. First at concentration of 6.9 mg/L, a 25% immobility response was noted which increased to 100% at all higher concentrations. The study met the acceptability criteria prescribed by OECD guideline (202) and was considered valid. Based on these data, the 48 -h EC50 value is determined at 7.7 mg/L (95% C.L. is 6.7 -9.5).