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Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016

Materials and methods

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

Test material

Constituent 1
Chemical structure
Reference substance name:
1-methyl-4-(4-methyl-3-pentenyl)cyclohex-3-ene-1-carbaldehyde
EC Number:
257-942-2
EC Name:
1-methyl-4-(4-methyl-3-pentenyl)cyclohex-3-ene-1-carbaldehyde
Cas Number:
52475-86-2
Molecular formula:
C14H22O
IUPAC Name:
1-methyl-4-(4-methylpent-3-en-1-yl)cyclohex-3-ene-1-carbaldehyde
Constituent 2
Chemical structure
Reference substance name:
1-methyl-3-(4-methyl-3-pentenyl)cyclohex-3-ene-1-carbaldehyde
EC Number:
257-941-7
EC Name:
1-methyl-3-(4-methyl-3-pentenyl)cyclohex-3-ene-1-carbaldehyde
Cas Number:
52474-60-9
Molecular formula:
C14H22O
IUPAC Name:
1-methyl-3-(4-methylpent-3-en-1-yl)cyclohex-3-ene-1-carbaldehyde
Specific details on test material used for the study:
Identification: Precyclemone B
Appearance: Clear colourless liquid (determined by Charles River Den Bosch)
Batch: SC00016633
Purity/Composition: 99.5% (sum of two isomers)
Test item storage: In refrigerator (2-8°C) protected from light
Stable under storage conditions until: 31 October 2016 (expiry date)
Chemical name (IUPAC): Reaction mass of 1-methyl-3-(4-methyl-3-pentenyl)cyclohex-3-ene-1-carbaldehyde and 1-methyl-4-(4-methyl-3-pentenyl)cyclohex-3-ene-1-carbaldehyde

Sampling and analysis

Details on sampling:
Samples for possible analysis were taken from all test concentrations and the control according to the schedule below. The method of analysis is described in the Analytical Report.

Frequency: At the start and the end of both 24-hour renewal periods
Volume: 15 mL from the approximate centre of the test vessels
Storage: Samples were stored in a freezer until analysis.

At the end of both refreshment periods, samples were taken from one of the replicates of each concentration.
Additionally, reserve samples of 15 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.

Test solutions

Details on test solutions:
Combined limit/range-finding test (static)
The study started with a combined limit/range-finding test. Twenty daphnids per concentration (four replicates, five daphnids per replicate) were exposed to a control and a WAF at 100 mg/L. Test procedure and conditions were similar to those applied in the final test with the following exceptions:
- The test was performed under static conditions, i.e. no renewal of test solutions during the test;
- Ten daphnids per concentration (two replicates, five daphnids per replicate) were exposed to WAFs at 1.0 and 10 mg/L;
- Dissolved oxygen concentrations and pH were only measured in the control and the highest test concentration;
- Samples for determination of actual exposure concentrations were taken at the start and the end of the test.

Final test (semi-static)
Test concentrations
Precyclemone B: 1.0, 2.2, 4.6, 10, 22 and 46% of a SS at 10 mg/L

A first final test was performed with the same range of concentrations but this test was not valid because too many daphnids were immobilized in the control group.
Control: Test medium without test item or other additives

Test organisms

Test organisms (species):
Daphnia magna
Details on test organisms:
Species Daphnia magna (Crustacea, Cladocera) (Straus, 1820), at least third generation, obtained by a cyclical 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).

Study design

Test type:
semi-static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h

Test conditions

Test temperature:
Temperature is recorded continuously in a temperature control vessel, beginning at the start of the test. 20 - 22°C
pH:
pH is recorded at the start and the end of both renewal periods, for all concentrations and the control. pH: 6.0-8.5,
Dissolved oxygen:
Dissolved oxygen is recorded at the start and the end of both renewal periods, for all concentrations and the control
Nominal and measured concentrations:
Nominal Test concentrations (final test): 1.0, 2.2, 4.6, 10, 22 and 46% of a SS at 10 mg/L
Measured test concentrations (final test): 0.045, 0.10, 0.23, 0.51, 1.1 and 2.4 mg/L

Results and discussion

Effect concentrations
Duration:
48 h
Dose descriptor:
EC50
Effect conc.:
0.15 mg/L
Nominal / measured:
meas. (TWA)
Conc. based on:
test mat.
Basis for effect:
mobility
Details on results:
Analysis of the samples taken at the start of the first renewal period showed measured concentrations of 0.045, 0.10, 0.23, 0.51, 1.1 and 2.4 mg/L in 1.0, 2.2, 4.6, 10, 22 and 46% of a SS at 10 mg/L, respectively. The two lowest concentration decreased to 69 and 37% of initial, respectively, at the end of the 24-hour renewal period. All higher concentrations remained stable (85-97% of initial). Analysis of the samples taken at the start of the second renewal period showed measured concentrations of 0.056, 0.12, 0.25, 0.58, 1.2 and 2.4 mg/L in 1.0, 2.2, 4.6, 10, 22 and 46% of a SS at 10 mg/L, respectively. These concentrations remained stable during the 24-hour renewal period (88-97% of initial at the end of the renewal period), except for the 0.25 mg/L concentration that slightly decreased to 77% of initial. Given this result, the EC50 values were expressed in terms of the average exposure concentrations that were calculated to be: 0.045, 0.086, 0.22, 0.53, 1.1 and 2.3 mg/L.

The study met the acceptability criteria prescribed by the study plan and was considered valid.

The 48h-EC50 was 0.15 mg/L based on average exposure concentrations (95% confidence interval between 0.12 and 0.18 mg/L).

Applicant's summary and conclusion

Validity criteria fulfilled:
yes
Conclusions:
The 48h-EC50 was 0.15 mg/L based on average exposure concentrations (95% confidence interval between 0.12 and 0.18 mg/L).

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