dimethyl 1,2-benzenedicarboxylate;reaction mass of: 1,2-dimethylpropylidene dihydroperoxide

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Test samples for analysis were taken from the approximate centre of the test vessels according to the following sampling schedule:

- Frequency
At t= 0 h from freshly prepared and at t= 24 h from 24h-old solutions of all test concentrations. At t =72 h from freshly prepared and at t=96 h from 24h-old solutions from 0, 10 and 18 mg/L.
- Volumes
0,10 and 18 mg/I: 6 ml
32 mg/I: 3 ml
56 mg/I: 2 ml
100 mg/I: 1 ml
- Storage
Samples were stored in a deep-freeze until analysis together with freshly taken samples at t=96 hours.
Additionally, reserve samples of 12 ml were taken from all test solutions at t=O and t= 24 hand from the test concentrations with surviving fish at t=72 and t=96 h for possible analysis. If not already used, these samples were stored in a freezer for possible analysis until delivery of the final report with a maximum of three months.

Vehicle:

no

Details on test solutions:

Preparation of test solutions started with stock solutions at nominally 100 mg/1. These solutions were magnetically stirred for 15 to 20 minutes following treatment with ultrasonic waves for 5 minutes during the range-finding test. The resulting, clear and colourless, stock solutions were then used to prepare the lower test concentrations by subsequent dilutions in test medium. The test solutions were daily renewed during the final test.

Test organisms (species):

Cyprinus carpio

Details on test organisms:

- Species: Carp (Cyprinus carpio, Teleostei, Cyprinidae) (Linnaeus, 1758)
- Source: Zodiac, proefacc, "De Haar Vissen", L.U. Wageningen, the Netherlands.
- Mean length: Static range-finding test: 2.5 ± 0.16 cm; Semi-static final test: 2.1 ± 0.18 cm
- Mean weight: Static range-finding test: 0.50 ± 0.12 g; Semi-static final test: 0.13 ± 0.04 g
- Characteristics: F1 from a single parent-pair bred in UV-treated water.
- Feeding: daily with Trouvit

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Hardness:

250 mg/L as CaCO3

Test temperature:

20.9 - 21.8 °C

pH:

7.4 - 7.8

Dissolved oxygen:

7.5 - 9.0 mg/L

Nominal and measured concentrations:

Nominal: 10, 18, 32, 56, 100 mg/L
Measured concentrations were close to nominals, within 80 to 120% of the nominals, therefore nominal concentrations are used.

Details on test conditions:

TEST SYSTEM
- Test vessel: 5.5 L all-glass
- Material, size, headspace, fill volume: 4 L test medium
- Aeration: no

- Renewal rate of test solution (frequency/flow rate): every 24h
- No. of organisms per vessel: 7
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: 0.23 g fish/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO-medium
- Ca/mg ratio: 20:3
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 16 h

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Mortality

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.8
- Range finding study
- Test concentrations: 0.1, 1.0, 10 AND 100 mg/L
- Results used to determine the conditions for the definitive study: All fish died within 3 hours at 100 mg/L and no fish died in the other test concentrations.

Reference substance (positive control):

yes

Remarks:

Pentachlorophenol

Key result

Duration:

96 h

Dose descriptor:

LC50

Effect conc.:

16 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 95% C.I. 14 - 21 mg/L

Key result

Duration:

96 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities:
18 mg/L: 24h and 72-96h swimming at the surface
32 mg/L: 3 3/4h swimming at the surface
56 mg/L: 3 3/4h immobile
- Other biological observations:
10 mg/L: 72-96h discoloured
18 mg/L: 24-96h discoloured
32 mg/L: 3 3/4h discoloured
56 mg/L: 3 3/4h discoloured
- Mortality of control: No mortality
- Other adverse effects control: No
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No
- Effect concentrations exceeding solubility of substance in test medium: No

Results with reference substance (positive control):

Results as expected

Reported statistics and error estimates:

the maximum likelihood estimation method with the probits of the percentages of dead fish as function of the logarithms of the corresponding concentrations (Finney, D.J., 1971: Probit analysis, Cambridge University Press, Cambridge, U.K., 3rd edition)

Sublethal observations / clinical signs:

Conc. (mg/L)	Initial no. fish	Cumulative mortality					Total mortality (%)
		3¾h	24h	48h	72h	96h
Control	7	0	0	0	0	0	0
10	7	0	0	0	0	0	0
18	7	0	1	5	5	5	71
32	7	0	7	7	7	7	100
56	7	2	7	7	7	7	100
100	7	7	7	7	7	7	100

Validity criteria fulfilled:

yes

Conclusions:

The 96h-LC50 was already reached within 48 hours of exposure being 16 mg/l with a 95% confidence interval between 14 and 21 mg/l.

Executive summary:

96-Hour Acute Toxicity Study in carp.

The study procedures described in this report were based on the EEC directive 92/69; Part C: methods for the determination of ecotoxicity, Publication No. L383, December 1992, C.1 . "Acute toxicity for fish", and the OECD guideline No. 203: "Fish Acute Toxicity Test", Adopted 17 July, 1992.

The test substance was completely miscible with test medium at the concentrations tested.

The project started with a static range-finding test exposing three carp per test group to nominal test substance concentrations of 0.1, 1.0, 10 and 100 mg/l. All fish exposed to nominally 100 mg/l died within 3 hours of exposure. No mortality was observed at the lower test concentrations. Hence, the 96h-LC50 for carp was expected to be between 10 and 100 mg/l. Analytical results showed that the measured concentrations of both main components decreased by more than 20% during the test period. It was decided to continue testing applying a semi-static test design with daily renewal of test solutions as concentrations did not decrease by more than 20% during the first 24-hour test period.

The project was continued with a final LC50 study exposing seven carp per concentration to test substance concentrations ranging from 10 to 100 mg/l, increasing with a factor of 1.8. All test solutions were daily renewed. Samples for analysis were taken at the start and after 72 hours of exposure from freshly prepared solutions and from 24-hour old test solutions at 24 and 96 hours of exposure.

Analysis of the samples taken during the final test showed that the measured concentrations (based on both components) were in agreement with nominal in the freshly prepared solutions at the start of exposure (88-101 %) and the freshly prepared solutions at 72 hours of exposure (93-99%). This indicated that preparation procedures were adequate and repeatable. During the 24-hour periods between renewals the concentrations measured did not decrease by more than 20% below initial. In addition, the average exposure concentrations all remained above 80% relative to nominal. Consequently, the calculated toxicity parameters were based on the nominal test concentrations. In the control group no fish died, and all test conditions (pH, oxygen and temperature) remained within the ranges prescribed by the protocol.

The test substance induced no mortality in carp at or below nominally 10 mg/l.

The 3%h-LC50 was 60 mg/l with a 95% confidence interval between 52 and 81 mg/l.

The 24h-LC50 was estimated to be 21 mg/l with 14% mortality at 18 mg/l and 100% mortality at 32 mg/l (regression line: log10(conc.)=1.01 +(probit-2.61 )/7.73).

The 96h-LC50 was already reached within 48 hours of exposure being 16 mg/l with a 95% confidence interval between 14 and 21 mg/l.

Description of key information

The 96h-LC50 was already reached within 48 hours of exposure being 16 mg/l

Key value for chemical safety assessment

Fresh water fish

Fresh water fish

Effect concentration:

16 mg/L