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

Short-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

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:

other: GLP study according to protocol. Not all details on chemical analysis are given but enough to validate the study.

Qualifier:

according to guideline

Guideline:

ISO 6341 (Water quality - Determination of the Inhibition of the Mobility of Daphnia magna Straus (Cladocera, Crustacea))

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

samples for analysis were taken from all tst concentrations and the control according to the following sampling schedule:

- Frequency
At t= 0 h from freshly prepared and at t= 24 h from 24h and 48h 24h-old solutions of all test concentrations.
- Volumes
0,10 and 18 mg/I: 6 ml
32 mg/I: 3 ml
56 mg/I: 2 ml
100 mg/I: 1 ml (only from fresh t-0h and old t-24h samples)
- Storage
Samples were stored in a deep-freeze until analysis.
Additionally, reserve samples of 12 ml were taken from all test solutions. 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/l. 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. Test solutions were renewed after 24 hours of exposure in the final test. Most test solutions used originated from the simultaneously performed study with carp (NOTOX Project 338761). The solutions of 32 and 56 mg/I were separately prepared for the renewal at 24 hours.

Test organisms (species):

Daphnia magna

Details on test organisms:

- Species: Daphnia magna (Crustacea, Cladocera) (Straus, 1820)
- Validity of batch: Frequent inspection of the cultures with respect to the number of young, appearance of young and parental daphnia and global feeding behaviour.
- Characteristics: For the test selection of young daphnia with an age of < 24 hours.
- Start of each batch: With new-born animals, Le. less than 3 days old, by placing about 250 of them into 10 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, constant within ± 1°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:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Hardness:

250 mg/l as CaCO3

Test temperature:

20.3-20.4 °C in the control

pH:

7.7-7.9

Dissolved oxygen:

8.5-8.9

Nominal and measured concentrations:

Nominal: 10, 18, 32, 56 and 100 mg/l
During the 24-hour periods, for and after renewal, the concentrations measured did not decrease by more than 20% below initial. In addition, the average exposure concentrations all remained well above 80% relative to nominal. Consequently, the calculated toxicity parameters were based on the nominal test concentrations.

Details on test conditions:

TEST SYSTEM
- Test vessel: 100 ml, all glass
- Material, size, headspace, fill volume: 80 ml test medium
- Aeration: no
- Renewal rate of test solution (frequency/flow rate): every 24h
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO medium
- Culture medium different from test medium: yes

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

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

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 daphnids died within 24h in 100 mg/l, no daphnids died in the other concentrations.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

34 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 95% C.I. 29-41 mg/l

Key result

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

18 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mobility

Results with reference substance (positive control):

Results as expected

Reported statistics and error estimates:

The ECso-value was calculated at 24 and 48 hours of exposure from the probits of the percentages of affected daphnia and the logarithms of the corresponding test substance concentrations using the maximum likelihood estimation method (Finney, D.J., 1971: Probit analysis, Cambridge University Press, Cambridge, U.K., 3rd edition).

Conc. (mg/l)	Vessel no.	No. Daphnia exposed	Response at 24h *		Response at 48h
			Number	%	Number	%
Control	A B	10 10	0 0	0 0	0 0	0 0
10	A B	10 10	0 0	0 0	0 0	0 0
18	A B	10 10	0 0	0 0	1 0	10 0
32	A B	10 10	1 0	10 0	3 5	30 50
56	A B	10 10	9 (3) 8 (1)	90 80	9 9	90 90
100	A B	10 10	10 10	100 100	10 10	100 100

* Between brackets the number of daphnids trapped at the surface

Validity criteria fulfilled:

yes

Conclusions:

The 48h-EC50 was 34 mg/l with a 95% confidence interval between 29 and 41 mg/l.

Executive summary:

Acute Toxicity Study in Daphnia magna.

The study procedures described in this report were based on the ISO International Standard 6341: 'Water quality - Determination of the inhibition of the mobility of Daphnia magna Straus - Acute toxicity test, Third edition, 1996-04-01. In addition, the procedures were designed to meet the test methods and validity criteria of the EEC directive 92/69, Part C: Methods for the determination of ecotoxicity, Publication No. L383, December 1992, C.2. "Acute Toxicity for Daphnia", and the OECD guideline No. 202 Part I: "Daphnia sp., Acute Immobilisation Test", Adopted April 4, 1984.

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

The project started with a static range-finding test with daphnia exposed to nominal concentrations of 0.1 to 100 mg/l, increasing with a factor of 10. After 24 hours of exposure all organisms exposed to 100 mg/l became immobilised. No further immobilisation occurred during the remaining test period in any of the lower test concentrations. The EC50 was expected to be between 10 and 100 mg/l.

Analysis of the samples taken during the range-finding test of the simultaneously performed study with carp (NOTOX Project 338761) showed that the measured concentrations of both major components present in the test substance decreased by more than 20% during the test period. It was decided to continue testing applying a semi-static test design with renewal of test solutions after 24 hours as concentrations did not decrease by more than 20% during the first 24-hour test period.

The range-finding test was followed by a semi-static final EC50 test with renewal of test solutions after 24 hours of exposure. In the final EC50 test Daphnia were exposed for a maximum of 48 hours to nominal concentrations of 10, 18, 32, 56 and 100 mg/l. A blank-control was also included. The test was performed in duplicate with 10 daphnia per vessel. Samples for analysis of actual exposure concentrations were taken from the freshly prepared solutions at the start and after 24 hours of exposure and from the 24-hour old solutions after 24 and 48 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 24 hours of exposure (93-98%).

This indicated that preparation procedures were adequate and repeatable.

During the 24-hour periods, for and after renewal, the concentrations measured did not decrease by more than 20% below initial. In addition, the average exposure concentrations all remained well above 80% relative to nominal. Consequently, the calculated toxicity parameters were based on the nominal test concentrations.

In the control, no daphnia became immobilised or trapped at the surface of the water. Further, all test conditions (pH, oxygen and temperature) remained within the ranges prescribed by the protocol.

The test substance did not induce acute immobilisation of Daphnia magna at nominally 18 mg/l after 48 hours of exposure (NOEC). Note that a maximum response of 10% is acceptable for the control and therefore not considered treatment related.

The 24h-EC50 was 45 mg/I with a 95% confidence interval between 40 and 51 mg/l.

The 48h-EC50 was 34 mg/l with a 95% confidence interval between 29 and 41 mg/l.

Description of key information

The 48h-EC50 was 34 mg/l

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Effect concentration:

34 mg/L