2-Butenedioic acid, 2-methyl-, 1,4-bis(2-ethylhexyl) ester, (2Z)-

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

2013

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Non-GLP screening test, no analytical verification of test concentrations; very poorly soluble substance.

Cross-reference

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

yes

Remarks:

the concentrations were not analytically verified and statistics not performed

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Deviations:

yes

Remarks:

the concentrations were not analytically verified and statistics not performed

Principles of method if other than guideline:

Acute toxicity of bis(2-ethylhexyl)citraconate was assessed with acute immobilisation test on Daphnia Magna, over an exposure period of 48 hours in a semi-static system. Four test concentrations in a geometric series with a separation factor of 10 an one treated control were tested in the experiment. The nominal concentrations of test item used in the experiment were 0.1; 1; 10 and 100 mg/L. Ten animals, divided into two groups (glass beaker) of five animals each were used at the test concentrations and at the control groups.

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

1,4-bis(2-ethylhexyl) (2Z)-2-methylbut-2-enedioate

EC Number:

807-715-4

Cas Number:

1354569-12-2

Molecular formula:

C21 H38 O4

IUPAC Name:

1,4-bis(2-ethylhexyl) (2Z)-2-methylbut-2-enedioate

Constituent 2

Reference substance name:

di(2-ethylhexyl) (2Z)-2-methyl-2-butenedioate

IUPAC Name:

di(2-ethylhexyl) (2Z)-2-methyl-2-butenedioate

Test material form:

other: liquid

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Batch B018-000-00001

Sampling and analysis

Analytical monitoring:

no

Test solutions

Details on test solutions:

According to the OECD Guidance Document 23 (Aquatic Toxicity Testing of Difficult Substances and Mixtures), supersaturated test item solution (nominal loading rate of 100 mg/L) was prepared by dispersing/dissolving the appropriate amount of test item into the test medium (ISO medium) two days before the start of the experiment. The solution was shaken for about 24 hours at approx. 30 oC and then equilibriated for about 24 hours at approx. 20 oC. The non-dissolved test material was removed by filtration frough a fine (0.22 µm) filter to give the 100 % saturated solution.

During the formulation procedure the nominal test solution 10 mg/L was prepared by the method described above. Nominal test concentrations of 1 mg/L and 0.1 mg/L were prepared by appropriate dilution of nominal test concentrations of 10 mg/L and 1 mg/L respectively.

The test solutions were distributed into the appropriate beckers.

The test was performed under semi-static conditions. The frequency of the reconstituted water renewal period was 24 hours. Prior to treatment of each renewal period the test item solutions were prepared by the method described above.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

Esperimental animals were provided by National Institute of Public Health, 1097 Budapest Gyali u. 2 - 6, Hungary.
They were less than 24 h old at the beginning of the test. No acclimatization as the water used was similar to culture water.
10 animals in test and control groups, dived into 2 replicates (5 animals/replicate).

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test conditions

Hardness:

The reconstituted water (ISO medium) had an approx. Theoretical total hardness of 249 mg/L (as CaCO3).

Test temperature:

20.0 - 20.3 oC measured in the test vessels (daily, old and fresh media where possible). Additionally, climate chamber temperature was between 19.8 and 20.2 oC.

pH:

In the range of 7.68 - 7.96 (measured in each vessel daily, old and fresh media where possible). Not adjusted, not varied by more than 1.5 units in any one test.

Dissolved oxygen:

In the range of 7.48 - 8.54 mg/L (measured in each vessel daily, old and fresh media where possible).

Nominal and measured concentrations:

Only nominal concentrations were applied, as described above.

Details on test conditions:

The test was carried out in 16-h light and 8-hour dark cycle.
For untreated controls, dilution water (ISO medium) was used without addition of the test item (Appendix 3 of the test report as attachment 1)

Reference substance (positive control):

yes

Remarks:

potassium dichromate, batch 0769128

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

> 100 other: % saturated solution

Nominal / measured:

nominal

Conc. based on:

dissolved

Basis for effect:

mobility

Effect concentrations 2

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 100 other: % of saturated solution

Nominal / measured:

nominal

Conc. based on:

dissolved

Basis for effect:

mobility

Effect concentrations 3

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

> 100 other: % saturated solution

Nominal / measured:

nominal

Conc. based on:

dissolved

Basis for effect:

mobility

Effect concentrations 4

Duration:

48 h

Dose descriptor:

LOEC

Effect conc.:

> 100 other: % saturated solution

Nominal / measured:

nominal

Conc. based on:

dissolved

Basis for effect:

mobility

Details on results:

Based on the results of this study, the test item bis (2-ethylhexyl)citraconate had no toxic effect at saturation on Daphnia. The EC50 results and the LOEC are higher than the solubility level of the test item in the test medium.
Details of the test conditions data are available in Appendix 1 of the test report (attachment 2).

Results with reference substance (positive control):

Reference control: 24-h EC50 was 0.82 mg/L (95 % confidence limits 0.76 . 0.87 mg/L).

Reported statistics and error estimates:

No statistical analysis was performed because of the lack of toxic effects. The EC50, NOEC, LOEC and EC100 were determined directly from th rae data.

Any other information on results incl. tables

Table.1 Number and percentage of immobilised animals.

Nominal (mg/L)	Number of	Number (percent) of immobilised animals
preparation	treated animals	24 h	48 h
Control	10	0 (0)	0 (0)
0.1	10	0 (0)	0 (0)
1	10	0 (0)	0 (0)
10	10	0 (0)	0 (0)
100	10	0 (0)	0 (0)

Data of immobility in each test vessel are detailed in Appendix 2 of the test report (attachment 3).

Applicant's summary and conclusion

Validity criteria fulfilled:

no

Remarks:

The nominal concentrations o.1, 1 10 and 100 mg/L were prepared but not verified analytically. The substance is very poorly soluble in water.

Conclusions:

The acute aquatic toxicity (EC50 48-h) to daphinds was > 100% of the saturated solution. (The nominal concentrations were 0.1, 1, 10 and 100 mg/L at 20 oC; the substance is very poorly soluble in water.)

Executive summary:

The acute aquatic toxicity (EC₅₀ 48-h) to daphinds was > 100% of the saturated solution in a non-GLP screening test (OECD 202). The nominal concentrations were 0.1, 1, 10 and 100 mg/L at 20 ^oC. Concentrations were not analytically measured and the substance is very poorly soluble in water. The results are therefore rated as reliable with restrictions (Klimisch 2).