Di-tert-butyl 1,1,4,4-tetramethylbut-2-yn-1,4-ylene diperoxide

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

Study period:

from 2001-06-09 to 2001-09-06

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Version / remarks:

(1984)

Deviations:

yes

Remarks:

see 'Principles of method it other than guideline'

Qualifier:

according to guideline

Guideline:

EU Method C.2 (Acute Toxicity for Daphnia)

Version / remarks:

(1989)

Deviations:

yes

Remarks:

see 'Principles of method it other than guideline'

Principles of method if other than guideline:

- The test was carried out with the water accommodated fraction prepared from 0.5 g/L test substance.
- The actual test temperature of the test room varied less than 2 °C instead of less than 1 °C.

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

At the start of the test samples of about 10 mL were taken from the test vessels with the undiluted WAF and the control, just before the distribution of the test solutions over the four batches and adding the test animals. At the end of the test the solutions from the four batches were combined and mixed and samples of about 10 mL from the undiluted WAF and the control were used for analysis. The samples were analyzed immediately after sampling. The concentration as measured in the WAF at the beginning of the test was used to calculate the concentrations of the dilutions.

Vehicle:

yes

Details on test solutions:

The test solution was a Water Accommodated Fraction (WAF) prepared from 1.50 g test substance in 3 liters of DSW. This solution was slowly stirred for approximately 48 hours, then transferred into two separation funnels and left at test temperature for phase separation. After approximately 2.5 hours, when phase separation was complete, the aqueous phase was withdrawn and used to prepare the following dilutions: 1:32 - 1:16 - 1:8 - 1:4 - 1:2 and the undiluted WAF. A control containing only test medium was included in the test.

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna (water fleas)
- Source: Derived from Aquasense in Amsterdam.
- Age at study initiation: The animals used were less than 24 hours and were obtained from parent animals having an age of 2 - 4 weeks.
- Feeding during test: The animals were not fed during the test.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Post exposure observation period:

none

Hardness:

12 °dH

Test temperature:

19.85 - 21.65 °C

pH:

8.0 - 8.3

Dissolved oxygen:

8.7 - 9.0

Salinity:

no data

Nominal and measured concentrations:

Nominal: 0, 1:2, 1:4, 1:8, 1:16, 1:32 and the undiluted WAF.
Measured: 0, 0.166, 0.332, 0.664, 1.33, 2.66 and 5.31 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel, size and fill volume: 400 mL glass beakers containing approximately 250 mL of test solution
- Type: closed
- No. of organisms per vessel: 5
- No. of vessels per concentration: 4
- No. of vessels per control: 4

TEST MEDIUM: As test medium so called Dutch Standard water was used, with a pH of approximately 8.2 and a hardness of approximately 12°dH. The test medium was aerated before being used in the test. The air was water-saturated and purified by an active coal and cotton filter.

OTHER TEST CONDITIONS
- Photoperiod: light regime of 16 h of ambient light per day, provided by fluorescent tubes.
- Aeration: The test solutions were not aerated.

EFFECT PARAMETERS MEASURED: Immobility and sub-lethal effects were recorded at approximately 24 and 48 hours. The daphnids were considered immobile when they were not able to swim for 15 seconds after gentle agitation of the test vessel. In addition to immobility, sub-lethal effects such as floating at the surface were recorded.

TEST CONCENTRATIONS
- Test concentrations (WAF): 0, 1:2, 1:4, 1:8, 1:16, 1:32 and the undiluted WAF.
- Results used to determine the conditions for the definitive study:
The results presented are the results of a second test. The first test was rejected because no test substance could be detected in the samples taken at the beginning and end of the test. The test results of this first test indicated immobility after 48 hours even at the highest dilution (1:16) and therefore an additional dilution of 1:32 was introduced for the second test. To be able to detect the test substance during the second test the composition and flow rate of the mobile phase was slightly adjusted.

Reference substance (positive control):

no

Key result

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

> 5.31 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

mobility

Details on results:

Sublethal effects, such as floating at the surface were however observed in all dilutions. The number of animals floating on the surface seems to be related to the concentration of the test substance in the test. In spite of these sublethal effects no immobility was observed at the undiluted WAF (5.31 mg/L) or in any of the dilutions except for one immobile daphnid, which was detected in the 1:8 dilution (0.664 mg/L) after 24 hours. This daphnid is probably accidentally immobilized because it was sticking to the wall of the test vessel somewhat above the test solution. Therefore, this immobility is not considered as an effect of the test substance.
Due to the absence of any lethal effects it was not possible to calculate the EC50 and EC100 values.

Chemical analyses:

A small peak was detected in the control, which was also detected in pure Dutch Standard water. The concentration in the WAF was therefore calculated taking account of this background peak in the test medium.

The results show that the undiluted WAF contains 5.31 mg/L of the test item at the beginning of the test and 0.38 mg/L at the end of the test, which is less than 80% of the initial concentration. The concentration in the WAF is calculated using the summed area of the two peaks related to the test substance.

These results show that the test substance is probably not stable under test conditions. The concentration of the WAF at the beginning of the test was used for the calculation of the toxicity.

Validity criteria fulfilled:

yes

Conclusions:

The test substance had no lethal effect on Daphnia magna under the chosen test conditions.

Executive summary:

The acute toxicity of the Water Accommodated Fraction (WAF) of the test item in solvent to Daphnia magna (water flea) was tested under static conditions for 48 hours in accordance with OECD guideline 202 and EU method C.2.

The test item in solvent is immiscible with water and therefore a WAF was prepared. During chemical analyses, which were performed after finishing the test, the test substance could not be detected in WAF samples from the beginning and the end of the test. Therefore the test was repeated using an adjusted analytical method. The following dilutions of the WAF were tested during the second test: 1:32 -1:16 - 1:8 - 1:4 - 1:2 and the undiluted WAF. Chemical analyses of the control and WAF showed that the test substance is probably unstable under test conditions. At the beginning of the test a concentration of 5.31 mg/L was measured. At the end of the test a concentration of 0.38 mg/L was measured. The concentration at the beginning of the test was used for the calculation of the effect concentrations.

Sublethal effects, such as floating at the surface were observed in all dilutions. The number of animals floating on the surface seems to be related to the concentration of the test substance. In spite of these sublethal effects no test substance related immobility was observed in the undiluted WAF (5.31 mg/L; average initial concentration) or in any of the dilutions.

Description of key information

The test substance had no lethal effect on Daphnia magna under the chosen test conditions.

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

EC50

Effect concentration:

> 5.31 mg/L

Additional information

The acute toxicity of the Water Accommodated Fraction (WAF) of the test item in solvent to Daphnia magna (water flea) was tested under static conditions for 48 hours in accordance with OECD guideline 202 and EU method C.2.

The test item in solvent is immiscible with water and therefore a WAF was prepared. During chemical analyses, which were performed after finishing the test, the test substance could not be detected in WAF samples from the beginning and the end of the test. Therefore the test was repeated using an adjusted analytical method. The following dilutions of the WAF were tested during the second test: 1:32 -1:16 - 1:8 - 1:4 - 1:2 and the undiluted WAF. Chemical analyses of the control and WAF showed that the test substance is probably unstable under test conditions. At the beginning of the test a concentration of 5.31 mg/L was measured. At the end of the test a concentration of 0.38 mg/L was measured. The concentration at the beginning of the test was used for the calculation of the effect concentrations. Sublethal effects, such as floating at the surface were observed in all dilutions. The number of animals floating on the surface seems to be related to the concentration of the test substance. In spite of these sublethal effects no test substance related immobility was observed in the undiluted WAF (5.31 mg/L; average initial concentration) or in any of the dilutions.