[1,3(or 1,4)-phenylenebis(1-methylethylidene)]bis[tert-butyl] peroxide

Long-term toxicity to aquatic invertebrates

Description of key information

Several OECD 211 studies (detailed in section 'Additional information') were performed in order to assess the toxicity of the substance 1,3(or 1,4)-Phenylenebis(1-methylethylidene)]bis[tert-butyl]peroxide on the reproductive capacity output of Daphnia magna.
Following several technical issues raised during the first studies, it was decided under the frame of the Corap process to perform a new OECD 211 study under semi-static conditions with a daily renewal and in a completely closed test vessels, no headspace, and light conditions reduced to the lower end of the optimal range. These tests conditions were chosen to minimise losses by volatilization and adsorption of the substance observed previously.

Under the optimized test conditions of this last study, [1,3(and/or 1,4)-Phenylenebis(1-methylethylidene)]bis[tertbutyl] peroxide did not affect reproduction, growth and survival of Daphnia magna up to and including the highest tested concentration of nominally 40 μg/L corresponding to an average measured concentration of 11.4 μg/L after 21 days of exposure.
It is important to note that due to the extended storage time of the analyzed reserved samples (e.g.) freshly prepared day 14 low result), the average measured concentration of 11.4 μg/L after 21 days of exposure would be a worst-case scenario and hence an underestimation of the real exposure of the test material during the test.
This new OECD 211 study met the acceptability criteria prescribed by the study plan and was considered valid.

Additional information

The influence of [1,3(or 1,4)-phenylenebis(1 -methylethylidene)]bis[tertbutyl] peroxide to Daphnia magna in a semi-static reproduction test (daily renewal) has been investigated through OECD Guideline 211 and according to GLP requirements (Hugill, 2015). Nevertheless, some deficiencies have been observed during the study making interpretation of the results difficult. The major issue was related to the concentration stability of the test solution during the test since wide fluctuations of fresh medium concentrations were observed between each renewal. Therefore, preliminary trials to prepare test solutions in better conditions have been done leading to the performance of a new test.

The toxicity of [1,3(or 1,4)-phenylenebis(1 -methylethylidene)]bis[tertbutyl] peroxide dissolved in fresh water on the reproductive efficacy to Daphnia magna STRAUS - clone 5, in a semi-static test (refreshment taking place 6 times a week) has been investigated through OECD Guideline 211 and according to GLP requirements (Kean, 2016).

The test organisms were exposed to the test substance at its maximum achievable solubility in test medium (generated from a 10 mg/L water soluble fraction stirred for 48 hours) during 21 days. All the validity criteria were fulfilled.

Analytical determinations were made on fresh and old solutions throughout the test. The concentration of the test substance in fresh test solutions were all found to be consistently close to the water solubility limit of the test material. The measured concentration of test material was however not stable during the period between solution refreshments. In the old solutions the test material could be detected after a simple extraction but was heavily influenced by the amount of algae present due to the substance adsorbing to the algae cells. Hence a decreasing concentration was measured in the old solutions as progressively more algae was grazed away by the test organisms as they grew during the test. For this reason a mean measured initial concentration was calculated. This was considered a more reliable indication of the exposure to the test substance.

The test substance at its maximum achievable solubility in test medium (generated from a 10 mg/L water soluble fraction) did not have any detectable effects on the reproduction or length of Daphnia magna in a 21 day chronic study under semi static conditions with refreshment 6 times per week. TheNOELR (No observed effect loading rate) can therefore be expressed as 10 mg/L. The NOEC of the test material may also be expressed as 40.6 µg/L based on the mean of the measured initial concentrations.

A loading rate of 10 times the classification cutoff for chronic toxicity was chosen in an effort to make the results as worst case as possible and to ensure that an excess of parent material was always present therefore allowing stabilization at the water solubility limit. In addition analytical data was generated indicating the strong tendency of the test material to bind to algae cells used to feed the test organisms. Although this reduces the dissolved test concentration and exposure via the water phase, the consumption of the algae would still result in an environmentally relevant exposure to the test organism. Such data is also useful when concluding the behavior of other similar organic peroxides in the environment. In addition hydrolysis data indicated better stability than was measured during this test. Indicating that the algae that was present is also likely to have influenced the test substance concentration significantly as well as the grazing behavior of the test organisms. The presence of peroxidase enzymes is widely documented in algae, bacteria and fungal species.

A new OECD 211 study was performed in the frame of the Corap assessment. The study was performed using semi-static conditions with daily renewals and in a completely closed test vessels, no headspace, and light conditions reduced to the lower end of the optimal range. These tests conditions were chosen to minimise losses by volatilization and adsorption of the substance observed previously. Actual concentrations deviated from nominal and, therefore, the effect parameters were expressed in terms of average measured concentrations. These corresponded to 2.8, 5.4, 6.4, 9.6, and 11.4 μg/L at nominally 10, 15, 20, 29 and 40 μg/L, respectively. Under the conditions of the present study [1,3(and/or 1,4)-Phenylenebis(1-methylethylidene)]bis[tertbutyl] peroxide did not affect reproduction, growth and survival of Daphnia magna up to and including the highest achievable concentration of nominally 40 μg/L corresponding to an average measured concentration of 11.4 μg/L after 21 days of exposure.
It is important to note that due to the extended storage time of the analyzed reserved samples (e.g.) freshly prepared day 14 low result), the average measured concentration of 11.4 μg/L after 21 days of exposure would be a worst-case scenario and hence an underestimation of the real exposure of the test material during the test.

This new OECD 211 study met the acceptability criteria prescribed by the study plan and was considered valid.