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Fish, acute

The acute toxicity of the test item DOBA to zebra fish (Brachydanio rerio) was determined in a 96‑hour semi-static test with daily test medium renewal according to the EU Commission Directive 92/69/EEC, Part C.1, the Commission Regulation (EC) No 440/2008, Part C.1 and the OECD Guideline for Testing of Chemicals No. 203 (1992), as well as according to the OPPTS Guideline No. 850.1075 (Public Draft, April 1996).

A limit test was performed in accordance with the test guidelines to demonstrate that the test item has no toxic effect on the test organisms up to the solubility limit of the test item in test water. Due to the low water solubility of the test item, a dispersion of the test item with the loading rate of 100 mg/L was continuously stirred at room temperature in the dark over 3 hours. Then, the dispersion was filtered. The undiluted filtrate of the dispersion was used as the test medium. Additionally, a control was tested in parallel.The preparation of the test medium was based on the OECD Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, 2000 and EPA Ecological Effects Test Guidelines OPPTS 850.1000 Special Considerations for Conducting Aquatic Laboratory Studies, 1996.

In the control and at the loading rate of 100 mg/L (mean measured concentration of 1.0 mg/L), no mortality or other visible abnormalities were determined during the test period of 96 hours.

In a second study the acute toxicity of the test item DOBA to fish (zebrafish) was determined according to OECD-Guideline for Testing of Chemicals No. 203. A static limit test with a saturated solution was conducted. The preparation method has been chosen with regard to the test item properties and in accordance with the relevant OECD guidance document (2000). Duration of the test was 96 h. 7 test organisms were exposed to the saturated solution and the control. Water quality parameters pH-value, temperature and oxygen-saturation measured after 0, 24, 48, 72 and 96 h were determined to be within the acceptable limits.

In this study DOBA did not cause any effects to zebrafish after 96 h when tested with the saturated solution ( 100 mg/L). The NOEC (0 – 96 h) is laid down as the saturated solution.

In conclusion, the test item DOBA had no acute toxic effects on zebra fish up to the loading rate of 100 mg/L.

Daphnia, acute

The acute toxicity of the test item DOBA toDaphnia magnawas determined in a 48 hour semi-static test according to the EU Commission Directive 92/69/EEC, Part C.2, the Commission Regulation (EC) No. 440/2008, Part C.2 and the OECD Guideline for Testing of Chemicals, No. 202 (2004), as well as according to the OPPTS Guideline No. 850.1010 (Public Draft, April 1996).  

A limit test was performed in accordance with the test guidelines to demonstrate that the test item has no toxic effect on the test organisms up to the solubility limit of the test item in test water. Due to the low solubility of the test item, a dispersion of the test item with the loading rate of 100 mg/L was continuously stirred at room temperature in the dark over 3 hours. Then, the dispersion was filtered. The undiluted filtrate of the dispersion was used as the test medium. Additionally, a control was tested in parallel. The preparation of the test medium was based on the OECD Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, 2000 and EPA Ecological Effects Test Guidelines OPPTS 850.1000 Special Considerations for Conducting Aquatic Laboratory Studies, 1996.

At the start of the test medium renewal periods, the analytically determined concentrations of DOBA in the undiluted filtrate were 2.23 and 2.13 mg/L (on Day 0 and 1, respectively). During the test medium renewal periods of 24 hours a decrease of the test item concentration in the test medium was determined. At the end of the test medium renewal periods, 1.90 and 1.99 mg/L were measured, respectively.

 

In the control and at the loading rate of 100 mg/L (mean measured concentration of 2.1 mg/L), no immobilized test organisms were observed during the test period of 48 hours.

 

In conclusion, the test item DOBA had no acute toxic effects on Daphnia magna up to the loading rate of 100 mg/L (mean measured concentration of 2.1 mg/L), its solubility limit in test water under the present test conditions.

Alga

The influence of the test item DOBA on the growth of the freshwater green algal speciesPseudokirchneriella subcapitatawas investigated in a 96 ‑hour static test according to the OECD Guideline 201 (2006), and the Commission Regulation (EC) No 761/2009, C.3, as well as according to the OPPTS Guideline No. 850.5400 (Public Draft, April 1996).

Due to the low solubility of the test item in test water, a dispersion of the test item with the loading rate of 100 mg/L was continuously stirred at room temperature in the dark over 3 hours. Then, the dispersion was filtered. The undiluted filtrate with the loading rate of 100 mg/L and dilutions 1:2, 1:4, 1:8 and 1:16 of the undiluted filtrate with the loading rate of 100 mg/L were used as test media. Additionally, a control was tested in parallel. The preparation of the test medium was based on the OECD Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, 2000 and EPA Ecological Effects Test Guidelines OPPTS 850.5400 Special Considerations for Conducting Aquatic Laboratory Studies, 1996.

At the start of the test, the analytically determined concentrations of DOBA in the test media (dilutions 1:16, 1:8, 1:4, 1:2 and the undiluted filtrate) were 0.571, 1.16, 2.23, 4.51 and 8.81 mg/L, respectively. During the test period a decrease of test item concentration in the test media occurred. After 24 hours test duration the measured concentrations were below LOQ (limit of quantification, LOQ=0.0174 mg test item /L) for the two lowest test concentrations and 0.485, 2.78 and 7.91 mg/L for the three highest test concentrations, respectively. On Day 2, 3 and at the end of the test, all measured concentrations were below LOQ.The biological results were related to the mean measured test item concentrations calculated as the time-weighted means over all measurements per test concentration.

 

Conclusion

The test item had no biological relevant inhibitory effect on the growth rateμ of the algae after the test period of 72 hours up to the solubility limit in the test medium (undiluted filtrate: loading rate 100 mg/L, mean measured concentration of 2.4 mg/L). The maximum inhibition of growth rate was 4.4 %.

Microorganisms

The inhibitory effect of the test item DOBA on the respiration rate of aerobic wastewater microorganisms of activated sludge was investigated in a 3-hour respiration inhibition test according to the EU Commission Directive 88/302/EEC, Part C.11, Commission Regulation (EC) No. 440/2008, Part C.11 and the OECD Guideline for Testing of Chemicals, No. 209 (1984).

Based on the EU testing guidelines, a limit test with a nominal test concentration of 1000 mg/L was performed

The test item had no inhibitory effect on the respiration rate of activated sludge after the incubation period of three hours at a loading rate of 1000 mg/L. The saturation concentration (water solubility limit of DOBA under the test conditions) was reached and no inhibitory effect was determined after three hours contact time. Thus, the 3-hour NOEC (EC10) of the test item to activated sludge microorganisms was at least 1000 mg/L (i.e. the saturation concentration of the test item). This value might even be higher but concentrations above 1000 mg/L were not tested. The 3-hour EC20, EC50, and EC80 could not be calculated but were clearly higher than 1000 mg/L.