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EC number: 246-807-3
CAS number: 25307-17-9
The effects of the test item 2,2’-(Octadec-9-enylimino)bisethanol (Batch-No. 1950627) on the early-life stage of fish (Danio rerio / Zebrafish) were determined according to OECD Guideline 210 under GLP conditions.
The test item is a yellow liquid UVCB substance which is dispersible/insoluble in water. Methanol was used as solvent with a concentration of 0.025 mL/L dilution water. Stock solutions in methanol with nominal concentrations of 100, 200, 400, 800 and 1600 mg/L were prepared in appropriate intervals of 7 days and continuously dosed to the dilution water in a flow-through system. Based on the results of a range finding test the test was conducted as a dose-response test with the nominal test item concentrations 2.50, 5.00, 10.0, 20.0 and 40.0 µg/L, corresponding to the overall arithmetic mean measured test item concentrations of 1.29 – 2.19 – 6.00 – 12.9 - 27.0 µg/L. The term overall arithmetic mean means that all measured samples (stabilized samples, from study days 0, 7, 14, 21 and 34) were taken into account for calculation of the arithmetic mean measured concentrations.
The test was started by placing fertilized eggs into the test vessels and it lasted 35 days (30 days post-hatch). 80 eggs of Danio rerio / zebrafish were exposed to each test concentration, the solvent control and the control (4 replicates with 20 eggs each).
The water quality parameters pH-value, oxygen concentration, temperature and total hardness were within the acceptable limits.
On study day 5, 95% of the control and 96% the solvent control larvae had hatched. Therefore, study day 5 was defined as post hatch day 0 (= PHD 0).
Different toxicological endpoints were determined: hatching success, fry growth (assessed via length and fresh weight measurements on PHD 30), morphological and behavioral effects, post-hatch survival and overall survival.
Specific analysis of various concentrations of 2,2’-(Octadec-9-enylimino)bisethanol in the test media and the controls was carried out via LC-MS/MS. The concentrations of the test item 2,2’-(Octadec-9-enylimino)bisethanol, the solvent control and the control were determined during exposure on days 0, 7, 14, 21 and 34. The constituent C18:1 Amine + 2EO was determined via LC MS/MS. The samples taken from the test vessels were completely clear and did not contain any particles or turbidity. These samples were subjected to the analytical procedure and the results are used for the calculation of the dose response.
The stock solutions prepared in methanol (see section 4.2 for details) were sampled and analyzed from the freshly prepared and corresponding 7 days aged stock solutions. Measured concentrations of the freshly prepared stock solution were 106 to 112% of the nominal values. Measured concentrations of the 7 days aged stock solution were in the range of 111 to 115% of the nominal values.
All effect values are given based on the nominal and the overall arithmetic mean measured concentrations of the test item 2,2’-(Octadec-9-enylimino)bisethanol.
Findings and Observations
The results of the parameters hatching success, fry growth (expressed as weight and length measurement at PHD 30), post-hatch survival and overall survival were checked for statistically significant differences. No statistically significant difference was detected between the dilution water control and the solvent control for all parameters (hatching success, fry growth expressed as length or weight on PHD 30, post-hatch survival and overall survival). Therefore, both controls were pooled for statistical analysis. The effect values NOEC, LOEC, ECx, LCx values were determined based on the statistical results.
NOEC, LOEC, ECx Values of Hatching Success and Fry GrowthBased on nominal and overall arithmetric mean measured test item concentrations [µg/L] with 95% Confidence intervals in brackets
Based on nominal test item concentrations [µg/L]
Based on overall arithmetric mean measured test item concentrations [µg/L]
Hatching success after 7 days1)
Fry Growth expressed as:
Hatching success after 6 days1)
3.54 (< 2.50 to > 20.0)
1.61 (< 1.29 to > 12.9)
1) = end of hatching periodn.d. = not determinable
NOEC, LOEC, LCx values of Post Hatch Survival and Overall SurvivalBased on nominal and overall arithmetic mean measured test item concentrations [µg/L] with 95% Confidence intervals in brackets
12.7(9.56 – 17.2)
15.6(9.63 – 20.0)
7.83(5.71 – 11.0)
10.0(5.76 – 20.2)
16.8(12.6 – 18.7)
18.3(10.9 – 30.8)
10.7(7.76 – 12.0)
11.7(6.60 – 21.0)
n.d. = not determinable
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