Decyloxirane

Toxicity to aquatic algae and cyanobacteria

Description of key information

The key values for algae were derived from a new guideline study(OECD 201)according to GLP with Pseudokirchneriella subspicatua

being EC50= 0.52 mg/L and the EC10 = 0.25 mg/L, respectively.

Key value for chemical safety assessment

EC50 for freshwater algae:

0.52 mg/L

EC10 or NOEC for freshwater algae:

0.25 mg/L

Additional information

Two guideline studies (OECD 201) according to GLP with Pseudokirchneriella subspicata are available for Decyloxirane. The EC50 of the first study was determined to be 0.0557 mg/l and the NOEC was determined to be 0.00416 mg/l based on mean measure concentrations. The EC50 of the second study was 0.52 mg/l and the EC10 0.25 mg/l related to growth rate based on initial measured concentrations.

Due to analytical inconsistencies of the first study and subsequent performed additional studies (hydrolysis: OECD 111 / second study OECD 201) the first study has to be re-evaluated in the light of the additional findings (justification see following passage). Therefore, the key values for algae were derived from the second study being EC50: 0.52 mg/l and EC10 0.25 mg/l, respectively.

Re-evaluation of algae growth inhibition studies performed with C12 -epoxides

 

1                   Algae studies performed at CERI

In 2015 CERI laboratories performed a test for growth inhibition of algae with C12 -epoxides.

The epoxide has very low water solubility, approx. 1.4 mg/l . Therefore, a stock solution was prepared with 100 mg/l of the epoxide and stirred for 24 hours. After stirring, the solution were filtrated and diluted with fresh medium to prepare the test solutions for the algae test. The concentration of epoxide was below the limit of determination (LOD approx. 0.003 mg/l) already 24 hours after start of incubation of the algae in most test solutions.

Based on the fact that at the end of incubation the epoxide was not detected and on the assumption that the epoxide would hydrolyse during incubation the results were based on geometric means of the test concentrations. For C12-epoxide an EC50value of 0.056 mg/l and an NOEC of 0.0042 mg/l were derived. In Table 1 the levels of growth rate inhibition by C12-epoxide are provided in relation to the concentrations measured at start of incubation as well as in relation to geometric means of measured concentrations at beginning and end.

Table 1: Inhibition of growth rate at different initial concentrations of C12-epoxide

 

Concentration [mg/l]		Inhibition of growth rate
Measured at beginning of incubation	Geometric means of measured concentrations	[%]
1.41	0.128	92
0.58	0.031	21
0.25	0.014	5.7
0.13	0.010	2.6
0.05	0.006	1.9
0.02	0.004	0.5

 

Providing that the epoxide would hydrolyse in algae medium it was assumed that during stirring of the concentrated stock solution (100 mg/l) for 24 hours an unknown concentration of unknown hydrolysis product(s) was formed and dissolved in the stock solution.

Thereupon some additional tests were performed at CERI beginning 2016 to identify the hydrolysis product(s) of C12-epoxide, however without success. Finally, a new, short test was performed by preparing a stock solution by stirring 100 mg/l C12-epoxide for 24 hours. After filtration algae were added and incubated. At the beginning of incubation the concentration of C12-epoxide amounted to

1.33 mg/l and that of dodecane-1,2-diol, the most likely hydrolysis product, amounted to 0.75 mg/l. At the end of incubation the concentrations amounted to < 0.03 mg/l and 0.79 mg/l, respectively. Analysis (GC/MS) of the extract of the algae medium at the end of incubation did not show any new peaks compared to the beginning of incubation. It was concluded “that it is not possible to identify the hydrolytic product at the present”.

Therefore, it was decided to perform a hydrolysis test with C12-epoxide as well as a new algae study with a different procedure of test substance preparation.

 

2                   Hydrolysis test performed at BASF

Hydrolysis testing was performed at BASF according to OECD 111. The half-lives at 20°C and pH 4, pH 7, and pH 9 were determined to be 118 h, 183 h, and 188 h, respectively. Dodecane-1,2-diol was identified as hydrolysis product.

 

3                   Algae study performed at IES

In the new algae study performed at IES test concentrations of nominally 1.3 mg/l, 0.65 mg/l and 0.33 mg/l were prepared by direct addition of C12-epoxide to the test medium, stirring for 24 h and filtration before addition of algae. Nominal concentrations of 0.16 mg/l and 0.08 mg/l were prepared by dilution of the filtrated 0.33 mg/l test concentration. The highest test concentration of nominal 1.3 mg/l was incubated in parallel without addition of algae. At the end of incubation the concentration of C12-epoxide in the presence of algae was below the limit of determination, i.e. < 0.03 mg/l. In contrast, in the samples incubated without algae the measured concentration of C12- epoxide remained constant over the incubation time. From this observation it can be concluded that C12-epoxide was stable in algae medium during the incubation period. This means that the algae were continuously exposed to C12-epoxide and not to unknown hydrolysis product(s). As at the end of incubation C12-epoxide was not detectable in the algae cultures it is concluded that C12-epoxide adsorbed to algae. For derivation of ECxvalues the concentrations measured at the beginning of incubation were taken into account. Results are summarised in Table 2.

Table 2: ECx, NOEC and LOEC values derived from the algae study performed at IES

 

EC values [mg/l] based on growth rate
EC10	0.25
EC20	0.32
EC50	0.52
NOEC	0.26
LOEC	0.39

 

In Table 3 the levels of growth rate inhibition by C12-epoxide are provided in relation to the concentrations measured at start of incubation.

Table 3: Inhibition of growth rate in relation to initial concentrations of C12-epoxide

 

Concentration at beginning of incubation [mg/l]		Inhibition of growth rate [%]
nominal	measured
1.3	0.70	68.8
0.62	0.59	60.9
0.33	0.39	32.8
0.16	0.26	8.3
0.08	0.14	3.0

 

4                   Assessment of results

Two tests for growth inhibition of algae were performed with C12-epoxide, one at CERI and one at IES laboratories. Both tests were performed with Pseudokirchneriella subcapitata according to OECD 201.

Based on the fact that C12-epoxide “disappeared” during incubation of algae and the assumption of hydrolysis of C12-epoxide in algae medium derivation of EC50 and NOEC values in the CERI study were based on geometric means of measured test substance concentrations at beginning and end of the incubation period. The respective values were EC50= 0.0557 mg/l and NOEC = 0.00416 mg/l.

Derivation of EC50 and NOEC values in the IES study were based on measured test substance concentrations at the beginning of incubation because it was demonstrated that C12-epoxide was stable during incubation in algae medium. The derived values were EC50= 0.52 mg/l and NOEC = 0.26 mg/l.

The differing bases used for derivation of EC50 and NOEC values lead to significant differences between the results obtained in one or the other study. In the light of the actual knowledge of stability of C12-epoxide in algae medium during the incubation period the measured concentrations at the beginning of incubation have to be used as basis for dose- effect correlations. The levels of growth rate inhibition at different test substance concentrations observed in the two independent algae studies can be directly compared, see Table 4.

Table 4: Comparison of levels of growth rate inhibition determined in the CERI and IES studies

 

CERI study		IES study
Concentration measured at beginning of incubation [mg/l]	Inhibition of growth rate [%]	Concentration measured at beginning of incubation [mg/l]	Inhibition of growth rate [%]
1.41	92	0.70	68.8
0.58	21	0.59	60.9
--	--	0.39	32.8
0.25	5.7	0.26	8.3
0.13	2.6	0.14	3.0
0.05	1.9	--	--
0.02	0.5	--	--

 

From the data shown in Table 4 it becomes obvious that the maximum concentration of C12-epoxide in saturated algae medium differed by a factor of 2 between the two studies. No final explanation for this difference is currently available. However, the levels of inhibition of algae growth are very well comparable between the two studies; at 0.25/0.26 mg/l and 0.13/0.14 mg/l the levels of inhibition are almost identical, in both studies being below 10%.

From this comparison of the effect levels of C12-epoxide on the growth rate of algae it can be concluded that the EC50 and NOEC values in both studies are very close when related to the concentrations measured at the beginning of incubation.

 

Based on the actual knowledge of stability of C12-epoxide in algae medium during the incubation period derivation of EC50 and NOEC values should be based on initially measured concentrations rather than on geometric mean measured concentrations since algae were exposed throughout the whole incubation period to C12-epoxide which very probably adsorbed to algae.

From the fact that the levels of growth rate inhibition are well comparable between the two studies it can be concluded that during preparation of the stock solution of C12-epoxide in algae medium by stirring 100 mg/l for 24 hours no toxic hydrolysis product(s) were formed from C12-epoxide in significant amounts. This is supported by analytical screening of test solutions after incubation of algae: no hydrolysis products were detected. In the hydrolysis test the half-lives of C12-epoxide at 20°C and pH 7 or pH 9 amounted to approx. 185 hours whereas the incubation phase takes 72 hours.

 

 

5                     Conclusion

Based on the information summarised above it is concluded that the algae study performed at CERIwith C12-epoxide should be re-evaluated and derivation of EC50 and NOEC values should be based on the initially measured concentrations.