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A Bacterial Reverse Mutation Assay was performed with methyl hexanoate (CAS No. 106-70-7) according to OECD Guideline 471 in Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 (Banduhn, 1992). Test substance concentrations of 0, 8, 40, 200, 1000 and 5000 µg/plate in DMSO were tested in triplicates in two independent experiments using the plate incorporation method with and without the addition of a rat liver homogenate metabolising system (S9-mix).

Cytotoxic effects were observed at concentration of 5000 ¿g test substance per plate. No increase in the frequency of revertant colonies compared to concurrent negative controls were observed in all tested strains, neither in the presence nor in the absence of metabolic activation. Thus, methyl hexanoate did not induce gene mutations in five tested Salmonella strains under the given test conditions.

 

A Bacterial Reverse Mutation Assay was performed with methyl laurate (CAS No. 111-82-0) according to OECD Guideline 471 (Banduhn, 1992). Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 were treated with methyl laurate diluted in acetone using theplate incorporation method. Two independent experiments were performed with identical dose levels of 0, 8, 40, 200, 1000 and 5000 µg/plate. Both experiments were performed in triplicates with and without the addition of a rat liver homogenate metabolising system (S9-mix).

Cytotoxic effects were observed at concentration of 5000 ¿g test substance per plate. No increase in the frequency of revertant colonies compared to concurrent negative controls were observed in all tested strains, neither in the presence nor in the absence of metabolic activation. Thus, methyl laurate did not induce gene mutations in five tested Salmonella strains under the given test conditions.

 

Methyl palmitate (CAS No. 112-39-0) was tested for mutagenicity in S. tyhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 according to OECD Guideline 471 (Banduhn, 1992). Test substance concentrations of 0, 8, 40, 200, 1000 and 5000 µg/plate in acetone were tested in triplicates in two independent experiments using the plate incorporation method with and without the addition of a rat liver homogenate metabolising system (S9-mix).

No cytotoxicity was observed. No increase in the frequency of revertant colonies compared to concurrent negative controls were observed in all tested strains, neither in the presence nor in the absence of metabolic activation. Thus, methyl palmitate did not induce gene mutations in five tested Salmonella strains under the given test conditions. 

 

Another Ames test was performed with methyl stearate (CAS No. 112-61-8) according to OECD Guideline 471 in S. typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 (Banduhn, 1992). Test substance concentrations of 0, 8, 40, 200, 1000 and 5000 µg/plate in acetone were tested in triplicates in two independent experiments using the plate incorporation method with and without the addition of a rat liver homogenate metabolising system (S9-mix).

Cytotoxic effects were observed at concentration of 5000 ¿g test substance per plate. No increase in the frequency of revertant colonies compared to concurrent negative controls were observed in all tested strains, neither in the presence nor in the absence of metabolic activation. Thus, methyl stearate did not induce gene mutations in five tested Salmonella strains under the given test conditions.

 

Methyl oleate (CAS No. 112-62-9) was tested for mutagenicity in S. typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 using a plate incorporation method equivalent to OECD Guideline 471 (Richold, 1982). Test substance concentrations of 0, 50, 150, 500, 1500 and 5000 µg/plate in DMSO were tested in triplicate with and without S9 mix.

Cytotoxicity was not observed. No increase in the frequency of revertant colonies compared to concurrent negative controls were observed in all tested strains, neither in the presence nor in the absence of metabolic activation. Thus, methyl oleate did not induce gene mutations in five tested Salmonella strains under the given test conditions.

 

A Bacterial Reverse Mutation Assay was performed with methyl linoleate (CAS No. 112-63-0) according to OECD Guideline 471 in Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 (Banduhn, 1993). Test substance concentrations of 0, 8, 40, 200, 1000 and 5000 µg/plate in acetone were tested in triplicates in two independent experiments using the plate incorporation method with and without the addition of a rat liver homogenate metabolising system (S9-mix).

Cytotoxic effects were not observed. No increase in the frequency of revertant colonies compared to concurrent negative controls were observed in all tested strains, neither in the presence nor in the absence of metabolic activation. Thus, methyl linoleate did not induce gene mutations in five tested Salmonella strains under the given test conditions.

 

The bacterial mutagenicity of Fatty acids, C6-10, Me esters (CAS No. 68937-83-7) was tested according to OECD Guideline 471 in Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 (Banduhn, 1988). Test substance concentrations of 0, 8, 40, 200, 1000 and 5000 µg/plate were used in the first experiment. In the second experiment 1.25, 5, 20, 80 and 360 µg/plate were used without S9-mix and 3.1, 12.5, 50, 200 and 800µg/plate were used with S9-mix. The test substance was dissolved in Tween 80/water.

Cytotoxicity occurred in the presence and absence of metabolic activation at test substance concentrations above 200 ¿g/plate. No increase in the frequency of revertant colonies compared to concurrent negative controls were observed in all tested strains, neither in the presence nor in the absence of metabolic activation. Thus, Fatty acids, C6-10, Me esters did not induce gene mutations in five tested Salmonella strains under the given test conditions.

 

Fatty acids, C14-18 and C16-18-unsatd., Me esters (CAS No. 67762-26-9) was tested according to OECD Guideline 471 in Salmonella typhimurium strains TA 1535, TA 1537, TA 1538, TA 98 and TA 100 (Banduhn, 1988). Test substance concentrations of 0, 8, 40, 200, 1000 and 5000 µg/plate in Tween 80/bidest. water were used in triplicates in two independent experiments.

Cytotoxic effects were not observed. No increase in the frequency of revertant colonies compared to concurrent negative controls were observed in all tested strains, neither in the presence nor in the absence of metabolic activation. Thus, Fatty acids, C14-18 and C16-18-unsatd., Me esters did not induce gene mutations in five tested Salmonella strains under the given test conditions

 

An in vitro mammalian chromosome aberration test was performed with methyl laurate (CAS No. 111-82-0) in primary human lymphocytes according to OECD Guideline 473 (Buskens, 2010). Duplicate cultures of human lymphocytes were evaluated for chromosome aberrations in the presence and absence of metabolic activation (rat liver S9-mix).

In the first experiment test substance concentrations of 33, 100 and 200 µg/mL in ethanol were used for 3 hours of exposure with and without metabolic activation. In the second experiment 100, 120 and 140 µg/mL were used for 24 hours exposure followed by 24 hours expression time and 30, 120 and 140 µg/mL for 48 hours exposure following 48 hours expression time without S9. 200 µg/mL was chosen as maximum dose due to limited solubility. Mitomycin C and cyclophosphamide were used as positive control substances. Evaluation of 100 well-spread metaphase cells from each culture for structural chromosomal aberrations revealed no increase in the frequency of chromosome aberrations and polyploid cells at any dose level in comparison to the negative controls. The test material demonstrated only modest cytotoxicity. All vehicle (solvent) controls had frequencies of cells with aberrations within the range expected for normal human lymphocytes. All the positive control materials induced statistically significant increases in the frequency of cells with aberrations indicating the satisfactory performance of the test and of the activity of the metabolising system.The test material did not induce a statistically significant increase in the frequency of cells with chromosome aberrations in either the absence or presence of a liver enzyme metabolising system in either of two separate experiments. The test material was therefore considered to be non-clastogenic to human lymphocytes in vitro.

A second, supporting in vitro mammalian chromosome aberration test was performed with methyl laurate (CAS No. 111-82-0) in chinese hamster cells similar to OECD Guideline 473 (National Institute of Health Sciences, 2000). The test material was evaluated for generation of chromosome aberrations in the presence and absence of metabolic activation (rat liver S9-mix). In the first experiment test substance concentrations of 0, 15, 30 and 60 µg/mL in acetone were used for 24 and 48 hours of exposure without metabolic activation. In the second experiment 0, 0.53, 1.1 and 2.1 mg/mL were used for 6 hours exposure followed by 12 hours expression time without metabolic activation and 0, 0.025, 0.05 and 0.1 mg/mL for 6 hours exposure following 12 hours expression time with metabolic activation. Mitomycin C and cyclophosphamide were used as positive control substances. 200 cells per dose were analysed for chromosome aberrations. All vehicle controls had frequencies of cells with aberrations within the expected range. All the positive control materials induced statistically significant increases in the frequency of cells with aberrations indicating the satisfactory performance of the test and of the activity of the metabolising system. The test material did not induce a statistically significant increase in the frequency of cells with chromosome aberrations in either the absence or presence of a liver enzyme metabolising system in either of the two separate experiments. The test material was therefore considered to be non-clastogenic to chinese hamster cells in vitro.

An in vitro mammalian cell gene mutation assay was performed with methyl laurate (CAS No. 111-82-0) according to OECD guideline 476 in L5178Y mouse lymphoma cells (Verspeek, 2010). In the first experiment, methyl laurate was tested up to concentrations of 95 and 200 ¿g/ml in the absence and presence of 8% (v/v) S9-mix, respectively. The incubation time was 3 hours. Methyl laurate was tested up to cytotoxic levels of 85 and 91% in the absence and presence of S9-mix, respectively. In the second experiment, methyl laurate was tested up to concentrations of 70 and 220 ¿g/ml in the absence and presence of 12% (v/v) S9-mix, respectively. The incubation times were 24 hours and 3 hours for incubations in the absence and presence of S9-mix, respectively. Methyl laurate was tested up to a cytotoxic level of 90% in the absence of S9-mix and up to 88% in the presence of S9-mix. The spontaneous mutation frequencies in the solvent-treated control cultures were between the minimum and maximum value of the historical control data range and within the acceptability criteria of this assay. Mutation frequencies in cultures treated with positive control chemicals were increased by 12- and 8.4-fold for MMS in the absence of S9-mix, and by 17- and 15-fold for CP in the presence of S9-mix. In the absence of S9-mix, methyl laurate did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the duration of treatment time. In the presence of S9-mix, methyl laurate did not induce a significant increase in the mutation frequency in the first experiment. This result was confirmed in an independent repeat experiment with modifications in the concentration of the S9 for metabolic activation. It is concluded that methyl laurate is not mutagenic in the mouse lymphoma L5178Y test system under the experimental conditions described.


Short description of key information:
Studies on the mutagenicity in bacteria were available for the following category members (CAS No.): 106-70-7; 111-82-0; 112-39-0; 112-61-8; 112-62-9; 112-63-0; 68937-83-7; 67762-26-9. In all studies no mutagenicity in bacteria was observed.
In vitro cytogenicity study in mammalian cells: CAS No. 111-82-0. No clastogenic effects on mammalian cells could be observed.
In vitro gene mutation study in mammalian cells: CAS No. 111-82-0 No mutagenic effects on mammalian cells could be observed.

Endpoint Conclusion: No adverse effect observed (negative)

Justification for classification or non-classification

All category members were judged to be of sufficient physico-chemical similarity to exert similar characteristics in points of genotoxic potential. Methyl laurate (C12) was therefore chosen for in vitro genotoxicity testing in mammalian cells as the representative of the category. Non of the available experimental data indicated a genotoxic potential.

According to EU classification criteria for genotoxicity, no classification is required.