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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
key study
Study period:
14th August 2018 - 13th September 2018
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Remarks:
OECD 471 Bacterial Reverse Mutation Test

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2018

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Remarks:
No deviations from regulatory requirements.
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
liquid

Method

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Additional strain / cell type characteristics:
other: histidine deficient
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
0.16, 0.31 0.63, 1.25, 2.5 and 5 µL/plate of 11-methyldodecyl laurate in absence and presence of the metabolic activation system (10% v/v S9 mix).
The test item was found insoluble in distilled water and in dimethyl sulfoxide, while it was found soluble in 95% ethanol. Hence, 95% ethanol was selected as the vehicle for treatment. A volume of 100 µL was added to 2 mL of top agar, to assess the precipitation. Precipitation was not observed at the tested concentration of 5 µL/plate. Hence, 5 µL/plate was selected as the highest concentration to be tested for initial toxicity-mutation test both in the absence and presence of metabolic activation.
Vehicle / solvent:
95% ethanol
Controls
Untreated negative controls:
yes
Remarks:
Untreated control
Negative solvent / vehicle controls:
yes
Remarks:
95% ethanol
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
2-nitrofluorene
sodium azide
mitomycin C
other: 2-Aminoanthracene
Evaluation criteria:
A result was considered positive if concentration-related increase over the range tested and/or a reproducible increase at one or more concentrations in the number of revertant colonies per plate in at least one strain with or without metabolic activation system. Biological relevance of the result was considered first.
Statistics:
Simple linear regression analysis for TA1537, TA1535, TA98, TA100 and TA102, separately, to assess the dose dependent nature of any increase in revertant colonies. A response that did not met the criteria of magnitude, concentration-responsiveness and reproducibility, was not evaluated as positive.

Results and discussion

Test results
Key result
Species / strain:
other: Salmonella typhimurium TA1537, TA1535, TA98, TA100 and TA102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
not examined
Untreated negative controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
11-methyldodecyl laurate was non-mutagenic to any of the five strains of Salmonella typhimurium (TA1537, TA1535, TA98, TA100 and TA102) when tested under the specified experimental conditions. Validity criteria were fully met.
Executive summary:

The study was performed to evaluate mutagenic activity of 11-methyldodecyl laurate by the bacterial reverse mutation test, using five histidine deficient mutant tester strains of Salmonella typhimurium (i.e., TA1537, TA1535, TA98, TA100 and TA102).

11-Methyldodecyl laurate was tested in two independent experiments, in the absence and presence of metabolic activation. Bacterial cultures were exposed to test item at 8 concentrations (two plates/concentration) between 0.0015 and 5 µL/plate in the initial toxicity-mutation test. Normal growth was observed up to the tested concentration of 5 µL/plate both in absence and presence of the metabolic activation (5% v/v S9 mix) in the tester strains TA1537, TA1535, TA98, TA100, and TA102. No increase in the number of revertant colonies (no mutagenic effect) was observed both in absence and presence of the metabolic activation system (5% v/v S9 mix) in all the tester strains. To confirm the negative results obtained in the initial toxicity mutation test, confirmatory mutation test was conducted with an increased S9 concentration i.e., 10% v/v S9 mix and modified concentration spacing. Bacterial cultures were exposed to 11-methyldodecyl laurate at 6 concentrations (three plates/concentration) between 0.16 and 5 µL/plate for the tester strains TA1537, TA1535, TA98, TA100, and TA102 both in absence and presence (10% v/v S9 mix) of the metabolic activation. After 48 hours of incubation at 37 ± 1°C, the revertant colonies were scored.

11-Methyldodecyl laurate did not induce any significant increase in the number of revertant colonies, in both trials, with and without S9 mix, in any of the five tester strains. All the values for the negative control (95% ethanol) in all strains were comparable with untreated control of respective strains and also with the laboratory historical data of distilled water and other solvents, which shows that the solvent does not have any impact on the growth of the test system, and positive controls showed an increase in the number of revertant colonies demonstrating the efficiency of the test system.

All criteria for a valid study were met as described in the study plan. From the results of this study, under the specified experimental conditions, test item is concluded to be non-mutagenic in the Bacterial Reverse Mutation Test using Salmonella typhimurium.