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Genetic toxicity: in vitro

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in vitro gene mutation study in bacteria
Type of information:
experimental study
Adequacy of study:
supporting study
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference Type:
A survey of chemicals for mutagenic action on E. coli
Demerec, M., Bertani, G. and Flint., J.
Bibliographic source:
The American Naturalist 85: 119-136

Materials and methods

Test guideline
equivalent or similar to guideline
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
GLP compliance:
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
Lactic acid
EC Number:
EC Name:
Lactic acid
Cas Number:
Molecular formula:
2-hydroxypropanoic acid


Target gene:
Species / strain
Species / strain / cell type:
E. coli, other: Strains B/Sd-4/1,3,4,5 and B/Sd-4/3,4
Details on mammalian cell type (if applicable):
- Type and source of cells: strain/cell type: Streptomycine-dependent strains B/Sd-4/1,3,4,5 and B/Sd-4/3,4:
Metabolic activation:
Test concentrations with justification for top dose:
0.01 and 0.02 %
Details on test system and experimental conditions:
Strains B/Sd-4/1,3,4,5 and B/Sd-4/3,4 of E. coli were used throughout this work. For every experiment, bacteria were grown for 24 hours at 37 °C in an aerated broth culture containing 10 micrograms of streptomycin per milliliter. They reached a saturation tier of approximately 2 to 3.0E+09 cells per milliliter. Each culture was started from an inoculum, usually large, taken from streptomycin-agar slants kept in a refrigerator. Before treatment the bacteria were washed in saline and resuspended in distilled water. A sample of the new suspension was added to the desired solution of chemical in distilled water and incubated at 37 °C for a certain period of time; no growth occurs under these conditions. Another sample of the same suspension was added to an equal amount of distilled water and incubated for the same period of time, as a control. At the end of the treatment period, both treated and control suspensions were assayed by plating suitable dilutions on streptomycin-agar plates. At the same time, they were plated (0.1 ml per Petri dish), either undiluted or diluted not more than 1:10 in plain broth, onto a number of streptomycin-free plates, using a glass spreader and a turntable. The assay plates were incubated for 48 hours, after which it was possible to count the colonies and calculate the tiers of the two suspensions at the end of treatment, and the percentage of survivors. The streptomycin-free plates (mutant plates) were incubated for at least six days. After this time the colonies were scored, and the frequency of mutants calculated by dividing the number of colonies by the number of (viable) bacteria plated.

Results and discussion

Test results
Species / strain:
E. coli, other:
Metabolic activation:
Cytotoxicity / choice of top concentrations:
other: 0.02 %
Additional information on results:
Some of the experiments with lactic acid indicated a weak mutagenic effect; but this is rendered doubtful by the fact that experiments with sodium lactate consistently gave negative results.

Applicant's summary and conclusion

Under the experimental conditions reported, lactic acid did not cause gene mutations. Therefore, the test item is considered to be non-mutagenic in this bacterial reverse gene mutation assay.
Executive summary:

Thirty-one chemicals, representing various organic and inorganic groups, were tested for ability to induce back-mutations from streptomycin dependence (Sd-4) to non-dependence in E. coli. Nineteen were found to be mutagens. It was demonstrated that mutagenicity is not a specific property of any group of chemicals, but appears among widely different groups. Chemicals having such different properties as boric acid, ammonia, hydrogen peroxide, copper sulfite, acetic acid, formaldehyde, and phenol were found to be mutagenic, indicating that genetic changes may be induced by many agents that are able to enter the living cell and upset its metabolic functions. Lactic acid was not found to be mutagenic and is therefore considered to be non-mutagenic in this bacterial reverse gene mutation assay.