Registration Dossier

Toxicological information

Genetic toxicity: in vitro

Currently viewing:

Administrative data

Endpoint:
in vitro gene mutation study in bacteria
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
17-Feb-2014 to 03-Mar-2014
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study according to OECD 471.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2014
Report Date:
2014

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: clear colourless liquid
Details on test material:
- Name of test material (as cited in study report): L(+)-lactic acid
- Physical state: liquid
- Expiration date of the lot/batch: 01-Jan-2015
- Stability under test conditions: valid expiry date
- Storage condition of test material: room temperature in the dark

Method

Species / strainopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Additional strain characteristics:
other: each strain contained the following additional mutations: rfa, gal, chl, bio, uvrB
Metabolic activation:
with and without
Metabolic activation system:
S9-mix
Species / strain:
E. coli WP2 uvr A
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9-mix
Test concentrations with justification for top dose:
strains TA100 and WP2uvrA: 3, 10, 33, 100, 333, 1000, 3330 and 5000 µg/plate
strains TA1535, TA1537 and TA98: 100, 333, 1000, 3330 and 5000µg/plate
Vehicle:
- Vehicle(s)/solvent(s) used: water
Controls
Negative controls:
yes
Remarks:
water
Solvent controls:
yes
Remarks:
solvent for test substance was water
True negative controls:
no
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
2-nitrofluorene
sodium azide
methylmethanesulfonate
other: ICR-191, 2-aminoanthracene
Remarks:
2-aminoanthracene was used for all strains, sodium azide used for TA1535, ICR-191 for TA1537, 2-nitrofluorene for TA98, mehtylmethanesulfonate for TA100 and 4-nitroquinoline N-oxide for WO2uvrA
Details on test system and conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 ± 4 h
Evaluation criteria:
A test substance is considered negative (not mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is not greater than two (2) times the concurrent control, and the total number of revertants in tester strains TA1535, TA1537, TA98 or
WP2uvrA is not greater than three (3) times the concurrent vehicle control.
b) The negative response should be reproducible in at least one independently repeated experiment.

A test substance is considered positive (mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is greater than two (2) times the concurrent control, or the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is greater than three (3) times the concurrent vehicle control.
b) In case a repeat experiment is performed when a positive response is observed in one of the tester strains, the positive response should be reproducible in at least one independently repeated experiment.

The preceding criteria were not absolute and other modifying factors might enter into the final evaluation decision.

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
no
Vehicle controls valid:
yes
Positive controls valid:
yes
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Water solubility:
- Precipitation: no

RANGE-FINDING/SCREENING STUDIES: Dose range finding test performed with TA100 and WP2uvrA. Doses tested: 0, 3, 10, 33, 100, 333, 1000, 3330, 5000 µg/plate.

COMPARISON WITH HISTORICAL CONTROL DATA: The negative control values were within the laboratory historical control data ranges, except for TA100 in the absence of S9-mix, second experiment. Evaluation: The mean plate count (146) was just outside the limit of the range (144) and clear negative results are observed in all experiments, therefore this deviation in the mean plate count of the solvent control had no effect on the results of the study.

The strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly, except the response for TA1535 in the absence of S9-mix, first experiment. Evaluation: The value (257) was just below the limit of the range (262). The purpose of the positive control is as a reference for the test system, where a positive response is required to check if the test system functions correctly. Since the value was more than 3 times greater than the concurrent solvent control values, this deviation in the mean plate count of the positive control had no effect on the results of the study.


ADDITIONAL INFORMATION ON CYTOTOXICITY:
No reduction of the bacterial background lawn and no biologically relevant decrease in the number of revertants were observed

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative without metabolic activation
negative with metabolic activation

L(+)-lactic acid is not genotoxic in the reverse mutation assay.
Executive summary:
In a reverse gene mutation assay in bacteria, S. typhimurium strains TA1535, TA1537, TA98, TA100 and E. coli strain WP2uvrA were exposed to L(+)-lactic acid at concentrations of 0, 100, 333, 1000, 3330 and 5000 µg/plate in the presence and absence of mammalian metabolic activation. The test with metabolic activation (10% S9) was a plate incorporation test.

L(+)-lactic acid was tested up to the limit concentration of 5000 µg/plate. The positive controls induced the appropriate responses in the corresponding strains. There was no evidence of induced mutant colonies over background.

This study is classified as acceptable and satisfies the requirement for Test Guideline OECD 471 for in vitro mutagenicity (bacterial reverse gene mutation assay).