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Toxicological information

Genetic toxicity: in vitro

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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP compliant guideline study, available as unpublished report, no restrictions, fully adequate for assessment.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2001
Report Date:
2001

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 (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Name: FAT 66'036/B
Batch number: 008494MOXK
Physical state: solid
Purity: >85%

Method

Target gene:
Strain Target gene
TA 1537 his C 3076
TA 98 his D 3052
TA 1535 his G 46
TA 100 his G 46
WP2 uvrA Trp
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Metabolic activation system:
Phenobarbital/p-Naphthoflavone induced rat liver S9
Test concentrations with justification for top dose:
33, 100, 333, 1000, 2500 and 5000 µg/plate
Vehicle / solvent:
Deionised water
Controlsopen allclose all
Untreated negative controls:
yes
Remarks:
untreated
Negative solvent / vehicle controls:
yes
Remarks:
deionised water
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
Strains TA1535 and TA100, without metabolic activation
Untreated negative controls:
yes
Remarks:
untreated
Negative solvent / vehicle controls:
yes
Remarks:
deionised water
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 4-nitro-o-phenylene-diamine
Remarks:
Strains TA1537 and TA98, without metabolic activation
Untreated negative controls:
yes
Remarks:
untreated
Negative solvent / vehicle controls:
yes
Remarks:
deionised water
True negative controls:
no
Positive controls:
yes
Positive control substance:
methylmethanesulfonate
Remarks:
Strain WP2 uvrA, without metabolic activation
Untreated negative controls:
yes
Remarks:
untreated
Negative solvent / vehicle controls:
yes
Remarks:
deionised water
True negative controls:
no
Positive controls:
yes
Positive control substance:
other:
Remarks:
All strains, with metabolic activation
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation) (experiment 1); preincubation (experiment 2)

EXPERIMENT 1
DURATION
- Preincubation period: -
- Exposure duration: 72 hours at 37°C in the dark

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth / reduction in the number of revertants

EXPERIMENT 2
DURATION
- Preincubation period: 60 minutes
- Exposure duration: 72 hours at 37°C in the dark

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth / reduction in the number of revertants
Evaluation criteria:
A test item is considered as a mutagen if a biologically relevant increase in the number of revertants exceeding the threshold of twice (strains TA 98, TA 100 and WP2 uvrA) or thrice (strains TA 1535 and TA 1537) the colony count of the corresponding solvent control is observed.
A dose dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration.
An increase exceeding the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.
A dose dependent increase in the number of revertant colonies below the threshold is regarded as an indication of a mutagenic potential if reproduced in an independent second experiment. However, whenever the colony counts remain within the historical range of negative and solvent controls such an increase is not considered biologically relevant.
Statistics:
No statistical evaluation of the data is required.

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
The plates incubated with the test item showed normal background growth up to 5000 µg/plate with and without metabolic activation in both independent experiments.
No relevant toxic effects, evident as a reduction in the number of revertants, occurred in the test groups with and without metabolic activation.
No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with the substance at any dose level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance.
Appropriate reference mutagens were used as positive controls and showed a distinct increase of induced revertant colonies.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion