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in vitro gene mutation study in bacteria
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Dec 2009
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP certified and acc. to OECD guideline

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
GLP compliance:
yes (incl. QA statement)
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
2,6-difluoro-3-(propane-1-sulfonamido)benzoic acid
EC Number:
Cas Number:
Molecular formula:
2,6-difluoro-3-(propane-1-sulfonamido)benzoic acid
Details on test material:
- Physical state: white powder
- Storage condition of test material: at room temperature, light protected

- Expiry Date: September 08, 2010


Target gene:
histidine mutation and tryptophan mutation
Strain Histidine mutation
TA1537 hisC3076
TA98 hisD3052/R-factor
TA1535 hisG46
TA100 hisG46/R-factor
*: R-factor = plasmid pKM101 (increases error-prone DNA repair)
Escherichia coli WP2uvrA strain
Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Test concentrations with justification for top dose:
3, 10, 33, 100, 333, 1000, 2500 and 5000 µg/plate
Vehicle / solvent:
- Vehicle used: DMSO (Merck, D-64293 Darmstadt; purity > 99%). The solvent was chosen because of its solubility properties and its relative non-toxicity to the bacteria (Maron et al., 1981 Compatibility of organic solvents with the Salmonella/Microsome Test, Mutation Res. 88, 343-350)
Untreated negative controls:
Negative solvent / vehicle controls:
Positive controls:
with and without metabolic activation
Positive control substance:
other: sodium azide, 4-NOPD, MMS, 2-AA
Details on test system and experimental conditions:
Method of application: in agar (plate incorporation)
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 (3).
A dose dependent increase is considered biologically relevant if the threshold is exceeded
at more than one concentration (2).
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.

Results and discussion

Test results
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
Untreated negative controls validity:
Positive controls validity:
Remarks on result:
other: all strains/cell types tested
Migrated from field 'Test system'.

Applicant's summary and conclusion

Interpretation of results (migrated information):

Based on the results of this study SAC-Sulfonamidsaure is considered to be non-mutagenic in
this Salmonella typhimurium and Escherichia coli reverse mutation assay.
Executive summary:

This study was performed to investigate the potential of SAC-Sulfonamidsaure to induce gene mutations in the plate incorporation test (experiment I) and the pre-incubation test (experiment 11) using the Salmonella typhimurium strains TA 1535, TA 1537, TA 98, and TA 100, and the Escherichia coli strain WP2 uvrA. The assay was performed in two independent experiments both with and without liver microsomal activation. Each concentration, including the controls, was tested in triplicate. The test item was tested at the following concentrations: Pre-Experiment/Experiment I and 11: 3; 10; 33; 100; 333; 1000; 2500; and 5000 !Jg/plate The plates incubated with the test item showed normal background growth up to 5000 !Jg/plate in nearly all strains with and without metabolic activation in both independent experiments. Only in experiment I reduced background growth was observed in strain WP2 uvrA from 1 000 !Jg/plate up to 5000 !Jg/plate in the absence of metabolic activation. No toxic effects, evident as a reduction in the number of revertants (below the indication factor of 0.5), occurred in nearly all test groups with and without metabolic activation. Only in experiment I a reduction in the number of revertants (below the indication factor of 0.5) was observed in strain WP2 uvrA at 5000 !Jg/plate in the absence of metabolic activation. No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with SAC-Sulfonamidsaure 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.