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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:
other: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
8 July to 23 July, 1988
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
guideline study with acceptable restrictions
Remarks:
study conducted on the analogue substance; the read across justification is detailed in section 13. The Reliability of the Source Study is 2 (the study was performed using 5 strains but all of Samonella typhimurium, since followed the OECD 471 of 1983, which was modified later in 1997).
Justification for type of information:
The read across justification is detailed in section 13.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1988
Report Date:
1988

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
1983
Deviations:
no
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Test material form:
solid: particulate/powder

Method

Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Additional strain / cell type characteristics:
not specified
Species / strain / cell type:
other: S. typhimurium TA 1538
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9
Test concentrations with justification for top dose:
10.0; 100.0; 333.3; 1000.0; and 5000.0 µg/plate

Dose selection was based on the results of the pre-experiment. The concentration range covered at least two decadic logarithms. Appropriate dose levels were selected regarding the characteristics of the test item. At least five different concentrations of the test item were tested. The highest dose level should prodece some toxic eggects. The maximum concentration is 5000 µg/plate, unless limited by toxicity or solubility of the test item.
In case the results of pre-experiment were in accordance with the these criteria, these data were reported as part of the main experiments.
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: water, DMSO
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
without S9, TA 1535 and TA 100
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 4-nitro-o-phenylene-diamine
Remarks:
without S9, TA1535, TA 1538 and TA 98
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene
Remarks:
with S9, TA 1535, TA 1537, TA 1538, TA 98 and TA 100
Details on test system and experimental conditions:
METHOD OF APPLICATION: in Agar
NUMBER OF REPLICATES: 3
Evaluation criteria:
A test item is considered a mutagen if, in strain TA 100, the number of reversions is at least twice as high and in strains TA 1535, TA 1537, TA 1538, and TA 98 it is at least three times higher, compared to the spontaneous reversion rate.
Also, a dose-dependent increase in the number of revertants is regarded as an indication of possibly existing mutagenic potential of the substance regardless of whether the highest dose induced the above described enhancement factors or not.
Statistics:
No valid statistical procedure can be recommended for analysis of data from the bacterial assays at this time

Results and discussion

Test resultsopen allclose all
Species / strain:
other: S. typhimurium TA 1537, TA 1538 and TA 100
Metabolic activation:
with and without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 98
Metabolic activation:
without
Genotoxicity:
positive
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
The test item induces point mutations by base pair changes or frameshifts in the genome of the Salmonella typhimurium strains TA1537, TA1538, TA98, and TA100, therefore, the test item is considered to be mutagenic in this reverse mutation (Ames) assay.
Executive summary:

The test item was evaluated for its ability to induce point mutations by base pair change or frameshift in an in vitro bacterial cell reverse mutation assay (Ames test), according to the plate incorporation method of the OECD Guideline 471 (1983). The assay used Salmonella typhimurium strains TA1535, TA1537, TA1538, TA98 and TA100 in two independent experiments, using identical procedures, both with and without liver microsomal activation (S9). Each concentration, including the controls, was tested in triplicate. The test item was tested at five concentrations from 10.0 to 5000 µg/plate. No toxic effects occurred in any of the test groups either in the presence or absence of metabolic activation, as evidenced by a reduction in the number of spontaneous revertants. The plates incubated with the test item showed normal background growth in concentrations up to 5000 ug/plate with and without S9 mix in all strains used. Appropriate reference mutagens were used as positive controls.

Up to the highest investigated dose, a significant and reproducible dose-dependent increase in revertant colony numbers was obtained in S. typhimurium strains TA1537, TA1538, TA98, and TA100. The presence of liver microsomal activation did not influence these findings. However, the positive response in strain TA98 was not reproduced in the independent experiment in the presence of S9 mix. Positive controls showed a distinct increase of induced revertant colonies.

Therefore, the test item is considered to be mutagenic in this reverse mutation (Ames) assay.