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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
Study period:
1988
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Although the test protocol was not described in the publication (in stead, reference to another publication was made), the results described in the paper combine the results of Ames testing of 300 substances by a renowed Research institute (National Institute of Environmental Health Sciences, Genetic toxicity department) in the framework of the National Toxicology Program.

Data source

Reference
Reference Type:
publication
Title:
Salmonella Mutagenicity Tests: IV. Results from the testing of 300 chemicals.
Author:
Zeiger E, Anderson B, Haworth S, Lawler T and Mortelmans K
Year:
1988
Bibliographic source:
Environmental and Molecular Mutagenesis 11 (Suppl. 12), 1-158

Materials and methods

Principles of method if other than guideline:
Ames test
GLP compliance:
not specified
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name: N,N'-di-sec-butyl-p-phenylenediamine
- Analytical purity: > 82%
- Source: Pfaltz & Bauer

Method

Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 97
Species / strain / cell type:
S. typhimurium TA 98
Species / strain / cell type:
S. typhimurium TA 100
Species / strain / cell type:
S. typhimurium TA 1535
Metabolic activation:
with and without
Metabolic activation system:
rat and hamster S-9 fractions
Test concentrations with justification for top dose:
0.000 - 0.330 - 1.000 - 3.300 - 10.000 - 33.000 - 100.000 - 200.000 - 333.000 µg/plate (as tested by lab. MIC)
0.000 - 0.300 - 1.000 - 3.000 - 10.000 - 16.000 - 33.000 - 66.000 - 100.000 - 166.000 - 333.000 µg/plate (as tested by lab. SRI)
Vehicle / solvent:
DMSO
Controls
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
9-aminoacridine
sodium azide
other: 4-nitro-o-phenylenediamine, 2-aminoanthracene
Details on test system and experimental conditions:
METHOD OF APPLICATION: preincubation assay

DURATION
- Preincubation period: 20 minutes
- Expression time (cells in growth medium): 2 days

NUMBER OF REPLICATIONS: 3

DETERMINATION OF CYTOTOXICITY
The toxicity assay was performed using TA100 or the system developed by Waleh et al (Waleh, N.S., Rapport, S.J., Mortelmans, K. 1982. Development of a toxicity test to be coupled to the Ames Salmonella assay and the method of construction of the required strains. Mutat. Res. 97:247-256.). Toxic concentrations were those that produced a decrease in the number of his+ colonies, or a clearing in the density of the background lawn, or both.
Evaluation criteria:
Evaluations were made at both the individual trial and overall chemicals. Individual trials were judged mutagenic (+), weakly mutagenic (+W), questionable (?) or non-mutagenic (-), depending on the magnitude of the increase of his+ revertants, and the shape of the dose-response. A trial was considered questionable (?) if the dose-response was judged insufficiently high to support a call of "+W", if only a single dose was elevated over the control, or if the increase seen was not dose related. It was not necessary for a response to reach twofold background for a chemical to be judged mutagenic.
A chemical was judged mutagenic (+) or weakly mutagenic (+W) if it produced a reproducible dose-related response over the solvent control in replicate trials. A chemical was judged questionable (?) if the results of individual trials were not reproducible, if increases in his+ revertants did not meet the criteria for a "+W" response, or if only single doses produced increases in his+ revertants in repeat trials. Chemicals were judged non-mutagenic (-) if they did not meet the criteria for a mutagenic or questionable response.

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 97
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Vehicle controls validity:
valid
Positive controls validity:
valid

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

The test substance was not mutagenic in the Ames/Salmonella assay to strains TA97, TA98, TA100 or TA1535 in the presence or absence of metabolic activation.
Executive summary:

The substance was tested in the framework of the National Toxicology Program and coordinated by the National Institute of Environmental Health Sciences, Genetic Toxicology Department, Cellular and Genetic Toxicology Branch. The test was performed in two independent laboratories (Microbiological Associates Inc. (MIC) and SRI International), which both concluded the test substance to be non mutagenic in this assay. As such, a good interlaboratory reproducibility of the assay was shown.