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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:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study
Remarks:
Read-across, Original study in Japanese

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1995

Materials and methods

Test guideline
Qualifier:
according to guideline
Guideline:
other: Guidelines for Screening Mutagenicity Testing of Chemicals (Japan) and OECD (471 and 472)
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Reference substance name:
6,6'-di-tert-butyl-4,4'-thiodi-m-cresol
EC Number:
202-525-2
EC Name:
6,6'-di-tert-butyl-4,4'-thiodi-m-cresol
Cas Number:
96-69-5
Molecular formula:
C22H30O2S
Specific details on test material used for the study:
SOURCE OF TEST MATERIAL
- Source and lot/batch No.of test material: Sumitomo Chemical Co., Ltd; 40701
- Purity: >98%

Method

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 and 5,6-benzoflavone-induced rat liver S9
Test concentrations with justification for top dose:
Dose-range finding: 0, 50, 150, 500, 1500, 5000 µg/plate.

Main tests:
-S9 mix, 0, 0.781, 1.56, 3.13, 6.25, 12.5, 25 and 50 μg/plate (TA100, TA1535 and TA1537), 0, 3.13 - 200 μg/plate (TA98) and 0, 313 - 5000 μg/plate (WP2)

+S9 mix, 0, 12.5, 25, 50, 100, 200, 400 and 800 μg/plate (TA100, TA1535, TA98 and TA1537) and 0, 313 - 5000 μg/plate (WP2)
Vehicle / solvent:
DMSO
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
True negative controls:
no
Positive controls:
yes
Remarks:
-S9 mix: 2-(2-Furyl)-3-(5-nitro-2-furyl)acrylamide (TA100, WP2, TA98), Sodium azide (TA1535)and 9-Aminoacridine (TA1537); +S9 mix, 2-Aminoanthracene (five strains)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 hours

NUMBER OF REPLICATIONS: 2
Evaluation criteria:
A concentration-related increase over the range tested and/or a reproducible increase at one or more concentrations in the number of revertant colonies per plate in at least one strain with or without metabolic activation system.

Results and discussion

Test resultsopen allclose all
Species / strain:
other: S. typhimurium TA98, TA100, TA1535, TA1537
Metabolic activation:
without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
>12.5 μg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Species / strain:
other: S. typhimurium TA98, TA100, TA1535, TA1537
Metabolic activation:
with
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
>400 μg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Species / strain:
E. coli WP2
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: In main test 1, precipitate was observed at 50μg/plate -S9 and at 400μg/plate +S9 in all S. typhimurium strains. Precipitate was observed at all concentrations in the E. coli strain.

In main test 2, precipitate was observed at 50μg/plate -S9 and at 200μg/plate +S9 in all S. typhimurium strains. Precipitate was observed at all concentrations in the E. coli strain.



Applicant's summary and conclusion

Conclusions:
Under the conditions described for this study, it is concluded that TBBC is non-mutagenic in Salmonella typhimurium TA100, TA1535, TA98, TA1537 and Escherichia coli WP2 uvrA.
Executive summary:

In a reverse gene mutation assay in bacteria, strains of Salmonella typhimurium TA100, TA1535, TA98, TA1537 and Escherichia coli WP2 uvrA were exposed to TBBC (>98%) in DMSO at concentrations of 0, 0.781, 1.56, 3.13, 6.25, 12.5, 25 and 50 μg/plate (TA100, TA1535 and TA1537), 0, 3.13 - 200 μg/plate (TA98) and 0, 313 - 5000 μg/plate (WP2) and 0, 12.5, 25, 50, 100, 200, 400 and 800 μg/plate (TA100, TA1535, TA98 and TA1537) and 0, 313 - 5000 μg/plate (WP2) in the absence and presence of mammalian metabolic activation respectively (Phenobarbital and 5,6-benzoflavone-induced rat liver S9).

Toxicity was observed at 12.5 μg/plate (TA100 and TA1537), 25 μg/plate (TA1535) and 100 μg/plate (TA98) without S9 mix, and 400 μg/plate (TA100, TA1535 and TA1537) and 500 μg/plate (TA98) with S9 mix. No toxicity was observed in WP2 either without S9 mix or with S9 mix. 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. This study satisfies the requirement for Test Guideline OECD 471 for in vitro mutagenicity (bacterial reverse gene mutation) data.