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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
Study period:
26-Oct-2005 to 30-Nov-2010
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: The study has been performed according to the guideline stipulated by the Japanese Ministry of health, labour and welfare.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
JAPAN: Guidelines for Screening Mutagenicity Testing Of Chemicals
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Substance type: yellow powder
- Lot/batch No.: 50162

Method

Target gene:
- S. typhimurium: Histidine gene
- E. coli: Tryptophan gene
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Species / strain / cell type:
E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Rat liver S9-mix induced by a combination of phenobarbital and ß-naphthoflavone
Test concentrations with justification for top dose:
Experiment 1:
Without and with S9-mix: 20, 78, 313, 1250 and 5000 µg/plate
Experiment 2:
TA100
Without S9-mix: 78, 156, 313, 625 and 1250 µg/plate
With S9-mix: 313, 625, 1250, 2500 and 5000 µg/plate
TA1535, TA1537, TA98 and WP2uvrA
Without S9-mix: 313, 625, 1250, 2500 and 5000 µg/plate
With S9-mix: 313, 625, 1250, 2500 and 5000 µg/plate
Experiment 2:
TA100
Without S9-mix: 78, 156, 313, 625 and 1250 µg/plate
With S9-mix: 313, 625, 1250, 2500 and 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: Test compound was soluble in DMSO and DMSO has been accepted and approved by authorities and international guidelines
Controlsopen allclose all
Negative solvent / vehicle controls:
yes
Positive controls:
yes
Positive control substance:
other: 2-(2-Furyl)-3-(5-Nitro-2-Furyl)acrylamide for TA98, TA100 and WP2uvrA
Remarks:
without S9
Positive control substance:
9-aminoacridine
Remarks:
without S9

Migrated to IUCLID6: for TA1537
Positive control substance:
N-ethyl-N-nitro-N-nitrosoguanidine
Remarks:
without S9

Migrated to IUCLID6: for TA1535
Positive control substance:
other: 2-aminoanthracene in DMSO for all tester strains
Remarks:
with S9
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (preincubation, 20 min 37 °C)

DURATION
- Exposure duration: 48 hour

NUMBER OF REPLICATIONS:
- Doses of the test substance were tested in duplo in each strain. Two independent experiments were conducted. An additional experiment with TA100 was performed.

NUMBER OF CELLS EVALUATED: 10E8 per plate

DETERMINATION OF CYTOTOXICITY
- Method: The reduction of the bacterial background lawn, the increase in the size of the microcolonies and the reduction of the revertant colonies.

OTHER EXAMINATIONS:
- The presence of precipitation of the test compound on the plates was determined.
Evaluation criteria:
A compound is deemed to provide evidence of mutagenic potential if:
(1) a statistically significant dose-related increase in the number of revertant colonies is obtained in two seperate experiments and
(2) the increase in the number of revertant colonies is at least twice the concurrent solvent control value and
(3) the reappearance of (1) and (2) is certificated

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
But tested beyond toxicity
Vehicle controls validity:
valid
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
But tested beyond toxicity
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: Precipitation was observed at dose levels of 78 µg/plate

RANGE-FINDING/SCREENING STUDIES:
- In tester strain TA100, toxicity was observed at dose levels of 1250 μg/plate and above in the absence of S9-mix. In the tester strains TA98 and TA1537, toxicity was observed at 5000 µg/plate in the absence of S9-mix.
EXPERIMENT 2
- In the tester strains WP2uvrA and TA1537, toxicity was observed at 5000 µg/plate in the absence of S9-mix. In tester strain TA1537, toxicity was observed at 5000 µg/plate in the presence of S9-mix.
EXPERIMENT 3
- In tester strain TA100, toxicity was observed at 1250 and 5000 µg/plate in the absence and presence of S9-mix, respectively.

COMPARISON WITH HISTORICAL CONTROL DATA:
- The negative and strain-specific positive control values were within our laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.


Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

It is concluded that the test substance shows no evidence of mutagenic activity when tested in this bacterial system.