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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:
2003
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study performed according to OECD guideline 471 in compliance to GLP.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2003
Report Date:
2003

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Principles of method if other than guideline:
N/A
GLP compliance:
yes (incl. certificate)
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
Test item : HC Blue No 2
EC number : 251-410-3
Batch number : 118pt 2

Method

Target gene:
Histidine gene
Species / strainopen allclose all
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98 and TA 100
Details on mammalian cell type (if applicable):
N/A
Additional strain / cell type characteristics:
other: see below
Species / strain / cell type:
S. typhimurium TA 102
Details on mammalian cell type (if applicable):
N/A
Additional strain / cell type characteristics:
not applicable
Metabolic activation:
with and without
Metabolic activation system:
rat liver S9
Test concentrations with justification for top dose:
Experiment 1 :
50, 150, 500, 1500 and 5000 ug/plate

Experiment 2 :
50, 150, 500, 1500 and 5000 ug/plate for all of the tester strains except TA98 which was allocated an amended dose range of 150, 500, 1500, 3000 and 5000 ug/plate. The intermediate dose level (3000 ug/plate) was included to achieve a better dose-response relationship.
Vehicle / solvent:
Solvent (DMSO)
Controlsopen allclose all
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
N-ethyl-N-nitro-N-nitrosoguanidine
Remarks:
TA100 and TA1535 (both without S9)
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
9-aminoacridine
Remarks:
TA1537 (without S9)
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
mitomycin C
Remarks:
TA102 (without S9)
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
4-nitroquinoline-N-oxide
Remarks:
TA98 (without S9)
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-aminoanthracene
Remarks:
TA100, TA1535, TA1537 (all with S9)
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
Remarks:
TA98 (with S9)
Untreated negative controls:
yes
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 1,8dihydroxyanthraquinone
Remarks:
TA102 (with S9)

Results and discussion

Test resultsopen allclose all
Species / strain:
S. typhimurium, other: TA1535, TA 1537, TA102, TA100
Metabolic activation:
with and without
Genotoxicity:
negative
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
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Species / strain:
S. typhimurium, other: TA1535, TA1537, TA102, TA100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
valid
Positive controls validity:
valid
Remarks on result:
other: other: experiment 1
Remarks:
Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
positive without metabolic activation

Under the experimental conditions used in this study, HC Blue No 2 was considered to be genotoxic (mutagenic) in Salmonella typhimurium strain TA98 in the absence of metabolic activation.
Executive summary:

Salmonella typhimurium strains TA1535, TA1537, TA102, TA98 and TA100 were treated with the test material HC Blue No 2 using the Ames plate incorporation method method at five dose levels, in triplicate, both with and without the addition of a rat liver homogenate metabolising system. The dose range was determined in a premiminary toxicity assay and was 50 to 5000 ug/plate in the first experiment. The experiment was repeated on a separate day using the same dose range as experiment 1 for all tester strains except TA98, fresh cultures of the bacterial strains and fresh test material formulations. The dose range for tester strain TA98 was amended slightly following information obtained from experiment 1 and was 150, 500, 1500, 3000 and 5000 ug/plate. The intermediate dose level of 3000 ug/plate was included in an attempt to establish a better dose-response relationship. The vehicle control plates gave counts of revertant colonies within the normal range. All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies, both with and without metabolic activation. The sensitivity of the assay and the efficacy of the S9 mix were validated. Statistically significant, dose related and reproducible increases in the frequency of revertant colonies were observed in tester strain TA98, without metabolic activation, at the upper dose levels of the test material. No significant increases in the frequency of revertant colonies were recorded for any of the remaining bacterial strains, with any dose of the test material, either with or without metabolic activation. Under the experimental conditions used in this study, HC Blue No 2 was considered to be genotoxic (mutagenic) in Salmonella typhimurium strain TA98 in the absence of metabolic activation.