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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
Study period:
2004-10-14 - 2004-11-12
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: GLP guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2005
Report Date:
2005

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Target gene:
Histidine operon for TA-1535, TA-1537, TA-98, TA-100 and tryptophan operon for WP2 uvrA (pKM101)
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 pKM 101
Metabolic activation:
with and without
Metabolic activation system:
S9-mix purchased from MolTox Inc.
Test concentrations with justification for top dose:
For initial dose finding experiments: 100, 200, 500, 1000, 2500, 5000 µg/plate with and without S9-Mix
For main experiment: 2, 5, 10, 20, 50, 100 µg/plate with and without S9-Mix
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: Dimethylsulfoxid (DMSO), water for sodium azide positive control only
- Justification for choice of solvent/vehicle: Solubility of substances
Controlsopen allclose all
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
DMSO 10 µL/mL culture volume
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: 2-Aminoanthracene (2AA), 1 µg/plate for TA-98 and TA-100, 2 µg/plate for TA-1535 and TA-1537
Remarks:
with S9-mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
DMSO 10µL/mL culture volume
True negative controls:
no
Positive controls:
yes
Positive control substance:
benzo(a)pyrene
Remarks:
with S9-mix Migrated to IUCLID6: 5 µg/plate for WP2 uvrA(pKM 101)
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
DMSO 10 µL/mL culture volume
True negative controls:
no
Positive controls:
yes
Positive control substance:
sodium azide
Remarks:
without S9-mix Migrated to IUCLID6: 2 µg/plate for TA-100 and TA-1535
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
DMSO 10 µL/mL culture volume
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: Acridine Mutagen ICR191, 2 µg/plate for TA-1537
Remarks:
without S9-mix
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
DMSO 10 µL/mL culture volume
True negative controls:
no
Positive controls:
yes
Positive control substance:
no
Remarks:
without S9-mix Migrated to IUCLID6: Daunomycin Hydrochloride (DR), 1 µg/plate for TA-98
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
Remarks:
DMSO 10 µL/mL culture volume
True negative controls:
no
Positive controls:
yes
Positive control substance:
N-ethyl-N-nitro-N-nitrosoguanidine
Remarks:
without S9-mix Migrated to IUCLID6: 1 µg/plate for WP2 uvrA(pKM 101)
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation),

DURATION
- Preincubation period: 1 h
- Exposure duration: 3 days

NUMBER OF REPLICATIONS: two independent experiments, in triplicates each

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth by colony count

Evaluation criteria:
A positive response in a (valid) individual experiment is achieved when one or both of the following criteria are met: a) a significant, dose-related increase in the mean number of revertants is observed; b) a two-fold or greater increase in the mean number of revertant colonies (over that observed for the concurrent solvent control plates) is observed at one or more concentrations
A negative result in a (valid) individual experiment is achieved when: a) there is no significant dose-related increase in the mean number of revertant colonies per plate observed for the test substance; and b) in the absence of any such dose response, no increase in colony numbers is observed (at any test concentration) which exceeds 2x the concurrent solvent control.
Statistics:
All derived calculations (i.e. mean colony count/plate; standard deviation, etc.) were carried out by computer, no further specification of methods

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:
≥200 µg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Species / strain:
E. coli WP2 uvr A pKM 101
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
cytotoxicity
Remarks:
≥200 µg/plate
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
- RANGE-FINDING/SCREENING STUDIES: In an initial experiment, significant toxicity was observed at concentrations of 200 µg/plate and above.
- The positive controls for each experiment induced the expected responses, indicating the strains were responding satisfactorily in each case. - COMPARISON WITH HISTORICAL CONTROL DATA: Data of positive and vehicle controls is range of experiments done between 12/1997 and 06/2004

Any other information on results incl. tables

In two subsequent experiments with each tester strain, the test substance did not induce any significant, reproducible increases in the observed numbers of revertant colonies with any of the tester strains in the presence or absence of S9-mix.

It is concluded that, under the conditions of this assay, ATMER 163 gave a negative, i.e. non-mutagenic, response in S. typhimurium strains TA1535, TA1537, TA98 and TA100 and E. coli strain WP2 uvrA (pKM101) in both the presence and absence of S9-mix.

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative