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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:
1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Justification for type of information:
None

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1994
Report date:
1994

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Version / remarks:
1983
Deviations:
no
Qualifier:
according to guideline
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Version / remarks:
1992
Deviations:
no
Qualifier:
according to guideline
Guideline:
EPA OTS 798.5265 (The Salmonella typhimurium Bacterial Reverse Mutation Test)
Version / remarks:
1987
Deviations:
no
Principles of method if other than guideline:
Guidelines followed
GLP compliance:
yes
Type of assay:
bacterial reverse mutation assay

Test material

Constituent 1
Chemical structure
Reference substance name:
[4-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-3-hydroxynaphthalene-1-sulphonato(3-)]chromium
EC Number:
271-352-2
EC Name:
[4-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-3-hydroxynaphthalene-1-sulphonato(3-)]chromium
Cas Number:
68541-72-0
Molecular formula:
C20H13CrN4O5S
IUPAC Name:
[4-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]-3-hydroxynaphthalene-1-sulphonato(3-)]chromium
Test material form:
solid: particulate/powder
Specific details on test material used for the study:
Test material: FAT 20042/D (Neolan Rosa BE ZP feucht)
Batch No.: 276
Purity: Approx. 50%
Appearance: Black-reddish humid pieces
Expiry date: October 1998
Storage: Room temperature
Material submitted by: CIBA-GEIGY Limited, Dyestuffs Division, Basle, Switzerland

Method

Target gene:
Histidine gene

Species / strain
Species / strain / cell type:
S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102
Additional strain / cell type characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
S9 mix
Test concentrations with justification for top dose:
Original and confirmatory experiment- 123.46, 370.37, 1111.11, 3333.33 and 10000.00 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: Dimethylsulfoxide (DMSO)
Controls
Untreated negative controls:
other: same as solvent control
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: all strains (except TA 1535) with S9: 2-aminoanthracene; for TA 1535 with S9: cyclophosphamide; for TA 100 and 1535 without S9: sodium azide, for TA 102 without S9: mitomycin-C; for TA 98 without S9: 2 nitrofluorene, for TA1537 without S9: 9-aminoacridine
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 h

NUMBER OF REPLICATIONS: 3 per test substance concentration and controls


OTHER EXAMINATIONS:
- Colonies were counted electronically using an Artek Colony Counter (Fisher Scientific), or manually where minor agar damage or test chemical precipitates might have interfered with automating counting. The results were sent on line to a computer. They were checked on a random basis by the operator. Observations indicating precipitates of the test substance in the top agar or a reduced or absent bacterial background lawn were registered additionally. Means for all mutagenicity assays were calculated and included in the Results section.
Rationale for test conditions:
None
Evaluation criteria:
Assay acceptance criteria: A test is considered acceptable if the mean colony counts of the control values of all strains are within the acceptable ranges and if the results of the positive controls meet the criteria for a positive response.
Statistics:
A statistical analysis of the test data was not performed.

Results and discussion

Test resultsopen allclose all
Key result
Species / strain:
S. typhimurium TA 1535
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 1537
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 100
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 102
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Key result
Species / strain:
S. typhimurium TA 98
Metabolic activation:
with and without
Genotoxicity:
positive
Remarks:
without metabolic activation only
Cytotoxicity / choice of top concentrations:
no cytotoxicity
Vehicle controls validity:
valid
Untreated negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
Range finding test
Normal background growth was observed. The numbers of revertant colonies were not reduced. From the results obtained, the highest concentration suitable for the mutagenicity test was selected to be 10000.0 µg/plate with and without metabolic activation (since the purity of the test substance is about 50%, this corresponds to a concentration of 5000 µg/plate).

Mutagenicity test, original experiment
In the experiment performed without metabolic activation, treatment of strain TA 98 with FAT 20042/D (Neolan Rosa BE ZP feucht) led to a slight increase in the number of back-mutant colonies at the highest concentration. A marginal increase at two intermediate concentrations was also registered with strain TA 102 in both, the experiment with and without metabolic activation. No effects were observed with the other strains

Mutagenicity test, confirmatory experiment
In the experiment performed without metabolic activation, again after treatment of strain TA 98 with FAT 20042/D (Neolan Rosa BE ZP feucht) a slight increase in the incidence of histidineprototrophic mutants was observed in comparison with the negative control. The slight increase in the number of revenants in the experiment without activation on strain TA 1535 at the concentration of 3333.3 µg/plate was not observed in the original experiment conducted on this strain and is attributed to spontaneously occurring back-mutants. No effects at all occurred and in the experiments with activation on strains TA 98 and TA 1535 and with the other strains.

The slight effect observed with strain TA 102 in the original experiment also is attributed to fluctuations in the incidence of spontaneously occurring back-mutant colonies.
In the mutagenicity tests normal background growth was observed with all strains at all concentrations. The numbers of revertant colonies were occasionally reduced at the upper concentrations.
There were no known circumstances or occurrences in this study that were considered to have affected the quality or integrity of the test data.

Any other information on results incl. tables

None

Applicant's summary and conclusion

Conclusions:
FAT 20042/D (Neolan Rosa BE ZP feucht) exerted a marginal mutagenic effect on strain TA 98 of Salmonella typhimurium without metabolic activation only.
Executive summary:

FAT 20042/D, identified as black-reddish humid pieces, purity approx. 50%, batch no. 276, was tested for mutagenic effects in vitro in histidine-requiring strains of Salmonella typhimurium. The following strains of Salmonella typhimurium were used: TA 98, TA 100, TA 102, TA 1535 and TA 1537. The test was performed with and without the addition of rat-liver post mitochondrial supernatant (S9 fraction) as an extrinsic metabolic activation system. The compound was dissolved in DMSO and tested at five concentrations in the range of 123.5 to 10000.0 µg/plate in the presence and absence of a metabolic activation system. In order to confirm the results, the experiments were repeated with and without metabolic activation at five concentrations in the range of 123.5 to 10000.0 µg/plate. Each strain was additionally tested in the presence and in the absence of a metabolic activation system with a suitable, known mutagen as positive control. FAT 20042/D led to a slight increase in the incidence of histidine-prototrophic mutants in comparison to the negative control in strain TA 98 at the highest concentration in both original as well as confirmatory experiments when tested in absence of metabolic activation. No relevant effects occurred with the other strains. Based on the results of these experiments and using standard evaluation criteria, it is concluded that FAT 20042/D (Neolan Rosa BE ZP feucht) exerted a marginal mutagenic effect on strain TA 98 of Salmonella typhimurium without metabolic activation only.