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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:
1992 - 1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
1992
Report Date:
1992
Reference Type:
other: amendment to CIBA-GEIGY924089 ["test dates"]
Title:
Unnamed
Year:
1994
Report Date:
1994
Reference Type:
other: amendment to CIBA-GEIGY924089 ["Purity" and "Expiration date"]
Title:
Unnamed
Year:
1994
Report Date:
1994
Reference Type:
other: amendment to CIBA-GEIGY924089 ["Appearance"]
Title:
Unnamed
Year:
1995
Report Date:
1995
Reference Type:
other: synopsis of CIBA-GEIGY924089
Title:
Unnamed
Year:
1992
Report Date:
1992
Reference Type:
other: expert statement to CIBA-GEIGY924089
Title:
Unnamed
Year:
1993
Report Date:
1993
Reference Type:
other company data
Title:
Unnamed
Year:
1991
Report Date:
1991

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 471 (Bacterial Reverse Mutation Assay)
Deviations:
no
Qualifier:
according to
Guideline:
EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)
Deviations:
no
Qualifier:
according to
Guideline:
EPA OTS 798.5265 (The Salmonella typhimurium Bacterial Reverse Mutation Test)
Deviations:
no
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

Method

Target gene:
In the Salmonella typhimurium strains (TA 1535, TA 100, TA 1537, TA 98) the amino acid histidine locus is the target gene, in which induced back mutations will transform the histidine auxotrophy (his-) to histidine prototrophy (his+). The Salmonella typhimurium histidine (his) reversion system measures his- => his+ reversions. The Salmonella typhimurium strains are constructed to differentiate between base pair (TA 1535, TA 100) and frameshift (TA 1537, TA 98) mutations. The strain Escherichia coli (WP2 uvrA) is a tryptophan-auxotrophic strain (base-pair substitution).
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:
S9 mix (from liver homogenate of Aroclor 1254 treated rats)
Test concentrations with justification for top dose:
0, 312.5, 625, 1250, 2500, and 5000 µg/plate.
Vehicle / solvent:
- Vehicle(s)/solvent used: DMSO (50 mg/mL, suspension)
- Justification for choice of solvent/vehicle: DMSO was chosen because of its solubility properties and its relative nontoxicity
- On the day of the experiment (immediately before the experiment), the test item was dissolved in DMSO
Controls
Untreated negative controls:
other: = vehicle control
Negative solvent / vehicle controls:
yes
Remarks:
DMSO
Positive controls:
yes
Positive control substance:
other: sodiuim azide, 4-NQO, 2-nitrofluorene (strains except TA 1537), 9-aminoacridine (TA 1537) - Without metabolic activation; 2-aminoanthracene (all strains), cyclophosphamide (all strains except TA 1537) - With metabolic activation
Remarks:
The solvent alone was used as the negative control.
Details on test system and experimental conditions:
METHOD OF APPLICATION: plate method, no preincubation
Evaluation criteria:
The generally accepted conditions for the evaluation of the results are:
- mean colony counts of the control values of all strains are within the acceptable ranges
- results of the positive controls meet the criteria for a positive response

The test item is considered as mutagen in case of:
- reproducible doubling of the mean number of revertants per plate above that of the negative control at any concentration; Strains: S. typhimurium TA 98, TA 1535, TA 1537 and E. coli WP2 uvrA
- reproducible increase of the mean number of revertants per plate for any concentration above that of the negative control by at least a factor of 1.5 for strain S. typhimurium TA 100
- a concentration-related effect should be demonstrable.
Statistics:
A statistical analysis was not performed.

Results and discussion

Test resultsopen allclose all
Key result
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:
no cytotoxicity nor precipitates, but tested up to recommended limit concentrations
Vehicle controls validity:
valid
Untreated negative controls validity:
other: = vehicle control
Positive controls validity:
valid
Key result
Species / strain:
E. coli WP2 uvr A
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:
other: = vehicle control
Positive controls validity:
valid
Additional information on results:
Concentration of test substance resulting in precipitation: approximately 600 - 5000.0 µg/plate.

Any other information on results incl. tables

Test item EC 411-380-6: Salmonella typhimurium/E.coli reverse mutation assay

Plate incorporation without S-9 mix: experiment I/II

Dose (µg/plate)

 TA 100

 TA 1535

 WP2 uvrA

 TA 98

 TA1537

 Negative control
(solvent alone)

131.7 / 139.3

12.0 / 10.3

17.0 / 14.7

21.7 / 13.0

6.0 / 7.0

 312.5

 112.7 / 129.3

 12.0 / 8.7

 17.0 / 16.7

 21.7 / 17.3

 4.3 / 

 625

 111.0 / 135.0

 11.3 / 9.0

 16 / 15.3

 21.7 / 12.7

 6.3 / 

 1250

 112.0 / 112.0

 11.3 / 7.0

 17 / 14.0

 18.0 / 12.3

 4.3 / 

 2500

 116.7 / 126.0

 8.3 / 7.3

 15 / 14.3

 19.0 / 17.0

 5.0 / 

 5000

 109.3 / 111.7

 10.3 / 7.3

 19 / 13.3

 21.7 / 14.0

 3.7 / 

Positive controls

 

 

 

 

 

- Sodum azide

 961.7 / 1092.3

 797.3 / 802.0

 

 

 

- 4-NQO

 

 

 758.0 / 935.7

 

 

2-nitrofluorene

 

 

 

 1209.7 / 1650.7

 

9-aminoacridine

 

 

 

 

 1436.3 / 1456.0

 

Plate incorporation with S-9 mix: experiment I/II

Dose (µg/plate)

 TA 100

 TA 1535

 WP2 uvrA

 TA 98

 TA1537

 Negative control
(solvent alone)

140.3 / 140.0

12.0 / 10.3

18.3 / 19.0

46.7 / 30.0

8.7 / 6.7

 312.5

 131.7/ 133.7

 12.0 / 9.3

 22.3 / 15.7

 37.3 / 27.3

 7.7 / 8.7

 625

 118.7 / 129.7

 11.7 / 14.0

 22.0 / 14.3

 36.0 / 28.3

 7.3 / 7.0

 1250

 120.0 / 112.0

 10.7 / 12.7

 21.3 / 13.3

 38.0 / 22.3

 7.3 / 6.7

 2500

 118.7 / 124.7

 13.0 / 10.3

 16.7 / 14.0

 43.0 / 24.7

 8.0 / 7.0

 5000

 129.0 / 96.7

 10.7 / 9.7

 16.7 / 20.3

 38.3 / 30.0

 5.0 / 4.3

Positive control

 

 

 

 

 

- 2-Aminoanthracene

 1650.7 / 1432.3

 

 1084.3 / 733.3

 1917.7 / 1785.7

 249.3 / 185.7

- Cyclophosphamide

 

 374.0 / 362.3

 

 

 

Revertants/plate (I / II): mean from three plates     

Applicant's summary and conclusion

Conclusions:
Interpretation of results: negative
Executive summary:

In a reverse gene mutation assay in bacteria, strains TA 1535, TA 1537, TA 98 and TA 100 of S. typhimurium and Escherichia coli strain WP2 uvrA were exposed to the test item (as described in section 1.2) dissolved in DMSO at concentrations of 312.5, 625, 1250, and 5000.0 µg/plate in the presence and absence of mammalian metabolic activation without pre-incubation. Concurrent positive and solvent controls were included. All concentrations were tested in triplicates. Concentrations of approximately 600 - 5000.0 µg/plate resulted in precipitation of the test item; no cytoxicity was noted up to the limit concentration. The positive controls induced the appropriate responses. No evidence of a mutagenic potential associated with the test substance was observed. This study is classified as acceptable and satisfies the requirement for Test Guideline OECD 471 for in vitro mutagenicity (bacterial reverse gene mutation) data.