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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:
August 18, 1992 - September 10, 1992
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: Acceptable, well documented study report which meets basic scientific principles

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1992
Report Date:
1992

Materials and methods

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

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Physical state: solid; white
- Storage condition of test material: room temperature

Method

Target gene:
his, trp
Species / strain
Species / strain / cell type:
other: S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2 uvr A
Metabolic activation:
with and without
Metabolic activation system:
Aroclor induced rat liver S9-mix
Test concentrations with justification for top dose:
Range in the cytotoxicity test: 20.6 - 5000 µg/plate
Range in the mutagenicity test: 185.2 - 5000 µg/plate
Vehicle / solvent:
- Vehicle(s)/solvent(s) used: DMSO
Controls
Untreated negative controls:
no
Negative solvent / vehicle controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
other: see below
Remarks:
with and without metabolic activation
Details on test system and experimental conditions:
METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 hours
- Expression time (cells in growth medium): 48 hours

NUMBER OF REPLICATIONS: two plates per test substance concentration

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth

POSITIVE CONTROLS
WITHOUT S9:
TA 100, TA 1535: sodium azide in bidist.water, 5.0 µg/plate
WP2 uvrA: 4-nitroquinoline-N-oxide in DMSO, 2.0 µg/plate
TA 98: 2-nitrofluorene in DMSO, 20.0 µg/plate
TA 1537: 9(5)-aminoacridine in DMSO, 150.0 µg/plate

WITH S9
TA 100, TA 98, TA 1537: 2-aminoanthracene in DMSO, 2.5 µg/plate
TA 1535: cyclophosphamide in bidist.water, 400.0 µg/plate
WP2 uvrA: 2-aminoanthracene in DMSO, 50.0 µg/plate
Evaluation criteria:
The test substance is considered to be mutagenic in this test system if:
- A positive effect is observed in one strain and the effect can be reproduced in a confirmatory experiment.
- A positive effect is observed in two or more strains.
A positive effect is defined as an increase of the mean number of revertants per plate for any concentration above that of the negative control by at least a factor of 2.0 with strains TA 98, TA 1535, TA 1537 and WP2 uvrA, or by a factor of at least 1.5 with strain TA 100. Generally a concentration-related effect should be demonstrable. If equivocal results are obtained, the final decision has to be based on scientific judgement.

Results and discussion

Test results
Species / strain:
other: S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2 uvr A
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
Additional information on results:
RANGE-FINDING/SCREENING STUDIES:
Six concentrations ranging from 20.6 to 5000 µg/plate were tested with strains S. typhimurium TA 100 and E.coli WP2 uvrA to determine the highest concentration to be used in the mutagenicity assay. The experiments were performed with and without metabolic activation. Normal background growth was observed with both strains. The numbers of revertant colonies were not reduced. From the results obtained, the highest concentration suitable for the mutagenicity test was selected to be 5000 µg/plate without and with metabolic activation.

COMPARISON WITH HISTORICAL CONTROL DATA:
Negative Controls were within the historical control range and positive controls produced the expected reactions.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
In the mutagenicity test normal background growth was observed with all strains at all concentrations. The numbers of revertant colonies were not reduced. The test substance exerted no toxic effect on the growth of the bacteria.
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.

Any other information on results incl. tables

Experiment with metabolic activation:

Treatment/Strain TA100 TA1535 WP2 uvrA TA98 TA1537
Negative control 100.0 10.0 15.5 41.5 7.0
185.2 µg/plate 117.5 17.0 20.5 40.5 7.0
555.6 µg/plate 115.0 9.5 15.5 41.5 5.5
1666.7 µg/plate 118.0 11.0 14.0 37.0 4.5
5000.0 µg/plate 89.0 12.0 16.5 44.0 7.0
2-aminoanthracene 841.5 573.5 1086.0 139.5
cyclophosphamide 431.5

Experiment without metabolic activation:

Treatment/Strain TA100 TA1535 WP2 uvrA TA98 TA1537
Negative control 85.5 13.0 14.0 17.0 6.0
185.2 µg/plate 72.0 16.0 17.0 18.0 5.0
555.6 µg/plate 84.0 12.0 17.5 19.5 3.5
1666.7 µg/plate 84.0 16.5 14.5 12.5 3.5
5000.0 µg/plate 67.5 17.5 13.5 12.5 4.5
sodium azide 1066.0 790.0
4-NQO 789.0
2-nitrofluorene 1230.5
9-aminoacridine 1549.5

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information):
negative

Based on the results of these experiments and on standard evaluation criteria, it is concluded that the test item and its metabolites did not induce gene mutations in the strains of S. typhimurium and E. coli used.