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Toxicological information

Genetic toxicity: in vitro

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Administrative data

Endpoint:
in vitro gene mutation study in mammalian cells
Remarks:
Type of genotoxicity: gene mutation
Type of information:
experimental study
Adequacy of study:
key study
Study period:
From 21 MAR 1994 to 14 JUN 1994
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Guideline study (OECD 476)

Data source

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

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)
Deviations:
no
GLP compliance:
yes (incl. certificate)
Remarks:
in accordance with Directive 88/320/EEC
Type of assay:
mammalian cell gene mutation assay

Test material

Reference
Name:
Unnamed
Type:
Constituent

Method

Target gene:
TK
Species / strain
Species / strain:
mouse lymphoma L5178Y cells
Details on mammalian cell lines (if applicable):
- Type and identity of media: RPMI 1640 medium supplemented with 10% donor horse serum and 20 mM HEPES buffer (R10)
- Properly maintained: yes
- Periodically checked for Mycoplasma contamination: no data
- Periodically checked for karyotype stability: no data
- Periodically "cleansed" against high spontaneous background: yes
Additional strain characteristics:
not specified
Metabolic activation:
with and without
Metabolic activation system:
rat liver S9 mix from Aroclor 1254 pretreated rats
Test concentrations with justification for top dose:
62.5, 125, 250, 500, 1000 µg/l
Vehicle:
- dimethyl sulphoxide
Controlsopen allclose all
Negative controls:
no
Solvent controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
cyclophosphamide
Remarks:
with metabolic activation
Negative controls:
no
Solvent controls:
yes
True negative controls:
no
Positive controls:
yes
Positive control substance:
ethylmethanesulphonate
Remarks:
without metabolic activation
Details on test system and conditions:

DURATION
- Exposure duration: 3 hrs with manual shaking at approximately 0.5 hour intervals
- Expression time (cells in growth medium): 24 hrs
- Selection time (if incubation with a selection agent): 10-14 days


SELECTION AGENT (mutation assays): trifluorothymidine (4 µg/ml)

NUMBER OF REPLICATIONS: 2 experiments, treatments performed in duplicate, both with and without metabolic activation

DETERMINATION OF CYTOTOXICITY
- Method: colony formation

Results and discussion

Test results
Species / strain:
mouse lymphoma L5178Y cells
Metabolic activation:
with and without
Genotoxicity:
negative
Cytotoxicity:
no
Vehicle controls valid:
yes
Negative controls valid:
not examined
Positive controls valid:
yes
Remarks on result:
other: all strains/cell types tested
Remarks:
Migrated from field 'Test system'.
Additional information on results:
TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: a precipitate of the test item was noted at all dose levels

RANGE-FINDING/SCREENING STUDIES:
- the maximum dose level of 1000 µg/ml was selected on the basis that it was the maximum practical dose level that could be achieved

Any other information on results incl. tables

- no marked reductions in viability of cells treated with the test item in the presence or absence of metabolic activation

- vehicle controls were within the normal range (10 - 125 x 10E-6)

- positive controls gave marked increases in mutant frequency, demonstrating that the test system was operating satisfactorily

- the test item did not induce any statistically significant or dose related increase in mutant frequency

- experiment 1 and 2 gave concordant results

Applicant's summary and conclusion

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

The test substance did not increase the mutant frequency at the TK+/- locus in L5178Y cells and is therefore considered to be non-mutagenic under the conditions of the test.
Executive summary:

Mouse Lymphoma cells (L5178Y TK+/-) were treated with the test substance at five concentrations in the range of 62.5 to 1000 µg/ml. A heavy precipitate of the test substance was noted at all dose levels. The solvent control gave mutant frequencies within the normal range. The positive controls gave marked increases in the mutant frequency, both in the presence or absence of metabolic activation, indicating the satisfactory performance of the test system. The test substance demonstrated no statistically significant or dose related increases in mutant frequency at any dose level, either with or without metabolic activation, thus indicating that the test item is not mutagenic under the conditions tested.