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EC number: 220-006-9
CAS number: 2601-33-4
method was designed to conform to the guidelines for bacterial
mutagenicity testing published by the major Japanese Regulatory
Authorities including METI, MHLW and MAFF. It also meets the
requirements of the OECD Guidelines for Testing of Chemicals No. 471
"Bacterial Reverse Mutation Test", Method B13/14 of Commission Directive
2000/32/EC and the USA, EPA (TSCA) OPPTS harmonised guidelines.
TA1535, TA1537, TA98 and TA100 and Escherichia
WP2uvrA· were treated with the test material using the Ames plate incorporation
method at up to seven dose levels, in triplicate, both with and without
the addition of a rat liver homogenate metabolising system (10% liver S9
in standard co-factors). The dose range for the range-finding test was
determined in a preliminary toxicity
assay and was 5 to 1500 µg/plate for the
Salmonella strains and 5 to 5000 µg/plate for Escherichia
coli strain WP2uvrA-. The experiment
was repeated on a separate day using fresh cultures of
the bacterial strains and fresh test
material formulations. The test material dose range was amended slightly, following the
results of the range-finding test, and was 5 to 1500 µg/plate for all of
the bacterial strains.
dose levels were included to allow for test material induced toxicity,
ensuring that at least four non-toxic doses were achieved.
vehicle (tetrahydrofuran) control plates gave counts of revertant
colonies within the normal range. All of the positive control chemicals used
in the test induced
increases in the
frequency of revertant colonies, both with or without metabolic
activation. Thus, the sensitivity
of the assay and the efficacy of the S9-mix were validated. At
500 µg/plate the test material caused a visible reduction in the growth
of the bacterial background lawn of all of the tester strains tested
either with or without S9-mix. The test material was, therefore, tested
up to the maximum recommended dose level of 5000 µg/plate or the toxic
limit, depending on bacterialstraintype
and experiment number. No test material precipitate was observed on the
plates at any of the doses tested in either the presence or absence of S9-mix.
significant increases in the frequency of revertant colonies were
recorded for any of the
bacterial strains, with any dose of the test material, either with or
without metabolic activation
test material was considered to be non-mutagenic under the
conditions of this test.
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