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EC number: 237-354-2
CAS number: 13760-80-0
Table 1: Summary of Experiment 1
± S9 Mix
Mean number of colonies/plate
Base-pair Substitution Type
Mean no. colonies/plate
ENNG = N-ethyl-N’-nitro-N-nitrosoguanidine
4NQO = 4-Nitroquinoline-1-oxide
9AA = 9-aminoacridine
2AA = 2-aminoanthracene
BP = benzo(a)pyrene
Table 2: Summary of Experiment 2
The genetic toxicity of the test material was investigated in accordance
with the standardised guidelines OECD 471, EU Method B13/14, EPA OCSPP870.5100
and the major Japanese Regulatory Authorities including
METI, MHLW and MAFF. The reverse mutation assay was performed under GLP
Salmonella typhimurium strains TA1535, TA1537, TA98 and TA100 and
Escherichia coli strain WP2uvrA were treated with suspensions of the
test material using both the Ames plate incorporation and pre-incubation
methods at up to eight 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 Experiment 1 was
predetermined and was 1.5 to 5000 µg/plate. The
experiment was repeated on a separate day (pre-incubation method) using
fresh cultures of the bacterial strains and fresh test material
formulations. The dose range was amended following the
results of Experiment 1 and was 15 to 5000 µg/plate. Six
test material concentrations were selected in Experiment 2 in order to
achieve both four non-toxic dose levels and the potential toxic limit of
the test material following the change in test methodology.
The vehicle (dimethyl sulphoxide) control plates gave counts of
revertant colonies within the normal range. All of the
positive control chemicals used in the test induced marked increases in
the frequency of revertant colonies, both with and without metabolic
activation. Thus, the sensitivity of the assay and the
efficacy of the S9-mix were validated.
The maximum dose level of the test material in the first experiment was
selected as the maximum recommended dose level of 5000 µg/plate. There
was no visible reduction in the growth of the bacterial background lawn
at any dose level, either in the presence or absence of metabolic
activation (S9-mix), in the first mutation test (plate incorporation
method) and consequently the same maximum dose level was used in the
second mutation test. Similarly, there was no visible
reduction in the growth of the bacterial background lawn at any dose
level, either in the presence or absence of metabolic activation
(S9-mix), in the second mutation test (pre-incubation method). No
test material precipitate was observed on the plates at any of the doses
tested in either the presence or absence of S9-mix.
There were no significant increases in the frequency of revertant
colonies recorded for any of the bacterial strains, with any dose of the
test material, either with or without metabolic activation (S9-mix), in
Experiment 1 (plate incorporation method). Similarly,
no 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 (S9-mix), in
Experiment 2 (pre incubation method).
Under the conditions of this study the test material was considered to
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