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EC number: 202-334-4
CAS number: 94-46-2
Gene mutation toxicity study was
performed to determine the mutagenic nature of Isoamyl benzoate. Genetic
toxicity test was performed on strain of Escherichia coli (strain
Sd-4-73) by paper disk method. Paper-disk method was performed to
check for the ability of E. coli Sd-4-73 to show reversion from
streptomycin dependence to independence. Overnight culture of
strain Sd-4-73 grown at 36°C in aerated nutrient broth containing 20
µg/ml of streptomycin was used as a inoculum. The culture was
centrifuged and washed at least twice with saline or distilled water to
remove the extraneous streptomycin, and was resuspended in saline to a
concentration of approximately 109 cells/ml. 0.1-ml. aliquot
of this suspension was mixed with 2.5 ml of molten soft nutrient agar
(0.7 per cent agar) and poured over a base of 20 ml of 1.5 per cent
nutrient agar. After the soft layer was solidified, the mutagen was
applied in form of small drops or crystal. Additional plates were
prepared with small inocula (one fifth and one twenty-fifth of the
original) so as not to miss the optimum cell density; the number of
cells per plate was rather critical, the yield of mutant colonies being
reduced either by crowding or by insufficient population size. After the
soft agar layer had set, the mutagen, in the form of a microdrop of
solution (0.01 to 0.025 ml.) or a small crystal, was applied to a small
filter-paper disk resting on the agar. To determine whether the
substance was inhibitory for the assay organism at the concentration
employed, the procedure was repeated on a nutrient agar plate containing
of streptomycin and seeded with approximately 107 bacteria.
Mutagenicity was manifested as a zone of streptomycin-independent mutant
colonies around a filter-paper disk saturated with the mutagenic agent
and resting on the surface of streptomycin-free nutrient agar seeded
heavily with a streptomycin-dependent parental population. Isoamyl
benzoate did not induce mutation from streptomycin dependence to
independence in Escherichia coli (strain Sd-4-73) and hence the chemical
is not likely to classify as a gene mutant in vitro.
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