Aflatoxin B 1 activation to a plasmid mutagen by a chemical model of cytochrome P-450

Abstract

Aflatoxin B 1 (AFB 1) was oxidised by a chemical model of cytochrome P-450 and the products obtained analysed by reversed-phase hplc. The oxidation system employs a water-insoluble iron(III)porphyrin catalyst (tetraphenylporphinatoiron(III) chloride; FeTPPCl) and an oxygen donor, iodosylbenzene (PhIO). The two AFB 1 products obtained from this reaction were derived by initial oxidation across the 8–9 double bond of AFB 1 and subsequent breakdown of AFB 1-8,9-epoxide, the compound postulated to be the ultimate carcinogenic and mutagenic derivative of AFB 1. Oxidation of AFB 1 by the porphyrin-catalysed system in the presence of calf-thymus DNA, resulted in the formation of AFB 1-DNA adducts identical to those formed in vivo and in vitro after liver mono-oxygenase activation. The chemical model system was therefore used on a microscale to react AFB 1 with plasmid pHR1800, a plasmid containing the genes for β-lactamase ( amp-r) and galactokinase ( galK). AFB 1-modified plasmid DNA was then transformed into E. coli AB1886 ( uvrA −) and the effects of the AFB 1-DNA adducts on plasmid survival and the ability of pHR1800-transformed bacteria to metabolise galactose, were then studied. The results showed an AFB 1 dose-dependent decrease in plasmid survival and increase in the mutation frequency of the galK gene only when the complete oxidation system was used to generate the reactive AFB 1 metabolite.