ROLE OF PHASE-II ENZYMES IN THE BIOACTIVATION OF 1,4-DICHLOROBENZENE AND 1,4-DIBROMOBENZENE

The use of sodium phenobarbital (PB, CYP2B1 inducer) combined with bet a-naphthoflavone (beta-NF, 1A1) to induce certain Phase I reactions in S9 liver fractions is a standard method for conducting short-term bio assays for genotoxicity. However, because post-oxidative enzymes are a lso able to activate many precarcinogens, we tested the possibility of adapting S9 liver fractions derived from Phase II-induced rodents to the field of genetic toxicology. Tn this study, S9 liver fractions der ived from Swiss albino CD1 mice fed 7.5g/kg 2-(3)-tert-butyl-4-hydroxy anisole (BHA; a monofunctional Phase II-inducer) for 3 weeks, show a c lear pattern of induction with an approximately 3.5-9.5-fold increase in glutathione S-transferase activity. In vitro DNA binding of the pro mutagenic agents, [C-14]-1,4-dichlorobenzene (DCB) and [C-14]-1,4-dibr omobenzene (DBB), is mediated by such metabolic liver preparations and showed a significant increase in covalent binding capability. In some instances, enzyme activity was more elevated when compared to that ob tained with traditional(Phase I induced) S9. Together with DNA binding , the genetic response of these chemicals in the diploid D-7 strain of Saccharomyces cerevisiae used as a biological test system, revealed t he ability of the BHA-derived preparations to activate the promutageni c agents, as exemplified by the significant enhancement of mitotic gen e-conversion (up to 5.2-fold for DCB and 3.4-fold for DBB) and reverse point mutation (up to 3.6-fold for DCB and 2.5-fold for DBB) at a 4mM concentration. This novel metabolising biosystem, with enhanced Phase II activity, is recommended together with a traditional S9, for detec ting unknown promutagens in genotoxicity studies. The routine use of e ither oxidative or post-oxidative S9 increases the responsiveness of t he test and can contribute to the identification of promutagens not de tected when using traditional protocols.