Lack of influence of modulators of epoxide metabolism on the genotoxicity of tans-anethole in freshly isolated rat hepatocytes assessed with the unscheduled DNA synthesis assay

Abstract

The aniseed food flavour trans-anethole) was implicated as a weak hepatocarcinogen only in female Sprague Dawley-CD rats administered high doses (1% in the diet for 121 wk). However, this substance is apparently non-genotoxic in a range of test systems. Anethole is metabolized in the rat along three primary pathways, one of which is epoxidation across the double bond of the side-chain. The epoxides of a number of the alkenylbenzene family of food flavours, of which anethole is a member, are putative genotoxins, being bacterial mutagens but not mammalian carcinogens. The authors have previously shown that the cytotoxicity of anethole is enhanced when the cellular epoxide defence mechanisms of conjugation with reduced glutathione and hydration by cytosolic epoxide hydrolase are severely compromised. They now report, however, that modulation of epoxide metabolism in cultured cells by the same mechanisms fails to induce unscheduled DNA synthesis (UDS) by anethole not was there a UDS response in hepatocytes of female rats dosed with anethole in vivo. The epoxide of anethole was synthesized for the first time in this investigation and tested directly. As expected, it was markedly cytotoxic but not genotoxic. Anethole epoxide has chemical characteristics that differ from those of other structurally similar epoxides being labile to hydrolysis in aqueous media at physiological pH and temperature. This gives greater relevance to tests of its genotoxicity after formation within the hepatocyte rather than by adding the epoxide extracellularly to the culture medium. The direct and indirect demonstration of the lack of induction of UDS by anethole epoxide provides further support for the hypothesis that marginal hepatocarcinogenicity observed in female rats given 1% anethole in the diet for 121 wk was not initiated by a genotoxic event.