Mechanism of H2S Formation from the Metabolism of Anetholedithiolethione and Anetholedithiolone by Rat Liver Microsomes

The drug anetholedithiolethione (ADT) and its analogs have been extensively used as H2S donors. However, the mechanism of H2S formation from ADT under biologic conditions remains almost completely unknown. This article shows that only small amounts of H2S are formed during incubation of ADT and of its metabolite anetholedithiolone (ADO) with rat liver cytosol or with rat liver microsomes (RLM) in the absence of NADPH, indicating that H2S formation under these conditions is of hydrolytic origin only to a minor extent. By contrast, much greater amounts of H2S are formed upon incubation of ADT and ADO with RLM in the presence of NADPH and dioxygen, with a concomitant formation of H2S and para-methoxy-acetophenone (pMA). Moreover, H2S and pMA formation under those conditions are greatly inhibited in the presence of N-benzyl-imidazole indicating the involvement of cytochrome P450-dependent monooxygenases. Mechanistic studies show the intermediate formation of the ADT-derived 1,2-dithiolium cation and of the ADO sulfoxide during microsomal metabolism of ADT and ADO, respectively. This article proposes the first detailed mechanisms for the formation of H2S from microsomal metabolism of ADT and ADO in agreement with those data and with previously published data on the metabolism of compounds involving a C=S bond. Finally, this article shows for the first time that ADO is a better H2S donor than ADT under those conditions. SIGNIFICANCE STATEMENT: Incubation of anetholedithiolethione (ADT) or its metabolite anetholedithiolone (ADO) in the presence of rat liver microsomes, NADPH, and O2 leads to H2S. This article shows for the first time that this H2S formation involves several steps catalyzed by microsomal monooxygenases and that ADO is a better H2S donor than ADT. We propose the first detailed mechanisms for the formation of H2S from the microsomal metabolism of ADT and ADO.