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

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.