INVITRO DIFFERENTIAL METABOLISM OF MERBARONE BY XANTHINE-OXIDASE AND MICROSOMAL FLAVOENZYMES - THE ROLE OF REACTIVE OXYGEN SPECIES

Merbarone (MB), a nonsedating derivative of thiobarbituric acid, wa re cently found to induce profound hypouricemia. When incubated with xant hine oxidase (XO) and hypoxanthine in vitro, MB is both an inhibitor o f XO and degraded by the XO-hypoxanthine interaction. Compared with al lopurinol (K(i) = 0.025 muM), MB is a very weak inhibitor of XO (K(i) = 51 +/- 8 muM). MB interacts with XO in the presence of hypoxanthine to yield three chromatographically separate products. One of these pro ducts has been identified by HPLC retention time and spectral characte ristics as 2-oxo-2-desthiomerbarone (2-oxo-MB). The other two products are thought to be S-oxide intermediates in the oxidative desulfuratio n of this drug. Formation of these products was blocked by catalase, s uggesting that the conversion was dependent on reactive oxygen species (especially H2O2) generated by the hypoxanthine-XO system. This sugge stion was confirmed by incubating MB with H2O2. In vitro studies with rat liver microsomes have documented the formation of 2-oxo-MB and 4'- OH-MB (4'-OH-MB), the latter being identified by the characteristic HP LC retention time of its acetylated derivative. The formation of 4'-OH -MB has many characteristics of a cytochrome P-450-dependent monooxyge nase reaction (NADPH requirement and SKF 525-A inhibition); formation of 2-oxo-MB occurs by a different mechanism that is, as yet, uncharact erized. Incubation of kidney microsomes with MB generated 2-oxo-desthi omerbarone but no detectable 4'-OH-MB. These results indicate that MB is a weak inhibitor of XO, and that reactive oxygen species generated by cytochrome P-450 and or by flavin-containing enzymes such as XO pla y a key role in the oxidative desulfuration to 2-oxo-MB. The hydroxyla tion of MB to 4'-OH-MB by liver microsomes is, however, mediated by NA DPH and cytochrome P-450-dependent metabolism only. The pertinence of these in vitro observations to the metabolism, antitumor, nephrotoxici ty, and hypouricemic effects of MB is discussed.