Production of OH-radical-type oxidant by lucigenin

In the presence of NADH- reductases (dihydrolipoamide: NAD oxidoreductase E. C.1.8.1.4 from pig heart or from Clostridium kluyveri; frequently also addressed as "diaphorases") and NADH lucigenin strongly increases ethylene production from a-keto-methylthiobutyrate (KMB) as an indicator for strong oxidants of the OH-radical type. These reactions are further stimulated in the presence of Fe3+ ions. With these NADH-"diaphorases", the structurally similar poison, paraquat, in the absence or presence of Fe3+ has no effect. With ferredoxin-NADP reductase (E. C.1.18.1.2.), however, paraquat reacts quasi identical to lucigenin. Superoxide dismutase, catalase, free radical- or OH-scavengers such as mannitol, propylgallate, DABCO, and desferal inhibit the reaction whereas EDTA (in the presence or absence of added Fe3+) is stimulatory. From these data we conclude that the superoxide--indicator LUC is redox-active after unspecific coupling to several almost ubiquitory NAD(P)H- reductases catalyzing monovalent oxygen reduction. Lucigenin thus should no longer be used as a "specific" superoxide indicator. This report is in agreement with very recent results by Liochev and Fridovich (Arch. Biochem. Biophys. 337, 115 [1997]) and Vasquez-Vivar et al. (FEBS Lett. 403, 127 (1997).