COPPER,ZINC-SUPEROXIDE DISMUTASE AND HYDROGEN-PEROXIDE - A HYDROXYL RADICAL GENERATING-SYSTEM

Because superoxide (O-2(radical-anion)) is a mediator of inflammation, Cu,Zn-superoxide dismutase (Cu,Zn-SOD) has been employed as an anti-i nflammatory compound. However, Cu,Zn-SOD can increase intra- and extra cellular H2O2. This may react with the Cu atom of SOD in a Fenton-type reaction producing the hydroxyl radical((OH)-O-.). With a non-physiol ogical concentration of H2O2 (0.8 mmol/l) to stimulate chemiluminescen ce (CL) at a level <2 mV, it was observed that the addition of Cu,Zn-S OD (100 mu g/ml) yielded an increase of 204.7 +/- 78.2 mV (P < 0.05). This increase in CL depended on the concentrations of H2O2 and Cu,Zn-S OD and was only seen with luminol (reacts with O-2(radical-anion) and (OH)-O-.) but not with lucigenin (reacts with O-2(radical-anion)). No CL was observed when Cu,Zn-SOD was heat inactivated, or when Mn-SOD wa s used. Dissipators of H2O2, copper chelators and (OH)-O-. scavengers attenuated this CL. In electron paramagnetic resonance, with the use o f the spin-trap dimethylpyrroline-N-oxide (DMPO), it was demonstrated that, in the reaction between H2O2 and Cu,Zn-SOD, (OH)-O-. was generat ed. The oxidation of keto-methylthiobutyric acid (KMB) to ethylene, as sessed by gas chromatography, demonstrated that H2O2/Cu,Zn-SOD-generat ed (OH)-O-. can react with KMB and not only with the SOD molecule itse lf. We conclude that H2O2 reduces SOD-bound CU2+ to Cu1+ which, in rea ction with H2O2 catalyses its reduction to (OH)-O-.. Whether this 'pro -inflammatory' reaction occurs in vivo remains to be established.