MODULATORY ROLE OF NITRIC-OXIDE ON SUPEROXIDE-DEPENDENT LUMINOL CHEMILUMINESCENCE

Reactive oxygen species are involved in luminol chemiexcitation induce d in biological systems, but the contribution of nitrogen-derived oxid ants in the process still remains unclear. Herein, we report that lumi nol chemiluminescence (LCL) induced by a superoxide (O-2(.-))- and hyd rogen peroxide (H2O2)-generating system (2-25 mU/ml xanthine oxidase p lus acetaldehyde and oxygen) was markedly inhibited by nitric oxide (( NO)-N-.) added either as bolus (0-10 mu M) Or a continuous flow (0-10 mu M/min). However, the inhibition of LCL was followed by an overshoot in light emission after most (NO)-N-. was consumed or the infusion st opped and was due to reactions of remaining peroxynitrite, the product of the reaction between O-2(.-) and (NO)-N-., with luminol. Nitric ox ide also inhibited peroxynitrite- and glucose oxidase-induced LCL, but no overshoot was observed. On the other hand, a continuous flux of pu re peroxynitrite, at 2 to 10 mu M/min, induced LCL with quantum yields close to those obtained by identical micromolar fluxes of O-2(.-), wh ile peroxynitrite formed from the decomposition of the sydnonimine SIN -1 yielded 76% of the chemiluminescence obtained with authentic peroxy nitrite. Peroxynitrite-induced LCL was 80 and 55% inhibitable by SOD a nd catalase, respectively, showing that there were O-2(.-) and H2O2-de pendent routes of chemiexcitation. The hydroxyl radical scavengers dim ethyl sulfoxide, mannitol, and ethanol and the metal chelator diethyle netriaminepentaacetic acid did not inhibit peroxynitrite-induced LCL w hile desferrioxamine was an efficient inhibitor of Light emission by r eaction with an activated state of peroxynitrous acid which is respons ible of performing the initial one-electron oxidation of luminol, Our results are consistent with a dual role of (NO)-N-. in O-2(.-)-induced LCL: (I) formation of peroxynitrite which in turn promotes the cal in termediates directing the system toward a dark pathway. These consider ations are of critical importance when analyzing cell- and tissue-deri ved LCL in (NO)-N-.-, O-2(.-), and peroxynitrite-producing systems. (C ) 1996 Academic Press, Inc.