DETECTION OF SUPEROXIDE RADICALS AND PEROXYNITRITE BY DROXY-4-PHOSPHONOOXY-2,2,6,6-TETRAMETHYLPIPERIDINE - QUANTIFICATION OF EXTRACELLULAR-SUPEROXIDE RADICALS FORMATION

The reactions of the new sterically hindered hydroxylamine droxy-4-pho sphonooxy-2,2,6,6-tetramethylpiperidine (PP-H) with superoxide radical and peroxynitrite have been studied. These reactions produce the nitr oxide 4-phosphonooxy-2,2,6,6-tetramethyl-piperidinyloxy. The rate cons tant for reaction of superoxide with PP-H is determined as (8.4+/-0.6) . 10(2) M-1 s(-1). It was found that PP-H provides almost the same sp in trapping efficacy as 1-hydroxy-3-carboxy-pyrrolidine (CP-H). The ba ckground oxidation of PP-H in blood is much less than for CP-H. The ex tremely slow PP-H penetration into the cells makes possible the study of extracellular formation of superoxide radical. The acute treatment of blood with nitroglycerin is shown to induce an extracellular supero xide radical formation. PP-H is more sensitive for detection of reacti ve oxygen species as compared with CP-H. PP-H is an effective scavenge r of superoxide radical and of peroxynitrite, and can be used to quant ify the extracellular formation of these reactive oxygen species. (C) 1998 Academic Press.