Mechanistic studies of the oxidation of pyridoxalated hemoglobin polyoxyethylene conjugate by nitrogen monoxide

Pyridoxalated hemoglobin polyoxyethylene conjugate (PHP), a modified human- derived hemoglobin, is currently in clinical trials as a nitrogen monoxide scavenger for the treatment of shock. Stopped-flow spectroscopy studies of the reaction between nitrogen monoxide and PHP indicate that at pH 7 the se cond-order rate constant is (88 +/- 3) x 10(6) M-1 s(-1), a value very simi lar to that for the unmodified human hemoglobin. At alkaline pH the reactio n proceeds via the intermediate peroxynitrito complex PHP-(FeOONO)-O-III, w hich rapidly decomposes to nitrate and the iron(III) form of PHP. The rate of decay of PHP-(FeOONO)-O-III increases significantly with decreasing pH s uch that it does not accumulate in concentrations large enough to be observ ed spectroscopically under neutral or acidic conditions. Ion chromatographi c analysis of the nitrogen-containing products of the NO.-mediated reaction of PHP shows that nitrate is formed quantitatively at both pH 7 and pH 9. (C) 1999 Academic Press.