On the irreversible destruction of reduced nicotinamide nucleotides by hypohalous acids

Degradation of the reduced pyridine nucleotides NMNH and NADH by HOCl invol ves two distinct stages: a fast reaction, k = 4.2 x 10(5) M-1 s(-1), leads to generation of stable pyridine products (Py/Cl) with a strong absorption band at 275 nm (epsilon = 12.4 x 10(3) M(-)1 cm(-1) in the case of NMNH); s econdarily, a subsequent reaction of HOCl, k = 3.9 x 10(3) M-1 s(-1), leads to a complete loss of the aromatic absorption band of the pyridine ring. H OBr and HOI(I-2) react similarly. Apparent rate constants of the primary re actions of HOX species with NMNH at pH 7.2 increase in the order HOCl (3 x 10(5) M-1 s(-1)) < HOBr(similar to 4 x 10(6) M-1 s(-1)) < HOI(I-2)(similar to 6.5 x 10(7) M-1 s(-1)). HOBr reacts fast also with the primary product P y/Br, k similar to 9 x 10(5) M-1 s(-1), while the reactions of HOI and I-2 with Py/I are slower, similar to 1.4 x 10(3) M-1 s(-1) and >6 x 10(3) M-1 s (-1), respectively. Halogenation of the amide group of NMN+ by HOX species is many orders of magnitude slower than oxidation of NMNH. Taurine inhibits HOCl-induced oxidation of NADH, but HOBr-induced oxidation is not inhibite d because the taurine monobromamine rapidly oxidizes NADH, and oxidation by HOI(I-2) is not inhibited because taurine is inert toward HOI(I-2), Also s ulfur compounds (GSH, GSSG, and methionine) are less efficient in protectin g NADH against oxidation by HOBr and HOI(I-2) than against oxidation by HOC l, The results suggest that reactions of HOBr and HOI(I-2) in a cellular en vironment are much more selectively directed toward irreversible oxidation of NADH than reactions of HOCl, It is noteworthy that the rather inert N-ch loramines react with iodide to generate HOI(I-2), i.e., the most reactive a nd selective oxidant of reduced pyridine nucleotides, NMR investigations sh ow that the primary stable products of the reaction between NMNH and HOCl a re various isomeric chlorohydrins originating from a nonstereospecific elec trophilic addition of HOCl to the C5=C6 double bond of the pyridine ring. T he primary products (Py/X) of NMNH all exhibit similar absorption bands aro und 275 nm and are hence likely to result from analogous addition of HOX to the C5=C6 bond of the pyridine ring. Since the Py/X species are stable and inert toward endogeneous reductants like ascorbate and GSH, they may gener ally be useful markers for assessing the contribution of hypohalous acids t o inflammatory injury. (C) 2000 Academic Press.