The kinetics of the reaction between NO and O2 as studied by a novel approach

The kinetics of the reaction between NO and O-2 was determined by measuring the time course of the decrease in tion of NO with a quench-flow technique. NO and O-2 were mixed rapidly and reacted for periods of time varying from 10 to 50 s. A second rapid mixing with a solution containing an excess of deoxyhemoglobin and sodium disulfite trapped free NO as nitrosylhemoglobin and reduced O-2. The spectrum of the mixture of deoxy and nitrosylhemoglobin was recorded within 30 s from the second mixing, before any appreciable dissociation of NO from the protein, by means of a flow-cell mounted on-line with the quench-flow apparatus. The amount of NO not consumed in the auto-oxidation reaction was calculated from the proportion of nitrosylhemoglobin in the mixture. As NO and O-2 bind deoxyhemoglobin at comparable rates and NO is oxidized to nitrate by oxyhemoglobin, the ratio of hemoglobin/(NO + O-2) had to be optimized to avoid the interference of this oxidation reaction. The kinetics was first and second order with respect to O-2 and NO, respectively and third order overall with a rate constant k = 4 X k(aq) = 4 X 2.23 (+/-0.26) X 10(6) M-2 s(-1) at 20 degrees C, invariant in the pH range 7-9, in agreement with published values obtained by different methodologies. (C) 1999 Elsevier Science B.V. All rights reserved.