Formation and Properties of Peroxynitrite as Studied by Laser Flash Photolysis, High-Pressure Stopped-Flow Technique, and Pulse Radiolysis

Abstract

Flash photolysis of alkaline peroxynitrite solutions results in the formation of nitrogen monoxide and superoxide. From the rate of recombination it is concluded that the rate constant of the reaction of nitrogen monoxide with superoxide is (l.9 ± 0.2) × 10¹⁰ M^-1 s^-1. The p<i>K</i>_a of hydrogen oxoperoxonitrate is dependent on the medium. With the stopped-flow technique a value of 6.5 is found at millimolar phosphate concentrations, while at 0.5 M phosphate the value is 7.5. The kinetics of decay do not follow first-order kinetics when the pH is larger than the p<i>K</i>_a, combined with a total peroxynitrite and peroxynitrous acid concentration that exceeds 0.1 mM. An adduct between ONOO- and ONOOH is formed with a stability constant of (1.0 ± 0.l) × 10⁴ M. The kinetics of the decay of hydrogen oxoperoxonitrate are not very pressure-dependent: from stopped-flow experiments up to 152 MPa, an activation volume of 1.7 ± 1.0 cm³ mol^-1 was calculated. This small value is not compatible with homolysis of the O-O bond to yield free nitrogen dioxide and the hydroxyl radical. Pulse radiolysis of alkaline peroxynitrite solutions indicates that the hydroxyl radical reacts with ONOO- to form [(HO)ONOO]•- with a rate constant of 5.8 × l0⁹ M^-1 s^-1. This radical absorbs with a maximum at 420 nm (ε = 1.8 × 10³ M^-1 cm^-1) and decays by second-order kinetics, <i>k</i> = 3.4 × l0⁶ M^-1 s^-1. Improvements to the biomimetic synthesis of peroxynitrite with solid potassium superoxide and gaseous nitrogen monoxide result in higher peroxynitrite to nitrite yields than in most other syntheses.