A kinetic study of the reaction of NO with O2 in aqueous solution has been carried out using a colorimetric method based on a pH indicator and a stopped-flow spectrophotometer. The reaction is second order in NO concentration and first order in O2 concentration; the overall third order rate constant is 6.3 (+/- 0.4) x 10(6) (M-1)2 s-1. Model calculations for the reaction at estimated physiological concentrations of NO and O2 indicate that the simple autoxidation of NO will not limit its diffusion from the site of production in endothelial cells to a spatially removed target molecule such as guanylate cyclase in myocytes and platelets.