The characteristics of the formation of the superoxide radical anion ([Formula: see text]) and hydrogen peroxide by xanthine oxidases isolated from microorganisms and from cow's milk were investigated. The increase in pH led to an increase in the rate of xanthine oxidation with oxygen by both xanthine oxidases. The functioning of xanthine oxidase from milk along with the two-electron reduction of O2 to H2O2 carries through the one-electron reduction of O2 to [Formula: see text], and the rate and the fraction of generation of [Formula: see text] increased with increasing pH. Under operation of the microbial xanthine oxidase, the [Formula: see text] radical was not detected in the medium. The results suggest a difference in the operation of active centers of enzyme from different sources.