The effect of bicarbonate on the oxidation rates and yields of the reaction of peroxynitrite with Ni-II(cyclam), ferrocyanide, and ABTS was studied using the stopped-flow technique. In the absence of bicarbonate, Ni-II(cyclam) is oxidized directly by peroxynitrous acid whereas ferrocyanide and ABTS are oxidized by intermediates that are formed during the self-decomposition of peroxynitrous acid. In the presence of sufficient concentrations of bicarbonate, the rate-determining step of these oxidation processes is the reaction of ONOO- with CO2. The latter reaction results in the formation of an adduct ONOOCO2-, which is incapable of oxidizing directly these substrates. It is shown that about 33% of this adduct decomposes into highly oxidizing species, most probably into (NO2)-N-. and CO3.- via the homolysis of the peroxo O-O bond. Under physiological conditions, where the most abundant species is ONOO- and the concentration of CO2 exceeds 1 mM, the half-life of peroxynitrite is reduced, and consequently its diffusion distance is shortened. Therefore, indirect oxidation by peroxynitrite through intermediates that are formed during its self-decomposition will not take place in vivo, whereas direct oxidation by peroxynitrite may take place provided that the rate of this reaction competes efficiently with that of peroxynitrite with CO2. We conclude that the toxicity of peroxynitrite in vivo is most probably governed by ONOOCO2-, which has a reactivity different from that of peroxynitrite.