The primary product of the interaction between nitric oxide (NO) and superoxide (O2/-) is peroxynitrite (ONOO-), which is capable of either oxidizing or nitrating various biological substrates. However, it has been shown that excess NO or O2/- can further react with ONOO- to form species which mediate nitrosation. Subsequently, the controlled equilibrium between nitrosative and oxidative chemistry is critically dependent on the flux of NO and O2/-. Since ONOO- reacts not only with NO and O2/- but also with CO2, the effects of bicarbonate (HCO3/-) on the biphasic oxidation profile of dihydrorhodamine-123 (DHR) and on the nitrosation of both 2,3- diaminonaphthalene and reduced glutathione were examined. Nitric oxide and O2/- were formed with DEA/NO [NaEt2NN(O)NO] and xanthine oxidase, respectively. The presence of HCO3/- did not alter either the oxidation profile of DHR with varying radical concentrations or the affinity of DHR for the oxidative species. This suggests that the presence of CO2 does not affect the scavenging of ONOOby either NO or O2/-. However, an increase in the rate of DHR oxidation by ONOO- in the presence of HCO3/- suggests that a CO2-ONOO- adduct does play a role in the interaction of NO or O2/- with a product derived from ONOO-. Further examination of the chemistry revealed that the intermediate that reacts with NO is neither ONOO- nor cis-HOONO. It was concluded that NO reacts with both trans-HOONO and a CO2 adduct of ONOO- to form nitrosating species which have similar oxidation chemistry and reactivity with O2/- and NO.