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Bonini, M G; Radi, R; Ferrer-Sueta, G; Ferreira, A M; Augusto, O
Journal of biological chemistry/The Journal of biological chemistry, 04/1999, Volume: 274, Issue: 16Journal Article
The biological effects of peroxynitrite have been recently considered to be largely dependent on its reaction with carbon dioxide, which is present in high concentrations in intra- and extracellular compartments. Peroxynitrite anion (ONOO â ) reacts rapidly with carbon dioxide, forming an adduct, nitrosoperoxocarboxylate (ONOOCO 2 â ), whose decomposition has been proposed to produce reactive intermediates such as the carbonate radical (COÂ·Ì 3 ). Here, by the use of rapid mixing continuous flow electron paramagnetic resonance (EPR), we directly detected the carbonate radical in flow mixtures of peroxynitrite with bicarbonate-carbon dioxide over the pH range of 6â9. The radical was unambiguously identified by its EPR parameters (g = 2.0113; line width = 5.5 G) and by experiments with bicarbonate labeled with 13 C. In this case, the singlet EPR signal obtained with 12 C bicarbonate splits into the expected doublet because of 13 C (a( 13 C)= 11.7 G). The singlet spectrum of the unlabeled radical was invariant between pH 6 and 9, confirming that in this pH range the detected radical is the carbonate radical anion (COÂ·Ì 3 ). Importantly, in addition to contributing to the understanding of nitrosoperoxocarboxylate decomposition pathways, this is the first report unambiguously demonstrating the formation of the carbonate radical anion at physiological pHs by direct EPR spectroscopy.
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