Direct EPR detection of the carbonate radical anion produced from peroxynitrite and carbon dioxide

Marcelo G. Bonini, Rafael Radi, Gerardo Ferrer-Sueta, Ana M.Da C. Ferreira, Ohara Augusto*

*Corresponding author for this work

Research output: Contribution to journalArticle

269 Scopus citations

Abstract

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-) reacts rapidly with carbon dioxide, forming an adduct, nitrosoperoxocarboxylate (ONOOCO2/-), whose decomposition has been proposed to produce reactive intermediates such as the carbonate radical (CO3-). 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 6-9. The radical was unambiguously identified by its EPR parameters (g = 2.0113; line width = 5.5 G) and by experiments with bicarbonate labeled with 13C. In this case, the singlet EPR signal obtained with 12C bicarbonate splits into the expected doublet because of 13C (a(13C) = 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 (CO3-). 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.

Original languageEnglish (US)
Pages (from-to)10802-10806
Number of pages5
JournalJournal of Biological Chemistry
Volume274
Issue number16
DOIs
StatePublished - Apr 16 1999
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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