The line asymmetry of electron spin resonance spectra as a tool to determine the cis:trans ratio for spin-trapping adducts of chiral pyrrolines N-oxides: The mechanism of formation of hydroxyl radical adducts of EMPO, DEPMPO, and DIPPMPO in the ischemic-reperfused rat liver

Marcel Culcasi, Antal Rockenbauer, Anne Mercier, Jean Louis Clément, Sylvia Pietri

Research output: Contribution to journalArticle

40 Citations (Scopus)

Abstract

Nonstereospecific addition of free radicals to chiral nitrones yields cis/trans diastereoisomeric nitroxides often displaying different electron spin resonance (ESR) characteristics. Glutathione peroxidase-glutathione (GPx-GSH) reaction was applied to reduce the superoxide adducts (nitrone/{radical dot}OOH) to the corresponding hydroxyl radical (HO{radical dot}) adducts (nitrone/{radical dot}OH) of two nitrones increasingly used in biological spin trapping, namely 5-diethoxyphosphoryl-5-methyl-1-pyrroline N-oxide (DEPMPO) and 5-ethoxycarbonyl-5-methyl-1-pyrroline N-oxide, and of 5-diisopropoxyphosphoryl-5-methyl-1-pyrroline N-oxide (DIPPMPO), a sterically hindered DEPMPO analogue. The method offered improved conditions to record highly resolved ESR spectra and by accurate simulation of line asymmetry we obtained clear evidence for the existence of previously unrecognized isomer pairs of cis- and trans-[DEPMPO/{radical dot}OH] and [DIPPMPO/{radical dot}OH]. Additional nitrone/{radical dot}OH generation methods were used, i.e. photolysis of hydrogen peroxide and the Fenton reaction. We developed a kinetic model involving first- and second-order decay and a secondary conversion of trans to cis isomer to fully account for the strongly configuration-dependent behavior of nitrone/{radical dot}OH. In the reductive system and, to a lower extent, in the Fenton or photolytic systems cis-nitrone/{radical dot}OH was the more stable diastereoisomer. In various biologically relevant milieu, we found that the cis:trans-nitrone/{radical dot}OH ratio determined right after the spin adduct formation significantly differed upon the GPx-GSH vs (Fenton or photolytic) systems of formation. This new mechanistic ESR index consistently showed for all nitrones that nitrone/{radical dot}OH signals detected in the postischemic effluents of ischemic isolated rat livers are the reduction products of primary nitrone/{radical dot}OOH. Thus, ESR deconvolution of cis/trans diastereoisomers is of great interest in the study of HO{radical dot} formation in biological systems.

Original languageEnglish
Pages (from-to)1524-1538
Number of pages15
JournalFree Radical Biology and Medicine
Volume40
Issue number9
DOIs
Publication statusPublished - May 1 2006

Keywords

  • DEPMPO
  • DIPPMPO
  • Diastereoisomers
  • EMPO
  • ESR
  • Free radical
  • Hydroxyl radical spin adducts
  • Kinetics
  • Nitroxides
  • Reperfused liver
  • Spin trapping

ASJC Scopus subject areas

  • Biochemistry
  • Physiology (medical)

Fingerprint Dive into the research topics of 'The line asymmetry of electron spin resonance spectra as a tool to determine the cis:trans ratio for spin-trapping adducts of chiral pyrrolines N-oxides: The mechanism of formation of hydroxyl radical adducts of EMPO, DEPMPO, and DIPPMPO in the ischemic-reperfused rat liver'. Together they form a unique fingerprint.

  • Cite this