Detection and identification of oxidants formed during •NO/O2•⁻ reaction: a multi-well plate CW-EPR spectroscopy combined with HPLC analyses. Free Radic Res 2014 Apr;48(4):478-86
Date
01/28/2014Pubmed ID
24460755Pubmed Central ID
PMC4780754DOI
10.3109/10715762.2014.886774Scopus ID
2-s2.0-84895739300 (requires institutional sign-in at Scopus site) 6 CitationsAbstract
New techniques and probes are routinely emerging for detecting short-lived free radicals such as superoxide radical anion (O₂(•-)), nitric oxide ((•)NO), and transient oxidants derived from peroxynitrite (ONOO(-)/ONOOH). Recently, we reported the profiles of oxidation products (2-hydroxyethidium, ethidium, and various dimeric products) of the fluorogenic probe hydroethidine (HE) in the (•)NO/O₂(•-) system (Zielonka et al. 2012). In this study, we used HPLC analyses of HE oxidation products in combination with continuous wave electron paramagnetic resonance (CW-EPR) spin trapping with 5-tert-butoxycarbonyl-5-methyl-1-pyrroline N-oxide (BMPO) to define the identity of the oxidizing species formed in the (•)NO/O₂(•-) system. EPR spin-trapping technique is still considered as the gold standard for characterization of free radicals and their intermediates. We monitored formation of BMPO-superoxide (BMPO-(•)OOH) and BMPO-hydroxyl (BMPO-(•)OH) radical adducts. Simultaneous analyses of results from EPR spin-trapping and HPLC measurements are helpful in the interpretation of the mechanism of formation of products of HE oxidation.
Author List
Koto T, Michalski R, Zielonka J, Joseph J, Kalyanaraman BAuthors
Balaraman Kalyanaraman PhD Professor in the Biophysics department at Medical College of WisconsinJacek M. Zielonka PhD Assistant Professor in the Biophysics department at Medical College of Wisconsin
MESH terms used to index this publication - Major topics in bold
Chromatography, High Pressure LiquidElectron Spin Resonance Spectroscopy
Free Radicals
Humans
Hydroxyl Radical
Nitric Oxide
Oxidants
Oxidation-Reduction
Phenanthridines
Superoxides
