Quantification of superoxide radicals and peroxynitrite in vascular cells using oxidation of sterically hindered hydroxylamines and electron spin resonance

Nitric Oxide : Biology and Chemistry
Sergey DikalovE Bassenge

Abstract

The reactions of two hydroxylamines, 1-hydroxy-3-carboxy-pyrrolidine (CP-H) and 1-hydroxy-2,2,6,6-tetramethyl-4-oxo-piperidine (TEMPONE-H), with superoxide radicals and peroxynitrite were studied. In these reactions corresponding stable nitroxyl radicals 3-carboxy-proxyl (CP) and 1-hydroxy-2,2,6,6-tetramethyl-4-oxopiperidinoxyl (TEMPONE) are formed and the amount of them can be quantified by electron spin resonance (ESR). It was found that CP-H and TEMPONE-H provide almost the same efficacy in assaying peroxynitrite by ESR in vitro at pH 7.4. The formation of superoxide radicals in suspensions of cells was discriminated from that of peroxynitrite using superoxide dismutase or dimethyl sulfoxide as competitive reagents. The stability of the radicals CP and TEMPONE in the presence of ascorbate or thiols was studied in vitro. The reduction rate of CP by ascorbate was 66-fold slower than the rate of reduction of TEMPONE. Therefore, the quantification of the formation of superoxide radicals and of peroxynitrite is much less affected by ascorbic acid when CP-H, but not TEMPONE-H, is used. Both TEMPONE-H and CP-H were used to determine the formation rates of superoxide radicals and peroxynitrite in suspensions of cultured aortic smoot...Continue Reading

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Citations

Jul 11, 2012·Chemical Research in Toxicology·Kevin M NashFrederick A Villamena
Feb 14, 2007·Hypertension·Sergey DikalovDavid G Harrison
Aug 2, 2005·Respiratory Research·Norbert WeissmannFriedrich Grimminger
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Sep 18, 2012·Antioxidants & Redox Signaling·Sergey I Dikalov, David G Harrison
Jul 31, 2013·Journal of Agricultural and Food Chemistry·Yu-Ting ZhouJun-Jie Yin
Sep 1, 2007·Cellular Physiology and Biochemistry : International Journal of Experimental Cellular Physiology, Biochemistry, and Pharmacology·Paul K WittingRoland Stocker
Dec 7, 2010·Free Radical Research·Sergey I DikalovIgor A Grigor'ev
Sep 15, 2010·Free Radical Biology & Medicine·Catherine A ShawAdriano G Rossi
May 6, 2009·Clinical and Experimental Pharmacology & Physiology·Darren C HenstridgeBronwyn A Kingwell
Feb 18, 2006·Clinica Chimica Acta; International Journal of Clinical Chemistry·Grzegorz Bartosz
Apr 10, 2013·Biochimica Et Biophysica Acta·Clare L Hawkins, Michael J Davies
Sep 12, 2015·Wiley Interdisciplinary Reviews. Nanomedicine and Nanobiotechnology·Jie Kai TeeDavid Tai Leong
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Jun 22, 2021·Antioxidants & Redox Signaling·Ken-Ichiro MatsumotoIchio Aoki

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