Pulse radiolysis and steady-state analyses of the reaction between hydroethidine and superoxide and other oxidants. Arch Biochem Biophys 2006 Dec 01;456(1):39-47
Date
11/04/2006Pubmed ID
17081495Pubmed Central ID
PMC2270254DOI
10.1016/j.abb.2006.09.031Scopus ID
2-s2.0-33751438500 (requires institutional sign-in at Scopus site) 54 CitationsAbstract
Hydroethidine (HE) is a cell-permeable probe used for the intracellular detection of superoxide. Here, we report the direct measurement of the rate constant between hydroethidine and superoxide radical anion using the pulse radiolysis technique. This reaction rate constant was calculated to be ca. 2 x 10(6) M(-1) s(-1) in water:ethanol (1:1) mixture. The spectral characteristics of the intermediates indicated that the one-electron oxidation product of HE was different from the one-electron reduction product of ethidium (E+). The HPLC-electrochemical measurements of incubation mixtures containing HE and the oxygenated Fenton's reagent (Fe2+/DTPA/H2O2) in the presence of aliphatic alcohols or formate as a superoxide generating system revealed 2-OH-E+ as a major product. Formation of 2-OH-E+ by the Fenton's reagent without additives was shown to be superoxide dismutase-sensitive and we attribute the formation of superoxide radical anion to the one-electron reduction of oxygen by the DTPA-derived radical. Addition of tert-butanol, DMSO, and potassium bromide to the Fenton's system caused inhibition of 2-OH-E+ formation. Results indicate that reducing and oxidizing radicals have differential effects on the formation of 2-OH-E+.
Author List
Zielonka J, Sarna T, Roberts JE, Wishart JF, 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
Hydrogen PeroxideIron
Kinetics
Oxidants
Oxidation-Reduction
Phenanthridines
Pulse Radiolysis
Superoxides