Use of 3,3,5,5-tetramethyl-1-pyrroline-1-oxide spin trap for the continuous flow ESR monitoring of hydroxyl radical generation in the ischemic and reperfused myocardium

Abstract

A new approach for in vivo spin trapping and quantitation of oxygen-derived free radicals has been developed using a continuous flow high speed ESR detection system. Spin adducts of OH were detected as 1:1:1:1:1:1 sextets (a N=15.2 G, a H=16.8 G, g=2.0055) in the isolated rat heart when perfused with 3,3,5,5-tetramethyl-1-pyrroline-1-oxide (40 mM) during a 10-min control pretreatment (14 ml/min) followed by 50 min of low-flow ischemia (1 ml/min), 30 min of global ischemia and subsequent reperfusion at 14 ml/min. The ESR signals appeared within 15–20 min of low-flow ischemia and grew moderately during the remaining 30 min at a rate of 2–6 nmoles of spin adduct released per minute. Post-ischemic reperfusion was characterized by a burst of spin adduct formation at 30 s−1 min, corresponding to 51.8 nmoles of spin adduct released between 30 s and 1 min.