ELECTRONIC SPIN-RESONANCE DETECTION OF SUPEROXIDE AND HYDROXYL RADICALS DURING THE REDUCTIVE METABOLISM OF DRUGS BY RAT-BRAIN PREPARATIONS AND ISOLATED CEREBRAL MICROVESSELS

A spin trapping technique was used to analyze by electron spin resonan ce (ESR) the formation of oxygen-derived free radicals during the cere bral reductive metabolism of xenobiotics able to undergo a single elec tron reduction, i.e. quinones, pyridinium compounds and nitroheterocyc lics. Paraquat, menadione and nitrofurazone were used as model compoun ds of these three classes of molecules. ESR spectra indicative of supe roxide and hydroxyl radical formation were obtained by incubation of b rain homogenates directly within the ESR cavity at 37 degrees C for ea ch of the three molecules tested. These signals were dependent on nucl eotide cofactors, and increased in a time-dependent manner. The NADPM and NADH dependent free radical production was further characterized i n brain microsomal and mitochondrial fractions, respectively. By using various combinations of reactive species inactivating enzymes (supero xide dismutase, catalase), a metal chelator (deferoxamine), and an hyd roxyl trapping agent (dimethylsulfoxide), it was shown that (1) the pr imary radical generated was the superoxide anion; and (2) a significan t production of Ihe hydroxyl radical also occurred, that was secondary to the superoxide anion production. Consistent signals indicative of the production of both oxygen-derived free radicals were obtained when isolated cerebral microvessels which constitute the blood-brain barri er were incubated with the model molecules. This is of particular toxi cological relevance, because this barrier represents a key element in the protection of the brain, and is in close contact with blood-born e xogenous molecules. (C) 1998 Elsevier Science Inc.