Spin adduct formation from lipophilic EMPO-derived spin traps with various oxygen- and carbon-centered radicals

Abstract

Free radicals are involved in the onset of many diseases, therefore the availability of adequate spin traps is crucial to the identification and localization of free radical formation in biological systems. In recent studies several hydrophilic compounds of 2-ethoxycarbonyl-2-methyl-pyrroline- N-oxide (EMPO) have been found to form rather stable superoxide spin adducts with half-lives up to twenty minutes at physiological pH. This is a major improvement over DMPO ( t 1/2 = ca. 45 s), and even over DEPMPO ( t 1/2 = ca. 14 min), the best commercially available spin trap for the unambiguous detection of superoxide radicals. In order to allow the detection of superoxide and also other radicals in lipid environment a series of more lipophilic derivatives of EMPO was synthesized and their structure unambiguously characterized by 1H and 13C NMR spectroscopy. In this way, six different compounds with a n-butyl group in position 5 and either an ethoxy- (EBPO), propoxy- (PBPO), iso-propoxy- ( iPBPO), butoxy- (BBPO), sec-butoxy- ( sBBPO) or tert-butoxycarbonyl group ( tBBPO) in position 5 of the pyrroline ring were obtained and fully analytically characterized (NMR, IR). The stability of the superoxide adducts of all investigated spin traps were comparable with EMPO ( t 1/2 = ca. 8 min), except for the two compounds bearing an additional methyl group in position 3 or 4 of the pyrroline ring, 5-butyl-5-ethoxycarbonyl-3-methyl-pyrroline- N-oxide (BEMPO-3) and 5-butyl-5-ethoxycarbonyl-4-methyl-pyrroline- N-oxide (BEMPO-4), of which the superoxide adducts were stable for more than 30 min. Spin adducts of other carbon- and oxygen-centered radicals were also investigated.