RESUMEN
General procedures for the synthesis and purification of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) and its synthetic intermediates for electron paramagnetic resonance (EPR) spin trapping is described. The synthetic methods are not new per se and are based on Todd's original scheme (1959). In contrast to his account, however, we provide a detailed description of the numerous procedures and precautions necessary to obtain DMPO of very high 'chemical' and (most importantly) 'EPR' purity for use in biological spin trapping.
Asunto(s)
Óxidos N-Cíclicos/síntesis química , Marcadores de Spin/síntesis química , Óxidos N-Cíclicos/aislamiento & purificación , Espectroscopía de Resonancia por Spin del ElectrónRESUMEN
The spin trap 5,5-dimethyl-1-pyrroline-N-oxide (DMPO), is commonly used for the detection of short-lived free radicals produced by neutrophils during their respiratory burst. The spin adducts of both the superoxide ion (O2-) and the hydroxyl radical (.OH) are detectable during this process. Since myeloperoxidase (MPO), which is also active during the respiratory burst, produces hypochlorous acid (HOCl) (HOCl) in the presence of chloride ions (Cl-) and hydrogen peroxide (H2O2), this species has been investigated as a possible source of the DMPO-OH adduct. At concentrations of hypochlorous acid between 0.1 and 0.7 mumol/ml the DMPO-OH spin adduct is detected using electron spin resonance (ESR) techniques. Two possible mechanisms for the formation of this adduct are proposed. These findings suggest that the product of MPO, namely hypochlorous acid, is a possible source of the hydroxyl spin adduct detected during the respiratory burst.