Neutrophil-mediated phospholipid peroxidation assessed by gas chromatography-mass spectroscopy

Disease pathophysiology frequently involves manifestations of the systemic inflammatory response syndrome. Oxyradicals represent key inflammatory mediators, and neutrophils are one important source of oxyradicals. This investigation examined neutrophil-mediated peroxidation of dilinoleoyl phosphatidyl-choline (DLPC) liposomes by monitoring the appearance of monohydroxyl linoleic acid with the use of gas chromatography-mass spectroscopy (GC-MS), compared with traditional assessment of thiobarbituric acid-reactive species (TBARS) and phosphatidylcholine-specific conjugated dienes. DLPC was peroxidized in a system using activated neutrophils in balanced salt solution containing chelated iron. 9-Monohydroxyl linoleic acid and 13-monohydroxyl linoleic acid were readily identified in neutrophil-mediated peroxidized DLPC with the use of GC-MS. Neutrophil NADPH oxidoreductase specific activity correlated highly with total ion current or specific ion monitoring of integrated peak areas for peroxidized linoleic acid but correlated poorly with DLPC-derived TBARS or conjugated dienes. These results ascertain that activated neutrophils mediate phosphatidylcholine lipid peroxidation to specific products, which may be precisely monitored with the use of GC-MS. The extent of this peroxidation is highly correlated with the magnitude of the neutrophil respiratory burst.