DEVELOPMENT OF A HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC MASS-SPECTROMETRIC TECHNIQUE, WITH AN IONSPRAY INTERFACE, FOR THE DETERMINATION OFPLATELET-ACTIVATING-FACTOR (PAF) AND LYSO-PAF IN BIOLOGICAL SAMPLES

An HPLC-mass spectrometric technique with an ionspray interface was de veloped for the determination of platelet-activating factor (PAF) and PAF-related compounds in biological samples. HPLC separations were per formed using a reversed-phase column. The mass spectra showed intense [M + H]+ ions. Collision-induced dissociation of protonated molecular ions gave characteristic daughter ions corresponding to the phosphoryl choline group. By selective-ion monitoring, a detection limit of 0.3 n g was obtained for all molecules; by multiple reaction monitoring, the same sensitivity was achieved for PAF whereas for lyso-PAF the limit was 3 ng. Finally, PAF was comparatively determined by bioassay and HP LC-MS after extraction from the cell pellets and the supernatants of h uman polymorphonuclear neutrophils unstimulated or stimulated with ops onized zymosan. The good correlation observed between these techniques indicated the reliability of HPLC-MS for biochemical studies on PAF a nd PAF-related molecules.