Analysis of effector cell-derived lyso platelet activating factor by electron capture negative ion mass spectrometry.

摘要: Quantification of 1-O-alkyl-2-lyso-sn-3-glycero-phosphocholine (lysoPAF) and determination the different molecular species released by cells has been hampered molecules's lack intrinsic bioactivity, unavailability a suitable internal standard, reliance on derivatives requiring electron impact techniques. We have synthesized trideuterated standards (labeled terminal carbon alkyl chain) for both C16:0 C18:0 lysoPAF. Using these standards, we isolated quantified lysoPAF from A23187-stimulated human neutrophils rat alveolar macrophages. Extracted was purified solid-phase extraction thin-layer chromatography. The polar phosphorylcholine group removed with 29 M HF or phospholipase C. two free hydroxyl groups were derivatized pentafluorobenzoyl chloride. resultant bis-pentafluorobenzoyl derivative, analyzed gas chromatography/electron capture negative ion mass spectrometry, underwent substantial fragmentation. Lowering source temperature resulted in dramatic increase signal-to-noise ratio, vast majority current carried anion. Stimulated 16.3 10.2 ng/10(6) lysoPAF, respectively. Rat macrophages 15.9 but variably detected at low levels. conclude that use bispentafluorobenzoyl ester derivative allows facile quantification this autacoid metabolite biological matrices.