Servillo L, Iorio E L, Quagliuolo L, Camussi G, Balestrieri C, Giovane A
Dipartimento di Biochimica e Biofisica, II Università degli Studi di Napoli, Italy.
J Chromatogr B Biomed Sci Appl. 1997 Feb 21;689(2):281-6. doi: 10.1016/s0378-4347(96)00319-2.
A high-performance liquid chromatography (HPLC) procedure for the separation of choline lysophospholipids including 1-acyl-lysophosphatidylcholines and 1-O-alkyl-lysophosphatidyl- cholines, like the lysoform of the platelet activating factor (2-lysoPAF), is described. The lysophospholipids are derivatized at the sn-2 position of the hydroxyl group by 7-diethylaminocoumarin-3-carbonylazide, which converts them into the corresponding carbamoyl derivatives. The derivatized compounds were well separated by reversed-phase HPLC and quantified by fluorimetric detection. This method shows a high sensitivity and allows the separation and quantification of mixtures of lysophospholipids at picomolar level. The method was applied to assay enzyme activities, like phospholipase A2 and PAF-acetylhydrolase, on single phospholipids or their mixtures.
本文描述了一种高效液相色谱（HPLC）方法，用于分离胆碱溶血磷脂，包括1-酰基溶血磷脂酰胆碱和1-O-烷基溶血磷脂酰胆碱，如血小板活化因子的溶血形式（2-溶血PAF）。溶血磷脂在羟基的sn-2位通过7-二乙氨基香豆素-3-羰基叠氮化物进行衍生化，将其转化为相应的氨基甲酰基衍生物。衍生化后的化合物通过反相HPLC得到良好分离，并通过荧光检测进行定量。该方法具有高灵敏度，能够在皮摩尔水平上分离和定量溶血磷脂混合物。该方法被应用于测定单一磷脂或其混合物上的酶活性，如磷脂酶A2和PAF-乙酰水解酶。
