High-performance liquid chromatographic analysis of arachidonic acid metabolites by pre-column derivatization using...

Arachidonic acid (AA) metabolites produced by washed human platelets and rat macrophages were analyzed by high-performance liquid chromatography (HPLC) using a pre-column derivatization method. The reagent, 9-anthryldiazomethane, used in this study and AA metabolites derivatized by the reagent were purified by gel permeation chromatography (PG-pak C column), prior to normal-phase HPLC analysis. A sample containing eleven derivatives (12-, 15- and 5-hydroxyeicosatetetraenoic acid, 12-L-heptadecatrienoic acid, leukotriene B4, prostaglandins B2, D2, E2 and F2 alpha, thromboxane B2 and 6-keto-prostaglandin F1 alpha) was separated on a normal-phase column (PG-pak B); the detection limit is better than 100 pg for all components.