High-performance liquid chromatographic method for the determination of bamifylline and its three metabolites in...

The performance of a gradient elution normal-phase system and of two isocratic elution reversed-phase ion-pair systems was investigated for the high-performance liquid chromatographic separation of bamifylline and its three metabolites. Three packings and mobile phases were tested and the best separation was achieved using a reversed-phase ion-pair system with Hypersil ODS 3 micron as stationary phase. The effects of alkyl chain length and concentration of alkyl sulfonates, salt concentration, column temperature and competing amine on separation and peak tailing are discussed. The determination of bamifylline and metabolites in human plasma using the defined optimal chromatographic conditions is reported.