Development of alternative HPLC method for determination of tirofiban in rat serum

Tirofiban hydrochloride is a reversible antagonist of fibrinogen binding to the GPIIb/IIIa receptor, used for the treatment of acute coronary syndrome. A novel RP-HPLC method has been developed and validated for the determination of Tirofiban in serum of Wistar rats with and without deep acute venous thrombosis. The chromatographic separation was carried out using a reverse-phase HPLC column Purospher® RP-18e (150 mm ´ 4.6 mm i.d.; 5 μm) coupled with a guard column LiChrosorb® (4 mm ´ 4 mm i.d.; 7 μm) and mobile phase consisting of the mixture of 1-octane sulfonic acid in water (pH 3.0, adjusted with orthophosphoric acid) and acetonitrile, with a ratio of 60 : 40 (v/v) and a flow rate of 1.0 ml/min, at a wavelength of 277 nm. The serum concentrations of Tirofiban in the group of rats with DVT were lower than those in the control group, and it could be explained with the binding of Tirofiban with the GPIIb/IIIa receptors.