HPLC-MS/MS Method for the Pharmacokinetic Study of Itraconazole a Low Uv Absorbable Drug Involving Wistar Rats

A new LC-MS/MS method was developed for the estimation of itraconazole, a weak UV absorbing drug, in rat plasma using verapamil as the internal standard. Chromatographic separation was achieved on an Agilent Zorbax Eclipse XDB-C18 column (50 mm × 4.6 mm, 1.8µ) as a stationary phase. A mixture of methanol-10 Mm ammonium formate pH 4.0 (90:10, v/v) was used as a mobile phase. Plasma samples were extracted with a mixture of diethyl ether and di-chloromethane (80:20, v/v) followed by subsequent reconstitution in the mobile phase. Treated plasma samples were analyzed by electrospray ionization method in selective reaction monitoring (SRM) mode in tandem mass spectrometry. The separated products were identified by characteristic peaks and specific fragment peaks at m/z 705.3 to 392 and m/z 455 to 149.8 for itraconazole and verapamil, respectively. The present method was found to be linear in the concentration range of 0.5 to 600 ng/mL for itraconazole. Pharmacokinetic study was performed after an oral administration of 30 mg/kg of itraconazole to Wistar rats.