Validated LC–MS/MS method for the determination of amlodipine enantiomers in rat plasma and its application to a stereoselective pharmacokinetic study

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Abstract

A rapid and sensitive method was established to determine amlodipine enantiomers using liquid chromatography-tandem mass spectrometry (LC–MS/MS). Stereoselective separation was performed on CHIRALCEL OZ-RH column (150mm×4.6mm i.d., 5μm) with acetonitrile-water (10mM ammonium acetate, 0.5% ammonia solution) (95:5, v/v) at a flow rate of 0.5mL/min. The substances were detected by mass spectrometer equipped with an electrospray ionization source interface in positive ion mode. Multiple reaction monitoring was selected with the transition of the m/z 409.1→238.0 for amlodipine enantiomers and m/z 237.0→194.1 for carbamazepine (IS) respectively. Calibration curves were linear at the range of 0.9375–120ng/mL for both isomers with r>0.99, while using a lower sample volume (50μL) compared with previously reported enantiospecific methods The accuracy was at the range of 84.1–119.0% for R-amlodipine, and 87.4–118.2% for S-amlodipine, respectively. The within- and between-run precision (CV%) was within 10% in all cases for both enantiomers. Enantiomers were stable under different conditions, e.g. processed sample, short-term, residue, long-term and freeze/thaw. The LC–MS/MS method was successfully applied in pharmacokinetic study of amlodipine enantiomers in rats. It was observed the concentration of the S- amlodipine was significantly higher than that of the R-amlodipine in racemate-treated group. And there was no significant difference in the pharmacokinetic profiles of the S-amlodipine between the 10mg/kg racemate- and 5mg/kg S-amlodipine-treated groups. In addition, it was the first time to find that the main pharmacokinetic parameters (AUC(0-t), AUC(0-∞) and Cmax) of R-amlodipine were significantly lower in the 5mg/kg R-amlodipine-treated group compared with the racemate-treated group.

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