BIOANALYTICAL METHOD DEVELOPMENT AND VALIDATION OF SITAGLIPTIN PHOSPHATE BY RP-HPLC AND ITS APPLICATION TO PHARMACOKINETIC STUDY

Abstract

A simple, rapid and selective HPLC method was developed for the estimation of Sitaliptin phosphate (SP) in human plasma by protein precipitation technique. The chromatographic separation was carried out on a reverse phase Phenomenex C 18 (250 × 4.6mm, 5µ) column. A mixture of 0.5% v/v of Triethylamine solution and acetonitrile (77:23 v/v) was used as mobile phase. The pH of 0.5% v/v Triethylamine was adjusted to 6.8 using orthophosphoric acid. The flow rate of mobile phase was set at 1.0 ml/min and the detection of SP was carried out at 267 nm by an UV detector. The retention time of Sitagliptin phosphate and internal standard was 6.1 and 7.7 min, respectively. The method was validated and found to be linear in the range of 10-1000ng/ml. the limit of detection (LOD) and limit of quantification (LOQ) were 1 ng/ml and 10 ng/ml. The results indicate the bio analytical method is linear, precise and accurate. The developed method was validated and found suitable for application in designing pharmacokinetic studies with simplified solvent system.