An accurate,precise,simple and rugged Chiral HPLC method has been developed and validated for the quantitation of (S)-isomer in Trelagliptin. The enantiomeric separation of Trelagliptin was achieved Chiral pak AD-3 (250×4.6mm, 3?m) column. The ratio of Hexane,ethanol and diethyl amine (70:30:0.1)in the mobile phase were optimized to obtain the best separation. UV detection was performed at 275 nm. The method is linear over a range of LOQ 150 % of (S)-isomer. The mean recovery of (S)-isomer was found to be in the range of 95105%. The method is simple, rapid, accurate, selective and precise, useful in the quality control of bulk manufacturing.