The study was aimed to evaluate various pharmacokinetic parameters of a commercially available fixed dose combination of oral antidiabetics (Metformin/Glibenclamide 500/5mg tablets) in plasma sample of normal healthy adult male volunteers by applying an accurate, selective, and reproducible HPLC-UV analytical method for quantification of Metformin HCL and Glibenclamide simultaneously in a single chromatographic run. Previously no HPLC-UV analytical method for simultaneous estimation of Metformin/Glibenclamide has been reported in Pakistan. The human plasma samples were evaluated by using an isocratic High Performance Liquid Chromatography (HPLC) system of Sykam consisted of a pump with a column of Thermo Electron Corporation USA (ODS hypersil C18 4.6 mm x 250 mm), a UV-detector with data processing Clarity software. The mobile phase of 0.040M Potassium dihydrogen phosphate containing 0.25mL/L triethylamine at pH 3.5 (adjusted with 1:1 phosphoric acid) and acetonitrile (465: 535v/v) was delivered with injection volume of 100μL at flow rate of 1 mL/min at 25°C temperature. The detection was performed at λmax230 nm. By applying this method, important pharmacokinetic parameters Cmax, Tmax, AUCo-oo, AUMCo-oo, t1/2, Ke, MRT, Vdand CIT are calculated. Maximum plasma concentrations Cmax was 131.856±8.050ng/ml for Glibenclamide (Mean ± SEM) and 511.106±12.675 ng/ml for Metformin HCl (Mean ±SEM).