A sensitive and selective liquid chromatographic--mass spectrometric (LC-MS) method for the determination of gliclazide (CAS 21187-98-4) in human plasma has been developed. Sample treatment was based on protein precipitation with acetonitrile. Analytical determination was carried out on a C18 column interfaced with a triple quadrupole mass spectrometer. Positive electrospray ionization was employed as the ionization source. The mobile phase was acetonitrile-water containing 10 mmol/l ammonium acetate, pH 3.5 adjusted with acetic acid (75:25) at the flow rate of 1.0 ml/min. Both the analyte and the internal standard glipizide (CAS 29094-61-9) were detected by use of selected ion monitoring mode. The method was linear in the concentration range of 0.025-2.5 microg/mL. The lower limit of quantification (LLOQ) was 0.025 microg/mL. The intra- and inter-day relative standard deviation across three validation runs over the entire concentration range was less than 9.8%. The accuracy determined at three concentrations (0.05, 0.2 and 1.5 microg/mL for gliclazide) was within +/- 10.11% in terms of relative error (RE). The method herein described was successfully applied for the evaluation of pharmacokinetic profiles of gliclazide sustained release tablets in 18 healthy volunteers. The results show that AUC, Tmax, Cmax and T1/2 between the test formulation and reference formulation have no significant difference (P > 0.05). Relative bioavailability is 96.7.4 +/- 12.9%.