There is interest in evaluating the efficacy of lower doses of certain thiazolidinedione family or insulin sensitizers for clinical care. We have developed a sensitive and rapid positive polarity electro-spray ionization with Tandem-mass spectrometry method was validated as per FDA guidelines for the determination of Pioglitazone (PGZ), their metabolites Keto-Pioglitazone (KPGZ) and Hydroxy-Pioglitazone (HPGZ) in human plasma samples. Glyburide was used as an internal standard. The chromatographic separation was achieved by Discovery C18 column using isocratic mobile phase consists a mixture of 5mM ammonium acetate (pH 6.4 ± 0.1) and acetonitrile (40:60 v/v). An aliquot of 150 µL plasma was used for the liquid- liquid plasma extraction technique for quantification of the analytes. The molecular ion Q1, product ion Q3 transitions for Pioglitazone, Keto-Pioglitazone and Hydroxy-Pioglitazone were found to be 357.2→134.1, 371.3→148.4 and 373.2→150.1 m/z respectively. The proposed method was validated in the concentration range of 10.0-3529.0, 5.0-1209.7 and 5.0-2029.7 ng/mL for PGZ, KPGZ and HPGZ respectively in multiple reaction monitoring mode. The pharmacokinetic parameters for Pioglitazone were found to be Tmax - 2.5 Hours, Cmax - 2088.2 ng/mL, T1/2 - 6.5 Hours, AUC (0-T) - 20925.9 ng/mL and AUC (0-∞) - 21276.2 ng/mL). The entire results obtained in the study were well within acceptance limits.