A simple and rapid ultra-performance liquid chromatographic tandem mass spectrometric assay for the measurement of bisphenol-A (BPA) in human urine was developed and validated. Sample preparation involved addition of 4-nitrophenol (as an internal standard, IS) to 1.0 ml human urine sample containing BPA, and extraction with ethyl acetate and hexane (6:4,v:v), evaporation, and dissolving the residue in the mobile phase. Analytes separation was performed using a reversed phase Atlantis dC18 column (2.1x 100 mm, 3 µm) protected by guard pre-filter and a mobile phase that consisted of 5.0 mM ammonium acetate: acetonitrile (20:80, v:v) and was delivered at a flow rate of 0.30 ml/min. The analytes were quantified in negative ion mode, using electrospray ionization (ESI) set at transitions of mass to charge (m/z): 226.87 → 212.01 and 137.84 → 107.86 for BPA and IS, respectively. The relationship between BPA concentration and peak area ratio (BPA /IS) was linear in the range 0.2 - 20 ng/ml. Mean extraction recovery of BPA and the IS was ≥ 90% and 97%, respectively. The method was validated in respect to accuracy, precision, linearity, and specificity. Stability of BPA in urine was determined under conditions generally encountered in the clinical laboratories. The method was successfully applied to determine BPA level in urine samples obtained from healthy volunteers