OBJECTIVE: To establish a LC-MS/MS method for determination of verapamil in human plasma. METHODS: Thermo Hypurity C 18 column (2.1 mm × 150 mm, 5 μm) was used in the experiment. The column temperature was set at 40°C. 10 mmol · L -1 ammonium (including 0.1% ammonium formate)-acetonitrile (35:65) was used as the mobile phase and the flow rate was 0.3 mL · min -1. The sample room temperature was set at 15°C and the injection volume was 10 μL. ESI source and positive ion scan were used, verapamil and the internal standard imipramine were monitored at (m/z) 455.0 → 165.0 and (m/z) 281→86.0, respectively. RESULTS: The calibration curve of verapamil was linear in the range of 0.4575-234.20 ng · mL -1, and the detection limit for verapamil was about 0.1 ng · mL -1. The method had high sensitivity, stability and specificity and had been used for the pharmacokinetic study of verapamil in vivo successfully. CONCLUSION: The method is simple, accurate and repetitive for the determination of verapamil in human plasma and suitable for pharmacokinetic study.