A sensitive and specific method using liquid chromatography-electrospray tandem mass spectrometry (LC-MS/MS) for the determination of pinaverium bromide in human plasma was developed and validated. Pinaverium bromide and an internal standard (paclitaxel) were isolated from plasma samples by precipitating plasma, and determined by LC-MS/MS in multiple-reaction monitoring mode. The main metabolite of pinaverium bromide and endogenous substances in plasma did not show any interference. The calibration curve was linear over the plasma concentration range of 10.0-10000.0 pg/mL with a correlation coefficient of 0.9979. The relative standard derivations intra- and inter-day at 30.0, 300.0 and 8000.0 pg/mL in plasma were less than 15%. The absolute recoveries of pinaverium bromide and the internal standard were 99.7-111.7 and 106.2%, respectively. The lower limit of quantitation was 10 pg/mL. The analytical method was successfully applied to study the pharmacokinetics of pinaverium bromide tablets in healthy Chinese volunteers.