Abstract: Objective To develop a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for detecting tenofovir in human plasma. Methods Twenty four healthy male volunteers received a single oral dose of 300 mg tenofovir disoproxil fumarate tablets under fasting and high-fat diet conditions in a randomized four-way crossover bioequivalence study with two preparations of tablets. Plasma samples were taken and analyzed using the LC-MS/MS method. The pharmacokinetic parameters of the two preparations were calculated and compared statistically to evaluate their bioequivalence using Phoenix Winnonlin6.3. Results Linear detection responses were obtained for tenofovir at the range from 3.13 to 500 ng/mL. The intra- and inter-day precisions were high, with lower than 5.43% 〔relative standard deviation (RSD)%〕, high recovery and good stability. The 90% confidence intervals of peak concentration (Cmax) of tenofovir and its area under the curve (AUC0-t and AUC0-∞ ) all fell within the bioequivalence limit 80.00%-125.00% under both fasting and high-fat diet conditions. No significant difference in peak time (Tmax) was demonstrated between the two preparations （P＞0.05）. Conclusion The LC-MS/MS method can be used for simultaneous determination of tenofovir in human plasma. The two preparations of tablets are bioequivalent.