Abstract: OBJECTIVE To develop a LC-MS/MS method for the determination of rivaroxaban in human plasma and apply it to the pharmacokinetics study. METHODS The plasma samples were precipitated by acetonitrile. The ZORBAX Eclipse XDB C₁₈ column(2.1 mm×100 mm, 3.5 μm) was adopted. The mobile phase was acetonitrile-10 mmol ammonium acetate (containing 0.1% formic acid) with the gradient elution at the flow rate of 0.30 mL·min^-1. Detection of the analyte was achieved by using positive ion electrospray ionization (ESI) in the multiple reaction monitoring (MRM) mode. The MS/MS ion transitions monitored were m/z 436.2→m/z 145.2 and m/z 256.1→m/z 165.2 for rivaroxaban and internal standard, respectively. RESULTS The linear range of rivaroxaban was 5.0-1 000 ng·mL^-1(r=0.995 8). The lower limit of quantitation was 5.0 ng·mL^-1. The recovery rate was 94.1%-106.7%, and the intra-day and inter-day relative standard deviations were <15%. The absolute recovery was 84.4%-91.3%, and no significant matrix effect was found. CONCLUSION This method is rapid, sensitive, accurate, specific, reliable, and suitable for the determination of rivaroxaban in human plasma and pharmacokinetic study.