Abstract: OBJECTIVE To set up a rapid and sensitive LC-MS/MS method for simultaneous quantitative determination of lopinavir(LPV) and ritonavir(RTV) in human plasma. METHODS LPV and RTV were extracted with ethyl acetate. The residues were analyzed with an Agilent Eclipse Plus CM₁₈ column(4.6 mm×150 mm, 3.5 μm) with the mobile phase consisted of acetonitrile with 0.005 mol·L^-1 ammonium formate(0.1% formic acid)=90∶10, with a flow rate of 0.5 mL·min^-1, temperature of 40 ℃. Selected reaction monitoring(SRM) using the precursor to product ion combinations of m/z 629.5→155.1, 721.4→296.1 and m/z 614.5→465.4 was performed to detect LPV, RTV and the internal standard(indinavir, IDV), respectively. RESULTS The average recovery rate RSD for LPV of three levels of concentration of high (10 000 μg·L^-1), medium (1 000 μg·L^-1) and low (40 μg·L^-1), were less than 15%. The calibration curves for LPV was Y=1.669 9X-0.001 3, r=0.998 4, over the range of 20-20 000 μg·L^-1. The limits of quantitation was 20 μg·L^-1. Meanwhile, the average recovery rate RSD for RTV of three levels of concentration of high (2 500 μg·L^-1), medium (250 μg·L^-1) and low (10 μg·L^-1), were also less than 15%. The calibration curves for RTV was Y=1.723 7X-3.274 8×10^-4，r=0.998 7(n=7), over the range of 5-5 000 μg·L^-1. The limits of quantitation was 5 μg·L^-1. CONCLUSION The method provides a sensitive, accurate, precise and reliable analytical procedure for the therapeutic drug monitoring of LPV and RTV simultaneously in clinic and phamacokinetic studies.