A selective, rapid and sensitive ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed for the determination and pharmacokinetic study of azithromycin in human plasma.With roxithromycin as internal standard, sample pretreatment involved a one-step extraction with diethyl ether of 0.5 mL plasma.The analysis was carried out on an ACQUITY UPLCTM BEH C18 column (50rnm× 2.1mm, i.d., 1.7μm) with gradient elution at flow-rate of 0.35mL/min.The mobile phase was 50mM ammonium acetate-water and acetonitrile.The detection was performed on a triple-quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode via Turbo ion spray ionization (ESI).Linear calibration curves were obtained in the concentration range of 1-1000 ng/mL, with a lower limit of quantification of l ng/mL.The intra-and inter-day precision (R.S.Ds.) values were below 15% and accuracy (RE) was-1.3 %~5.7 % at all three QC levels.The method was utilized to support clinical pharmacokinetic studies of azithromycin in healthy volunteers following oral administration.