Objective To develop a method for determining the contents of dendrobine, dendrophenol, moscatilin, chrysotoxine, and erianin in dendrobium by ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS).Methods The samples were ultrasonically extracted with 70% methanol (by volume) for 40 min. The extract was filtered through a 0.22 μm organic membrane and then determined by UPLC-MS/MS. Gradient elution was performed using methanol and 0.1% formic acid water as the mobile phase. The determination was carried out in MRM mode with an electrospray ionization (ESI) source in positive ion mode, and quantification was done by the external standard method.Results Dendrobine, dendrophenol, chrysotoxine, moscatilin, and erianin show a good linear relationship in the range of 50.0~500.0 ng/mL (R²>0.999). The detection limits of the five substances are 0.15~0.71 μg/kg, and the quantification limits are 0.50~2.37 μg/kg. The average recovery rates of the five substances at three spiked concentrations (50.0, 100.0, and 150.0 μg/kg) are in the range of 95.34%~105.00%, with relative standard deviations of 0.80%~1.88% (n=3). Six batches of commercially available dendrobium samples were determined. The contents of dendrobine, dendrophenol, moscatilin, and erianin are in the range of 0.046%~0.318%, 0.024%~0.170%, 0.001%~0.099%, and 0.013%~0.099%, respectively. Chrysotoxine is detected in three batches, with contents in a range of 0.001%~0.004%.Conclusion A method for determining the contents of dendrobine, dendrophenol, moscatilin, chrysotoxine, and erianin in dendrobium is developed by UPLC-MS/MS. This method is accurate, has a short analysis time and high sensitivity, and is suitable for evaluating the quality of dendrobium.