Abstract: A method was established for the simultaneous determination of 3-methoxytyramine and adrenaline in dairy products by high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) compled with isotope dilution. Sample was extracted by perchloric acid solution, cleaned by weak cationic solid phase extraction column. The analyte was separated by ACQUITY UPLC BEH C₁₈ (100 mm×2.1 mm, 1.7 μm) column and agradient elution of mobile phase with 0.1% formic acid aqueous solution-methanol. The detection was achieved by HPLC-MS/MS with electrospray ion source and positive ion scanning mode as well as multiple reaction monitoring (MRM) mode. The quantification was accomplished by the internal standard method. It was demonstrated that the linear relationships of 3-methoxytyramine and adrenaline were good in the range of 0.5~12.0 ng/mL and 50~1200 ng/mL (r>0.99), respectively. The detection limits and the limit of quantification of 3-methoxytyramine were 0.2 and 0.5 μg/kg, respectively. At the concentrations of 0.5, 1.0 and 3.0 μg/kg, the recoveries of the 3-methoxytyramine ranged from 76.97% to 119.20%, with the corresponding precision (n=6) of 1.53%~9.61%. While the recoveries of adrenaline at 50, 100 and 300 μg/kg were 90.65%~108.55% and the relative standard deviations were in the range of 1.29%~3.95%. The method would be accurate in results and good in repeatability, and it can be an important measure to monitor 3-methoxytyramine and adrenaline in dairy products.