Abstract: Matrix effect exists widely in mass spectrometry and is considered to be one of the key factors affecting the quantitative accuracy of high-performance liquid chromatography-tandem mass spectrometry Based on isotope dilution mass spectrometry, this study systematically studied the influence of chromatographic conditions on the matrix effect of salbutamol and clenbuterol mass spectrometry in pork, including the comparison of matrix effect and its correction factors under three chromatographic separation conditions, and the influence of different calibration solutions on the quantitative results. The results showed that the influence of the matrix on the target substance is significantly weakened when the chromatographic conditions with the best separation effect are adopted. The isotopic dilution mass spectrometry correction factors of salbutamol and clonbuterol are 1.01 and 1.00, respectively. There is no significant difference between the measurement results of the solvent calibration solution and the matrix calibration solution. The influence of matrix effect on mass spectrometry can be effectively eliminated by optimizing the analytical conditions of liquid chromatography and the accuracy of quantitative results can be improved