Abstract: The feed sample (2.50 g) was taken and placed in a 50 mL-centrifuge tube. 20 mL of extract solvent methanol solution containing 0.25% (volume fraction) formic acid was added, and the mixture was shaken for 15 min and centrifuged for 3 min. The supernatant was taken and placed in a 50 mL-centrifuge tube. The extraction of the residue was repeated with 20 mL of extract solvent. The two supernatants were combined, and the solution was made its volume up to 50 mL with extract solvent and mixed well. 2.0 mL of the solution was taken and passed through an EMR solid-phase extraction column (300 mg/3 mL), and 1 mL of methanol was added for rinsing. The column extraction solution and the eluent were collected and blown to dryness by nitrogen at 50 ℃. 4 mL of the mixed solution of 0.1% (volume fraction, the same below) formic acid solution and methanol at a volume ratio of 9∶1 was added and vortexed. The solution was passed through a 0.22 μm organic filter membrane, and tulathromycin (calculated based on tulathromycin A) in the filtrate was determined by ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Acquity UPLC BEH C₁₈ was used as the stationary phase and the mixed solution composed of 0.1% formic acid solution and methanol at different volume ratios was used as the mobile phase for gradient elution. Electrospray ion (ESI) source was used for mass spectrometry. The targets were analyzed by multiple reaction monitoring (MRM) mode in positive ion (ESI⁺) scanning mode. As shown by the results, linear relationship between the corresponding peak areas of the quantitative ion pair and mass concentrations of tulathromycin was kept in the range of 0.50-100 μg·L⁻¹, with detection limits (3S/N) in the range of 3.0 μg·kg⁻¹. Test for recovery was made by the standard addition method, giving results in the range of 93.0%-106%, with RSDs (n=5) of the determined values less than 10%. This method was used for the analysis of 38 actual feed samples, and tulathromycin was not detected.