Abstract: The concentrations of tilmicosin residues in swine tissues were determined by HPLC. Swine tissues were extracted with acetonitrile and potassium dihydrogen phosphate buffer. The extracts were eluted by the C18 solid-phase extraction (SPE) cartridge. The residues were eluted from SPE cartridge with methanolacetonitrileammonium acetate solution, and the eluate was determined by HPLC at 290 nm. The limits of quantitation (LOQ) were 0.02 μg/g and 0.05 μg/g for tilmicosin in muscle and liver, respectively. The limits of detection (LOD) were 0.01 μg/g and 0.025 μg/g for tilmicosin in muscle and liver, respectively. Recoveries of tilmicosin in muscle fortified at 0.02~2.0 μg/g and in liver fortified at 0.05~5.0 μg/g were in the range of 84.2%~96.6% and 82.0%~92.8%, respectively. Coefficients of variation were in the range of 5.0%~8.2% and 5.8%~11.3%, respectively. The method can meet the requirement for tilmicosin residue analysis.