Abstract: A method based on solid-phase extraction and high performance liquid chromatography coupled with electrospray ionization tandem mass spectrometry(SPE-HPLC-MS/MS) had been proposed for the determination of chloramphenicol, thiamphenicol, florfenicol and florfenicol amine residues in beeswax. The sample was pre dissolved with n-hexane and then extracted with water, purified the Waters Oasis MCX micro-column and separated on the InertSustain Swift C₁₈ column (100 mm×2.1 mm, 1.9 μm). The determination was carried out with eletrospray ion source and multiple reaction monitoring (MRM) mode and the corresponding quantification was performed by employing the method of internal standards. The calibration curve of mixed standard solution showed a good linearity in the concentration rang from 0.1 to 10 ng/mL with the correlation coefficients (r) more than 0.99. The limits of quantification (S/N>10) for antibiotics were 0.1 μg/kg. The recovery was 65.0% to 113.20% in the spiked concentration 0.1, 0.3 and 1.0 μg/kg, with the relative standard deviations (RSD, n=6) lower than 20%.The method was applied to detect 15 beeswax samples, and the results showed that chloramphenicol residues were detected in 4 batches of samples. The method is rapid, sensitive and accurate, and could be applied for the qualitative and quantitative analysis of chloramphenicol, thiamphenicol, florfenicol and florfenicol amine residues antibiotics in beeswax.