Objective To determine chloramphenicol and metronidazolel in honey by isotope-labelled internal standards ultra performance liquid chromatography-mass spectrometry. Methods Samples were extracted with ethyl acetate solution, and cleaned up on a MCS cartridge. The target analytes were separated on a ZORBAX SB-C18 column with gradient elution using a mobile phase made up of methanol and 5 mmol/L ammonium acetate solution (containing 0.05% formic acid). Detection was carried out using positive and negative electrospray ionization and multiple reaction monitoring (MRM), and quantified with isotope internal standardmethod . Results The chloramphenicol and metronidazolel showed good linearity in the range of 0.05-5.00 ng/ml. The recovery at three spiked levels of 0.5,2.0 and 5.0 μg/kg were in the range of 79.3%-96.7%. The relative standard deviation (RSD) was 5.5%-14.8%. The limits of quantitation were 0.15 μg/kg, the limits of detection were 0.05 μg/kg. Conclusion The method is sensitive and accurate. It could be applied to the high-throughput analysis of chloramphenicol and metronidazolel.