Abstract: A method for simultaneous determination of residues of 6 antibacterial drugs including flufenicol, sulfamethoxazole, carbendazim, norfloxacin, amikacin, and cefazolin sodium in poultry breeding wastewater by high performance liquid chromatography-tandem mass spectrometry with online solid phase extraction was proposed. The poultry breeding wastewater sample was taken and passed through gauze and filter paper. 10.0 mL of the filtrate was taken and performed online solid phase extraction using Waters Oasis HLB solid phase extraction column. Capcell PAK AG ODS C₁₈ column was used as the stationary phase and the mixed solution composed of 0.1% (volume fraction) formic acid solution and acetonitrile 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 relationships between the corresponding peak areas and mass concentrations of 6 antibacterial drugs were found within 0.005-5.0 mg·L⁻¹, with detection limits (3S/N) in the range of 0.018 3-0.072 2 μg·L⁻¹. Test for recovery was made by the standard addition method, giving results in the range of 83.8%-105%, with RSDs (n=6) of the determined values less than 5.0%. This method could be used to determine the residues of 6 antibacterial drugs in 5 actual poultry breeding wastewater, and the detection amounts were in the range of 16.09-277.14 μg·L⁻¹.