Abstract: A method of QuEChERS-online gel permeation chromatography gas chromatography-mass spectrometry (QuEChERS-GPC-GC/MS) was established for 9 hydroxy veterinary drug residues in fish. GPC system was adopted for online purifying pretreatment to remove most of the fat in fish so as to effectively reduce the time of sample pretreatment. At the same time, the selective ion monitoring acquisition mode could greatly reduce the interference and provide selectivity. Extracted with acetonitrile and purified by dispersive solid phase extraction method, the analytes were detected and determined under SIM scan type with tandem mass analyzer. The analytical method was well validated, and the results were obtained at the respect of precision, repeatability and spiked recovery. The detection limits of 9 hydroxy veterinary drug residues are 0.3-1.0 μg/kg with the linear range of 0.5-20.0 μg/L. The overall recoveries are between 63.5% and 90.2% with the relative standard deviations (RSDs) of 3.6%-15.4%. Finally, the validated method was carried out in total of 10 samples to evaluate 9 hydroxy veterinary drug residues, and one sample has chloramphenicol positive, and the content is 6.1 μg/kg. This method is suitable for the determination of 9 hydroxy veterinary drugs in fish.