As a non-traditional sample matrix, feather samples can be used to effectively monitor antibiotic addition and organismal residue levels in poultry feeding. Therefore, an ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS) method was developed to simultaneously determine the residue levels of 26 quinolones in poultry feathers. The feather samples were extracted by sonication with a 1% formic acid and acetonitrile mixture in a water bath at 50 °C for 30 min, purified by the adsorption of multiple matrix impurities, dried with nitrogen, redissolved, and analyzed by UPLC-MS/MS. The linearity, limit of detection (LOD), limit of quantification (LOQ), recovery and precision were calculated. The 26 antibiotics demonstrated good linearity in the linear range. The recoveries and coefficients of variation were 78.9–110% and <13.7% at standard spiked levels of 10, 100 and 200 μg/kg, respectively. The LOD and LOQ were 0.12–1.31 and 0.96–2.60 μg/kg, respectively. The method also successfully identified quinolone residues in 50 poultry feather samples. The results showed that quinolones can accumulate and stabilize for a certain period of time after transferring from the body to the feathers of poultry.
