A simple and sensitive method is proposed for the simultaneous determination of quinolones (marbofloxacin, ciprofloxacin, danofloxacin, enrofloxacin, sarafloxacin, difloxacin, oxolinic acid and flumequine) in different fish samples. Ultra-high performance liquid chromatography using a partially porous C18 column coupled to fluorescence detection with a wavelength excitation/emission program was used. The sample treatment consisted of extraction and clean-up using the QuEChERS methodology, showing a high efficiency without interferences in the chromatographic determination. The method was characterized in fish tissue in terms of linearity, precision, trueness and limits of detection and quantification. Limits of detection between 0.1 and 4.7 μg/kg were obtained, with recoveries between 72 and 108%. The proposed method has been tested in bass, trout, panga and sturgeon, showing its simplicity, sensitivity and suitability for routine analysis in that complex matrix.