Fluoroquinolones (FQs) are among the most important antibiotics used in human and veterinary medicines. A simple and effective analytical method based on reversed-phased liquid chromatography with fluorescence was developed and validated for the simultaneous determination of eight FQs in wastewater at trace level. Aqueous samples were extracted using AnpelTM MEP cartridges where they were subsequently eluted by formic acid in methanol. The aqueous extracts were analyzed by gradient elution Liquid Chromatography with Fluorescence Detection (LC–FLD), whose mobile phase was composed of acetonitrile and 10 mM tetrabutyl ammonium bromide (TBAB). The limits of detection (LOD) and the relative standard deviation (RSD) were as low as 0.11–1.06 μg/L and 2–5%, respectively. The presented method was successfully applied to quantify FQs in the influent and effluent of several typical Sewage Treatment Plants (STPs) in Shanghai. For the extraction of 100 mL influent and 500 mL effluent sewage water samples, recoveries obtained were between 79–109% and 80%–105%, respectively. 7 FQs were occurred and identified in the STPs with the concentrations varying from 7 ng/L to 1 μg/L. Norfloxacin, ciprofloxacin and lomefloxacin were the most frequently detected antibiotics occurring in the wastewaters. The analytical procedure developed may be used for more in-depth studies on the occurrences and the fate of these commonly used pharmaceuticals in the sewage treatment plants and in the aquatic environment.