Abstract: A high-performance liquid chromatography-tandem mass spectrometry method was developed for simultaneous determination of 18 sulfonamide drugs residues in fish. The samples were extracted by 1% formic acid acetonitrile and the extracts were purified on enhanced solid-phase extraction column. The 18 sulfonamide drugs were analyzed by CNW Athena C₁₈-WP column (2.1 mm×100 mm, 3 μm) using methanol and 0.1% formic acid as the mobile phases, the sample was analyzed under the multiple reaction monitoring mode with positive electrospray ionization(ESI⁺) and quantify by internal standard. The results showed that good linearities were obtained for the 18 sulfonamides in 5~200 ng/mL (r≥0.9962). The limits of detection and quantification for sulfonamides were 0.11~0.47 and 0.37~1.57 μg/kg, respectively. The recoveries at three concentrations ranged from 73.30%~116.9%, with the RSD from 0.29%~7.74% (n=6). The method was simple, accurate, rapid, and highly sensitive. The method was successfully used for measurement of multiple sulfanilamide drugs residues in fish.