Abstract: The current determination methods of perfluorinated and polyfluoroalkyl substances (PFASs) did not involve the new PFASs, and were difficult to meet the requirements of simultaneous determination of PFASs, especially the low recoveries of long-chain compounds. In order to solve the above problems, the method mentioned by the title was proposed. The fish sample (2.000 g) was taken and mixed well with 20 μL of 100 μg · L⁻¹ 13 PFASs mixed isotopic internal standard solution and 2.0 mL of water, and then 10.0 mL of acetonitrile solution containing 1%(volume fraction) formic acid was added. The mixture was vortex oscillated for 30 min, and centrifuged for 10 min. The supernatant was taken and purified by pass-through Anavo HMR-Lipid solid phase extraction column. The purified solution obtained was blown to dryness by nitrogen at 40 ℃. The residue was redissolved with 0.4 mL of methanol. After swirl mixing and centrifugation for 5 min, the supernatant was taken, in which 23 PFASs including 2 new PFASs of 6∶2 chloropolyfluoroalkyl ether sulfonate (6/2F-53B) and 8∶2 chloropolyfluoroalkyl ether sulfonate (8/2F-53B) were simultaneously determined by ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) using Acquity UPLC BEH C₁₈ column as the stationary phase, and a mixture of methanol and 2 mmol · L⁻¹ ammonium formate solution at different volume ratios as the mobile phase for gradient elution. In mass spectrometry analysis, the negative ion mode of the electrospray ion source was used for scanning. The isotope internal standard method was used for quantification. As shown by the results, the linear ranges of 23 PFASs standard curves were all 0.2-50.0 μg · L⁻¹, with detection limits in the range of 0.01-0.02 μg · kg⁻¹. Test for recovery was made by the standard addition method, giving results in the range of 75.8%-110%, with RSDs (n=6) of the determined values less than 7.0%. This method was used for the analysis of actual fish samples, and the total detection amounts of PFASs were in the range of 0.039-1.310 μg · kg⁻¹.