Abstract: After the fingerprints were taken using the cotton swab and filter paper, the cotton swab head and the filter paper were cut and palced in 1.5 mL plastic centrifuge tubes, respectively. Then 0.25, 0.50 mL of methanol were added into the centrifuge tubes, respectively. The mixture was ultrasonically oscillated for 3 min, centrifuged for 3 min. After the supernatant was filtered through a 0.22 μm filter, glibenclamide, glimepiride, glipizide and gliclazide in the filtrate were separated on an ACQUITY UPLC HSS C₁₈ column (2.1 mm×50 mm, 1.8 μm), with a mixture of acetonitrile and 0.1% (φ) formic acid solution as the mobile phase for gradient elution. Electrospray positive ion source and multiple reaction monitoring mode were used in mass spectrometry analysis. Linearity ranges of the four sulfonylurea hypoglycemic agents were same in 0.25-250 ng/swab (patch), with same detection limit (3S/N) of 0.125 ng/swab (patch). Values of recovery obtained by standard addition method were in the range of 68.0%-123%, and RSDs (n=5) were in the range of 1.2%-9.5%.