Abstract: With the concept of green analytical chemistry, an experiment on the determination of sulfonamide drugs in water using liquid-liquid microextraction combined with high-performance liquid chromatography(HPLC) was designed for undergraduate experimental teaching. With a small amount of organic solvents in liquid-liquid extraction and ethanol as the green organic mobile phase for chromatographic separation, the extraction and separation conditions were optimized, and a new method of liquid-liquid microextraction combined with HPLC has been developed for determining sulfonamide drugs (sulfaguanidine (SG), sulfamethoxazole (SD), sulfamethoxazole (SDM), sulfamethoxazole (SMZ) and sulfamethoxazole (PST)) and trimethoprim (TMP, a commonly used compatible drug for sulfonamide drugs). The results indicated that the linear measurement ranges were 10−1000 μg/L for SG, SD, SDM and SMZ, 100−1000μg/L for TMP and 50−1000 μg/L for PST, respectively. The detection limits were in the ranges of 50−100 ng/L (S/N=3). The recovery rates of the lake water sample were between 93.6% and 105.2%. This experiment is based on the forefront of analytical chemistry and is moderately difficult. It is suitable as a researching experiment for senior undergraduate students majoring in chemistry, pharmacy and environment, which is beneficial for students to improve their innovation ability, low-carbon awareness and comprehensive quality.