In order to meet the teaching needs of the course of modern analytical methods for natural products of forestry chemical industry in our school, we designed a comprehensive experiment of “high performance liquid chromatography (HPLC) for the determination of flavonoids in Zanthoxylum bungeanum Maxim from different origins” in conjunction with the construction of “double first-class” majors. The content of quercetin and kaempferol in Zanthoxylum bungeanum Maxim varieties from different origins was determined by high performance liquid chromatography (HPLC) using methanol extraction and hydrochloric acid hydrolysis, and the results showed that the analytical method was accurate and reliable, and the detection limit was 0.125 μg/g with RSD less than 2%. The teaching practice showed that the experiment deepened students’ understanding of theoretical knowledge, improved the fusion of students’ theoretical knowledge and practical ability, promoted the scientific literacy and comprehensive thinking ability of students, and cultivated students’ creative thinking and teamwork.