Abstract: There are some problems in the teaching experiment of yeast alcohol dehydrogenase (YADH) extraction and purification. In order to obtain YADH with high purity, high activity, and high recovery, a single factor experiment is applied to optimize the separation and purification process of enzyme protein, including hot extraction, thermal denaturation precipitation, and organic solvent precipitation. which will improve the extraction and purification efficiency of YADH. The results show that the best effect of separation and purification is achieved when the solid-liquid ratio is 1 : 6, the crude extraction temperature is 25 ℃, the crude extraction time is 1.5 h, the thermal denaturation temperature is 65 ℃, the thermal denaturation time is 15 min, the organic solvent precipitant is ethanol, the adding proportion of precipitated impure protein was 0.5 times of the supernatant, and the adding proportion of precipitated enzyme protein is 0.45 times of the supernatant. Through the optimization of the above conditions, the effect of teaching experiment is improved. It is also helpful to exercise students’ practical skills, innovative thinking, and scientific research abilities.