Abstract: The hydrolysis process of bioactive peptidase with cholate adsorption effect was investigated using quinoa protein as the research object. The degree of hydrolysis (DH) and cholate adsorption effect were adopted as the assessment criteria for screening of protease used for enzymatic hydrolysis reaction. Enzymatic hydrolysis process was further improved using orthogonal test based on single-factor experiment. Ultimately, the flavourzyme was used as the best reactive enzyme, and the optimum enzymatic hydrolysis process conditions were as follows:pH7.0, substrate concentration 3% (w/v), enzymatic hydrolysis temperature 55℃, enzyme addition amount 6000 U/g, and enzymatic hydrolysis time 2 h. DH was detected to be 38.22%±0.65% under these conditions, and the amount of cholate adsorbed by active peptide was (7.56±0.11) mg/mL. These results indicate that the enzymatic hydrolysis process is stable and feasible and can be used for preparation of bioactive peptides with better cholate adsorption.