Abstract: Objective: To optimize process conditions for the hydrolysis of small yellow croaker by trypsin (PTN 6.0S) to prepare peptides with high immuno-activity. Methods: the proliferative effect (expressed as stimulation index, SI) of small yellow croaker hydrolysate on mouse spleen cells was assessed by the MTT assay. Based on the index, single-factor experiments followed by response surface analysis were carried out for the optimization of hydrolysis conditions. Results: The optimal hydrolysis conditions for preparing immune peptides were hydrolysis at 41.66 ℃ for 18.24 h at a substrate concentration of 6 g/100 mL and an enzyme-to-substrate mass ratio of 3.13‰. Under these conditions, the predicted SI of small yellow croaker hydrolysate was 0.387. When the temperature, time and substrate concentration were modified as 42 ℃, 18.2 h and 3‰, the actual SI was 0.369± 0.003, which was slightly lower than the predicted value. Thus, appropriate process conditions were achieved.

Key words: small yellow croaker, immune peptides, response surface methodology, MTT assay