The present study was conducted to determine the activity of enzymes in intestine, stomach, and hepatopancreas of puffer (Fugu obscurus) and whether corn gluten meal(CGM) partially replaced fish meal could affect the activity of trypisn, chymotrypsin, carboxypeptidase A, carboxypeptidase B, pepsin, aminopeptidases, lipase and amylase. Five isonitrogenous and isoenergetic experimental diets were formulated. Diets contained corn gluten meal from 0% to 20%. After 60 d, the activities of the enzymes were measured. The results showed that:(1) The enzymes activity in the intestine were significantly higher than those in stomach and hepatopancreas except pepsin. The enzymes activity of stomach were lower,except pepsin; (2) The corn gluten meal impacts the enzymes activity of Fugu obscurus. Trypsin activity was highest in 10% treatment. The activity of chymotrypsin, carboxypeptidase A and aminopeptidases were linearly decreased with increasing CGM percentage from 5 to 20%, and it was significantly lower than that of the control in 20% treatment. Pepsin activity of 10% treatment was significantly higher than that of other groups. Lipase and amylase activities were increased in the trial groups, and it is significantly higher in 10% treatment than that of the others(P<0.05). It was concluded that the activity of digestive enzymes was highest in intestine, and the upper limit of corn gluten meal for optimal digestive enzymes activity in puffer feed was 10%.