Abstract: A large amount of byproduct is produced in the process of tilapia fillet processing. Tilapia byproduct is low cost, rich in protein and contains abundant nutritional components. The hydrolysis of tilapia byproduct by the neutral protease and flavorzyme was studied. The enzymatic hydrolysate was obtained by centrifuging and filtering and the polypeptides solution was prepared by ultrafiltration membrane, and the polypeptides beverage was prepared. The anti-oxidation and anti-fatigue effects of the polypeptides beverage were investigated. The results showed that polypeptides beverage was rich in nutrient and performed the good palatability. Its polypeptide content reached 18.7 g/L, and the amino nitrogen content was 1.2 g/L. The analytic results of heavy metal and microorganism determination showed that the product conformed to the national hygienically standard for feeds. The animal experiment proved that anti-oxidation activity and the time span of weight-loaded swimming of mice had increased. It showed that the tilapia polypeptides beverage had effects of relieving fatigue and anti-oxidation.