The invention belongs to the technical field of biological preparation, and relates to a production method of turtle peptide, which comprises the following steps: slaughtering adult live turtles, cleaning, removing bones, and crushing into turtle meat paste slurry; mixing the tortoise shell extract and the tortoise meat paste slurry, performing ultrasonic homogenization, blending with ultrapure water to obtain a substrate solution, and adding protease for enzymolysis; and carrying out enzyme deactivation, sterilization, ultrafiltration separation and vacuum freeze drying on the reaction product obtained by enzymolysis to obtain the soluble powdery turtle peptide. According to the turtle peptide produced by taking multiple turtle compound components as raw materials and applying an enzymolysis method, active peptide molecules in adult turtles are extracted to the maximum extent, meanwhile, other bioactive components in the raw materials are preserved, and multiple bioactive components supplement one another. The turtle peptide can effectively regulate immunity, resist oxidation, resist tumors, resist fatigue, regulate blood fat, improve sleep and delay senescence, so that the effects of nourishing yin, tonifying yang, building bodies, maintaining beauty, keeping young and prolonging life are achieved.