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As a high-quality renewable protein source, the deep processing utilization of bone protein is beneficial to the consummation of the efficient utilization of animal bone resources, the increase of the processing by-products value-added, and the solution of serious environmental issues owing to the improper handling. However, due to the complex enzymolysis processes, the ossein peptides often suffer from poor dimension uniformity and poor performance stability of raw material, which will limit their materialized utilization. Therefore, it is of great significance to explore the effects of different enzymatic hydrolysis processes on the material properties of ossein peptides. The effects of different kinds of proteases, mainly including alkaline lipase, flavorzyme, protamex, neutral protease, and alkaline protease on the enzymolysis efficiency of bovine waste proteins were investigated, with the degree of hydrolysis (DH) as the response variable. The optimum enzymolysis process for ossein peptide with low molecular weight was determined by systematically analyzing the improvement of enzymatic hydrolysis of different kinds of proteases. Furthermore, the mechanisms were explored by analyzing the changes of various properties of the products in the enzymatic hydrolysis. It was found that the ossein peptides prepared by the process of lipase pretreatmentalkaline proteaseprotamex had the best overall performance. The enzymatic hydrolysis process was optimal as follows, at 0.09g/mL substrate mass concentration and initial pH value 7.5, with 0.08% lipase, followed by 4h incubation at 40℃, followed by alkaline protease(addition of enzyme was 0.36U/g) at 60℃ for 5h, and finally, protamex (addition of enzyme was 0.36U/g) was incubation at 55℃ for 5h. The DH reached 16.12%, and the content of total amino acids (TAA) reached 171.571mg/g. Meanwhile, the value of Mw, Mw/Mn, and the Mz/Mw was 883Da, 9.73 and 7.91, respectively. However, after the process of lipase pretreatment-alkaline protease-flavorzyme, the DH and FAA reached the maximum of 19.02% and 257.760mg/g. In addition, the cost input was the least when the physicochemical properties and molecular weight distributions (MWD) of ossein peptide samples were not significantly different from other samples. The results showed that finding the equilibrium point between the DH and the physicochemical properties of ossein peptides was conducive to the economy of protein recycling, cleaner production, and the security and stability of the new composites.