Abstract: To achieve the high-value utilization of squid processing by-products, specifically squid skin, this study focused on the extraction of angiotensin-converting enzyme (ACE) inhibitory peptides from the skin of Dosidicus gigas through enzymatic hydrolysis. Optimal conditions for enzymatic hydrolysis were established through single-factor experiments and response surface methodology, using the ACE inhibitory rate and the degree of hydrolysis as the evaluation metrics. The study assessed the ACE half-inhibitory concentration, molecular weight distribution, and DPPH radical scavenging activity of the hydrolyzed peptides. The results revealed that neutral protease was the most effective enzyme for this process, with optimal conditions established as follows: Hydrolysis temperature at 45 ℃, duration of 2.6 hours, substrate concentration at 2.0%, protease concentration at 7200 U/g, and a pH of 8.0. Under these parameters, the ACE inhibitory rate of the hydrolyzed squid skin product reached 88.22%±1.28%, closely with the predicted value. The IC₅₀ for the ACE inhibitory rate of the squid skin peptides was calculated to be 2.839 mg/mL, with a molecular weight distribution predominantly below 3 kDa, accounting for 93.68% of the total. Additionally, the IC₅₀ for the DPPH radical scavenging activity was found to be 6.183 mg/mL. These results provided a theoretical foundation for the development of bioactive peptides from squid skin and underscored the potential for high-value applications of squid processing by-products.