In this study, the purpose was to screen novel angiotensin converting enzyme inhibitory peptides (ACEIPs) from tuna muscle taking two-steps enzymatic hydrolysis (Neutrase and Alkaline). Following isolation and purification by ultrafiltration, the Sephadex G-15 gel chromatography and reversed-phase high-performance liquid chromatography based on active-guide, the amino acid sequence was identified using Q-Orbitrap-MS/MS. Five peptides were chose synthesized based on the in silico screening methods. Among these, the two novel ACEIPs LTGCP and YPKP showed better inhibitory ability, and their corresponding IC