In order to utilize the hairtail resources effectively and prepare high-quality angiotensin-converting enzyme (ACE) inhibitory peptide， the most suitable protease for the preparation of hairtail ACE inhibitory peptide was screened using ACE inhibition rate and hydrolysis degree as evaluation indexes. The enzymatic process conditions were optimized by single factor and response surface. The optimized hairtail ACE inhibitory peptide was analyzed for molecular weight and amino acid composition. The ACE inhibitory peptide was further purified by ultrafiltration and Prep-HPLC， and the active fraction with the highest ACE inhibition rate was obtained. The results showed that the protemax was the best protease for the preparing ACE inhibitory peptide， and the optimized process parameters were obtained as follows: enzymatic hydrolysis time of 3 h， hydrolysis temperature of 45 °C， enzyme dosage 5 000 U/g and pH 7.0. Under this condition， the theoretical ACE inhibition rate was 80.15%， the actual value was 80.43%， and the IC50 was 0.33 mg/mL. The relative molecular weight of the ACE inhibitory peptide was mainly distributed below 1 000 u. The amino acid composition pattern of the hairtail ACE inhibitory peptide was reasonable， with high content of fresh and hydrophobic amino acids. The six fractions were separated by ultrafiltration and Prep-HPLC， and the highest activity was obtained for fraction 5 with an IC50 of 0.11 mg/mL. This study provides a theoretical basis for the application of hairtail in functional foods for hypertension.