Abstract:In order to prepare high quality and efficient small fish protein antibacterial peptide，Zhoushan lowvalue small hairtail was taken as the research object，escherichia coli was used as an indicator，peptide and papain were preferred for single enzymatic hydrolysis by single factor experiments，purified by Sephadex G-25 gel chromatography and reversed-phase high performance liquid chromatography，the optimal conditions for complex enzymatic hydrolysis were as follows：the hydrolysis time of compound enzyme was 4.2 h，the temperature of complex enzymatic hydrolysis was 36.5℃，and the pH of complex enzymatic hydrolysis was 4.4.The effects of three factors were：hydrolysis time of complex enzyme＞complex solution pH＞complex hydrolysis temperature.Under this condition，the theoretical maximum inhibition rate of enzymatic hydrolysate was44.87%，and the actual inhibition rate was 44.48%，the difference from the response surface optimization prediction result was 0.8%，compared with the single enzymatic hydrolysate，the inhibition rate was 24.28%，it shows that the effect of complex enzymatic hydrolysis was significantly improved.At this time，the degree of hydrolysis of the enzymatic hydrolysate was 32.64%，and the protein concentration was 0.45 mg/mL.After further separation and purification，the enzymatic hydrolysate was 78.56%，which was 77.61%higher than that of the en－zyme preparation before purification.The antibacterial effect was significantly improved after purification.The results show that the squid protein antibacterial peptide prepared by the complex enzyme method has good antibacterial effect.This technology has certain theoretical significance for further research and application of lowvalue squid preparation antibacterial peptide products.