海鲈鱼胶原蛋白肽制备工艺优化及其促进成纤维细胞增殖活性比较

Optimization of Preparation Process of Collagen Peptides from Byproducts of Sea Bass and Comparison of the Activity of Promoting Fibroblasts Proliferation

目的:建立海鲈鱼胶原蛋白肽酶法制备的工艺条件,明确不同酶法制备的海鲈鱼胶原蛋白肽促进成纤维细胞增殖的活性。方法:采用海鲈鱼加工副产物为原料,以胶原蛋白肽提取率为优化指标,通过单因素结合响应面优化酶法制备海鲈鱼胶原蛋白肽工艺,建立碱性蛋白酶与底物浓度比、温度、时间和p H值的四因子回归模型;以成纤维细胞NIH/3T3增殖率为指标,比较碱性蛋白酶、胰蛋白酶和木瓜蛋白酶制备的胶原蛋白肽促进NIH/3T3细胞的增殖活性。结果:碱性蛋白酶制备鲈鱼胶原蛋白肽的最佳酶解工艺条件为:碱性蛋白酶与底物浓度比2.33%、温度47℃、时间2.5 h和p H值9.0,在此条件下鲈鱼胶原蛋白肽提取率为48.65%;各因子对胶原蛋白肽提取率的影响顺序为时间>E/S>温度>p H值。通过比较发现碱性蛋白酶制备的胶原蛋白肽促进NIH/3T3细胞增殖的活性最强。结论:通过碱性蛋白酶酶解海鲈鱼加工副产物,可制备具有促进成纤维细胞增殖活性的胶原蛋白肽,为开发创伤患者专用临床营养品提供胶原蛋白肽配料。

Objective： to establish an optimal enzymatic process of peptides from sea bass collagen（SBCP） and clarify the activity of peptides prepared by different enzmes to promote fibroblast proliferation. Method ： sea bass processing by-products were used as raw material. The conditions of enzymatic preparation SBCP process were optimized through the single factor experiment with Box-Benhnken central composite experimental design based on the indicators of SBCP extraction rate. The regression model of four factors included the enzyme and substrate concentration ratio（E/S）, temperature, time and p H was established. The activity of peptide prepared by different enzymes to promote fibroblasts NIH/3T3 proliferation were compared. Results： the preparation process of SBCP by alkaline protease was optimized, the optimum conditions were E/S 2.33%, temperature 48.65 ℃, time 2.5 h and p H 9.0. The SBCP extraction rate was 48.65% under the optimum condition. These four factors which impacted on SBCP extraction rate in descending order were time, E/S,temperature and p H. The activity of peptide prepared by alkaline protease was better than that of trypsin and papain.Conclusion： the SBCP to promote fibroblast proliferation can be obtained by hydrolysis of sea bass processing by-products using alkaline protease. This study could provide a theoretical guide for the development of collagen peptides ingredients in clinical nutrition.