双酶法制备具有促皮肤成纤维细胞活性的马鹿茸胶原蛋白肽

Preparation Collagen Peptides from Antlers of Red Deer (Cervus elaphus) with the Activity of Promoting Proliferation of Human Skin Fibroblasts by Bi-enzymatic Hydrolysis

摘要:

胶原蛋白作为细胞外基质的主要成分与细胞的增殖、分化等密切相关。为探究制备具有促进皮肤成纤维细胞增殖功能的马鹿茸胶原蛋白肽的适宜工艺条件，本研究以马鹿茸为原料，采用Alcalase和胰蛋白酶进行酶解，以水解度、多肽得率和正常人皮肤成纤维细胞（NHDF细胞）增殖率为目标值进行单因素实验和正交实验，确定适宜的工艺条件。实验结果表明，马鹿茸胶原蛋白肽适宜制备条件为：先在底物浓度1%，酶底比2%，pH 9.0，温度60 ℃条件下采用Alcalase进行酶解；再在底物浓度1%，酶底比6%，pH 8.0，温度42 ℃条件下采用胰蛋白酶酶解，酶解产物最终水解度为21.7%，多肽得率是86%，细胞增殖率为22.75%。Alcalase-胰蛋白酶双酶分步水解马鹿茸胶原蛋白可获得促进正常人皮肤成纤维细胞增殖率较高的胶原蛋白肽。

Abstract:

As the main component of the extracellular matrix, collagen was closely related to the proliferation and differentiation of the cells. In order to explore the suitable preparation conditions of the collagen peptides from red deer (Cervus elaphus) antler with the function of promoting the proliferation of the skin fibroblasts, the antler was used as the raw material and hydrolyzed by alcalase and trypsin . The degree of hydrolysis (DH), peptides yield and the proliferation rate of normal human dermal fibroblasts (NHDF) were used as the target to perform the single factor experiments and orthogonal experiments and determine the appropriate process conditions. The results showed that the optimal preparation conditions were as follows: alcalase was used for enzymolysis under the conditions of substrate concentration of 1%, enzyme substrate ratio of 2%, pH 9.0 and temperature of 60 ℃, and then trypsin was used for enzymolysis under the conditions of substrate concentration of 1%, enzyme substrate ratio of 6%, pH 8.0 and temperature of 42 ℃. The final hydrolysis degree of the hydrolyzate, the yield of peptides and proliferation rate of cells were 21.7%, 86% and 22.75%, respectively. Collagen peptide with promoting the proliferation rate of NHDF could be obtained from red deer antler by alcalase-trypsin bi-enzyme hydrolysis.