中华鳖酶解产物的理化性质及促成骨细胞增殖活性分析

本文使用碱性蛋白酶、胃蛋白酶和胰蛋白酶水解中华鳖骨和肉、壳和裙边制备中华鳖功能性肽，并对其主要构象变化、粒度等理化性质进行分析。同时通过测定各酶解物对MC3T3-E1细胞增殖情况的影响对其潜在的骨密度调节活性进行评价。结果显示，中华鳖壳和裙边干粉蛋白含量为54.18%，骨和肉干粉蛋白质含量为80.38%。酶解2 h后，碱性蛋白酶和胰蛋白酶的酶解产物分子量集中分布于1000?Da以下，胃蛋白酶酶解产物分子量分布较为均匀。酶解产物的结构以无规则卷曲为主，粒径集中分布于20和200 nm区域，说明酶解较为充分。酶解产物中赖氨酸、缬氨酸、苯丙氨酸、亮氨酸和苏氨酸等必需氨基酸的含量较高，非必需氨基酸中甘氨酸、丙氨酸和脯氨酸的含量相对较高。其中，胃蛋白酶酶解中华鳖骨和肉后必需氨基酸的总量达39.81%。细胞实验结果显示，六种酶解产物均有一定的促成骨细胞增殖活性，壳和裙边酶解物的增殖活性强于骨肉酶解物，其中胃蛋白酶酶解的壳和裙边产物活性最强，说明中华鳖酶解产物具有潜在的骨密度调节活性。本研究对于中华鳖等产品的深加工提供了可行性分析和理论参考。

Analysis of the Physicochemical Properties and Osteocyte Proliferative Activity of the Enzymatic Hydrolysis Products of Chinese Soft-shelled Turtle

Alkaline protease, pepsin and trypsin were used to hydrolyze the bones and meat, shell and skirt of Chinese soft-shelled turtle to prepare functional peptides, and their physical and chemical properties such as main conformational changes and particle size were analyzed. The potential bone mineral density-regulating activity of each enzymatic hydrolyzate was evaluated by measuring the effect on proliferation of MC3T3-E1 cells. The results showed that the protein content of Chinese soft-shelled turtle shell and skirt dry powder was 54.18%, and the protein content of bone and meat dry powder was 80.38%. After 2 h of enzymatic hydrolysis, the molecular weights of alkaline protease and trypsin hydrolyzed products were concentrated below 1000 Da, and the molecular weight distribution of pepsin hydrolyzed products was relatively uniform. The structure of the enzymatic hydrolysis products was dominated by random coil, and the particle sizes were concentrated in the 20 and 200 nm regions, indicating that the enzymatic hydrolysis was sufficient. The content of essential amino acids such as lysine, valine, phenylalanine, leucine and threonine in the enzymatic hydrolysis products was relatively high, and the content of glycine, alanine and proline in non-essential amino acids was relatively high. Among them, the total amount of essential amino acids after enzymatic hydrolysis of Chinese soft-shelled turtle bones and meat by pepsin reached 39.81%. The results of cell experiments showed that the six enzymatic hydrolysates had a certain pro-osteogenic activity, and the proliferative activities of the enzymatic hydrolysates of shell and skirt were stronger than those of bone meat, and the activity of the shell and skirt products of pepsin enzymatic hydrolysis was the best. The above data showed that the enzymatic hydrolysate of Chinese soft-shelled turtle had potential bone density regulation activity. This research provides feasibility analysis and theoretical reference for deep processing of Chinese soft-shelled turtle products.