复合酶水解蚕蛹蛋白制备功能性寡肽的工艺研究

Optimization of Enzymatic Hydrolysis Conditions for Preparation of Functional Oligopeptide from Silkworm Pupa Protein

摘要:

为了制备分子量低、肽含量高的蚕蛹功能性寡肽，本文采用单素实验与响应面分析法，对不同蛋白酶及其不同组合、温度、pH值、底物浓度、酶的添加量等因素对蚕蛹分离蛋白水解工艺的影响进行了研究。研究结果表明，多酶复合水解可显著提高蚕蛹蛋白的水解度和寡肽得率，其中胰酶、风味蛋白酶与中性蛋白酶是最佳多酶组合，其最佳水解条件为底物浓度7.33%、酶添加量[E]/[S]3.62%、pH7.38、水解温度54.6 ℃、水解时间6 h。在此条件下，蚕蛹蛋白的水解度高达29.2%，寡肽得率高达为81.14%。酶解后产品数均分子量为665.5 Da，重均分子量为726.9 Da，肽含量高达74.6%，而游离氨基酸含量仅为7.33%，表明复合酶解虽然提高了蚕蛹蛋白的水解度，但主要产物仍然是低分子寡肽，并没有大量生成游离氨基酸。

Abstract:

In order to optimize the production process of functional polypeptides from silkworm pupa protein by enzymatic hydrolysis, the effects of different proteases, temperature, pH, substrate concentration and enzyme amount on the degree of hydrolysis (DH) and the trichloroacetic acid nitrogen solubility index (TCA-NSI) were explored using response surface methodology. It was proved that trypsinase, flavourzyme and neutral protease were the most effective enzymes for the hydrolysis of silkworm pupa protein. The optimal conditions were found to be hydrolysis temperature of 54.6℃, pH 7.38, substrate concentration of 7.33%, protease dosage of 3.62% and hydrolysis time of 6 h. Under these optimized conditions, DH of 29.2 and TCA-NSI of 81.14% were obtained. In the obtained functional polypeptides, the content of peptide reached 74.6%, with a number average molecular weight of 665.5 Da and a weight average molecular weight of 726.9 Da, while the content of free amino acid was only 7.33%, which indicated that the low molecular peptide instead of free amino acids was the main ingredient, although the silkworm pupa protein was enzymatic hydrolysis by depth.