基于NE-4C细胞模型制备猪脑促增殖肽

Proliferation Peptide Preparation of Porcine Brain Based on NE-4C Cell Model

以新鲜猪脑为原料,分别采用胃蛋白酶、胰蛋白酶及脂肪酶酶解得到猪脑多肽,研究用其培养小鼠神经干细胞NE-4C的条件,分析不同酶解液的促增殖活性。通过高效凝胶渗透色谱法比较猪脑多肽的分子质量分布。采用正交试验优化猪脑多肽提取工艺中胰蛋白酶的添加量、酶解温度、酶解pH值和酶解时间等工艺参数。结果表明:与完全培养基相比,添加5 mg/mL胰蛋白酶酶解物能促进NE-4C细胞增殖。胰蛋白酶酶解液中分子质量500 u以下小多肽占总多肽的72.09%,高于胃蛋白酶及复合酶的,说明胰蛋白酶猪脑酶解产物的促增殖活性可能与其较高比例的低分子质量肽相关。最佳酶解工艺为加酶量25 mg,酶解时间4 h,酶解温度45℃,酶解pH 7.5,在最适条件下,NE-4C促增殖活力为0.161。

Fresh porcine brain was used as raw material to prepare porcine brain polypeptide using enzymatic hydrolysis of pepsin,trypsin and lipase.Culture conditions of NE-4C for porcine brain polypeptide and its proliferation were studied.The molecular weight distribution of porcine brain polypeptide was compared by efficient gel permeation chromatography.Orthogonal test was used to study the effects of trypsin addition amount,enzymolysis temperature,pH value and time on proliferation activity of porcine brain polypeptide.The results showed that proliferative activity of NE-4C was enhanced by 5 mg/mL trypsin hydrolysate.Small molecular weight polypeptides below 500 u trypsin hydrolysate accounted for 72.09%in total peptides,higher than that of pepsin and compound enzyme hydrolysate.Proliferative activity of porcine brain products hydrolyzed by trypsin may be related to its higher proportion of low molecular weight peptides.The optimal enzymatic hydrolysis parameters of porcine brain polypeptide were the enzyme dosage 25 mg,enzymolysis time 4 h,enzymolysis temperature 45℃,pH 7.5.Under optimal conditions,the proliferative activity of NE-4C was 0.161.