基于酶解-膜分离集成的胶原蛋白肽分子量控制技术

发布时间：2018-04-29 04:01

本文选题：胶原蛋白肽 + 酶解　；参考：《山东农业大学》2017年硕士论文

【摘要】：胶原蛋白肽是以动物胶原为原料通过酶法、酸碱法或热降解法制备而成的小分子多肽混合物,在功能食品、美容护肤品、药物控释材料和诊断辅料等领域应用广泛。胶原蛋白肽有着抗氧化、防治骨质疏松和抑制血管紧张素转化酶(ACE)活性等许多优秀的生物活性,因此胶原蛋白肽的制备成为近年的研究热点,具有重要的意义。对于传统的胶原蛋白制备技术而言,其存在分子量范围宽、工艺条件不完善和降解过程不可控等缺点存在。本研究以胶原蛋白的不同蛋白酶的传统酶解反应过程研究为基础,考察了基于超滤膜分离技术对于胶原蛋白肽的提取工艺,采用酶解-膜分离集成工艺提取分子量均一可控的的胶原蛋白肽,并采用高效凝胶色谱对制备的胶原蛋白肽分子量范围进行检测,以牛明胶为原料探索了分子量均一多肽的制备方法。具体内容如下:1.酶解工艺研究:实验采用牛明胶为原料,利用不同蛋白酶对其进行酶解,探究其酶解动力学,对酶解速率及酶解程度进行探索,经实验所得,经酶解60min后10k Da以下的小分子多肽所占比例分别为:碱性蛋白酶44.7%、胰蛋白酶39.8%、木瓜蛋白酶35.7%和中性蛋白酶15.3%。2.胶原多肽膜分离研究:膜分离过程中多肽的透膜行为是进行酶解-膜分离集成工艺的基础,实验研究了明胶酶解产物超滤处理过程中溶液体积、循环流量等对不同分子量多肽跨膜行为的影响。实验对1.2L、600m L、300m L的明胶溶液进行比较,60min内3k Da左右的多肽组分变化量分别为分别为7.84%、16.99%、21.05%。3.酶解-膜分离集成过程研究:酶解膜分离集成过程采用连续酶解反应,在酶解的过程中通过中空纤维膜进行多肽过滤。实验采用了碱性蛋白酶酶解,使用300m L明胶溶液进行循环反应。在特定条件下经3k Da中空纤维膜的酶解-膜分离集成反应60min后,相对比传统酶解反应酶解速率提高了15%,所得多肽组分分子量集中在1700与600Da左右。
[Abstract]:Collagen peptide is a small polypeptide mixture prepared from animal collagen by enzymatic, acid-base or thermal degradation. It is widely used in functional foods, cosmetic products, drug controlled release materials and diagnostic excipients. Collagen peptide has many excellent biological activities such as antioxidation, prevention and treatment of osteoporosis and inhibition of angiotensin converting enzyme (ACE) activity. Therefore, the preparation of collagen peptide has become a hot topic in recent years and has important significance. For the traditional collagen preparation technology, there are some disadvantages such as wide molecular weight range, imperfect process conditions and uncontrollable degradation process. Based on the traditional enzymatic hydrolysis process of different proteases of collagen, the extraction process of collagen peptides based on ultrafiltration membrane separation was investigated. Collagen peptides with uniform molecular weight and controllable molecular weight were extracted by the integrated process of enzymatic hydrolysis and membrane separation, and the molecular weight range of the prepared collagen peptides was determined by high performance gel chromatography. The preparation method of homogenous polypeptide with molecular weight was studied by using bovine gelatin as raw material. The details are as follows: 1. Study on enzymatic hydrolysis technology: bovine gelatin was used as raw material to hydrolyze it with different proteases, and the kinetics of enzymatic hydrolysis was studied. The rate and degree of enzymatic hydrolysis were explored. The proportion of small polypeptides less than 10 kDa after enzymatic hydrolysis of 60min were: alkaline protease 44.7, trypsin 39.8, papain 35.7% and neutral protease 15.30.2. Study on the Separation of Collagen Peptide membrane: the permeation behavior of polypeptide in the process of membrane separation is the basis of the integrated process of enzymatic hydrolysis and membrane separation. The volume of solution during ultrafiltration treatment of gelatin hydrolysate was studied experimentally. Effects of cyclic flow rate on the transmembrane behavior of polypeptides with different molecular weight. The changes of polypeptide components about 3kDa in 1.2L ~ 600m / L ~ (300ml) gelatin solution in 60 min were 7.84 ~ 16.995% and 21.05% 路3 ~ (-1), respectively. Studies on the Integrated process of Enzymatic Hydrolysis and membrane Separation: the integrated process of enzymatic membrane separation was carried out by continuous enzymatic hydrolysis and polypeptide filtration was carried out by hollow fiber membrane in the process of enzymatic hydrolysis. The enzymatic hydrolysis of alkaline protease was carried out, and the reaction was carried out by using 300 mL gelatin solution. Under certain conditions, the enzymatic hydrolysis and membrane separation integrated reaction of 60min with 3kDa hollow fiber membrane increased the hydrolysis rate by 15%, and the molecular weight of the peptides was about 1700 and 600Da.
【学位授予单位】：山东农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：TQ936.16

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