固相法合成丙型肝炎疫苗多肽工艺研究

发布时间：2018-03-28 23:31

本文选题：固相法　切入点：丙型肝炎疫苗多肽　出处：《西北大学》2007年硕士论文

【摘要】： 本文以工业化生产为目的，研究了用Fmoc固相法人工合成丙型肝炎疫苗多肽(以下简称疫苗多肽)的生产工艺，比较了Wang树脂和二氯三苯甲基树脂(以下简称二氯树脂)在固相合成中的的优势和劣势，以及粗产物肽和目标肽的产率和纯度；创造性的引入N~α-Dmb基团，提高了困难氨基酸Val的连接率；优化了脱保护的策略以及切割试剂的选择；通过质谱鉴定分析所得目标多肽的分子量，用RP-HPLC进行粗肽的分析和纯化，系统完整的建立了整套合成纯化工艺路线及相关的工艺参数，为后续的工业化生产奠定了坚实的基础。采用该合成纯化工艺路线，以二氯树脂为载体，合成的粗肽产率达到了90.76％，纯化后的目标肽产率为63.02％纯度达到了98.5％，并实现了18克每批的合成量。二氯树脂以其高效率、低成本、产品的高纯度完全符合工业化生产疫苗多肽的各项经济技术指标。论文包括以下四个部分： 1文献综述部分，简述了丙型肝炎疫苗的研究现状，详细介绍了多肽固相合成技术中的合成方法、分离纯化与鉴定技术，最后概述了多肽类药物的应用情况与研究展望。 2采用Fmoc固相法人工合成了疫苗多肽，比较了两种树脂在合成过程中的异同点，，对合成过程中的几个重要步骤和难点进行了优化和改进，如针对困难氨基酸Val的连接，设计了引入N~α-Dmb基团的独特合成策略，提高了连接率及切割效率，确定了合成工艺中相关的各项工艺技术参数。最后用二氯树脂做为载体进行了18克每批的疫苗多肽合成。 3用RP-HPLC进行粗肽的分析和纯化，根据疫苗多肽的理化性质采用乙腈—水的流动相体系，并确定了RP-HPLC分析和纯化的条件。 4总结了实验结果，确定了疫苗多肽合成、纯化工艺路线及相关的工艺技术参数。
[Abstract]:In this paper, the production process of hepatitis C vaccine polypeptide (hereinafter referred to as hepatitis C vaccine polypeptide) was studied by using Fmoc solid phase method for the purpose of industrial production. The advantages and disadvantages of Wang resin and dichlorotriphenylmethyl resin (hereinafter referred to as dichloromethyl resin) in solid phase synthesis, the yield and purity of crude product peptide and target peptide were compared, and the N- 伪 -Dmb group was introduced creatively. The binding rate of difficult amino acid Val was improved, the deprotection strategy and the selection of cutting reagent were optimized, the molecular weight of the target peptide was identified by mass spectrometry, and the crude peptide was analyzed and purified by RP-HPLC. A complete set of synthetic and purification process routes and related process parameters were established, which laid a solid foundation for subsequent industrial production. The synthesis and purification process route was adopted, and dichlororesin was used as carrier. The crude peptide yield reached 90.76%, the purity of the purified peptide was 63.02%, and the yield of 18 g / batch of dichlororesin was achieved by its high efficiency and low cost. The high purity of the product fully accords with the economic and technical indexes of the industrialized production of vaccine peptides. The thesis includes the following four parts:. 1 in the part of literature review, the research status of hepatitis C vaccine was briefly introduced, the synthesis methods, separation, purification and identification of peptide solid phase synthesis technology were introduced in detail. Finally, the application and prospect of polypeptide drugs were summarized. The peptide of vaccine was synthesized by Fmoc solid phase method. The similarities and differences between the two resins in the process of synthesis were compared. Several important steps and difficulties in the process of synthesis were optimized and improved, such as the connection of difficult amino acid Val. The unique synthesis strategy of introducing Na--Dmb group was designed, the connection rate and cutting efficiency were improved, and the relevant technological parameters were determined. Finally, 18 grams of vaccine polypeptides were synthesized with dichlororesin as carrier. 3The crude peptides were analyzed and purified by RP-HPLC. The mobile phase system of acetonitrile-water was used according to the physicochemical properties of vaccine peptides, and the conditions for RP-HPLC analysis and purification were determined. 4 the experimental results were summarized, and the synthesis, purification process and related technological parameters of vaccine polypeptide were determined.
【学位授予单位】：西北大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R392

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