糖基化胶原蛋白改性聚己内酯在离子液体中的酶促合成及性能研究

As a kind of implantable medical material, the synthesis of polycaprolactone was primarily through the inorganic salt catalytic. However, the polycaprolactone has some disadvantages, such as high hydrophobicity, low cell affinity, slow biodegradation rate, trigger reject reaction in the body. In this project, the glycosylated collagen which can be used as the cell growth factore,is grafted with ε-caprolactone by lipase catalyzation in ionic liquids system, and the modified polycaprolactone, which has biocompatibility and biodegradability properties, will be prepared. First, the aminocarbonyl reaction mechanism between lactose and collagen in the ionic liquid homogeneous system will be studied. In order to initate the lactone ring-opening polymerization through lipase catalyzation, the modified collagen which structure containing a specific primary hydroxyl will be prepared.Secondly, the project will study the ring-opening polymerization reaction mechanism between the modified collagen and ε-caprolactone by lipase catalyzation in the ionic liquid. The amphiphilic glycosylation collagen that has polycaprolactone-containing side chains will be obtained by the reaction, and its biocompatibility and biodegradability will be detailed studied. To solve the problems of the low molecular weight of the side chain and the low utilization rate of the enzyme-catalyzation, the project will adopt some special methods to control the reaction, such as ionic liquids design, control of water activity, immobilized lipase with pH-dependent autoprecipitating carrier. The project is very important for the synthesis of novel aliphatic polyester material that has the functionality of the end groups.

作为一种可植入体内的医用材料，聚己内酯主要通过无机盐催化合成，疏水性较高，细胞亲和性差，用作支架材料时降解速率较慢，易引发体内排斥。为此，本课题拟利用离子液体作为脂肪酶催化酯化反应的非水相体系，将具有导向作用的细胞生长因子- - 糖基化胶原蛋白与ε-己内酯进行酶催化聚合，制备生物相容性和降解性能较好的改性聚酯。课题将首先研究离子液体均相体系中乳糖与胶原蛋白间的羰氨反应机制，制备结构中含有酶催化特异位点-伯羟基的改性蛋白，为引发内酯开环聚合做准备。其次，研究离子液体非水相环境中由糖基化改性蛋白引发的脂肪酶催化ε-己内酯本体开环聚合机制，制备含聚己内酯侧链的两亲性糖基化胶原蛋白材料，研究其生物相容性和降解性。为解决侧链分子量不高、酶催化利用率低等问题，课题将采用离子液体再设计、水活度控制、pH响应型载体固定化脂肪酶等措施加强反应调控。该课题对于合成具有端基功能的新型脂肪族聚酯材料具有重要借鉴意义。

聚己内酯是一种重要的生物医用组织材料，其合成条件比较苛刻，容易产生有机金属催化剂残留，而且合成的聚己内酯疏水性过强，导致细胞亲和性不佳。为此，本项目采用离子液体非水相体系中的酶促开环聚合技术，通过引发剂方法，将具有生物活性的还原糖及其改性糖蛋白接枝到聚己内酯末端，改善其细胞亲和性能。. 首先，本项目以小分子还原糖和Ɛ-聚赖氨酸为底物研究了离子液体中的美拉德反应，建立了一种以离子液体为溶剂的美拉德反应方法。研究表明：糖以及聚赖氨酸等物质能高效溶解于[Bmim]Cl离子液体，形成均一体系，加热时，还原糖的羰基会优先与赖氨酸的自由氨基缩合生成希夫碱，继而重排形成酮糖基胺无色物质，继续反应会形成棕色大分子产物。经过美拉德反应制得的糖化胶原蛋白，由于多羟基糖分子的引入，其肽链变得更为舒展，同时空间位阻增大，其溶解稳定性和乳化性能都得到了提升。. 其次，本项目对离子液体中的酶促开环聚合过程进行了研究。结果表明，脂肪酶能够在[Bmim]PF6中成功催化Ɛ-己内酯开环聚合，其主要产物为聚己内酯的线性聚合物，其末端分别为伯羟基和羧基，数均分子量介于4000-7000之间。温度、时间、水活度和给酶量对ε-己内酯开环聚合的单体转化率和产物分子量都有很大的影响。优化后的反应条件为：温度60℃、反应时间为24-48h，体系水活度0.33，给酶量为50 mg/g己内酯。. 最后，分别以乳糖及糖化胶原蛋白为引发剂，通过脂肪酶催化制备了末端官能化改性聚己内酯并研究了其产物性能。红外和核磁显示，在[Bmim]PF6离子液体中，脂肪酶能够成功催化两种产物接枝聚己内酯，实现其末端官能化改性。改性聚己内酯的亲水性得到改善，但是糖化胶原蛋白的引入则降低了聚己内酯的结晶度，改变了其溶解性，其在二氯甲烷和丙酮中的溶解性明显下降；细胞毒理性测试结果表明NIH/3T3细胞在糖化胶原蛋白改性聚己内酯上有更高的存活率，说明其引入可以改善聚己内酯的细胞亲和性。