含益生因子的静电纺乳铁蛋白结肠靶向控释体系的构建及其作用机制

Lactoferrin is a kind of important functional protein, and has many biological activities. However, its processing stabilityn is poor, in addition, it is easy to be decomposed by proteases presented in stomach and intestine during absorption which leads to its inactivation. To improve lactoferrin’s stability and bioavailability, the colon-specific nanofilm containing lactoferrin and prebiotics will be prepared by electrospinning in this study. The influence rule of electrospinning conditions on the lactoferrin molecular structure and its bioactivity will be explored. The release kinetics of lactoferrin will be investigated by establishing corresponding methmatic model, by which, the colon-specific release mechanism of lactoferrin will be clarified. The in vitro and in vivo experiments will be carried out, and the influence rule of the prebiotics on the structure of colon bacteria will be studied by the methods of megagenomics and real-time PCR. Combined the variation of lactoferrin absorption rate, the mechanism regarding the control of the structure of colon batearia by prebiotics and its influence on lactoferrin absorption will be elucidated. This study will provide the theoretical base and scientific methods for the colon-targeted delivery and utility of lactoferrin, in addition, it will also offer the reference in developing electrospun colon-specific controlled release system of other similar functional proteins.

乳铁蛋白是一种重要的功能蛋白，具有多种生理活性。然而，乳铁蛋白的加工稳定性差，且消化吸收时易被胃和小肠中存在的蛋白酶分解而失活。为提高乳铁蛋白的稳定性及生物利用度，本项目采用静电纺丝技术构建负载乳铁蛋白和益生因子的结肠靶向纳米纤维膜，探讨静电纺丝条件对乳铁蛋白分子结构及生物活性的影响规律；通过构建数学模型研究乳铁蛋白的释放动力学，阐明乳铁蛋白结肠靶向释放的机理；对纳米纤维膜进行体外益生实验和体内动物实验，采用宏基因组学及荧光定量PCR等技术研究益生因子对结肠微生物菌群结构的影响规律，结合乳铁蛋白在体内吸收率的变化，阐明益生因子调控结肠微生物菌群结构变化与其影响乳铁蛋白吸收之间的机制。本研究将为乳铁蛋白结肠靶向运输及利用奠定理论基础和提供相关的科学方法，并对构建其他类似功能蛋白的静电纺结肠靶向控释体系具有借鉴作用。

功能蛋白是一种重要的食品功能因子,具有多种生理活性。然而,功能蛋白的加工稳定性差,且消化吸收时易被胃和小肠中存在的蛋白酶分解而失活。为提高功能蛋白的稳定性及生物利用度,本项目首先以牛血清白蛋白（BSA）为模型，采用同轴静电纺丝技术构建多糖基酶触发型结肠靶向纳米纤维体系,蛋白质凝胶电泳及圆二色谱分析表明静电纺丝条件对BSA分子结构无显著性影响;通过构建数学模型研究BSA的释放动力学,揭示了BSA的释放是通过一种复杂的机制介导，主要包括扩散、溶胀和溶蚀;以藻蓝蛋白等为阴离子型功能蛋白模型，探究了所构建的多糖基结肠酶触发型纳米纤维体系对藻蓝蛋白等的口服靶向控释规律，发现益生元的加入可以促进藻蓝蛋白在结肠的释放速率，提高了其抑制结肠癌的活性。针对结肠酶触发型纳米纤维体系对阳离子型乳铁蛋白的结肠靶向递送效率不高的问题，本项目首次采用一步法静电纺丝技术构建了亲水性-高疏水性转变的纳米纤维体系，约92.3%的乳铁蛋白实现了结肠靶向释放。同时，揭示了高疏水性纤维对乳铁蛋白的结肠靶向递送机制，即主要是通过结肠菌群对高疏水性纤维的粘附及其产生的代谢产物特别是短链脂肪酸的协同作用所介导的。本研究为乳铁蛋白等功能蛋白结肠靶向运输及利用提供了理论依据，并将推动静电纺丝技术在食品领域的应用。