DHPM驱动脂质体二次组装机理及其结构与热力学稳定性关系

As a functional components delivery system, liposomes, which are self-assembled and cell-resembled, have been remarkable developed in pharmaceutical biology and food chemistry in recent years. In our previous studies,liposomes encapsulated different hydrophilic and hydrophobic contents were prepared by dynamic high pressure microfluidization (DHPM). The liposomal structures play important role in their physicalchemical properties and functinal properties. However, the relationships between preparation method and the formation mechanism of membrane, and liposomal structures and thermodynamic stability need to be further studied. Thus, the present study focuses on fluorescence labeled blank nanoliposomes formed by second-assembling forced by DHPM. The physicochemical properties of liposomes, including diameter, size distribution, zeta potential, phase-transition temperature, surface tension and relative turbidity, are measured by high-tech approaches such as laser scattering technique. The structure, distance, thickness, integrity and permeability of both knid of liposomal membrane are measured according to morden technology such as electron microscope. The second-assembling model of liposomes forced by DHPM could be established and the mechanism would be clarified, according to the changes of liposomal properties and membrane structures. Besides, the thermodynamic stability such as physical, chemical as well as long-term stability of liposomes before or after treated with DHPM is studied. Based on the multi-factors correlation analysis, the relationship between membrane structure and thermodynamic stability would be revealed. Our study may provide some useful technical guidance for the formation mechanism of liposomal second-assembling and quality control.

自组装形成类似细胞膜的脂质体，作为一种技术前沿的功能成分运载体是生物医药、食品化工等领域的研究热点。课题组前期发现，采用动态高压微射流（DHPM）技术制备各种亲水亲脂性芯材的脂质体结构差异与其理化和功能性质密切相关，但脂质体制备与膜结构的形成机理、结构与热力学稳定性的关系有待深入。为此，本项目采用DHPM驱动荧光标记空白脂质体二次组装成纳米脂质体，借助激光散射等技术测定DHPM处理前后脂质体的物化特性(粒径与分布、表面电荷、相变温度、表面张力和相对浊度)的影响；采用现代显微镜等技术表征脂质体膜的结构特性(层数、间距、厚度、完整性和通透性等)的变化，构建DHPM驱动脂质体二次组装的动力学模型并阐析其机理；研究脂质体的热力学(物理、化学和长期)稳定性变化，结合结构特性差异进行多因素相关性分析，揭示脂质体结构与热力学稳定性的作用关系，为脂质体二次组装理论和结构与性质有效调控等研究提供理论借鉴。

自组装形成类似细胞膜的脂质体，作为一种技术前沿的功能成分运载体是生物医药、食品化工等领域的研究热点。课题组前期发现，采用动态高压微射流（DHPM）技术制备各种亲水亲脂性芯材的脂质体结构差异与其理化和功能性质密切相关，但脂质体制备与膜结构的形成机理、结构与热力学稳定性的关系有待深入。为此，本项目借助激光散射等技术对DHPM处理前后脂质体的物化特性（粒径与分布、表面电荷、相变温度、表面张力和相对浊度）进行研究；采用现代显微镜等技术研究两种脂质体膜的层数、间距、厚度、完整性和通透性等结构特性变化，建立DHPM驱动脂质体二次组装的动力学模型并揭示其机理；研究脂质体的热力学(物理、化学和长期)稳定性变化，结合结构特性差异进行多因素相关性分析，揭示脂质体结构与热力学稳定性的作用关系根据实验结果推导出脂质体形成机理可能的三种模式为：（1）、膨胀-自组装模型；（2）、出芽-分裂模型；（3）、薄片-弯曲-闭合模型。在DHPM的驱动下，脂质体形成碎片，然后发生自装机，自动弯曲闭合，形成新的，颗粒较小的脂质体，即薄片-弯曲-闭合模型。结合DHPM处理后的脂质体结构性质和稳定性结果，阐析了结构与稳定性的关系：（1）制备方法不同影响了脂质体的磷脂排列方向和膜的空间结构；（2）脂质体的存在环境改变脂质体磷脂极性头端与膜表面的角度，从而改变脂质体的表面电荷，进而影响了脂质体的热稳定性。上述工作为脂质体二次组装理论和结构与性质有效调控等研究提供理论借鉴。.此外在项目执行过程中，也开展了一些原计划没有列入的工作：以层层修饰技术在脂质体表面修饰聚合物（海藻酸钠、壳聚糖、果胶等），能提高脂质体的物化稳定性，消化稳定性，生物利用率等；以脂质体为载体，包载茶多酚、表儿茶素没食子酸酯、丁香酚和姜黄素，研究结果表明脂质体包载后能提高多酚的溶解度、理化稳定性和生物活性；制备得到双亲性壳聚糖，双亲性壳聚糖与磷脂形成的杂合脂质有较高的稳定性和细胞摄取率，海藻酸钠修饰的双亲性壳聚糖脂质体具有非常好的消化稳定性。这些工作为突破制约脂质体制备与应用的瓶颈，为脂质体的品质调控提供技术指导。