基于食品蛋白—多配体复合物的活性成分载体的研究

It has been recently suggested that ligand-binding proteins could be potentially used as carriers or components of complex carrier systems for protection and delivery of bioactive components. To clarify the mechanism of protein-ligand interactions is of utmost importance for the development of carrier systems based on proteins. Most of the studies on protein-ligand interactions have focused on the formation and dissociation of the complexes of single ligand with different proteins. However, a single component may usually not meet the requirements in the real-world applications. Nowadays more and more proteins with multiple ligand-binding sites are being reported, which, together with interactions between different proteins, makes it possible to prepare protein-multiligand complexes. In this study, beta-lactoglobulin, casein and their mixtures will be used as model proteins to interact respectively with hydrophobic alfa-tocopherol, amphiphilic resveratrol and hydrophilic folic acid to reveal the mechanism of a single ligand binding to different proteins. On this basis, protein-multiligand complexes will be prepared by binding of multiple ligands simultaneously to the same proteins and influence of the complexes on the ligand hydrosolubility and stability will be analyzed, to discuss the characteristics of the complexes as carriers for active compounds. Finally, the complex nanoparticles will be prepared by interacting ligand-bound proteins with polysaccharides. We will analyze comprehensively influence of the protein-ligand-polysaccharide nanoparticles on the accessibility of the ligands to media and the ligand solubility and stability, in comparison with that of the protein-ligand complexes, to discuss the potential of the nanoparticles used as effective carriers of environmental stimuli-sensitive component payloads. The data gathered from the project could provide more theoretical and scientific basis for further study on the binding mode and interaction mechanism of ligands on proteins, and should help guide the development of stabilization technology based on proteins for the fortification of food and functional food with bioactive components.

近年来，蛋白质和配体复合物已被建议可作为活性成分的载体或复杂载体体系的组成部分。阐明它们的结合特性是基于蛋白质的载体系统开发的关键。目前，关于蛋白质和配体的研究多集中在蛋白质-单配体复合物。然而，单一成分通常不能满足实际应用的需要。随着具有多个配体结合位点的蛋白质不断被发现和不同蛋白质间相互作用的广泛研究，使得制备蛋白质-多配体复合物成为可能。本研究将选择疏水性维生素E、两亲性白藜芦醇和亲水性叶酸作为模型配体，乳球蛋白、酪蛋白和两者混合物作为模型蛋白，在对蛋白质和配体结合机制研究的基础上，制备可同时结合不同配体的蛋白质-多配体复合物，揭示复合物作为载体的优缺点；然后，通过蛋白质与多糖再组装，制备基于蛋白质-配体复合物的多功能性纳米载体。本研究结果将为蛋白质和配体作用模型和机理提供更多科学理论依据，将对基于食品蛋白的稳定化技术和活性成分在食品和保健食品领域应用具有重要的指导作用和现实意义。

由于活性成分的环境敏感性和低溶解度，随着具有多种有益健康功能的食品和功能食品的研发，使得制备可同时包埋不同活性成分的可食用载体体系成为必然。本项目利用蛋白质的配体结合特性，选择疏水性维生素E和维生素A、两亲性白藜芦醇、亲水性叶酸和茶多酚作为活性成分模型，在对蛋白质-单配体复合物、蛋白质和活性成分作用机制研究基础上，成功制备了蛋白质-多配体复合物和蛋白质-配体-多糖复合粒子；活性成分的加入顺序影响复合物和复合粒子的形成，如beta-乳球蛋白-三配体复合物的两种最优加入顺序为：1）白藜芦醇/维生素E/叶酸，2）维生素E/叶酸/白藜芦醇；蛋白质复合物的形成改善了维生素E、维生素A、叶酸的稳定性，但是却加速白藜芦醇的降解，而果胶复合粒子则可以延缓和抑制beta-乳球蛋白对白藜芦醇稳定性的影响。研究结果表明了基于食品蛋白的非乳状液型多活性成分载体粒子开发的可行性，为基于蛋白质的稳定化技术提供了新的思路，对多活性成分在食品工业等领域应用具有重要的指导作用和现实意义。本项目研究成果已发表论文论著7篇。