EGCG交联乳铁蛋白-多糖纳米载体构建及其对姜黄素递送的协同增效机制

Bioactive components have a variety of health effects, but their low stability and bioavailability limit their use in the food industry. Embedding and delivering the active substance is an effective way to increase its absorption and utilization. However, the available materials and the preparation methods of carriers show certain limitations. Therefore, the construction of food-grade carriers, the relationship between their structures and the sustained-release properties of bioactive compounds have become interesting scientific issues. In this project, curcumin will be hired as the functional bioactive ingredient, and lactoferrin, EGCG and different polysaccharide (sugar beet pectin, chitosan or hyaluronic acid) will be utilized as the wall-materials. The curcumin-loaded ternary complex nanoparticles will be firstly constructed by electrostatic interaction and heat-induced method, and the best forming conditions of nanoparticles will be optimized to find out the key regulation point of the microscopic structure, and then on this basis, curcumin emulsion will be prepared using layer-by-layer self-assembly method, its physicochemical stability and structural characteristics will be analyzed, and in vitro digestion model and Caco-2 cell model will be used to evaluate the sustained release property and bioavailability of curcumin. Furthermore, we will elucidate the relationship between the molecular interaction, the structural composition of the carrier, and the bioavailability of curcumin as well as the synergistic anticancer mechanism of curcumin and EGCG. Through the project, it is expected to theoretically reveal the structure-activity relationship, the stabilization, the release mechanism. In addition, it can provide useful reference for designing a safe delivery carrier with highly bioavailable bioactive ingredient.

生物活性成分具有多种健康功效，但较低的生物利用率限制了其在食品中的应用。将活性成分进行包埋和递送，是提高其吸收的有效途径。然而现有材料和载体制备方法具有一定局限性。因此，构建食品级载体、研究其结构与活性成分缓释特性的关系成为近年来备受关注的科学问题。本项目拟以姜黄素为活性成分，以乳铁蛋白、EGCG和多糖为材料，首先基于静电作用和热诱导方法，构建三元复合物纳米颗粒，确定其最佳形成条件、探明颗粒微结构的调控点；在此基础上，通过层层自组装法构建姜黄素纳米乳液，对其界面结构和稳定性进行分析；并利用体外消化模型和细胞模型评价姜黄素缓释效果和生物利用率，阐明载体组分间相互作用、结构组成和姜黄素生物利用率的内在联系及姜黄素与EGCG协同抗癌机制。通过本项目研究，有望在理论上阐明载体的构效关系、稳态化、控释与作用机制，在应用上为构建安全、高生物利用率和可控释放的活性成分输送载体提供有益参考。

疏水性生物活性成分由于难溶于水、稳定性差以及生物利用率低等问题限制了其在食品工业中的应用，借助递送体系可以有效提高这类活性成分的稳定性和生物利用率。本项目致力于探索并构建蛋白质-多酚-多糖递送体系并系统开展体系中不同组分间相互作用、载体的形成机制、活性成分稳定性及生物利用率研究。主要发现包括：1）采用非酶法（碱处理法和自由基接枝法）或酶促法（漆酶和酪氨酸酶）可诱导乳铁蛋白（LF）与表没食子儿茶素没食子酸酯（EGCG）发生共价结合，该共价复合物（LF-EGCG）显著提高了LF的抗氧化活性和乳化稳定性，并降低了LF对免疫球蛋白E和免疫球蛋白G的结合能力。2）LF与透明质酸（HA）的结合是一个自发放热的过程，主要驱动力是静电吸引和氢键。HA与LF的共价或物理结合均改善了LF的乳化性能和抗氧化活性。3）LF-EGCG-HA三元复合体系在pH2.0时形成宏观水凝胶，在pH3.0时形成微凝胶分散体，在pH4.0-5.0时形成可溶复合物。三元复合物的抗氧化活性在pH5.0时明显高于pH3.0，这是由于微凝胶能够抑制水相中产生的自由基与EGCG反应。4）与LF、LF-EGCG或加热的LF-EGCG二元复合物稳定的单层乳液相比，通过热诱导结合层层自组装法制备的LF-EGCG-多糖三元复合物稳定的双层乳液更有效地阻止了液滴的破裂，抑制了姜黄素的降解，提高了姜黄素的生物利用率和贮藏稳定性。5）LF-EGCG-HA三元复合物纳米颗粒具有较高的姜黄素包埋率（85.77%），其包封的姜黄素具有良好的稳定性和生物可及性。细胞实验证明LF-EGCG-HA三元复合物纳米颗粒促进了姜黄素的Caco-2细胞吸收以及HT-29和CT-26结肠癌细胞系的凋亡，且姜黄素和EGCG在促进HT-29细胞凋亡方面具有协同效应。.该项目完成了项目计划书的主要研究内容，达到了预期研究目标。截至2021年12月，标注本项目基金资助并在食品类高质量期刊上发表的学术论文达15篇，其中3篇为封面论文。代表性研究成果发表在Trends in Food Science and Technology、Journal of Agricultural and Food Chemistry、Food Chemistry、中国食品学报等期刊。该项目的完成为构建安全、高生物利用率和具有抗癌功效的活性物质输送载体提供了有益参考。