硅基磁性生物印迹材料原位分离/固载柚苷酶的行为与机制研究

As an important food enzyme, naringinase is commonly used for citrus juice debittering and taste improving, which can enhance the production quality. However, there are multi-steps in the process of naringinase separation, and it also produces large amount of salty effluent. In addition, the reuse of naringinase is very hard. To solve these problems, a novel method of simultaneous in-situ separation and immobilization of naringinase is proposed in this project. The magnetic molecular-imprinted biomaterials, with silica-based magnetic cores and core-shell-shell structure, will be designed and synthesized based on the configuration of naringinase. The magnetic adsorbents combined four factors of biomaterial, magnetic separation, silica-based magnetic core and molecular imprinting. Furthermore, based on the characterization of microstructure of the adsorbent, the layer-by-layer assembly mechanisms of the adsorbent will be illustrated, the correlation between binding sites of functional groups, pore structure and adsorption selectivity, immobilized naringinase activity will be analyzed, and the possible synergistic and spatial effects between binding sites of functional groups and the structure of adsorbent will be discussed. It will provide a theoretical foundation and feasible way for the combination of in-situ separation and immobilization of naringinase and its industrial application.

作为一种重要的食品酶，柚苷酶主要用于柚橘类果汁的脱苦和风味改善，进而提高产品品质。针对其分离纯化过程中步骤繁琐复杂、产生大量含盐废水，并且在使用过程中难于重复利用的问题，本项目提出利用磁性生物材料原位分离/固载柚苷酶的新工艺。结合生物材料、磁助分离、硅基磁核和分子印迹四个要素，设计合成绿色高效的基于柚苷酶构型的核-壳-壳结构硅基磁性生物印迹材料。在分离/固载实验中，通过表征吸附剂微观结构性质，阐明材料的层层组装机理，解析材料的官能团结合位点、孔结构与分离选择性和固载酶活力之间的关系，进一步探讨吸附剂官能团结合位点与吸附剂结构可能存在的协同效应和空间效应，为柚苷酶的原位分离耦合固定化技术及产业化应用奠定理论基础并提供可行性途径。

柚苷酶是柚柑果汁加工工业常用的食品酶。本项目针对柚苷酶分离纯化过程中存在的操作步骤复杂、高盐废水难于处理且游离酶难以回收利用等问题，结合生物材料、磁助分离、硅基磁核和分子印迹四个要素，设计合成绿色高效的基于柚苷酶构型的核-壳-壳结构硅基磁性生物印迹材料，并建立了磁助化学分离与分子印迹耦合的柚苷酶分离/固载研究方法。采用多种检测手段进行了材料的表征，验证了官能团在材料上的成功修饰；研究了不同环境条件下材料对发酵液中柚苷酶分离/固载性能的相关规律，进一步分析了固定化柚苷酶的酶活，并探讨了固定化柚苷酶的动力学和热力学过程，为实际发酵液体系中柚苷酶的原位分离/固载提供了基础数据和理论指导。通过本项目研究制备的磁性硅基固定化柚苷酶具备了固定化酶活高、磁场存在时固液快速分离和多次重复使用等性能，通过选择合适的材料制备和环境条件，可以用于实际发酵体系中柚苷酶的快速高效分离/固载，同时也为柚苷酶的产业化应用提供了新的研究思路。研究具有较好的理论研究意义和实际应用价值。