乳源两亲性抗氧化肽的可控制备及其调控乳液氧化稳定性的构效机制研究

Lipid oxidation usually occurs in oil-in-water emulsions, and the oxidation reaction is often initiated and propagated in the interfacial region that separates the oil phase from the aqueous phase. Due to their amphiphilic nature and antioxidant capacities, amphiphilic antioxidant peptides have great potential to inhibit lipid oxidation in emulsions. However, it is difficult to prepare amphiphilic antioxidant peptides from food protein. In addition, the relationship between the structure of amphiphilic antioxidant peptides as well as their self-assembled complexes in the interfacial layer and the oxidation stability of emulsions is still unknown. Therefore, milk protein will be hydrolyzed by an integrated approach of in silico and in vitro limited hydrolysis to release amphiphilic antioxidant peptides in this project. Subsequently, the amphiphilic peptides will be enriched by the formation and dissociation of aggregates from amphiphilic peptides. Then, the key amphiphilic antioxidant peptides in regulating the oxidation stability of emulsions will be identified by LC-MS/MS and recombination model. In addition, amphiphilic peptides with different structure will be synthesized and the varied conformation of their self-assembled complexes in the interfacial layer will be induced by controlling the environment factors. Finally, the structure-activity relationship of amphiphilic antioxidant peptides on regulating the oxidation stability of emulsions will be elucidated. The successful implementation of this project is of great significance to the development of amphiphilic antioxidant peptides in improving the oxidative stability of emulsion-based food products and also provides new ideas for the controlled release of self-derived amphiphilic antioxidant peptides in complex emulsion-based food systems.

油脂氧化普遍存在于水包油乳液体系，其油水界面是油脂氧化最活跃的部位。两亲性抗氧化肽兼具良好的亲水亲油和抗氧化能力，具有极大潜力用于调控乳液氧化稳定性。但是如何定向制备食源两亲性抗氧化肽是该领域的研究难点。此外，两亲性抗氧化肽的分子结构及其界面组装体结构与乳液氧化稳定性之间的关系亦不明确。为解决上述问题，本项目以乳源蛋白为原料，结合计算机辅助水解及蛋白限制性酶解技术靶向释放目标肽，并通过调控多肽的聚集和解离定向富集两亲性多肽；在此基础上，通过液质联用技术及重组试验鉴定具有关键调控作用的两亲性抗氧化肽，然后合成构建不同分子结构的两亲性抗氧化肽，改变环境因素诱导其在界面上形成不同结构的组装体，探讨二者对乳液氧化稳定性的影响规律，阐明其构效关系。本项目的顺利实施对开发具有显著提升乳液氧化稳定性的两亲性抗氧化肽具有重要的理论指导意义，同时为食品乳液体系中自源性两亲性抗氧化肽的可控释放提供新思路。

油脂氧化普遍存在于水包油乳液体系，其油水界面是油脂氧化最活跃的部位。两亲性抗氧化肽兼具良好的亲水亲油和抗氧化能力，具有极大潜力用于调控乳液氧化稳定性。但是如何定向制备食源两亲性抗氧化肽是该领域的研究难点。此外，两亲性抗氧化肽的分子结构与乳液氧化稳定性之间的关系亦不明确。本项目以酪蛋白为原料，通过探讨酪蛋白抗氧化肽的酶解特性与其调控O/W乳状液氧化稳定性的规律，进一步通过UPLC-MS/MS和生物信息学挖掘在乳液氧化体系中具有关键调控作用的两亲性抗氧化肽，明确提高O/W乳状液稳定性的酪蛋白肽的理化性质和肽段组成特征，解析其构效关系，并通过控制不同浓度的酪蛋白水解物与低或高浓度的表面活性剂调控乳液界面组成，探讨界面竞争性吸附对其乳状液氧化稳定性的影响机制。研究结果表明较宽DH范围的酪蛋白trypsin水解物具有更高的S0、更高的Zeta电位和最小的粒径尺寸，这支持了它具有优异的乳液形成能力。无论Tween 20是否存在,低DH的酪蛋白trypsin水解物都能较好抑制O/W乳液的脂质氧化，肽谱鉴定结果表明其可能含有更多分子量较均一且具有阳离子两亲性的活性肽。在界面调控上，低浓度的Tween 20制备的乳液的物理稳定性和氧化稳定性与CH含量呈剂量依赖关系，表明酪蛋白酶解产物作为助乳化剂促进吸附，进一步抑制脂质氧化。但是，高浓度的Tween 20制备的乳液脂质氧化抑制程度随酪蛋白酶解产物浓度呈先增大后减小的趋势。这可能是由于肽的相互作用导致部分肽从界面解离发生聚集，或者是较多的肽被Tween 20胶束包裹。本项目的顺利实施对开发具有显著提升乳液氧化稳定性的两亲性抗氧化肽具有重要的理论指导意义，同时为食品乳液体系中自源性两亲性抗氧化肽的可控释放提供新思路。