黑豆膳食纤维结合的多酚构成、胃肠道释放规律及其抗氧化效应研究

Bound phenolics are believed to be an important part of phenolic composition in plant. Apart from protein, black bean is also rich in phenolic compounds, particularly those bound phenolics binding to dietary fibre. To date, regarding phenolics in black bean, all studies focused on their free phenolic, but ignores the contribution of bound phenolic, and lack of available information on their gastrointestinal tract release behavior, antioxidant effects in vitro and in vivo. Therefore, in the present study, we will explore their bound phenolic characteristic by separating major macromolecular carrier compound, dietary fibre, that probably binds with bound phenolic, as well as by optimizing of enzyme hydrolysis method, and further comparing with traditional approaches analyzing bound phenolic including acid and hydrolysis alkaline hydrolysis; the release behavior of bound phenolic binding to dietary fibre in different phase of gastrointestinal tract will be explored by establishing in vitro digestion model and the human fecal fermentation model, and their relation with antioxidant property in vivo will be investigated by using rat model. The proposed project expects to comprehensive assess the contribution of bound phenolic to total phenolic composition and antioxidant capacity in vivo in black bean from the perspective of dietary fibre-bound phenolic，thus providing scientific evidence for deep understanding of the nutritional value of legume, and proper guidance on nutritious dietary.

结合态多酚是植物中酚类化合物构成不可忽视的重要组成部分。黑豆除富含蛋白质外，还富含多酚，特别是与膳食纤维结合多酚。 目前，国内外关于黑豆酚类化合物有限的研究基本集中在游离态多酚，而忽略了对结合态多酚的探讨，且对结合态酚类化合物胃肠道释放规律、及其对抗氧化效应的贡献研究甚少。本课题拟分离黑豆中主要与结合态多酚联接的膳食纤维大分子，优化酶完全水解方法，并与酸法和碱法进行比较研究结合态多酚，了解黑豆中与膳食纤维结合的多酚构成；建立胃、小肠体外消化模型和人源粪便结肠微生物体外发酵模型，探究黑豆中与膳食纤维结合多酚在胃肠道不同阶段的释放规律，并利用动物实验研究黑豆中与膳食纤维结合多酚在胃肠道释放与体内抗氧化特性的关系。通过本项目的实施，希望从与膳食纤维结合的多酚这个切入点，较系统评估结合态酚类化合物对黑豆酚类化合物构成和发挥体内抗氧化效应的贡献，为科学认识豆类营养价值、指导人们合理膳食提供科学数据。

以黑豆为代表的豆类的营养价值除体现在蛋白质和膳食纤维外，与其富含的天然多酚抗氧化物密切相关。目前，国内外关于豆类酚类化合物的研究基本集中在游离态酚类化合物方面，而忽略了对结合态多酚的探讨，且对不同结合型态酚类化合物胃肠道释放规律、体内外抗氧化特性及其可能的健康效应研究甚少。本课题分离结合态多酚的主要大分子载体物质，并采用酸、碱、酶水解三种分析方法研究结合态多酚构成；建立胃、小肠体外消化模型和人源粪便结肠微生物体外发酵模型，探究黑豆等豆类结合态多酚在胃肠道不同阶段的释放规律，并利用动物实验研究黑豆结合态多酚在胃肠道释放与体内抗氧化特性的关系。通过本项目的实施，希望从结合态多酚这个切入点，较系统评估黑豆中结合态多酚的贡献，为深度发掘植源性食物潜在的营养价值和体内生物活性提供理论支持。具体结果如下：1）利用HPLC-ESI-QTOF-MS定性黑豆结合型酚类组成，共鉴定出88种酚类化合物，包括23种酚酸及其酯类衍生物；8种异黄酮；10种黄酮醇；6种黄烷酮；5种黄烷醇；10种花青素；2）96孔板法测定了可萃取型和结合型酚类提取物(SPE)的总酚TPC和抗氧化活性(DPPH,ABTS，FRAP)。黑豆种皮与子叶中可萃取型总酚含量分别为38.04和10.60mgGAE/gDW。种皮与子叶样品总抗氧化活性；DPPH分别为119.73和21.93，FRAP分别为251.84和19.94μmolFE/gDW，ABTS分别为165.86和8.43μmolTE/gDW。表明黑豆种皮的可萃取型酚类提取物高于子叶，抗氧活性也远远高于子叶，总酚与抗氧化活性间呈现明显的正相关性，说明酚类化合物是最主要的抗氧化活性物质；不同水解方法处理得到的种皮结合型总酚分别为9.33GAE/gDW(酸处理结合酚提取物,A-BPE)，1.63GAE/gDW(酸-碱处理结合酚提取物,A-B-BPE)，8.84GAE/gDW(碱处理结合酚提取物,B-BPE)和3.01GAE/gDW(碱-酸处理结合酚提取物,B-A-BPE)；子叶结合型总酚分别为3.67mgGAE/gDW(A-BPE),1.23mgGAE/gDW(A-B-BPE),4.19mgGAE/gDW(B-BPE)和1.54mgGAE/gDW(B-A-BPE)。这些不同水解方法处理得