四种深色农产品抗氧化协同作用及其分子生物学协同机制的研究

In recent years, increasing attention has been paid to plants and foods rich in natural phytochemicals, particularly colored agricultural products, which have been associated with reduced risks of a number of chronic diseases in human. There is synergistic effect among different phytochemicals. So far, the mechanism of synergistic effect was explained by chemical structure and electric potential other than the theory of molecular biology. The main objective of this study is therefore to identify and evaluate the in vitro, cell-based and in vivo antioxidant activities and the synergistic effects of several standards (lycopene, β-carotene, petunidin and delphinidin) and colored agricultural products (tomato, carrot, purple potato and eggplant). The groups with great synergistic effects are suppose to picked up using effective equivalent diagram method, which is much more effective and scientific than others. A cell culture-based model system, using cultured rat cardiomyocyte H9c2 cells exogenously exposed to hydrogen peroxide (H2O2) will also be used to evaluate the protective effect of the groups in PI3K/Akt、NF-κB and Caspase signal paths. The mechanisms of the synergistic effects in the antioxidant activities are explained by the changes of enzyme/protein in these signal paths. Results from this study will aid the evaluation of synergistic effects in different agriculture products and will provide insight into the mechanisms of the synergistic effects in such phytochemicals.

农产品中富含的植物化学物具有抗氧化等多种有益于人体健康的功效，而且不同的植物化学物之间存在着抗氧化协同作用。目前国内外有限的研究都是从化学结构或电位角度解释抗氧化物之间的协同效应，评价方法存在缺陷，而且没有从生物体内抗氧化协同的分子生物学水平进行探讨。本项目拟以深色农产品番茄、胡萝卜、紫薯和茄子中主要的天然植物化学物番茄红素、β-胡萝卜素、牵牛花色素和飞燕草素为研究对象，分别比较标准品和农产品提取物两两协同前后的体外、细胞内和体内抗氧化能力，采用等效图数学评价方法，从中准确筛选出具有抗氧化协同作用的组合。并采用H2O2诱导H9c2细胞产生损伤，研究这些组合协同前后对细胞损伤过程中PI3K/Akt、NF-κB和Caspase信号传导途径上酶/蛋白表达的影响。通过准确评价农产品的抗氧化协同作用，找出与协同效果有关的信号通路，阐明增效作用与细胞内酶/蛋白变化的关系，从本质上解释抗氧化协同机制。

农产品中富含的植物化学物之间具有抗氧化协同作用。本项目以深色农产品番茄、胡萝卜、紫薯和茄子中主要的天然植物化学物番茄红素、β -胡萝卜素、牵牛花色素和飞燕草素为研究对象，采用等效图数学评价方法，分别比较标准品和农产品提取物两两复配前后的抗氧化能力。并从中准确筛选出具有抗氧化协同作用的组合。用H2O2 诱导H9c2 细胞产生损伤，研究这些组合对细胞损伤过程中信号传导途径上酶/蛋白表达的影响，从本质上解释植物化学物之间存在的抗氧化协同现象。获得的主要结果如下：.1、采用超声波辅助提取，并采用响应曲面法（RSM）对茄子和紫薯花色苷提取进行优化，得到最佳工艺条件。优化后，茄子花色苷和紫薯花色苷含量分别为5.18 mg/g DW和0.356 mg/g DW。.2、测定茄子和紫薯花色苷含量与结构鉴定，通过质谱并对其主要成分进行定量研究。茄子和紫薯的主要抗氧化成分为花色苷。番茄中主要含有番茄红素，胡萝卜中主要含有β-胡萝卜素。.3、对番茄、胡萝卜、紫薯和茄子中植物化合物以不同比例组合后的抗氧化相互作用进行评价。结果表明水溶性酚类化合物和脂溶性类胡萝卜素以合适的比例混合后，能不同程度地表现出抗氧化协同效果。.4、对四种标准品以不同比例组合后的抗氧化相互作用进行评价。除飞燕草素和β-胡萝卜素组合在ABTS中表现拮抗作用外，其他组合均有比例表现出抗氧化协同效果。所得结果与农产品提取物的抗氧化相互作用基本一致。动物体内，协同作用能提高机体的抗氧化能力，降低MDA和PCO含量，提高GSH、SOD、GSH-Px 和T-AOC的活力。.5、采用H2O2 诱导H9c2 细胞产生损伤，研究不同农产品或标准品组合协同前后对细胞损伤过程中信号传导途径上酶/蛋白表达的影响。结果表明：含花青素含量高、类胡萝卜素含量低的比例大都表现出细胞内的协同作用，反之则表现出拮抗作用。协同组能保护H2O2 诱导产生的细胞损伤，并显著提高细胞内抗氧化酶包括SOD、CAT、GPH、GSH-Px的表达水平，提高细胞存活率、降低细胞内ROS水平，并提高信号传导途径上p-AKT、NQOH1、Nrf2等的表达水平。