油菜籽中canolol的形成机制及其抗氧化构效关系研究

Among commercial oilseeds, rapeseed contains the highest amount of phenolic, which had been revealed as an important function in antioxidant, anti-tumor, lowering blood sugar etc. Sinapic acid and sinapine are the most important kinds of free and esterified phenolic, and sinapine is one of the main anti nutritional factor in rapeseed meal. Thermal pretreatment of rapeseed results in the production of canolol, formed by the decarboxylation of sinapic acid. Canolol is a highly potent antioxidant and can increase oil stability.The project team found that the amount of canolol generated was much higher than the loss of sinapic acid, and it was significantly correlated with the loss of sinapic acid and sinapine during roasting. According to this result,we conjectured it was possible that canolol was also formed from sinapine by different pathways. Study on formation mechanism of canolol to make a large number of sinapine converting to canolol is an important strategy for improving the utilization efficiency of rapeseed processing. In this project,on the basis of preliminary studies, rapeseed was treated with microwave, and oil was extracted by press and solvent, and HPLC-LIT-MS and NMR will be applied to investigate the variation of phenolic in rapeseed, and key influencing factors on generation and change of canolol during processing to reveal the chemical basis of formation of canolol. Preparative HPLC will be applied to purify sinapic acid and sinapine to build model systems, the structural changes of sinapine and sinapic acid will be evolute to reveal the formation mechanism of canolol. Otherwise, vitro evaluation platform will be applied to compare antioxidant activity of sinapic acid, sinapine, and canolol to clear their structure-antioxidant and dose-antioxidant relationships. The findings will provide the powerful theory basis and support for improving processing technologies of rapeseed and producing high-value rapeseed based products rich in phenolic for health promotion and disease prevention.

油菜籽富含多酚，芥子酸和芥子碱分别是菜籽中最主要的游离和酯化多酚，其中芥子碱是限制饼粕高效利用的主要抗营养因子之一。对菜籽进行热处理可使芥子酸转化为抗氧化活性更高的canolol，但canolol的增加量远高于芥子酸减少量，表明有未知成分如芥子碱通过不同途径转化成了canolol。探明canolol的形成机制并定向调控使大量芥子碱转化为canolol是提高菜籽加工利用效率的重要途径。本项目拟在前期研究基础上，采用微波技术对油菜籽进行热处理，再进行压榨和浸出制油，阐明菜籽多酚在加工过程中的变化规律与分布规律，明晰canolol形成的物质基础；通过构建芥子酸和芥子碱低水分固相模拟体系，揭示其在微波处理过程中的结构演变和canolol的形成机制；利用体外评价平台，研究菜籽多酚的抗氧化活性，明确其构效和量效关系，为菜籽高值化加工技术发展和菜籽加工产品功能化提供理论基础和科学依据。

大宗商品油料中，油菜籽的多酚含量最高，热处理油菜籽可使其中天然存在的芥子酸脱羧转化为新型菜籽多酚canolol。研究表明，热处理过程中，Canolol的增加量远高于芥子酸的减少量，因此推测，可能有其他芥子酸衍生物转化为canolol。采用高分辨质谱鉴定出5种新的芥子酸衍生物，重点剖析了10个油菜籽品种的游离多酚、酯化多酚、不溶性结合多酚和单宁含量，研究了微波-亚临界萃取过程中，总酚、芥子酸、芥子碱和canolol在油菜籽、菜籽油、压榨饼和脱脂粕中的变化与分布，结果表明，随微波时间的延长，油菜籽和菜籽油中总酚含量增加，芥子酸和芥子碱含量降低，Canolol含量增加；制油过程中，芥子酸从籽到油的转移率不超过3%，芥子碱的转移率不超过0.2%，Canolol的转移率为23%-77%，Canolol成为菜籽油中最主要的多酚。. 根据芥子酸衍生物和canolol在微波前后的含量变化，采用向后法进行回归分析，获得芥子碱、芥子酸马来酸和芥子酰葡萄糖苷关于canolol的线性回归方程，表明除芥子酸外，芥子酸马来酸和芥子酰葡萄糖苷也是canolol形成的重要前体物质。以SiO2为固相载体，构建体外微波模拟体系，研究菜籽多酚浓缩物、高纯度芥子酸、芥子碱和不含芥子酸芥子碱的菜籽多酚在微波处理过程中的热降解程度和产物状况。结果表明，芥子酸的热解速率是芥子碱的3.5倍，新鉴定的5种芥子酸衍生物热解速率是芥子碱的2倍多；从产物结果可以推测，在微波过程中，芥子酸衍生物可能首先热解为芥子酸和对应的配对基团，继续受热时，芥子酸再热脱羧生成canolol。. 采用DPPH自由基清除活力、FRAP总抗氧化能力等方法评价微波前后油菜籽、菜籽油、压榨饼和脱脂粕的抗氧化能力，结果表明，微波处理显著提高了菜籽油的抗氧化能力，其DPPH自由基清除活力、FRAP总抗氧化能力与油中总酚、酚酸和canolol含量呈显著正相关关系。细胞层面的研究表明，Canolol细胞抗氧化能力较强，且细胞毒性较低。构建canolol氧化应激保护模型，Canolol通过调节Bax和Bcl-2及caspase-3的表达而保护细胞免受H2O2诱导的凋亡，Canolol是一种非常有前景的天然抗氧化剂。