【作者】 丁兆堂；

【导师】 王秀峰； 沈生荣；

【作者基本信息】 山东农业大学 ， 园艺学， 2005， 博士

【摘要】 绿茶多酚是茶的主要成分,约占总干物质的18%~36%,其主要组成成分是儿茶素类。它清除超氧化物自由基和羟自由基的功效与茶保健和治疗某些疾病的作用机制是密切有关的。其氧化产物茶黄素是多酚物质氧化形成的一类能溶于乙酸乙酯的、具有苯骈卓酚酮的化合物的总称,是茶色素中生物学活性较强的物质,其保健功效、提取和分离技术等受到中外学者的高度重视。在茶黄素的组成、结构、性质、形成机理、分析技术及方法、与红茶品质的关系及茶黄素形成的影响因子和调控等方面,进行了广泛深入的研究和探讨,取得了一系列可喜的进展。实验证明,茶黄素具有高抗癌、抗心脑血管疾病、抗菌、抗病毒等多种活性,其效果明显优于茶多酚。茶黄素被广泛认定为茶叶中活性利用潜力最大的物质之一,而红茶中其含量低、难于实现工厂化生产是当前存在的主要难题。体外酶促氧化制取茶黄素是利用多酚氧化酶的氧化特性定向生产茶黄素。本实验利用纳米碳酸钙作为固定化材料,对茶多酚氧化酶进行固定化,研究了温度、pH值、反应时间、底物浓度对固定化酶活性的影响,确定了纳米碳酸钙多酚氧化酶体外氧化茶多酚的最佳工艺参数为:37。C、pH5.0、氧化10min,纳米碳酸钙固定化多酚氧化酶的高温耐受性及稳定性均较游离酶强,对pH的稳定性也显著增加,并证实纳米碳酸钙固定化多酚氧化酶的活性有显著提高。本实验以红茶中提取的茶黄素粗提物和氧化前的茶多酚为对照,利用HPLC技术分析了绿茶多酚纳米碳酸钙固定化多酚氧化酶氧化产物的茶黄素组成。结果发现,纳米碳酸钙固定化酶体外氧化产物中茶黄素组成以TF2A为主,而茶黄素粗提物中四种茶黄素单体均占一定的比例,说明纳米碳酸钙固定化多酚氧化酶对茶多酚的氧化有一定的定向性。以绿茶多酚、红茶中提取的茶黄素粗提物为对照,利用紫外扫描技术、UV-Vis法及CuSO4-Vc-H2O2-酵母体系和Fe2+-H2O2-酵母体系研究了绿茶多酚纳米固定化多酚氧化酶体外氧化产物的发光强度、颜色稳定性及对自由基的清除能力。结果发现,在不同pH值条件下,茶黄素粗提物和绿茶多酚纳米固定化酶体外氧化产物的紫外吸收光谱的变化趋势相似,碱性增加,吸光度增加;绿茶多酚、茶黄素粗提物及绿茶多酚纳米固定化酶体外氧化产物对DPPH?　自由基均有明更多还原

【Abstract】 Green-tea polyphenols are the main element in tea leaves, constituting 18%-36% of total dry matter, the key component of which is catechins. Their efficiency for cleaning up suporoxid radical and hydroxyl radical is closely related with the mechanism of tea in human health protection and treatment of some diseases. Theaflavins, the oxide of green-tea polyphenols, are formed after oxidation of polyphenols, can dissolve in ethyl acetate and contain benzotropolones,which owns greater bioactivity than other members in tea pigments and their health function, extraction and isolation have been concerned worldwide. Theaflavins have been widely and deeply studied and got a number of encouraging progress in knowledge or practice of their composition, structure, properties, formation mechanism, analyzing technique, relation to black tea quality, factors affecting theaflavin formation, control, etc. Experiments testified that theaflavins own activities to effectively resist bacteria, viruses, cancers, and cardiovascular and cerebrovascular diseases, etc., with the efficiency being over tea polyphenols. Theaflavins have been widely recognized as a matter which possesses the greatest potential of useful activities in tea leaves, but its content is low in black tea, so it is difficult to realize factorial production.Extraction by in vitro enzymatic oxidation is a method for directional production of theaflavins relying upon the oxidation property of polyphenol oxidase. Using nano-CaCO₃ as immobilizing material, the author immobilized the tea polyphenol oxidase , and investigated the effect of temperature, pH value, reaction duration, and substrate concentration on the activity of immobilized enzyme. As a result, a best technology for in vitro oxidation of tea polyphenols was established: the best parameters for oxidation were 37oC, pH 5.0, and 10 min. In comparison with free enzymes, the nano-CaCO₃ immobilized polyphenol oxidase owns higher activity, has greater resistance to high temperature and more significant stability to pH values.Components of theaflavins in the oxide of green-tea polyphenols were analyzed by HPLC, taking the crude theaflavin extracts from black tea and the unoxidated更多还原