College of Bioengineering, Sichuan University of Science and Engineering, Yibin, 644000, Sichuan, People's Republic of China.
Bioprocess Biosyst Eng. 2022 Jun;45(6):1047-1055. doi: 10.1007/s00449-022-02723-x. Epub 2022 Apr 30.
Theaflavin (TF), a chemical component important in measuring the quality of fermented tea, has a strong natural antioxidant effect and many pharmacological functions. Enzymatic oxidation has become a widely used method for preparing TFs at the current research stage. Using plant exogenous polyphenol oxidase (PPO) to enzymatically synthesize TFs can significantly increase yield and purity. In this study, tea polyphenols were used as the reaction substrate to discuss the optimal synthesis conditions of potato PPO enzymatic synthesis of theaflavins and the main products of enzymatic synthesis of TFs. The optimal enzymatic synthesis conditions were as follows: pH of the reaction system was 5.5, reaction time was 150 min, substrate concentration was 6.0 mg/mL, reaction temperature was 20 °C, and the maximum amount of TFs produced was 651.75 μg/mL. At the same time, high-performance liquid chromatography was used to determine the content of theaflavins and catechins in the sample to be tested, and the dynamic changes and correlations of the main catechins and theaflavins in the optimal enzymatic system were analyzed. The results showed that epicatechin (EC), epigallocatechin (EGC), epicatechin gallate (ECG), and epigallocatechin gallate (EGCG) are all the main substrates synthesis of TFs. The main substrate of TFs and its strongest enzymatic catalytic effect on EGCG make theaflavin-3,3'-digallate (TFDG) the most important synthetic monomer. In this study, theaflavins were synthesized by polyphenol oxidase catalysis, which laid a foundation for industrialization of theaflavins.
茶黄素（TF）是衡量发酵茶品质的重要化学物质成分，具有很强的天然抗氧化作用和多种药理功能。在现阶段的研究中，酶促氧化已成为制备 TF 的一种广泛应用的方法。利用植物外源多酚氧化酶（PPO）酶促合成 TF 可以显著提高产率和纯度。本研究以茶多酚为反应底物，探讨了马铃薯 PPO 酶促合成茶黄素的最佳合成条件及酶促合成 TF 的主要产物。酶促合成的最佳条件为：反应体系 pH 值为 5.5，反应时间为 150 min，底物浓度为 6.0 mg/mL，反应温度为 20°C，TF 最大产量为 651.75 μg/mL。同时，采用高效液相色谱法测定待测样品中茶黄素和儿茶素的含量，分析最佳酶促体系中主要儿茶素和茶黄素的动态变化及相关性。结果表明，表儿茶素（EC）、表没食子儿茶素（EGC）、表儿茶素没食子酸酯（ECG）和表没食子儿茶素没食子酸酯（EGCG）均为 TF 的主要合成底物。TF 及其对 EGCG 最强的酶促催化效应的主要底物使茶黄素-3,3′-二没食子酸酯（TFDG）成为最重要的合成单体。本研究通过多酚氧化酶催化合成茶黄素，为茶黄素的工业化生产奠定了基础。
