双酶偶联一锅法高效合成虎杖苷

Efficient synthesis of polydatin by a two-enzyme coupled with one-pot method

摘要:

虎杖是传统中药之一，其主要活性成分虎杖苷具有调节糖代谢、脂代谢、镇咳平喘、抗菌抗炎等药理作用，但传统的虎杖苷制备方法不足以满足当下市场需求。本研究聚焦于绿色高效的虎杖苷制备方法，在已获得的糖基转移酶UGT_BS三联突变体IGW (Y14I/I62G/M315W)基础上，以白藜芦醇为底物，通过双酶偶联一锅法生物催化高效制备虎杖苷，同时实现尿苷二磷酸葡萄糖(uridine diphosphate-glucose,UDPG)的循环再生。对双酶偶联催化体系进行条件优化，在35℃，pH为8.0，IGW:AtSuSy1酶活比3:4，二甲基亚砜(dimethyl sulfoxide,DMSO)体积比5%，尿苷二磷酸(uridine diphosphate,UDP)浓度0.10 mmol/L，蔗糖浓度0.6 mol/L的条件下，1 h内2 mmol/L白藜芦醇的转化率可达80.6%，虎杖苷的产物占比可达90%以上。本研究通过双酶偶联体系实现了UDPG的循环再生，缩短了反应时间，同时探究了分批补料策略，在一锅法偶联反应中24 h虎杖苷产量可达到6.28 g/L。本研究为绿色高效制备虎杖苷提供了新的策略。

Abstract:

Traditional Chinese medicine of Polygonum cuspidatum has been utilized in China for thousands of years. Its primary active compound, polydatin, exhibits a variety of pharmacological effects including the regulation of glucose and lipid metabolism, suppression of cough and asthma, as well as antibacterial and anti-inflammatory properties. However, conventional methods for polydatin production are inadequate to satisfy the market demand. This study aims to explore the green and efficient preparation of polydatin. With resveratrol as the substrate, we efficiently synthesized polydatin by using the triple mutant IGW (Y14I/I62G/M315W) of the glycosyltransferase UGT_BS based on a strategy of two-enzyme coupled with one-pot and realized the recycling of uridine diphosphate-glucose (UDPG). The conditions of the two-enzyme reaction were optimized. Under the conditions of 35 ℃, pH 8.0, IGW:AtSuSy1 activity ratio of 3:4, dimethyl sulfoxide (DMSO) volume fraction of 5%, uridine diphosphate (UDP) concentration of 0.10 mmol/L, and sucrose concentration of 0.6 mol/L, the conversion of 2 mmol/L resveratrol reached 80.6% within 1 h, and the proportion of polydatin was over 90%. This study achieved the recycling of UDPG via a two-enzyme coupling system and shortened the reaction time. At the same time, the fed-batch strategy was adopted, and the yield of polydatin reached 6.28 g/L after 24 h in the one-pot coupling reaction, which provided a new strategy for green and efficient preparation of polydatin.