伊卡瑞酮是淫羊藿中黄酮苷类的无糖苷基成分。在临床试验中，它在肝细胞癌治疗方面表现良好。然而，淫羊藿中自然存在的伊卡瑞酮含量很低。目前，伊卡瑞酮主要是通过α-L-鼠李糖苷酶和β-葡萄糖苷酶在两步催化过程中由黄酮苷类制备的。然而，一锅式伊卡瑞酮生产需要报告的酶被固定或者双功能酶来水解底物以及长时间的反应时间，这导致了复杂的操作和高成本。为了提高生产效率并降低成本，我们研究了α-L-鼠李糖苷酶SPRHA2和β-葡萄糖苷酶PBGL，直接将淫羊藿苷水解为伊卡瑞酮的一锅式方法，并开发了整细胞催化的高效伊卡瑞酮生产方法。SPRHA2和PBGL分别在大肠杆菌中表达。通过SPRHA2和PBGL的协同催化实现了一锅式的伊卡瑞酮生产。此外，我们还开发了一种整细胞催化的淫羊藿苷水解方法。在优化的反应条件下，在整细胞催化条件下，SPRHA2和PBGL的混合物将200 g/L的淫羊藿苷转化为103.69 g/L的伊卡瑞酮（收率为95.23%）需要4小时。为了进一步提高生产效率和简化操作，我们还构建了共表达SPRHA2和PBGL的重组大肠杆菌株。整细胞催化系统还能高效地水解淫羊藿提取物。与以往关于伊卡瑞酮生产的报告相比，此研究中整细胞催化展示了更高效的伊卡瑞酮生产能力。这项研究为未来工业生产伊卡瑞酮提供了有希望的方法。©2023. 作者。

Icaritin is an aglycone of flavonoid glycosides from Herba Epimedii. It has good performance in the treatment of hepatocellular carcinoma in clinical trials. However, the natural icaritin content of Herba Epimedii is very low. At present, the icaritin is mainly prepared from flavonoid glycosides by α-L-rhamnosidases and β-glucosidases in two-step catalysis process. However, one-pot icaritin production required reported enzymes to be immobilized or bifunctional enzymes to hydrolyze substrate with long reaction time, which caused complicated operations and high costs. To improve the production efficiency and reduce costs, we explored α-L-rhamnosidase SPRHA2 and β-glucosidase PBGL to directly hydrolyze icariin to icaritin in one-pot, and developed the whole-cell catalytic method for efficient icaritin production.The SPRHA2 and PBGL were expressed in Escherichia coli, respectively. One-pot production of icaritin was achieved by co-catalysis of SPRHA2 and PBGL. Moreover, whole-cell catalysis was developed for icariin hydrolysis. The mixture of SPRHA2 cells and PBGL cells transformed 200 g/L icariin into 103.69 g/L icaritin (yield 95.23%) in 4 h in whole-cell catalysis under the optimized reaction conditions. In order to further increase the production efficiency and simplify operations, we also constructed recombinant E. coli strains that co-expressed SPRHA2 and PBGL. Crude icariin extracts were also efficiently hydrolyzed by the whole-cell catalytic system.Compared to previous reports on icaritin production, in this study, whole-cell catalysis showed higher production efficiency of icaritin. This study provides promising approach for industrial production of icaritin in the future.© 2023. The Author(s).