微生物转化实现双氢青蒿素C-14位惰性甲基C-H键功能化

Functionalization of Unactivated Methyl C-H Bonds in Position C-14 of Dihydroartemisinin via Microbial Transformation

惰性C-H键,尤其是青蒿素类化合物中C-14位孤立惰性甲基C-H键的选择性羟化在有机化学反应中极为困难。课题组前期研究发现微生物转化可选择性羟化双氢青蒿素C-2和C-9位C-H键。在此基础上,该研究对灰色链霉菌ATCC13273生物转化双氢青蒿素的条件进行考察,结合色谱分离和结晶纯化,分离获得3个转化产物。经LC-MS和NMR鉴定,转化产物分别为2α-羟基双氢青蒿素(1)、9α-羟基双氢青蒿素(2)和14-羟基双氢青蒿素(3),其中化合物3为双氢青蒿素C-14位孤立惰性甲基C-H键羟基化的新化合物。微生物转化动态监测和转化p H条件优化结果显示,9α-羟基双氢青蒿素和14-羟基双氢青蒿素为双氢青蒿素生物转化的主要产物。在转化p H为5.5时,有利于转化产物14-羟基双氢青蒿素的积累,当底物浓度为1.2 g/L时,转化50 h后,14-OHDHA的产量可达267 mg/L。该研究首次通过微生物转化实现双氢青蒿素C-14位甲基C-H键的区域选择性羟化,为双氢青蒿素类化合物的结构修饰提供新的修饰位点,同时也为结构复杂化合物中孤立惰性C-H键的选择性羟化提供新思路。

Dihydroartemisinin is a compound with complex structure and high antimalarial activity.In its structure,the positions C-2,C-9 and C-14 belong to the sp3 unactivated C-H bonds.The functionalization of these C-H bonds is of great significance for the development of dihydroartemisinins lead compounds.But,it is still difficult to selectively hydroxylate of these unactivated C-H bonds,especially for the isolated unactivated methyl C-H bonds in position C-14 by current chemical synthesis.The authors’previous study showed that microbial transformation could selectively hydroxylate C-H bondsin positions C-2 and C-9 of dihydroartemisinin.However,it is unknown whether the isolated unactivated methyl C-H bonds in position C-14 of dihydroartemisinin can be hydroxylatedvia microbial transformation.In this study,the biotransformation process and conditions of dihydroartemisininby Streptomyces griseus ATCC13273 were investigated.The biotransformation products of dihydroartemisinin were separated and purified by chromatography and crystallization.The results are that three compounds were obtained.They were identified as 2α-hydroxydihydroartemisinin(1),9α-hydroxydihydroartemisinin(2)and 14-hydroxydihydroartemisinin(3)by LC-MS and NMR,respectively.Among them,compound 3 was a new compound of hydroxylationdihy droartemisinin in position C-14.The dynamic biotransformation of dihydroartemisinin and optimization of biotransformation condition p H shows that 9α-hydroxydihydroartemisinin and 14-hydroxydihydroartemisinin were the main products.And they were formed at the same time during the biotransformation process.Interestingly when the biotransformation condition p H was 5.5,substrates are preferred the conversion to 14-hydroxydihydroartemisinin,and when the substrate feed concentration is 1.2 g/L,the yield of 14-hydroxydihydroartemisinin could reach 267 mg/L after biotransformation for 50 hours.In conclusion,the region-selective hydroxylation of methyl C-H bondsin position C-14 of dihydroartemisinin is realized for the first time,which provides a new position for the structural modification of dihydroartemisinin.Meanwhile,it provides a new strategy for the selective hydroxylation of isolated unactivate C-H bond in complex compounds.