Enzymatic synthesis of α-flavone glucoside via regioselective transglucosylation by amylosucrase from Deinococcus geothermalis

α-Flavone glycosides have beneficial properties for applications in the pharmaceutical, cosmetic, and food industries. However, their chemical syntheses are often limited by a low efficiency or scarcity of substrates. In this study, α-flavone glucosides were enzymatically synthesized by amylosucrase from Deinococcus geothermalis (DGAS) using sucrose and various flavones as a donor for glucosyl units and acceptors, respectively. Luteolin was the most effective acceptor in the transglucosylation reaction using DGAS among nine flavone materials (apigenin, chrysin, 6,7-dihydroxyflavone, homoorientin, 7-hydroxyflavone, isorhoifolin, luteolin, luteolin-3′,7-diglucoside, and orientin). The highest production yield of luteolin glucoside was 86%, with a 7:1 molar ratio of donor to acceptor molecules, in 50 mM Tris-HCl buffer (pH 7) at 37°C for 24 h using 2 U of DGAS. The synthesized luteolin glucoside was identified as luteolin-4′-O-α-D-glucopyranoside with a glucose molecule linked to the C-4′ position on the B-ring of luteolin via an α-glucosidic bond, as determined by 1H and 13C nuclear magnetic resonance. This result clearly confirmed that the glucosylated luteolin was successfully synthesized by DGAS and it can be applied as a functional ingredient. Furthermore, this approach using DGAS has the potential to be utilized for the synthesis of various glucosylated products using different types of polyphenols to enhance their functionalities.α-黄酮糖苷（α-Flavone glycosides）在医药、化妆品及食品工业中具备有益的应用特性。然而其化学合成常受限于反应效率低下或底物来源稀缺。本研究采用来自地热异常球菌（Deinococcus geothermalis）的淀粉蔗糖酶（amylosucrase, DGAS），分别以蔗糖作为葡糖基供体、多种黄酮作为受体，通过酶法合成α-黄酮葡萄糖苷。在九种黄酮类物质——芹菜素（apigenin）、白杨素（chrysin）、6,7-二羟基黄酮（6,7-dihydroxyflavone）、异荭草素（homoorientin）、7-羟基黄酮（7-hydroxyflavone）、异瑞福林（isorhoifolin）、木犀草素（luteolin）、木犀草素-3′,7-二葡萄糖苷（luteolin-3′,7-diglucoside）及牡荆素（orientin）——中，木犀草素是DGAS催化转糖基化反应（transglucosylation reaction）的最有效受体。在50 mM Tris-HCl缓冲液（pH 7）、37℃条件下，使用2 U DGAS反应24小时，当供体与受体摩尔比为7:1时，木犀草素糖苷的最高产率可达86%。通过氢谱（¹H NMR）与碳谱（¹³C NMR）分析，鉴定所得合成产物为木犀草素-4′-O-α-D-吡喃葡萄糖苷，葡萄糖分子通过α-糖苷键连接于木犀草素B环的C-4'位。本结果明确证实，通过DGAS可成功合成糖基化木犀草素，其可作为功能性成分加以应用。此外，该基于DGAS的策略有望通过使用不同种类的多酚类物质合成多种糖基化产物，以增强其功能活性。