Department of Experimental Biology, Faculty of Science, Palacky University, Olomouc, Czech Republic.
Department of Chemical Biology, Faculty of Science, Palacky University, Olomouc, Czech Republic.
Sci Rep. 2022 Apr 29;12(1):7011. doi: 10.1038/s41598-022-11086-y.
Members of the Viola genus play important roles in traditional Asian herbal medicine. This study investigates the ability of Viola odorata L. extracts to inhibit Na,K-ATPase, an essential animal enzyme responsible for membrane potential maintenance. The root extract of V. odorata strongly inhibited Na,K-ATPase, while leaf and seeds extracts were basically inactive. A UHPLC-QTOF-MS/MS metabolomic approach was used to identify the chemical principle of the root extract's activity, resulting in the detection of 35,292 features. Candidate active compounds were selected by correlating feature area with inhibitory activity in 14 isolated fractions. This yielded a set of 15 candidate compounds, of which 14 were preliminarily identified as procyanidins. Commercially available procyanidins (B1, B2, B3 and C1) were therefore purchased and their ability to inhibit Na,K-ATPase was investigated. Dimeric procyanidins B1, B2 and B3 were found to be inactive, but the trimeric procyanidin C1 strongly inhibited Na,K-ATPase with an IC of 4.5 µM. This newly discovered inhibitor was docked into crystal structures mimicking the NaE∼P·ADP and KE·P states to identify potential interaction sites within Na,K-ATPase. Possible binding mechanisms and the principle responsible for the observed root extract activity are discussed.
堇菜属植物在传统亚洲草药中起着重要作用。本研究调查了紫花堇菜提取物抑制 Na,K-ATPase 的能力，Na,K-ATPase 是一种维持膜电位的重要动物酶。紫花堇菜的根提取物强烈抑制了 Na,K-ATPase，而叶和种子提取物则基本没有活性。采用 UHPLC-QTOF-MS/MS 代谢组学方法鉴定了根提取物活性的化学原理，检测到 35292 种特征。通过将特征面积与 14 个分离馏分的抑制活性相关联，选择候选活性化合物。这产生了一组 15 个候选化合物，其中 14 个初步鉴定为原花青素。因此，购买了市售的原花青素（B1、B2、B3 和 C1）并研究了它们抑制 Na,K-ATPase 的能力。发现二聚原花青素 B1、B2 和 B3 没有活性，但三聚原花青素 C1 强烈抑制 Na,K-ATPase，IC 为 4.5µM。这种新发现的抑制剂被对接入模拟 NaE∼P·ADP 和 KE·P 状态的晶体结构中，以确定 Na,K-ATPase 内的潜在相互作用位点。讨论了可能的结合机制和观察到的根提取物活性的原理。
