Jiangsu key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, China.
School of Science, China Pharmaceutical University, Nanjing 211198, China.
Molecules. 2018 Oct 11;23(10):2612. doi: 10.3390/molecules23102612.
Enzyme inhibitors from natural products are becoming an attractive target for drug discovery and development; however, separating enzyme inhibitors from natural-product extracts is highly complex. In this study, we developed a strategy based on tyrosinase-site blocking ultrafiltration integrated with HPLC-QTOF-MS/MS and optimized molecular docking to screen tyrosinase inhibitors from Radix extract. Under optimized ultrafiltration parameters, we previously used kojic acid, a known tyrosinase inhibitor, to block the tyrosinase active site in order to eliminate false-positive results. Using this strategy, puerarin, mirificin, daidzin and genistinc were successfully identified as potential ligands, and after systematic evaluation by several docking programs, the rank of the identified compounds predicted by computational docking was puerarin > mirificin > kojic acid > daidzin ≈ genistin, which agreed with the results of tyrosinase-inhibition assays. Structure-activity relationships indicated that C-glycosides showed better tyrosinase inhibition as compared with O-glycosides, with reduced inhibition achieved through the addition of glycosyl, which provides ideas about the screen of leading compounds and structural modification.
天然产物中的酶抑制剂正成为药物发现和开发的一个有吸引力的目标；然而，从天然产物提取物中分离酶抑制剂是非常复杂的。在本研究中，我们开发了一种基于酪氨酸酶位点阻断超滤与 HPLC-QTOF-MS/MS 相结合的策略，并对优化的分子对接进行了优化，以从葛根提取物中筛选酪氨酸酶抑制剂。在优化的超滤参数下，我们以前使用曲酸（一种已知的酪氨酸酶抑制剂）来阻断酪氨酸酶的活性位点，以消除假阳性结果。使用这种策略，我们成功地鉴定出了葛根素、米丽菲啶、大豆苷元和染料木素作为潜在的配体，并且经过几个对接程序的系统评估，通过计算对接预测的鉴定化合物的排名为葛根素>米丽菲啶>曲酸>大豆苷元≈染料木素，这与酪氨酸酶抑制试验的结果一致。构效关系表明，C-糖苷与 O-糖苷相比，显示出更好的酪氨酸酶抑制作用，通过糖苷基的添加抑制作用降低，这为先导化合物的筛选和结构修饰提供了思路。
