College of Food Science, Sichuan Agricultural University, private bag 625014, Yaan, Sichuan, China.
College of Food Science, Sichuan Agricultural University, private bag 625014, Yaan, Sichuan, China.
Food Res Int. 2022 Aug;158:111497. doi: 10.1016/j.foodres.2022.111497. Epub 2022 Jun 10.
The present study aimed to investigate the inhibition mechanism of polyphenols from Phyllanthus emblica Linn. fruit (PEF, family Euphorbiaceous) on acetylcholinesterase (AChE). Interaction assay, enzyme kinetics, spectroscopic methods, and molecular simulations were performed. Results showed that myricetin, quercetin, fisetin, and gallic acid were the most active components in PEF, because of their low docking scores and strong inhibition ability on AChE with IC values of 0.1974 ± 0.0047, 0.2589 ± 0.0131, 1.0905 ± 0.0598 and 1.503 ± 0.0728 mM, respectively. Among them, the results of kinetic study showed that myricetin, quercetin, and fisetin reversibly inhibited AChE in a competitive manner, while gallic acid inhibited it through a noncompetition type. The interaction assay implied that a combination of the four polyphenols at the selected concentrations manifested a synergistic inhibition effect on AChE in a mixed inhibition type. Fluorescence and UV-vis spectrophotometry revealed that the active PEF polyphenols could strongly quench the intrinsic fluorescence of AChE via a static quenching mechanism. Circular dichroism spectroscopy analysis indicated that the active PEF polyphenols gave rise to the secondary structure changes of AChE by increasing the content of α-helix and reducing β-sheet and random coil conformation. The molecular dynamics simulation results validated that all the four docked polyphenol-AChE complexes were relatively stable according to their root-mean-square distance, root-mean-square fluctuations, solvent accessible surface area, radius of gyration values and hydrogen bonds evaluations during the whole simulation process. Overall, our study provides a creative insight into the further utilization of PEF polyphenols as functional components in exploring natural AChE inhibitors.
本研究旨在探讨余甘子果（Phyllanthus emblica Linn.，大戟科）多酚对乙酰胆碱酯酶（AChE）的抑制机制。进行了相互作用测定、酶动力学、光谱学方法和分子模拟。结果表明，由于杨梅素、槲皮素、漆黄素和没食子酸与 AChE 的结合评分较低且对 AChE 具有较强的抑制能力，因此它们是 PEF 中最活跃的成分，其 IC 值分别为 0.1974±0.0047、0.2589±0.0131、1.0905±0.0598 和 1.503±0.0728 mM。其中，动力学研究结果表明，杨梅素、槲皮素和漆黄素以竞争性方式可逆地抑制 AChE，而没食子酸则通过非竞争型抑制。相互作用测定表明，四种多酚在选定浓度下的组合对 AChE 表现出协同抑制作用，属于混合抑制类型。荧光和紫外可见分光光度法表明，活性 PEF 多酚通过静态猝灭机制强烈猝灭 AChE 的固有荧光。圆二色性光谱分析表明，活性 PEF 多酚通过增加α-螺旋含量和降低β-折叠和无规卷曲构象来引起 AChE 的二级结构变化。分子动力学模拟结果验证了所有四个对接的多酚-AChE 复合物在整个模拟过程中根据其均方根距离、均方根波动、溶剂可及表面积、旋转半径值和氢键评估，都是相对稳定的。总的来说，我们的研究为进一步利用 PEF 多酚作为功能成分来探索天然 AChE 抑制剂提供了新的思路。
