Department of Nutrition, Dietetics and Food, School of Clinical Sciences at Monash Health, Faculty of Medicine, Nursing and Health Sciences, Monash University, Notting Hill BASE Facility, 264 Ferntree Gully Road, Notting Hill, VIC 3168, Australia.
Food Chem. 2021 May 1;343:128423. doi: 10.1016/j.foodchem.2020.128423. Epub 2020 Oct 26.
The aim was to determine inhibition of human α-amylase activity by (poly)phenols using maltoheptaoside as substrate with direct chromatographic product quantification, compared to hydrolysis of amylose and amylopectin estimated using 3,5-dinitrosalicylic acid. Acarbose exhibited similar IC values (50% inhibition) with maltoheptaoside, amylopectin or amylose as substrates (2.37 ± 0.11, 3.71 ± 0.12 and 2.08 ± 0.01 µM respectively). Epigallocatechin gallate, quercetagetin and punicalagin were weaker inhibitors of hydrolysis of maltoheptaoside (<50% inhibition) than amylose (IC: epigallocatechin gallate = 20.41 ± 0.25 µM, quercetagetin = 30.15 ± 2.05 µM) or amylopectin. Interference using 3,5-dinitrosalicylic acid was in the order punicalagin > epigallocatechin gallate > quercetagetin, with minimal interference using maltoheptaoside as substrate. The main inhibition mechanism of epigallocatechin gallate and punicalagin was through complexation with starch, especially amylose, whereas only quercetagetin additionally binds to the α-amylase active site. Interference is minimised using maltoheptaoside as substrate with product detection by chromatography, potentially allowing assessment of direct enzyme inhibition by almost any compound.
本研究旨在采用直链七糖作为底物，通过直接色谱产物定量，测定（多）酚对人α-淀粉酶活性的抑制作用，并与使用 3,5-二硝基水杨酸估计的淀粉和支链淀粉水解进行比较。阿卡波糖对直链七糖、支链淀粉或淀粉的 IC 值（50%抑制）相似（分别为 2.37±0.11、3.71±0.12 和 2.08±0.01μM）。表没食子儿茶素没食子酸酯、槲皮素和安石榴甙对直链七糖水解的抑制作用较弱（<50%抑制），而对淀粉的抑制作用较强（IC：表没食子儿茶素没食子酸酯=20.41±0.25μM，槲皮素=30.15±2.05μM）或支链淀粉。使用 3,5-二硝基水杨酸的干扰顺序为安石榴甙>表没食子儿茶素没食子酸酯>槲皮素，而使用直链七糖作为底物的干扰最小。表没食子儿茶素没食子酸酯和安石榴甙的主要抑制机制是与淀粉（尤其是直链淀粉）形成复合物，而槲皮素除了与α-淀粉酶的活性位点结合外，还与其他化合物结合。使用直链七糖作为底物，通过色谱法检测产物，可最大限度地减少干扰，这种方法可能允许对几乎任何化合物的直接酶抑制作用进行评估。
