摘要
The development of natural xanthine oxidase(XO)inhibitors is an effective strategy for the treatment of hy-peruricemia.In this study,the inhibition mechanism of phloretin on XO and its joint inhibition with clinical drugs(allopurinol and febuxostat)were investigated,using multispectroscopic and molecular simulation tech-niques.The results showed that phloretin reversibly inhibited XO in a mixed type with an IC_(50) value of 31.83±0.32μM.Phloretin quenched the fluorescence of XO with a static mechanism and hydrophobic interactions were the key binding force of phloretin binding to XO.Furthermore,the interaction between phloretin and XO resulted in a more relaxed structure and less stable of the enzyme with a reduction in theα-helix content from 21.5%to 11.3%.The heatmap analysis showed that a combination of phloretin(15μM)and allopurinol(10μM)or phloretin(30μM)and febuxostat(0.01μM)exhibited the strongest synergistic effect on XO inhibition in a mixed manner.The combination of phloretin and allopurinol/febuxostat enhanced the binding affinity with XO compared to the inhibitors individually.Molecular docking indicated that phloretin,allopurinol and febuxostat bound to different sites of XO to produce a synergistic inhibition by preventing substrate access to the Mo activity center and affecting electron transfer.This study may provide theoretical references for the development of phloretin and its formulation with clinical drugs as uric acid-lowering food functional factor or drug.
基金
financial support provided by the National Natural Science Foundation of China(No.22078143)
the Innovation and entrepreneurship training program for college students(No.202310403046).
