Preparation of Magnetic Microsphere-Gold Nanoparticle-Immobilized Enzyme Batch Reactor and Its Application to Enzyme Inhibitor Screening in Natural Extracts by Capillary Electrophoresis

Abstract

A novel strategy for screening the enzyme inhibitors from the natural products by capillary electrophoresis (CE) with a pre column enzyme batch reactor prepared by magnetic microsphere (MB)-gold nanoparticles (AuNPs) is reported. The enzyme batch reactor was prepared by immobilizing the enzymes on the MB conjugated AuNPs (MB-AuNPs). To demonstrate this strategy, xanthine oxidase (XOD) was employed as a model for the activity of the enzyme, inhibition study, and inhibitor screening. With the developed CE method, the enzyme activity was determined by the quantification of the peak area of the product. Enzyme inhibition can be read out directly from the reduced peak area of product in comparison to a reference electropherogram obtained in the absence of any inhibitor. A statistical parameter Z‘ factor was recommended for evaluation of the accuracy of a drug screening system. In the present study, it was calculated to be 0.7, implying a good accuracy. The screening of two natural extracts from Cortex Phellodendri and Rhizoma Galangae showed that they were positive for XOD inhibition by the present method. Using this immobilized enzyme technology combined with CE separation not only provides the advantages such as convenience, rapidity and low cost, but also provides a new platform for discovering enzyme-inhibitor drug lead compounds.