An MnO2-ZIF-67 immobilized acetylcholinesterase method for acetylcholinesterase activity assay and inhibitor screening from Inula macrophylla based on capillary electrophoresis

Abstract

The inhibition of acetylcholinesterase (AChE) from decomposing acetylcholine (ACh) is regarded as a common therapeutic strategy for Alzheimer's disease (AD). Screening AChE inhibitors is of great significance for AD treatment. In this work, AChE was physically absorbed on the synthesized MnO2-ZIF-67 through electrostatic interaction for the first time, and the fabricated MnO2-ZIF-67 immobilized AChE coupled with capillary electrophoresis (CE) was applied to screening AChE inhibitor. The established MnO2-ZIF-67 immobilized AChE with excellent pH, thermal and organic solvents tolerance stability was evaluated by Ellman's method. Huperzine A was used as model inhibitor to investigate the inhibition kinetics of the immobilized AChE, which possess the feasibility and applicability for its inhibitor screening. The screening mechanism of AChE inhibitor was based on the Ellman's method and measured by the changes of 5-thio-2-nitrobenzoic acid (TNB), which was produced by the reaction of thiocholine (TCh) with 5,5-dithiobis-(2-nitrobenzoic acid) (DTNB). Benefiting from the advantages of convenience and high sensitivity, the proposed platform was employed to screen AChE inhibitors from 11 compounds isolated from Inula macrophylla, and macrophyllic acid A (compound 1), dibutyl phthalate (compound 2) exhibited strong AChE inhibitory activity. Thus, this work not only provides a promising platform for monitoring AChE activity, but also holds great potential for the further exploration of active components with AChE inhibitory activity from Inula macrophylla.