Rapid screening of monoamine oxidase B inhibitors in natural extracts by capillary electrophoresis after enzymatic reaction at capillary inlet

Abstract

A facile capillary electrophoresis (CE) method was developed for the screening of monoamine oxidase B (MAO-B) inhibitors in natural extracts. In this method, the enzymatic reaction occurred at the capillary inlet during a predetermined waiting period, followed by the electrophoretic separation of the reaction compounds, and detected by their UV absorbance at 280 nm. Conditions for the separation of substrates, products and enzyme were optimized. The optimal buffer composition was 50 mM N-2-hydroxyethyl-piperazine-N'-2-ethane sulphonic acid (HEPES) solution containing 10 mM SDS (pH=7.4). Under the optimal condition, the baseline separation of substrates, products and enzyme was achieved within 2 min. The present method was used to determine MAO-B kinetic constants, K(i), K(m) and IC₅₀ based on quantitative of the substrate peak area compared with the reference electropherogram obtained from without the inhibitor. A validation study shows good reproducibility for both migration time (RSD=1.8%) and peak area (RSD=3.9%). Finally, the screening of 16 natural extracts was performed, and 2 natural extracts from Fructus crataegi and Radix polygoni multiflori were identified to be positive for MAO-B inhibition.