Ligand Fishing Study on Puerariae Lobatae Radix Extract Based on α-Amylase Functionalized MIL-53(Al)-NH

OBJECTIVE To screen the substances with α-amylase inhibitory activity from 95% ethanol extract of Puerariae Lobatae Radix using ligand fishing strategy. METHODS α-Amylase was immobilized on the surface of MIL-53(Al)-NH₂, which was a water-stable Metal-Organic Frameworks(MOFs), by glutaraldehyde to obtain the immobilized enzyme. According to the ligand fishing strategy, the immobilized enzyme interacted with the aqueous solution of 95% ethanol extract of Puerariae Lobatae Radix to extract those chemical components that could interact with α-amylase. RESULTS α-Amylase was successfully immobilized on the surface of MIL-53(Al)-NH₂ and had enzymatic activity. The compounds screened by fishing were separated by HPLC, and 11 active compounds were determined by the comparison with reference substances. Besides, the binding of these 11 active compounds to α-amylase was further confirmed by molecular docking technology. CONCLUTION α-Amylase is immobilized on MIL-53(Al)-NH₂ for the first time, and establish a ligand-fishing method based on MOFs-immobilized enzymes, which can be used to screen bioactive components from complex mixtures. In addition, this study provides a new idea for more use of MOFs for enzyme immobilization, and a reference for the specific identification and screening of active components from traditional Chinese medicine based on ligand-fishing of enzyme-functionalized MOFs.