Strategic Immobilization of α-Amylase on ZIF-8 Nanoplatform for Targeted Bioactivity Screening in Gynura medica Extracts

Abstract

This study presents an enzyme immobilization strategy utilizing zeolitic imidazolate framework-8 (ZIF-8), a metal-organic framework (MOF) material synthesized in-house, for α-amylase immobilization, establishing a novel solid-phase extraction platform for targeted isolation of enzymatic inhibitors from Gynura medica aqueous extracts. The ZIF-8@α-amylase nanoparticles was prepared using in-situ encapsulation method, which was exhibited 91% enzyme encapsulation efficiency and enhanced aqueous stability. ZIF-8@α-amylase exhibited a broader operational range, retaining more than 50% activity across 20-70°C and pH 4.0-9.0 (compared to 20-65°C and pH 5.5-7.5 for the free enzyme). This immobilized system also demonstrated a high binding capacity of 189.5 mg/g for acarbose. Using a mild alkaline buffer for elution, a concentrated fraction showing 18.1% α-amylase inhibition was obtained, which mainly contained phenolic acids, flavonoids, and hydrolyzable tannins as identified by LC-MS/MS. Dose-response analyses demonstrated significant pancreatic amylase inhibition by identified compounds: ellagic acid-4-O-glucoside: 98.86±2.3% (1.0 g/L), quercetin-3-O-rhamnoside: 66.32±1.9% (1.0 g/L), methyl gallate: 33.56±1.2% (1.0 g/L). Overall, ZIF-8@α-amylase is a promising solid-phase extraction platform for selectively enriching bioactive amylase inhibitors from complex botanical matrices, with potential in Pharmaceutical discovery and diabetes management.