Immobilized enzyme reactor coupled with UHPLC-QTOF-MS for screening acetylcholinesterase inhibitory leads from Aconitum carmichaelii aerial parts

The aerial parts of Aconitum carmichaelii Debx. (AAL), an abundant but discarded agricultural byproduct, represent an unexplored source of acetylcholinesterase (AChE) inhibitors. However, the direct screening of bioactive leads from such a complex matrix poses significant analytical challenges. Herein, we report an integrated platform coupling a magnetic immobilized enzyme reactor with ultra-high performance liquid chromatography–quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) for effective screening of AChE inhibitory leads. The AChE was immobilized on magnetic chitosan microbeads (AChE@MCM) to create a reusable and stable bioreactor with an impressive 80.9% activity recovery and the ability to be reused over 8 times, along with enhanced pH stability. When applied to AAL extracts, this strategy efficiently fished out five potential AChE inhibitory leads with diverse structures, directly unveiling the untapped medicinal potential of this waste resource. Our work not only provides a sustainable approach to utilizing agricultural byproducts but also validates a powerful tool for rapid bioactivity-guided screening in complex natural products.