Exploration of carbonic anhydrase inhibitors from Ginkgo biloba leaves: Fast screening with ligand-incorporated cross-linked enzyme aggregates followed by inhibitory mechanism study

Acute mountain sickness (AMS) is a common syndrome at high altitudes that can become life-threatening if not managed properly, and carbonic anhydrase (CA) is a promising target for treating AMS. This study reports the CA inhibitory activity of Ginkgo biloba leaves (GBL) for the first time, expanding the potential of this important industrial crop with extensive medicinal applications. To systematically identify its specific active components, a novel, highly sensitive fast-screening method based on ligand-incorporated cross-linked enzyme aggregates was developed, which facilitated the discovery of two new inhibitor classes, i.e., ginkgo biflavones and ginkgolic acids. Biflavones have been found to exhibit higher affinity and bioactivity than monomeric flavones and flavonol glycosides, while a distinct, hydrophobic-interaction-dominated mechanism is observed for ginkgolic acids. Consequently, a synergistic effect between biflavones and ginkgolic acids was identified. This study elucidated the potential mechanism of GBL against AMS, offering valuable insights for the rational formulation and high-value industrial valorization of GBL and derived products.