Metabolite-Based Network Pharmacology, Molecular Docking, and Dynamics Simulations to Preliminarily Verify Treating...

Kuwanon G (KWG), a bioactive flavonoid from mulberry, exhibits potential neuroprotective effects against diabetic encephalopathy (DE), yet its metabolic fate and therapeutic mechanisms remain unclear. This study integrated ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS), network pharmacology, molecular docking, and dynamics simulations to characterize KWG's metabolic profile and evaluate its anti-DE activity. In vivo and in vitro analyses identified 56 metabolites in rats, predominantly formed via oxidation, dehydrogenation, methylation, and glucuronidation. Nine metabolites with high intestinal absorption and pharmacophore compatibility were selected using Swiss ADME. Network pharmacology revealed core targets (AKT1, TNF, SRC, EGFR, ESR1) linked to DE, while molecular docking demonstrated strong binding affinities (-4.87 to -43.41 kcal/mol) between active metabolites (N1, N4, N6, N8) and these targets. Dynamics simulations confirmed stable interactions, highlighting metabolites' roles in modulating PI3K-Akt signaling and neurodegeneration pathways. Notably, KWG itself exhibited negligible binding, suggesting its metabolites are the primary bioactive forms. These findings underscore the importance of gut microbiota-mediated biotransformation in enhancing KWG's bioavailability and neuroprotective efficacy. This work provides critical insights into the metabolic activation of natural products and advances their application in functional foods or therapeutics for diabetes-related complications.