Piperlonguminine inhibits hen egg white lysozyme amyloid fibril formation and mitigates amyloid fibril-induced hepatocellular damage.

Protein misfolding and aggregation into amyloid fibrils (AFs) are critical issues associated with protein stability during food processing and storage, as well as with pathological damage in organisms. Hen egg white lysozyme (HEWL), a widely used protein in egg products, meat processing, and food preservation, serves as a classical model for investigating amyloidogenesis. Piperlonguminine (PPR), a low-toxicity bioactive alkaloid from Piperis Longi Fructus (a medicinal and food-homologous spice), has not yet been reported to regulate amyloid fibrillation in food proteins. This study explored the inhibitory activity of PPR against HEWL-AF formation and its protective effect on AF-induced hepatocellular damage, with a focus on its application potential in food systems. Spectroscopic and microscopic analyses confirmed that PPR effectively inhibited HEWL-AF formation by maintaining α-helical structures, reducing β-sheet transitions, and stabilizing hydrophobic regions of HEWL. Molecular docking and dynamics simulations revealed that PPR interacts with HEWL-AF through hydrogen bonding, π-π stacking, and van der Waals interactions, thereby disrupting the food protein aggregation pathway. Cellular experiments confirmed that HEWL-AF induced membrane damage and cytotoxicity in NCTC1469 cells, whereas PPR significantly mitigated these adverse effects. Collectively, these findings indicate that PPR is a promising natural inhibitor of food protein amyloid fibrillation with hepatoprotective activity, providing a theoretical foundation for its application in functional food development and the improvement of protein stability in food processing systems.