Mechanism of liquiritin from Codonopsis pilosula-Glycyrrhiza uralensis extract in alleviating piglet weaning stress via binding to alpha-1-microglobulin/bikunin precursor by machine learning and molecular dynamics simulation.

Weaning stress is a prevalent pathological condition in pig farming, primarily triggered by factors such as nutritional imbalance, oxidative stress, inflammation, and bacterial infections. In this study, response surface methodology was first employed to optimize the preparation process of Codonopsis pilosula-Glycyrrhiza uralensis extract (CGe). Dietary supplementation with CGe for 28 days revealed that 1% CGe significantly increased the growth performance of weaned piglets, enhanced antioxidant capacity, and promoted intestinal microbiota balance, demonstrating a favorable safety profile. Subsequently, by integrating Gene Expression Omnibus (GEO) database mining with machine learning, we identified cathepsin D (CTSD), alpha-1-microglobulin/bikunin precursor (AMBP), and prosaposin (PSAP) as potential biomarkers of weaning stress. Among these biomarkers, AMBP showed significant positive correlations with M0 macrophages, mast cells, and neutrophils, and a negative correlation with M1 macrophages, suggesting its involvement in macrophage polarization during weaning stress. Molecular docking and molecular dynamics simulations further revealed that liquiritin, a key bioactive component of CGe, formed a stable interaction with AMBP and exhibited low binding energy. This interaction likely underpins its potential to mediate anti-inflammatory and anti-oxidant effects. These results indicate that AMBP is a promising target for alleviating weaning stress, and CGe has potential as an eco-friendly feed additive for improving the health of weaned piglets.