Protective effect of Chlorogenic Acid complex against intestinal oxidative stress in broilers and screening of its key isomer

Oxidative stress triggers physiological and pathological damage in animals. As a natural polyphenol, chlorogenic acid (CGA) is a potential candidate for alleviating such damage, while discrepancies in antioxidant activity among its isomers and underlying mechanisms remain unclear. This study aimed to clarify the protective effects of the CGA complex on broiler intestinal oxidative stress and identify the key active isomers by integrating in vivo animal models, network pharmacology, molecular docking, and in vitro antioxidant assays. In vivo, the CGA complex effectively reversed DSS-induced reductions in broiler body weight, slaughter performance, and alterations in jejunal glutathione peroxidase (GSH-px) and superoxide dismutase (SOD) activities (P < 0.05). Network pharmacology analysis identified 4,5-dicaffeoylquinic acid (4,5-CQA) as a key isomer (Betweenness 1549.69), and molecular docking results showed that it can stably bind to the core targets CASP3, SRC and MMP9. 4-CQA exhibited stronger free radical scavenging activity in ABTS/DPPH/FRAP assays (P < 0.05); 4,5-CQA displayed superior cytoprotection, enhanced Nrf2, HO-1 protein expression (P < 0.05) and inhibited Caspase-3 protein expression (P < 0.05) in tBHP-stimulated intestinal epithelial cells. In conclusion, the CGA effectively mitigates DSS-induced intestinal oxidative stress in broilers via regulating multiple signaling pathways, and 4,5-CQA serves as the pivotal bioactive isomer in complex physiological processes.