Clarification of Mechanism of Anti-fatigue Active Components Derived from Earthworms Based on UHPLC-Q-TOF-MS/MS and Network Pharmacology

MENG Xuan; WANG Mengjiao; ZHAO Tianyang; ZHANG Guozhong; ZHAO Delong; HE Xihong; ZHAO Huabing; JIA Liming; JIA Jinghua; ZHANG Liming

doi:10.13982/j.mfst.1673-9078.2025.3.0001

Abstract：The anti-fatigue mechanism of the active constituents of earthworm was clarified using ultra-performance liquid chromatography-quadrupole-time of flight mass spectrometry/mass spectrometry(UHPLC-Q-TOF-MS/MS and network pharmacology.UHPLC-Q-TOF-MS/MS was used to identify chemical constituents of earthworm extract.The targets of these constituents and fatigue and anti-fatigue associated genes were collected to identify the common targets.Gene Ontology and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analyses were performed.KEGG Mapper was utilized to preliminarily elucidate the anti-fatigue mechanism,and animal experiments were performed to verify the anti-fatigue efficacy of earthworm extract.Network analysis showed that the anti-fatigue effects of earthworm extract mainly function via positive regulation of phosphorylation,NF-κB signaling pathway,serotonergic synapse signaling,JAK-STAT pathway,and arginine and proline metabolic pathways.As shown in the results of animal experiments,the time of weight-bearing swimming in mice in the middle-dose earthworm extract group was 34.07 min,significantly longer than that(9.78 min)of the CK group(P＜0.001).Hepatic glycogen content in the high-dose earthworm extract group was 2 591.21 mg/g,which was significantly higher than that in the CK group(P＜0.001).Urea nitrogen and blood lactic acid contents in the high-dose earthworm extract group were 3.65 nmol/L and 182.21 nmol/L,respectively,which was significantly lower than those of the CK group(P＜0.01).In conclusion,the anti-fatigue effect of earthworm extract was exerted through synergistic effects of multiple constituents,targets,and pathways,providing valuable insights and methods for further research into anti-fatigue mechanisms.