To explore the anti-rheumatoid arthritis mechanism of methotrexate by using metabonomic techniques.Methods:The model of rheumatoid arthritis in rats was replicated by adjuvant arthritis (AIA).The replication degree of the model was evaluated by classical foot swelling index and treated with methotrexate.Serum samples were collected to characterize the occurrence and development of diseases and the metabolic level after drug intervention by metabolic profiling technology.Biological information of each group of rats was obtained by MSE data acquisition mode during the experiment.Core metabolic enzymes related to rheumatoid arthritis were obtained by pattern recognition，enrichment analysis and other methods.The pathogenesis of rheumatoid arthritis and the action mechanism of methotrexate were further elucidated.Results:Non-targeted metabolic profile analysis showed that methotrexate had an obvious intervention effect on the overall metabolism of rheumatoid arthritis rats，and the metabolic profiles shifted from model state to blank group state.It was proved that methotrexate had a good effect on rheumatoid arthritis rats from the point of view of overall metabolism.The blood-related metabolic abnormality markers such as guanosine Pentaphosphate adenosine，glucuronic acid bilirubin，guanosine diphosphate，stearic acid，in rheumatoid arthritis rats were obtained by S-map focusing in orthogonal partial least squares discriminant analysis.Pathway analysis showed that fatty acid biosynthesis，starch and sucrose metabolism，pyrimidine metabolism，pentose and glucuronidation，purine metabolism were mainly involved in rheumatoid arthritis rats.Conclusion:Methotrexate has a good therapeutic effect on rheumatoid arthritis rats.The regulation of key metabolic enzymes and metabolic markers in metabolic pathways such as fatty acid biosynthesis，starch and sucrose metabolism，pyrimidine metabolism may play a role in inhibiting rheumatoid arthritis.