Magnoflorine (Mag) has multiple pharmacological activities for the prevention and treatment of prostatitis. However, its molecular mechanisms andpharmacological targets are not clear. In this study, the ultra-performance liquid tandem mass spectrometry-based metabolomics method was used to clarify the intervention of Mag against prostatitis and the biological mechanism. A total of 25 biomarkers associated with the prostatitis model were identified by metabolomics, and a number of metabolic pathways closely related to the model were obtained by MetPA analysis. After given Mag treatment, the results of each indicator were shown that Mag alkaloid could inhibit the development of prostatitis effectively. We found that Mag had regulative effects on potential biomarkers of prostatitis model, which can regulate them to the control group. Our results indicated that alkaloids have an effective intervention therapy for prostatitis, and five types of metabolic pathways closely related to prostatitis model were obtained, including phenylalanine, tyrosine and tryptophan biosynthesis, phenylalanine metabolism, tyrosine metabolism, arginine and proline metabolism, glycine, serine and threonine metabolism, alanine, aspartate and glutamate metabolism. This study has provided the basic experimental data for the development of Mag in the prevention and treatment of prostatitis. CI - Copyright (c) 2021 Yuan, Dong, Liu, Xiao and Zhou.