Abstract: To explore the protective effect of protropine in Corydalis humosa Migo. on lipopolysaccharide-induced acute kidney injury in mice (AKI), an approach that used ultra-performance liquid chromatography quadrupole time-of-flight tandem mass spectrometry (UHPLC-Q/TOF-MS) coupled with a multivariate analytical platform was established. The BALB/c mice were divided into normal group (CON), model group (LPS), and protropine group (PRO). Mice were injected intraperitoneally with lipopolysaccharide solution to replicate the AKI model. Three hours after modeling, mice were given the protropine solution by gavage. Protropine was a monomer compound isolated in the laboratory, and protropine solution was prepared by dissolving protropine in sterilized distilled water. Administration was performed twice a day for three days. After modeling and administration, serum samples were collected. UHPLC-Q/TOF-MS was used to generate metabolomics data. Multivariate statistical analysis and online databases were used to screen potential biomarkers and enrich metabolic pathways. The heatmap of relative quantitative biomarker data was generated through Mev software. Animal experiments were approved by the Animal Experimentation Ethics Committee of Henan University of Chinese Medicine (No. SYXK2015-0005). The results show that the metabolic profile of mice in the LPS group was significantly altered by intervention with protropine, and clustered towards the CON group. 70 biomarkers were identified from the CON group vs LPS group (35 in positive source mode, 35 in negative source mode), and 67 biomarkers were identified from the LPS group vs PRO group (37 in positive source mode, 30 in negative source mode). A total of 34 common markers (18 in positive source mode, 16 in negative source mode) were obtained from the two comparison groups. The enrichment of all biomarkers resulted in 8 metabolic pathways including linoleic acid metabolism, D-glutamine and D-glutamate metabolism, arginine and proline metabolism, and arachidonic acid metabolism. The results show that protropine in Corydalis rhizoma ameliorates the kidney damage, insufficient energy supply, and inflammation in AKI mice by regulating amino acid metabolism, energy metabolism, and lipid metabolism in AKI mice.