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OBJECTIVE To identify the potent metabolic biomarkers of cauda equina injury (CEI). MATERIAL AND METHODS A total of 50 Sprague-Dawley (SD) rats were used to establish the CEI model in this study. The serum was collected 12 h, 1 day, 2days and 7 days after surgery. Ultra-performance liquid chromatography, coupled with quadrupole time-of-flight mass spectrometry (UHPLC/Q-TOF-MS) was performed to analyze the metabolic changes in the serum of the CEI rats from different groups. The differences between the metabolic profiles of the rats in 5 groups were analyzed using partial least squares discriminant analysis (PLS-DA). RESULTS Metabolic profiling showed remarkable differences between the sham operation and other groups. 57 potential CEI metabolite biomarkers were identified between the sham operation group and the model groups of different time-point. Principal component analysis (PCA) and Partial least squares discriminant analysis (PLS-DA) analyses revealed clear segregation between CEI versus sham operation group. These potential biomarkers appear in 15 metabolic pathways. CONCLUSION By using metabolomic analysis we were able to identify the novel serum biomarkers of CEI. They might be useful for the diagnosis and prognosis of CEI patients. In addition, pathway analysis provided important insights into the etiology of CEI and a basis for clinical diagnosis, locating biomarkers in the early stages of the pathological process of CEI.