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Ultraperformance liquid chromatography coupled with quadrupole time-of-flight synapt high-definition mass spectrometry metabolomics was used to characterize the urinary metabolic profiling of diet-induced hyperlipidaemia in a rat model. Analysis was done by orthogonal partial least squares discriminant analysis, correlation analysis, heat map analysis, and KEGG pathways analysis. Potential biomarkers were chosen by S-plot and were identified by accurate mass, isotopic pattern, and MS/MS fragments information. Significant differences in fatty acid, amino acid, nucleoside, and bile acid were observed, indicating the perturbations of fatty acid, amino acid, nucleoside, and bile acid metabolisms in diet-induced hyperlipidaemia rats. This study provides further insight into the metabolic profiling across a wide range of biochemical pathways in response to diet-induced hyperlipidaemia.