The invention discloses a metabonomics-based application of endogenous small-molecular substances in rapid detection of hepatotoxicity. The endogenous small-molecular substances are 12 hepatotoxicity biomarkers, i.e., L-alanine, L-phenylalanine, L-carnitine, L-carnitine palmitoyl, tryptophan, arachidonate, cholic acid, lysophosphatidylcholine (14:0), lysophosphatidylcholine (16:1), lysophosphatidylcholine (18:2), lysophosphatidylcholine (20: 3) and hemolysis phosphatidyl ethanolamine (18:2); and the endogenous small-molecular substances also refers to 3 special hepatotoxicity biomarkers, i.e. L-carnitine, cholic acid and lysophosphatidylcholine (14:0). By adopting the hepatotoxicity biomarkers, the discovery and diagnosis of the liver injury can be earlier than those of the existing biochemical detection indexes, the hepatotoxicity biomarkers can be well used for preventing, discovering and treating the hepatotoxicity, the limitation of the existing indexes can be overcome, and accurate detection information can be provided in time.