The study aimed, by integrating transcriptomics and metabolomics, to reveal novel biomarkers caused by overdosed acetaminophen (APAP) and liver protection substances procured by pre-administration of ginseng shoots extract (GSE). Totally 4918 genes and 127 metabolites were identified as differentially expressed genes and differential metabolites, respectively. According to KEGG (Kyoto Encyclopedia of Genes and Genomes) enrichment, such pathways as primary bile acid biosynthesis, bile secretion, retinol metabolism, histidine and several other amino-related metabolism were significantly altered by GSE and disturbed by subsequent overdosed APAP at the transcriptomic as well as metabolomic levels. Fifteen key biomarker metabolites related to these pathways were up-regulated in APAP-treated vs GSE-pretreated liver tissues, and were reported exerting anti-oxidant, anti-inflammatory, anti-apoptotic and/or immunomodulate functions, three of which even possessed direct hepatoprotection effects. Twenty five vital unigenes modulating these metabolites were further verified by correlation analysis and expression levels of fifteen of them were examined by qRT-PCR. Our findings indicate that GSE may be an effective dietary supplement for preventing the liver damage caused by the overdosed APAP.