Background: Frequent overdose of paracetamol (APAP) has become the major cause of acute liver injury. The present study was designed to evaluate the potential protective effects of ginsenoside Rk1 on APAPinduced hepatotoxicity and investigate the underlying mechanisms for the first time. Methods: Micewere treated with Rk1 (10 mg/kg or 20 mg/kg) by oral gavage once per d for 7 d. On the 7th d, all mice treatedwith250mg/kgAPAPexhibitedsevere liver injuryafter24h, andhepatotoxicitywas assessed. Results: Our results showed that pretreatment with Rk1 significantly decreased the levels of serum alanine aminotransferase, aspartate aminotransferase, tumor necrosis factor, and interleukin-1b compared with the APAPgroup.Meanwhile, hepatic antioxidants, including superoxide dismutase andglutathione,were elevated compared with the APAP group. In contrast, a significant decrease in levels of the lipid peroxidation product malondialdehyde was observed in the ginsenoside Rk1-treated group compared with the APAP group. These effectswere associatedwith a significant increase of cytochromeP450 E1 and 4-hydroxynonenal levels in liver tissues. Moreover, ginsenoside Rk1 supplementation suppressed activation of apoptotic pathways by increasing Bcl-2 and decreasing Bax protein expression levels, which was shown using western blotting analysis.Histopathological observation also revealed that ginsenoside Rk1 pretreatment significantly reversed APAP-induced necrosis and inflammatory infiltration in liver tissues. Biological indicators of nitrative stress, such as 3-nitrotyrosine, were also inhibited after pretreatment with Rk1 compared with the APAP group. Conclusion: The results clearly suggest that the underlying molecular mechanisms in the hepatoprotection of ginsenoside Rk1 in APAP-induced hepatotoxicity may be due to its antioxidation, antiapoptosis, anti-inflammation, and antinitrative effects.