Acetaminophen is an analgesic antipyretic commonly used. Hepatotoxicity is one of the most common obstacles to acetaminophen therapy. Eucalyptus oil is an antioxidant with potent free radical-capturing activities. This research was designed to evaluate the inhibitory impact of Eucalyptus oil versus acetaminophen-triggered hepatotoxicity. The composition of Eucalyptus was detected utilizing gas chromatography mass spectroscopy. Forty-eight rats were assigned into six groups; control group, acetaminophen group (500mg/kg I.P twice on the 17th and, 20th of the experiment), silymarin group (50mg/kg, I.P once daily for three weeks), Eucalyptus oil group (30mg/kg orally once daily for three weeks), acetaminophen +silymarin group and Eucalyptus +acetaminophen group. gas chromatography mass spectroscopy discovered four compounds in Eucalyptus with eucalyptol representing the main compound. revealed that acetaminophen remarkably elevated serum alanine transaminase, aspartate aminotransferase, alkaline phosphatase, and total bilirubin levels, whereas it declined serum albumin and total protein levels. In addition, hepatic oxidant/antioxidant imbalance was evident in acetaminophen-intoxicated rats by the rising of lipid peroxidation biomarker; malondialdehyde and the downregulation of nuclear factor erythroid 2-related factor 2 and its transcriptional mediators; superoxide dismutase, glutathione peroxidase, and reduced glutathione. Furthermore, following acetaminophen injection, there was a remarkable increase in transforming growth factor-β gene expression, tumor necrosis factor-α and interleukin1-β levels along with a decline in interleukin-10 levels. Immunohistochemical and histopathological examinations were in parallel with the abovementioned results. However, all these abnormalities were significantly abrogated in rats pretreated with Eucalyptus. We concluded that prior administration of Eucalyptus oil counteracted acetaminophen-mediated hepatotoxicity via powerful antioxidant, anti-inflammatory, and anti-apoptotic impacts