Simvastatin is an established anti-hyperlipidemic drug and few studies have indicated its role in the mitigation of oxidative stress. However, a systematic study considering molecular binding/interaction of simvastatin with anti-oxidant enzymes followed by confirmational in vitro and in vivo studies have never been done. We investigated the molecular binding of simvastatin with multiple anti-oxidant enzymes and assessed their levels after the treatment of simvastatin in vitro and in vivo. This study is the first to show the molecular binding of simvastatin to catalase through molecular docking analysis. Moreover, the anti-oxidative properties of simvastatin have not been studied in Lipopolysaccharide (LPS) induced oxidative stress in HepG2 cells. We found that simvastatin effectively attenuated oxidative stress in LPS induced HepG2 cells and high-fat diet (HFD) fed hyperlipidemic rats by increasing the levels of antioxidant enzymes. The activity of catalase and superoxide dismutase (SOD) both increased significantly in oxidatively stressed HepG2 cells after the treatment with simvastatin (10 ​μM, 24 ​h). In addition to this, he original cell morphology of oxidatively stressed cells was restored by simvastatin, and an increase in antioxidant enzymes, catalase (0.08 U/cells to 0.12 U/cells), and SOD (0.57 U/cells to 0.74 U/cells) was also noted in HepG2 cells. Furthermore, a significant increase in the antioxidant enzymes such as Catalase, SOD, and reduced glutathione (GSH) was noted after simvastatin treatment in the HFD model. Moreover, we also observed degradation of by-products of lipid peroxidation thiobarbituric acid reactive substances (TBARs), nitric oxide (NO), and protein carbonyl levels. This indicates that simvastatin enhances anti-oxidant enzyme activities and can be repurposed for the treatment of oxidative stress in liver diseases in humans after extensive clinical trials.