Objective To investigate the preventive effect of stevia rebaudiana polysaccharide (SRP) on non-alcoholic fatty liver disease (NAFLD) and its therapeutic effect on cell fat accumulation. Methods Cultivation of LO2 cells and measurement of their proliferative activity, treatment of LO2 cells with stevia rebaudiana root polysaccharides at different concentrations to determine whether stevia rebaudiana cells have toxicity to LO2 cells, induction of LO2 cells with oleic acid (OA), development of a cell model of NAFLD in vitro, Use of cck8 to measure cell activity, and with oil red O staining to observe fat accumulation.Treatment with oleic acid (OA) in different concentrations of Stevia Rebaudiana polysaccharide induced LO2 cells and oily red O staining was performed to observe the improvement of Stevia Rebaudiana polysaccharide on LO2 fat accumulation. Results After SRP treatment, SRP significantly reduced the accumulation of fat around LO2 cells. Conclusion Stevia rebaudiana polysaccharide can prevent non-alcoholic fatty liver disease (NAFLD) by regulating lipid metabolism disorders and reducing the accumulation of fat around cells.Therefore, stevia root polysaccharide plays an important role in the prevention and treatment of NAFLD.