Ethnopharmacological relevance: Hugan Qingzhi Tablet (HQT) is composed of five Chinese herbs,i.e.Rhizoma alismatis,Fructus Crataegi,Cattail Pollen,Lotus Leaf,Panax notoginseng.It has been used in prevention and treatment of non-alcoholic fatty liver (NAFLD).Aim of the study: This manuscript aims to evaluate the effect of HQT medicated serum on hepatic steatosis by using in vitro experiments on cells and explore the relevant mechanisms with method of serum pharmacology.Materials and method: The L-02 cells and HepG2 cells with FFA-induced hepatic steatosis model were used to investigate whether HQT could attenuate lipid accumulation and oxidative stress and explore the possible mechanisms.High-performance liquid chromatography (HPLC) was used to study components in serum contained HQT.Intracellular lipid droplets and triglyceride (TG) were detected to evaluate the lipid-lowing effect of HQT.Lactate dehydrogenase (LDH),aspartate aminotransferase (AST),alanine aminotransferase (ALT) in culture media were measured to evaluate protective effects of HQT on hepatic cells.Total antioxidant capacity (T-AOC),malonaldehyde (MDA),superoxide dismutase (SOD) and glutathione (GSH) were detected to measure antioxidant effects of HQT.Adiponection expression was investigated to evaluate the effect of HQT on adipocytokines.Furthermore,western blot and quantitative reverse transcription polymerase chain reaction (qRT -PCR) were performed to explore the relevant mechanisms of lipid metabolism of HQT.The phosphorylation levels of AMP-activated protein kinase (AMPK) and sterol regulatory element-binding protein 1 (SREBP-1) expression were measured to evaluate the effect of HQT on hepatic lipogenesis.Peroxisome proliferator activated receptor-α (PPARα),carnitine palmitoyltransferase 1 (CPT-1A) and acetyl-CoA oxidase 1 (ACOX1) expression were measured to evaluate the effect of HQT on hepatic lipolysis.Results: HQT medicated serum alleviated the accumulation of lipid droplets and decreased the cellular TG content in L02 cells and HepG2 cells.They caused significant reductions in contents of LDH,AST,ALT and MDA as well as significant increase in T-AOC activity in the culture media.They also caused increase in levels of GSH and SOD in FFA-induced steatosis L-02 and HepG2 cells.Furthermore,HQT medicated serum enhanced adiponection expression with a concentration-dependent manner and reversed the FFA-induced decrease in the phosphorylation levels of AMPK and PPARα,CPT-1,ACOX1 as well as decreased the expression of SREBP-1.Conclusion: The HQT medicated serum inhibited lipid accumulation and oxidative stress which might be through the activation of AMPK and PPARα pathway.