Non-alcoholic fatty liver disease is caused by excessive lipid accumulation in hepatocytes. Although trans-anethole (TAO) affects hypoglycemia and has anti-immune activity and anti-obesity effects, its role in non-alcoholic fatty liver disease remains unknown. This study aimed to evaluate the effects of TAO on cellular senescence, lipid metabolism, and reinforcement of microenvironments in HepG2 cells. To analyze the lipid metabolic activity of TAO, PCR analysis, flow-cytometry, and Oil Red O staining were performed, and mitochondrial membrane potential (MMP) and cellular senescence kits were used for assessing the suppression of cellular senescence. At 2000 μg/mL TAO, the cellular viability was approximately 99%, and cell senescence decreased dose-dependently. In the results for MMP, activity increased with concentration. The levels of lipolytic genes, CPT2, ACADS, and HSL, strongly increased over 3 days and the levels of lipogenic genes, ACC1 and GPAT, were downregulated on the first day at 1000 μg/mL TAO. Consequently, it was found that TAO affects the suppression of cellular senescence, activation of lipid metabolism, and reinforcement of the microenvironment in HepG2 cells, and can be added as a useful component to functional foods to prevent fatty liver disease and cellular senescence, as well as increase the immunoactivity of the liver.