Background The Atractylenolide I (AO-I) is one of the main bioactive components of Atractylodes macrocephala Koidz (AMK), which is a healthy food in China with long history. This study aims to investigate the effects and mechanism of AO-I on estradiol (E 2) secretion and lipid metabolism induced by Dihydrotestosterone (DHT) in human granulosa-like tumor cells (KGN). Methods and results E 2 secretion and lipid metabolism of KGN cells were evaluated with CCK-8, Enzyme‐linked immunosorbent assay, lipid droplets assay, western blot and immunofluorescence. The results showed that DHT significantly decreased the cell viability, increased the E 2 secretion, the protein levels of CYP19, and lead to lipid metabolism disorder in KGN cells, which was characterized by lipid droplets (DLs) accumulation, decreased expression of PPARγ and increased the protein levels of FABP4. Notably, AO-I could significantly reverse the adverse effect induced by DHT and alleviated the impaired E 2 secretion and lipid metabolism through regulation of PPARγ in KGN cells. Conclusion Consumption of AO-I is beneficial to DHT induced ovarian granulosa cell dysfunction and disorder of lipid metabolism via the regulation of PPARγ. This research may provide the theoretical basis for AO-I as a dietary supplement to alleviating ovarian dysfunction.