It has been demonstrated from previous studies about the killing effect of dihydroartemisinin (DHA) on glioblastoma, which involves multiple aspects: cytotoxicity, cell cycle arrest and invasion inhibition. DHA has the advantages of low cytotoxicity to normal cells, selective killing effect and low drug resistance, making it one of the popular anti-tumor research directions. Ferroptosis is a newly discovered form of cell death characterized by iron dependence and lipid reactive oxygen species (ROS) accumulation. In the present study, we found differences in the expression of transferrin receptors in normal human astrocytes (NHA) and glioblastoma cells (U87 and A172), which may be one of the mechanisms of DHA selective killing effect. Through the determination of ferroptosis-related protein expression, we found that the significant decrease of GPX4, accompanied by the constant expression of xCT and ACSL4, suggesting GPX4 was a pivotal target for DHA-activated ferroptosis in glioblastoma. Total and lipid ROS levels were increased and all these results could be reversed by the ferroptosis inhibitor, ferrostatin-1. These findings demonstrated ferroptosis would be a critical component of cell death caused by DHA and GPX4 was the main target. All these results provide a novel treatment direction to glioblastoma. The association between ferroptosis and polyamines is also discussed, which will provide new research directions for ferroptosis caused by DHA in glioblastoma.