Sulforaphane (SFN) is considered an antioxidant agent, but the biological effects on hypoxia-treated osteoblasts remain unclear. Therefore, the aims of this study were to investigate the effects of SFN on the activity and mineralization of osteoblasts in hypoxia. Osteoblasts were treated with hypoxia with or without SFN, and apoptosis was assayed with caspase 3 Activity Assay Kit and flow cytometer. The levels of reactive oxygen species (ROS) were measured with DCFH-DA. The levels of glutathione (GSH) and glutathione disulphide were determined by the o-phthalaldehyde fluorimetric assay. Mineralization of Osteoblasts was detected by Alizarin red staining and alkaline phosphatase (ALP) staining, and the relative proteins levels were examined by Western blotting. Our results showed that SFN reduced the hypoxia-mediated apoptosis and ROS levels in osteoblasts. The utilization of SFN improved the inhibitory effect of osteoblast mineralization by hypoxia. Additionally, the effect of alleviating hypoxia by SFN will be an increase in osteoblast activity. These findings clarify the effects of SFN on hypoxia-treated osteogenesis and will help identify novel therapeutic strategies for the protection of skeletal health.