Background: To investigate the potential effects of molecular hydrogen on ovarian cancer. Methods: The in vivo study was performed using Hs38.T xenografted BALB/c nude mice. The mean tumor volume was monitored during 6 weeks of hydrogen inhalation. Immunohistochemistry (IHC) staining was performed to determine the Ki67 and CD34 expression. The in vitro effects of molecular hydrogen on proliferation of Hs38.T and PA-1 were determined by Cell Counting Kit (CCK)-8 assay. Matrigel invasion assay was performed to determine the cell invasion ability. Wounding assay was employed to examine the motile nature of ovarian cancer cells. Colony formation assay was performed to investigate the effect of molecular hydrogen in tumorigenicity. To further investigate the effects of molecular hydrogen on cancer stem cells (CSCs) properties, we performed sphere-formation assays. Results: The in vivo study demonstrated that 6 weeks of hydrogen inhalation significantly inhibited tumor growth, as evidenced by decreased mean tumor volume (32.30%) and Ki67 expression (30.00%). Hydrogen treatment decreased the expression of CD34 (74.00%) demonstrating its anti-angiogenesis effects. The in vitro study showed that hydrogen treatment significantly inhibits cancer cell proliferation, invasion, migration and colony formation both in Hs38.T and PA-1 cells. An important finding in this study was that molecular hydrogen could also markedly inhibit sphere-forming ability of both PA-1 and Hs38.T cells. Conclusions: Molecular hydrogen may exert anti-tumor role in ovarian cancer through suppressing the proliferation of CSCs-like cells and angiogenesis.