Hematopoietic stem cells (HSCs, CD34⁺ cells) have shown therapeutic efficacy for transplantation in various hematological disorders. However, a large quantity of HSCs is required for transplantation. Therefore, strategies to increase HSC numbers and preserve HSC functions through ex vivo culture are critically required. Here, we report that expansion medium supplemented with ASPP 049, a diarylheptanoid isolated from Curcuma comosa, and a cocktail of cytokines markedly increased numbers of adult CD34⁺ cells. Interestingly, phenotypically defined primitive HSCs (CD34⁺CD38^-CD90⁺) were significantly increased under ASPP 049 treatment relative to control. ASPP 049 treatment also improved two functional properties of HSCs, as evidenced by an increased number of CD34⁺CD38^- cells in secondary culture (self-renewal) and the growth of colony-forming units as assessed by colony formation assay (multilineage differentiation). Transplantation of cultured CD34⁺ cells into immunodeficient mice demonstrated the long-term reconstitution and differentiation ability of ASPP 049-expanded cells. RNA sequencing and KEGG analysis revealed that Hippo signaling was the most likely pathway involved in the effects of ASPP 049. These results suggest that ASPP 049 improved ex vivo expansion and functional preservation of expanded HSCs. Our findings provide a rationale for the use of ASPP 049 to grow HSCs prior to hematological disease treatment.