Postmenopausal osteoporosis (PMO) is a progressive bone disease characterized by the over-production and activation of osteoclasts in elderly women. In our study, we investigated the anti-osteoclastogenic effect of evodiamine (EVO) in vivo and in vitro, as well as the underlying mechanism. By using an in vitro bone marrow macrophage (BMM)-derived osteoclast culture system, we found that EVO inhibited osteoclast formation, hydroxyapatite resorption and receptor activator of NF-κB ligand (RANKL)-induced osteoclast marker gene and protein expression. Mechanistically, we found that EVO inhibited the degradation and RANKL-induced transcriptional activity of IκBα. RANKL-induced Ca2+ oscillations were also abrogated by EVO. In vivo, an ovariectomized (OVX) mouse model was established to mimic PMO, and OVX mice received oral administration of either EVO (10 mg/kg) or saline every other day. We found that EVO can attenuate bone loss in OVX mice by inhibiting osteoclastogenesis. Taken together, our findings suggest that EVO suppresses RANKL-induced osteoclastogenesis through NF-κB and calcium signalling pathways and has potential value as a therapeutic agent for PMO.