Osteoclasts play an important role in maintaining the relative stability of bone mass. Abnormal number and function of osteoclasts are closely related to osteoporosis and osteolytic diseases. Thiaplakortone B (TPB), a natural compound derived from the Great Barrier Reef sponge Plakortis lita , has been reported to inhibit the growth of the malaria parasite, Plasmodium falciparum , but its effect on osteoclastogenesis has not been previously investigated. In our study, we found that TPB suppresses the receptor activator of nuclear factor‐κB (NF-κB) ligand (RANKL)-induced osteoclast formation and resorption activity by tartrate‐resistant acid phosphatase (TRAcP) staining, immunofluorescence staining of F-actin belts and hydroxyapatite resorption assay. Furthermore, using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting analysis, we discovered that TPB inhibits osteoclast-specific genes and proteins expression. Mechanistically, TPB blocks multiple upstream pathways including calcium oscillation, NF‐κB, mitogen-activated protein kinase (MAPK) and nuclear factor of activated T cells 1(NFATc1) signaling pathways. In vivo , TPB could dampen bone loss in an ovariectomy (OVX) mouse model by micro-CT assessment and histological staining. Therefore, TPB may serve as a potential therapeutic candidate for the treatment of osteoporosis and osteolysis.