This study systematically investigated the effects of phytoestrogen glabrene on postmenopausal osteoporosis in an ovariectomy (OVX) rat model. Glabrene administration (25, 50, and 100 mg/kg) for 13 weeks can significantly slow down the body weight gain and slightly increase the uterus weight of OVX rats. The increased levels of U‐Ca, U‐P levels, urine DPD/creatinine, serum ALP, OCN, triglycerides, and total cholesterol induced by OVX were dramatically inhibited in rats, whereas no difference occurred for S‐Ca and S‐P in all groups. Furthermore, glabrene can enhance bone mineral density of the right femur, fourth‐lumbar vertebra and tibia and improve biomechanical parameters, such as femoral neck loading force, three‐point bending of the tibia, and vertebral compression in OVX rats. Moreover, glabrene greatly suppressed the expression of TRAP protein but increased OPG and BGP protein expression in tibia tissue of OVX rats. In addition, OVX‐induced reduction of Lrp‐5, β‐catenin, Runx2, and Osx protein expression was all restored by glabrene treatment. The present study indicated that glabrene might be a potential alternative medicine for the prevention and treatment of postmenopausal osteoporosis via activation of the Wnt/β‐catenin signaling pathway.