Tibolone (Org OD14) is a tissue-specific steroid with estrogenic effects on the bone and vagina but not endometrium or breast and has been shown to prevent ovariectomy-induced bone loss in young and old rats. We evaluated the effect of long-term tibolone treatment on bone parameters in mature ovariectomized (OVX) rats. Six-month-old rats were allotted to one of six groups (n = 8). Sham-operated and control OVX groups received vehicle, whereas other groups (all OVX) received tibolone (125, 250, or 500 μg/day orally) or 17α-ethinylestradiol (EE; 24 μg/day orally) for 16 months. Treatment with tibolone prevented ovariectomy-induced bone loss in peripheral (femur and tibia) and axial (L1-L2 and L4) skeleton. In peripheral skeleton, tibolone and EE prevented loss of bone mass and quality to a similar extent. Tibolone dose-dependently inhibited trabecular bone volume loss in L1-L2 and tibia, and at 500 μg/day it inhibited 88% of L1-L2 and 55% of tibial volume loss (p ≤ 0.05 in each case). Tibolone, 500 μg, resulted in 10% greater cortical strength of femur (p ≤ 0.05) and 60% greater compressive strength of L4 (p ≤ 0.05) compared with vehicle-treated OVX rats. Tibolone and EE inhibited bone resorption and turnover, assessed by urinary deoxypyridinoline/creatinine and plasma osteocalcin, respectively. We conclude that 16 months of tibolone treatment prevents ovariectomy-induced deterioration of axial and peripheral skeleton and preserves cortical and trabecular bone strength by reducing bone resorption.