Deficiency in Vitamin D and its metabolites leads to a failure in bone formation primarily caused by dysfunctional mineralization, suggesting that Vitamin D analogs might stimulate osteoblastic bone formation and mineralization. In this study, we compare the effect of selected Vitamin D analogs and active metabolite, 1α,25-dihydroxyvitamin D 3, 19-nor-1α, 25-dihydroxyvitamin D 2, and 1α-hydroxyvitamin D 2 or 1α,25-dihydroxyvitamin D 2 on bone formation and resorption. In a mouse calvariae bone primary organ culture system, all Vitamin D analogs and metabolite tested-stimulated collagen synthesis in a dose-dependent manner and 19-nor-1α, 25-dihydroxyvitamin D 2 was the most efficacious among three. 19-nor-1α, 25-dihydroxyvitamin D 2 and 1α,25-dihydroxyvitamin D 2 showed similar potencies and 1α,25-dihydroxyvitamin D 3 was less potent than others. Osteocalcin was also up-regulated in a dose-dependent manner, suggesting that the three Vitamin D analogs have the equal potencies on bone formation. 25-Hydroxyvitamin D-24-hydroxylase expression was induced in a dose-dependent manner and 19-nor-1α, 25-dihydroxyvitamin D 2 was less potent than other two compounds. In a mouse calvariae organ culture, all induced a net calcium release from calvariae in a dose-dependent manner, but the potency is in the order of 1α,25-dihydroxyvitamin D 2 ≅ 1α,25-dihydroxyvitamin D 3 > 19-nor-1α, 25-dihydroxyvitamin D 2. In a Vitamin D/calcium-restricted rat model, all caused an elevation in serum calcium in a dose-dependent manner. There is no significant difference between 1α,25-dihydroxyvitamin D 3 and 1α-hydroxyvitamin D 2 in potencies, but 19-nor-1α, 25-dihydroxyvitamin D 2 is at least 10-fold less potent than the other two compounds. Our results suggest that Vitamin D analogs have direct effects on bone resorption and formation, and 19-nor-1α, 25-dihydroxyvitamin D 2 may be more effective than 1α,25-dihydroxyvitamin D 3 and 1α-hydroxyvitamin D 2 on stimulating anabolic bone formation.