Reduced bone density is a well-known feature of osteoporosis, yet little is known about the changes in the chemical composition of bone or the impact of such chemical changes on fracture risks. Using ovariectomized cynomolgus monkeys (Macaca fascicularis) as a model for the menopausal onset of osteoporosis, we examined the microscopic chemical changes of bone measured by synchrotron infrared microspectroscopy as a function of time after ovariectomy. The results demonstrate that cortical bone formed 1 or 2 years after ovariectomy, as identified by fluorochrome labeling, has a higher phosphate content (PO4(3-)/matrix ratio), a lower carbonate content (CO3(2-)/matrix ratio), and more mature collagen cross-links (nonreducible cross-link/reducible cross-link ratio) than that formed in sham-operated controls. Trabecular bone after ovariectomy shows no changes in phosphate content, a lower carbonate content, and immature collagen cross-linking. Treatment with a bone turnover suppressor, (nandrolone decanoate) reverses most of the ovariectomy-induced chemical changes in the cortical bone to the levels of the ovary-intact controls, but has little effect on the trabecular bone. These results demonstrate that bone newly synthesized after ovariectomy is chemically different from healthy bone within specific bone regions, which may contribute to reduced bone quality in osteoporosis.