Osteoarthritis (OA) is not only related to the degradation of articular cartilage, but also possibly to the changes of subchondral bone. The purpose of this study was to assess whether specific differences could be resolved from bone composition, as also contributed to OA. These differences were assessed by using Fourier transform infrared spectroscopy (FTIRS). The main parameters including mineral content, carbonate content, crystallinity, collagen cross-linking ratio (XLR) and acid phosphate content were represented with characteristic peak integration. It was found that mineral and carbonate content varied significantly with depths at different OA stages. Mineral content increased with depth in healthy samples, while carbonate content showed opposite trend. The mineral content reduced obviously with OA duration, which was different with carbonate decreasing only at early stage of OA. In addition, the content of acid phosphate, collagen maturity (XLR) and crystallinity slight varied with the OA aggravation. Therefore, the changes in subchondral bone were significantly associated with cartilage degeneration and OA, the associated parameters should be targeted for OA therapies.