Structure, composition, and maturation of newly deposited calcium-phosphate crystals in chicken osteoblast cell cultures

Characterization of the very early calcium phosphate (CaP) crystals deposit ed in bone or in osteoblast cell cultures has been hampered by the overwhel ming presence of organic matrix components and cells that obscure spectral analyses. We have overcome this problem using isolated protein-free crystal s and have obtained new data including P-31 nuclear magnetic resonance (NMR ) spectra for the first time from mineral crystals deposited during osteobl ast calcification in culture. Crystals were isolated from cultures at two t ime points: (a) at first calcium accumulation (day 8-10) and (b) after 60 d ays of culture, to assess maturational changes. The analyses show that the chemical composition overall and short range order of the early and mature crystals are characteristic of the apatite crystals found in young embryoni c chick bone in vivo. No mineral phase other than apatite was detected by a ny of the methods used. P-31 NMR spectroscopy identified the HPO4 groups as those present in bone apatite, Similar to bone apatites, no OH groups were detected by Fourier transform infrared (FTIR) spectroscopy. The temporal m aturational changes in composition and structure of the mineral phase were difficult to assess because of the continuous deposition of crystals throug hout culturing. The pathway of the maturational changes observed were simil ar to those occurring in chick bone in vivo and synthetic apatite crystals in vitro although to a much smaller extent.