The carbonate and phosphate vibrational modes of different synthetic and biological carbonated apatites were investigated by Raman microspectroscopy, and compared with those of hydroxyapatite. The nu1 phosphate band at 960 cm-1 shifts slightly due to carbonate substitution in both A and B sites. The spectrum of type A carbonated apatite exhibits two nu1 PO43- bands at 947 and 957 cm-1. No significant change was observed in the nu2 and nu4 phosphate mode regions in any carbonated samples. The nu3 PO43- region seems to be more affected by carbonation: two main bands were observed, as in the hydroxyapatite spectrum, but at lower wave numbers. The phosphate spectra of all biominerals apatite were consistent with type AB carbonated apatite. In the enamel spectrum, bands were observed at 3513 and at 3573 cm-1 presumably due to two different hydroxyl environments. Two different bands due to the carbonate nu1 mode were identified depending on the carbonate substitution site A or B, at 1107 and 1070 cm-1, respectively. Our results, compared with the infrared data already reported, suggest that even low levels of carbonate substitution induce modifications of the hydroxyapatite spectrum. Increasing substitution ratios, however, do not bring about any further alteration. The spectra of dentine and bone showed a strong similarity at a micrometric level. This study demonstrates the existence of acidic phosphate, observable by Raman microspectrometry, in mature biominerals. The HPO42- and CO32- contents increase from enamel to dentine and bone, however, these two phenomena do not seem to be correlated.

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This study investigates the adhesion, cytoskeletal changes, and resorptive activity of disaggregated rat osteoclasts cultured on polished slices of three biomaterials: crystalline synthetic hydroxyapatite (HA), carbonated hydroxyapatite (C-HA), and natural calcium carbonate (C). The surface chemistry of each substrate was defined by X-ray diffraction and IR spectroscopy, surface wettability by the dispersive, and the polar components of the surface energies. Osteoclast adhesion was modulated by the polar component of the surface energy: fewer (p < 0.01) osteoclasts adhered to C-HA (97 +/- 20/slice, surface energy 9 +/- 5 mJ/m2) than to HA (234 +/- 16/slice, surface energy 44 +/- 2 mJ/m2) or to C (268 +/- 37/slice, surface energy 58 +/- 0.5 mJ/m2). Actin rings, which are the cytoskeletal structure essential for resorption, developed on all three materials. The area of the actin ring, which is resorbed by local acidification, and the osteoclast area, which reflects osteoclast spreading, were both greater in osteoclasts cultured on HA and C-HA than in those cultured on C. C was resorbed, but HA and C-HA were not. Thus, the surface energy plays an essential role in osteoclast adhesion, whereas osteoclast spreading may depend on the surface chemistry, especially on protein adsorption and/or on newly formed apatite layers. Resorption may be limited to the solubility of the biomaterial.

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Visible Raman and infrared microspectrometry studies performed on fluorapatite and hydroxyapatite powders have shown similar results. Small modifications of the nu2 and nu4 PO(3-)4 tetrahedra bending modes are observed. A small frequency shift of the nu1 mode and modifications on the nu3 mode region accompanied with a simplification of the hydroxyapatite and fluorapatite respective spectra from seven to four bands were observed. A broad and weak band which could be attributed to the Ca-F bond is detected at 311 cm(-1) on the Raman fluorapatite spectra. The phosphate vibration modes are little disturbed by fluoride substitution. This could indicate that phosphate groups interact strongly between themselves and weakly with substituted atoms (i.e. hydroxyle and fluoride atoms). Whatever crystallographic model is considered, the number of bands observed is always lower than the number of calculated ones, even for hydroxyapatite, whose symmetry is lower than that of fluorapatite.

Knowledge of the organization of the components of bone is of primary importance in understanding how this tissue responds to stresses and provides a starting point for the design and development of biomaterials. Bone structure has been the subject of numerous studies. The mineralized fiber arrangement in cortical bone is either a twisted or orthogonal plywood structure. Both mineral models coexist in compact bone. Raman polarized spectroscopy offers definite advantages in the study of biological samples, enabling the simultaneous analysis of mineral and organic components and the determination of molecular orientation through the polarization properties of the Raman scattering. In this study, we used the Raman polarization approach to simultaneously investigate the orientation of collagen fibrils and apatite crystals in human cortical bone. Raman bands ratios were monitored as a function of sample orientation. Specific ratios were chosen--such as nu(3) PO(4)/nu(1) PO(4), amide III (1271 cm(-1))/amide III (1243 cm(-1)), and amide I/amide III (1243 cm(-1))--due to their sensitivity to apatite-crystal and collagen-fibril orientation. Based on this original approach, spatial changes were monitored as a function of distance from the Haversian canal. The results revealed simultaneous tilting in intra-lamellar collagen-fibril and mineral crystal orientations. These results are consistent with a twisted plywood organization in the Haversian bone structure at the lamellar level. But at molecular level, the co-alignment of the collagen fibrils and the apatite crystal is observed in the innermost lamellae and becomes gradually less ordered as the distance from the Haversian canal increases. This work highlights the interest of Raman spectroscopy for the multiscale investigation of bone structure.

Despite numerous studies of human biominerals, some problems still remain concerning the relationship between their composition and their structure. For a better understanding of this problem, full spectra of the internal vibrations of human tooth enamel crystallites were obtained through polarized Raman microspectrometry and these are published for the first time. The micro-Raman technique is nondestructive and enables micrometric-scale examination of all the samples with a minimum of artifacts. The spectra show some variation from the predicted bands and many similarities with fluorapatite spectra. The mineral part of enamel is initiated in an organic environment and contains carbonate ions. Despite the carbonation, the crystal structure is preserved. Based on these results, a new description of the structure of apatite crystal is proposed. A box of Ca2+ ions surrounds and isolates the PO43– ions from one another, decreasing the influence of substitutions.