Sonochemical synthesis, characterisation and biological evaluation of nano hydroxyapatite for potential hard tissue engineering applications

Today, there is a great demand for advanced biosynthetic bone like materials for the development of biomedical devices and implants for use in a number of tissue engineering applications. During the first part of this thesis research project, the development of a new chemical route using calcium nitrate [Ca (NO3)2 .4H2O] and potassium dihydrogen phosphate [KH2PO4] as the main reactants was initiated. The synthesis process was carried out under the influence of low powered ultrasound irradiation, while the Ca:P ratio was maintained at 1.67 and the pH was maintained at 9. The resultant white precipitate was then thermally treated in a conventional tube furnace to produce the nanometre sized hydroxyapatite powder. The manufactured nanometre sized hydroxyapatite powders were then characterised using X-ray diffraction (XRD), field emission scanning electron microscopy (FESEM) and Fourier transform infrared spectroscopy (FT-IR). The XRD powder diffraction data was then used to model the crystal structure of the hydroxyapatite using the Endeavour® and the Materials Studio® software packages. Based on the successful first step, (totally inorganic ceramic), the procedure was adapted to manufacture a series of gelatine/hydroxyapatite composites that attempts to replicate the complex organic-inorganic ceramic composite of natural bone tissue, which consists of organic collagen fibrils with embedded inorganic nanometre, sized crystalline hydroxyapatite plates. The high degree of biological functional groups found in collagen based gelatine was found to be a suitable additive for a series of low concentration organic-inorganic ceramic composites. The addition of low concentrations of gelatine was found to act as a template, which formed organicinorganic ceramic composites similar to natural bone tissue. The composites were then characterised using X-ray diffraction (XRD), field emission scanning electron microscopy (FESEM) and Fourier transform infrared spectroscopy (FT-IR). To optimise the manufacture of nanometre sized hydroxyapatite based ultra fine powders, the influence of the thermal techniques used (Furnace heating & Microwave heating) and the use of low power ultrasound irradiation used during the synthesis process was investigated. The refined and optimised manufacturing process was then used to synthesize a 30 ± 5 nanometre sized powder. The powder was then compressed into a series of pellets before being sintered at temperatures ranging from 650 to 1250 ˚C to form ceramics of varying porosity and mechanical strength. The crystal size, crystalline structure and morphology of ultra fine powders and ceramics pellets were investigated using both X-ray diffraction (XRD) and Field Emission Scanning Electron Microscopy (FESEM). Further characterisation techniques such as Brunauer-Emmett- Teller (BET) particle surface area, porosity, bulk density, Vickers hardness and yield strength of the pellets sintered at various temperatures was evaluated. The ceramic pellets were then used in an in vitro biodegradability assessment and an in vivo (Ovine) study. The in vitro biodegradability assessment revealed a very small release of Ca2+ ions during the degradation of hydroxyapatite in a phosphate buffer solution. This release rate was found to be much smaller than the larger quantities of Ca2+ ions that are normally involved in natural bone remodelling processes. The in vivo testing revealed that the ceramic pellets were extremely biocompatible and the matrix porosity promoted both cell migration and colonisation throughout pellet. At the end of week 13, the presence of new collagen and large areas of bone matrix was seen throughout the pellet without any significant inflammatory response present in any of the samples. This highly positive biological response in the in vivo study indicates that the sonochemically assisted synthesis of a 30 ± 5 nanometre hydroxyapatite powders used to produce ceramic pellets has the potential to be used as tissue scaffold in hard tissue engineering applications