Study On Synthesis Of Multi-morphological Hydroxyapatite And Its Cytoicompatibility In Vitro

Hydroxyapatite (HAP) is highly similar to natural bone and tooth in the chemicalcomposition and microstructure, so it has been widely used as hard tissue restorative materials.Origniating from biomineralization, HAP crystals in natural bone and tooth were nano-flakesand hexagonal rods repectively, which are different from those of conventional synthetic HAP.In this study, on the basis of biomineralization, the effect of small molecules on themorphology of hydroxyapatite (HAP) crystals was studied via hydrothermal process withglycine, ethylenediamine, glutamic and sodium citrate working as model moluecules. XRD,FT-IR, Laman Spectrem, SEM, TEM, HRTEM and SAED were used to characterize theproducts. A typical plate-like HAP with a width of50-100nm and a ratio aspect of3-5wasobtained due to the presence of ethylenediamine. Glycine with carboxyl and amide terminalinduced the formation of hexagonal rod-like crystals with a length of500-1500nm in adiameter of50-100nm; and Glutamic induced the formation of plate-like crystals with a lengthof500-1500nm in a diameter of50-200nm, which were organized by nano-rod HAP withabout10nm in diameter; while HAP in a relatively unified size with a flake-like shape of200-500nm in length and20-50nm in width occurred in the presence of sodium citrate. Blankexperiment showed that these features only form in the presence of the model molecules. It wassuggesting that a distinct interaction of polar group with HAP crystal via electrostatic force ormolecular occupying would occur during either HAP nucleation or growth. The positivelycharged amino may occupy the position of Ca2+in the HAP cell, especially on the (001) plane,so the growth along the (001) plane would be restricted and HAP crystal would grow along theside plane, finally a plake-like crystal was obtained. The negatively charged carboxyl grouphas strong complexation with Ca2+resulted in glycine,glutamic and citrate molecules havingdifferent adsorption capacity of HAP surface, and further modulated the morphology of HAPcrystals, altogether with the action between molecule. On the (100) plane, carboxylate groups ofGlutamic acid (terminal carboxylate or side chain carboxylate) were adsorbed mainly on Casites. The amino groups on both sides of the carboxylate layer formed a C–N–C layeredstructure, while further the carboxylate bound Ca2+in another HAP crystals and organized thenano-rod HAP into parally assemabled plate-like shape. In the presence of citrate with3 carboxylate groups, the strong complexation would prevent HAP crystals from thickening. Thestudy indicates citrate’s critical role in interfering with crystal thickening and stabilizing theapatite nanocrystals in bone.The study on cytotoxicity of the different morphologies of nano-hydroxyapatite wasdeveloped by co-culture method. The results showed no obvious cytotoxicity has been detectedat the particle concentration of10μg/ml. No difference in cell activity attributed to morphologyshowed at the detectable concentration range (50-5μg/ml). For evaluating the biocompatibilityand bioactivity of different morphologies of nano-hydroxyapatite crystals with osteoblast, theproliferation of osteoblasts, alkaline phosphatase (ALP) expression and morphologicalobservation were carried out. MTT statistical results showed that the osteoblast activity wasgradually decreased with the increase in the concentration of nano-hydroxyapatite. When theconcentration was lower to10μg/ml, the cell activity kept the same level as the blank one. Asthe nano-particle size increased, osteoblast activity decreased obviously. Nano-hydroxyapatiteinhibits the growth of osteoblasts, but ALP results showed that nano-HAP particles to promotethe expression of alkaline phosphatase. SEM and confocal laser scanning microscopy resultsshowed that no significant difference of cell morphology attributed to the morphologiesoccurred.