Recent research on pseudobiological hydroxyapatite crystal growth and phase transition mechanisms

We have studied the growth and phase transition mechanisms of hydroxyapatite (HAP) and its interaction with a growth factor protein in a simulated physiological environment. Using atomic force microscopy (AFM) and real-time phase shift interferometry, we performed in situ observations of growth in simulated human body fluid solutions seeded with millimeter-sized HAP single crystals produced by hydrothermal synthesis, and we measured the normal growth rate. The step kinetic coefficient (derived from the velocity of growth steps) and the edge free energy (calculated from the variation in the normal growth rate with the degree of supersaturation) both deviated greatly from the standard values for typical inorganic salt crystals and were found to be close to those of protein crystals. This suggests that the growth units of HAP crystals are clusters rather than simple ions and that growth proceeds through the accumulation of these clusters. Observations using dynamic light scattering confirmed the presence of clusters with a diameter of about 1 nm in simulated body fluids. Ab initio analysis of the cluster energy stability indicated that calcium phosphate clusters based on Ca3(PO4)2 units achieve an energy minimum for clusters of the form [Ca3(PO4)2]3. These clusters have S6 symmetry, and, when they are used to build a HAP crystal structure, their structure is likely to become slightly modified, resulting in the formation of C3 structures.Since these clusters would also be the building blocks of amorphous calcium phosphate (ACP), they provide vital clues to the phase transition from ACP to HAP. Using time-resolved static light scattering, we tracked the ACP–HAP phase transition process and found that the degree of coarseness inside a cluster aggregate changes abruptly within a specific time interval and HAP is formed and deposited in the final stages. This suggests that an ACP aggregate changes into HAP as its internal structure becomes regularized. Using this phase transition process, we produced a complex of HAP and the FGF-2 growth factor protein. This complex releases the FGF-2 into a physiological saline solution over a period of at least a week, meaning that it can be used as a pharmacologically active third-generation biomaterial.