Octacalcium Phosphate Overgrowth on β-Tricalcium Phosphate Substrate in Metastable Calcium Phosphate Solution
The effects of the particle size of β-tricalcium phosphate (β-Ca3(PO4)2; β-TCP) on octacalcium phosphate (Ca8(HPO4)2(PO4)4·5H2O; OCP) overgrowth on a β-TCP substrate were evaluated under physiological conditions by using two types of substrate; one composed of micrometer-sized particles (micro-TCP substrate) and one composed of nanometer-sized particles (nano-TCP substrate). When the β-TCP substrate was immersed in a simple calcium phosphate solution, it was quickly covered with OCP. The morphology and size of the OCP crystals, as well as the structure, thickness, and crystal density of the overgrown OCP layer, depended on the β-TCP particle size. In case of the micro-TCP substrate, OCP crystals grew directly on the micrometer-sized particles. In case of the nano-TCP substrate, string-like (S) precipitates initially deposited, and then flake-like (F) crystals formed on them. Plate-like (PL) OCP crystals grew on the flake-like crystals; as a result, a three-layer structure (S-layer/F-layer/PL-layer) was formed. Small amounts of tiny OCP crystals and HAp-nanofibers precipitated in the micro-TCP substrate, whereas only HAp-nanofibers precipitated in the nano-TCP substrate. Thus, various types of OCP-overgrown layers were fabricated on β-TCP scaffold. These findings will facilitate the structural design of OCP-coating layers on a β-TCP scaffold.
KeywordsOctacalcium phosphate Coating Wet chemical method Overgrowth β-tricalcium phosphate Scaffold Particle size
Both β-TCP and OCP have been proven to have promising osteoconductive characteristics. β-TCP has been applied in the form of granules and three-dimensional (3D) scaffolds (Hench and Polak 2002; Karageogiou and Kaplan 2005; Wang et al. 2015). The biocompatibility and degradability of conventional micrometer-sized β-TCP powder have been improved by the use of nanometer-sized β-TCP (Zhang et al. 2008; Kato et al. 2016). On the other hand, OCP is a metastable phase of HAp and tends to transform into HAp spontaneously (Brown et al. 1962). The intrinsic properties of OCP are hypothesized to be responsible for its excellent performance in vivo; moreover, implanted OCP granules have provided cores for nucleating multiple osteogenic sites (Suzuki et al. 1991, 2006). The combined usage of β-TCP with its better formability and OCP with its better osteoconductivity would boost the potential of both materials as a bone graft substitute. A practical way to achieve this is coating β-TCP scaffolds with OCP.
The purpose of the present study was to examine OCP formation on a β-TCP substrate in a simple calcium phosphate solution and to evaluate the effect of the particle size of the β-TCP substrate on the overgrowth of OCP under physiological conditions. To achieve this, micrometer- and nanometer-sized β-TCP particles were used to form β-TCP substrate.
28.2 Materials and Methods
Each substrate section was immersed in a calcifying solution (5 mM CaCl2, 5 mM K2HPO4+KH2PO4, 50 mM CH3COONa, pH 6.2, 37 ± 0.5 °C) for required periods. After the reaction terminated, the substrate was rinsed in Mill-Q water and in 99.5% ethanol and dried in air. The crystals in the overgrown layer and inside the substrate were characterized using powder XRD, microbeam XRD, thin-film in-plane XRD, FE-SEM, and TEM.
28.3 Results and Discussion
28.3.1 Early Stage of Overgrowth
28.3.2 Later Stage of Overgrowth
On both substrates, OCP crystals grew in the same manner. As the immersion period increased, the length of OCP crystals in the c-axis direction increased: 4–6 μm after 60 min, 10 μm after 3 h, 15–20 μm after 5 h, and 66–70 μm after 20 h. In the case of the nano-TCP substrate, plate-like OCP crystals (PL) grew on the layer of small flake-like crystals (F-layer), under which the layer of string-like precipitates (S-layer) had formed, and as a result, a three-layer structure was observed (Fig. 28.3d3). On the contrary, in the case of the micro-TCP substrate, OCP crystals grew directly on β-TCP particles, and no such structure was observed (Fig. 28.3c3).
Thus, varying the particle size of the β-TCP had great effect on the early stage of overgrowth and precipitates inside the substrates. Various types of OCP-coating layers were formed on β-TCP substrate. There is a general consensus that the physical properties of the coating layer, i.e., its thickness and topography, affect the in vivo performance of the coated material (Curtis and Wilkinson 1997; Anselme and Bigerelle 2011). These findings will facilitate the structural design of OCP-coating layers on a β-TCP scaffold.
This study was supported by a Grant-in-Aid for Scientific Research from the Japan Society for the Promotion of Science (JSPS KAKENHI C; 16K04954).
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