Method of fabrication of submicron composite microparticles of hydroxyapatite and ferromagnetic nanoparticles for a protein drug carrier

Composite microparticles of hydroxyapatite (HAp) and ferromagnetic nanoparticles were prepared using algenic acid gel cross-linked by bivalent calcium ions. The resultant size of the composite microparticle could be reduced to the submicron scale using extremely vigorous stirring by a homogenizer at 16 000 r.p.m. The instantaneous gellation induced by the bivalent ion cross-linking was revealed to be advantageous for combining the constituent nanoparticles of HAp and ferromagnetic nanoparticles. Since the structural arrest occurs instantly at the moment of cross-linking by the bivalent calcium ion, the added constituent HAp/ferromagnetic nanoparticles do not have time to be segregated apart from the binding gel; consequently, the submicron-scale composite state of these constituents could be instantaneously immobilized as in the dispersed state in liquid by the applied high shear. Thus, the instantaneous gellation characteristic of the bivalent ion cross-linking is preferable for incorporating different nanoparticles. The retention ability of the test protein (cytochrome c) was examined using the bincinchoninic assay method. Comparison of the protein retention ability with the control sample without HAp nanoparticles revealed that the incorporated HAp predominantly contributes to the retention ability of cytochrome c.