Thermosensitive chitosan/collagen hydrogels are osteoconductive and injectable materials. In this study, we aimed to improve these properties by adjusting the ratio of nanohydroxyapatite particles to calcium carbonate microcapsules in a β-glycerophosphate-crosslinked chitosan/collagen hydrogel. Two hydrogel systems with 2% and 5% nanohydroxyapatite particles were studied, each of which had varying microcapsule content (i.e., 0%, 1%, 2%, and 5%). Quercetin-incorporated calcium carbonate microcapsules were prepared. Calcium carbonate microcapsules and nanohydroxyapatite particles were then added to the hydrogel according to the composition of the studied system. The properties of the hydrogels, including cytotoxicity and biocompatibility, were investigated in mice. The calcium carbonate microcapsules were 2-6 µm in size, spherical, with rough and nanoporous surfaces, and thus exhibited a burst release of impregnated quercetin. The 5% nanohydroxyapatite system is a solid particulate gel that supports homogeneous distribution of microcapsules in the three-dimensional matrix of the hydrogels. Calcium carbonate microcapsules increased the mechanical and physical strength, viscoelasticity, and physical stability of the nanohydroxyapatite hydrogels while decreasing their porosity, swelling, and degradation rates. The calcium carbonate microcapsules-nanohydroxyapatite hydrogels were noncytotoxic and biocompatible. The properties of the hydrogel can be tailored by adjusting the ratio of calcium carbonate microcapsules to the nanohydroxyapatite particles. The 1% calcium carbonate microcapsules containing 5% nanohydroxyapatite particle-chitosan/collagen hydrogel exhibited mechanical and physical strength, permeability, and prolonged release profiles of quercetin, which were superior to those of the other studied systems and were optimal for promoting bone regeneration and delivering natural flavonoids.
Keywords: bone substitutes; calcium carbonate microcapsules; controlled release; nanohydroxyapatite particles; quercetin; thermosensitive chitosan/collagen hydrogel.