Eggshell-derived amorphous calcium phosphate for upcycled bone repair: An In-Vivo study

Critical-sized bone defects require grafting materials that support early osteogenesis and predictable defect bridging. In this study, we evaluated eggshell-derived amorphous calcium phosphate (ACP) granules in a rat calvarial defect model and compared them with ACP obtained from commercial hydroxyapatite (HAp) and with cross-linked hyaluronic acid (HA) gel alone. Eggshell-derived and HAp-derived ACP were synthesized by the same dissolution–precipitation route, using formulations previously characterized in detail for composition, amorphous phase stability, and in vitro bioactivity. Bilateral 5 mm calvarial defects in rats were filled with HA gel (INT/HA), HA + HAp-derived ACP (Control ACP), or HA + eggshell-derived ACP (Eggshell ACP). Grafts were mixed immediately before implantation using a dual-syringe system. Bone regeneration at 2 and 4 weeks was assessed by micro-computed tomography and histology. Both ACP-containing groups enhanced bone volume fraction and reduced trabecular separation compared with HA alone, particularly in the central region of the defect. Eggshell ACP promoted more consistent defect bridging, with a continuous lamellar bone-like layer and more mature collagenous and mineralized tissue at the bone–graft interface. Within the limitations of a non-load-bearing small-animal model, these findings indicate that eggshell-derived ACP combined with HA supports early bone regeneration and exhibits higher osteogenic activity than HA alone and HAp-derived ACP, warranting further investigation as a bone graft candidate.