Synthesis and in vitro characteristics of biogenic-derived hydroxyapatite for bone remodeling applications

The inorganic component of bone matrix, hydroxyapatite (HAp) (with formula Ca₁₀(PO₄)₆(OH)₂), can be obtained from inexpensive waste resources that serve as excellent calcium precursors. In the present study, HAp nano-powder was synthesized from eggshells (ES) and crab shells (CS) by wet chemical precipitation method. Also, a hybrid sample was considered which is a mixture of HAp nano-powder synthesized from eggshells (25%) and crab shells (75%) (EC). The presence of phosphate, carbonate, and hydroxyl groups in the synthesized powder was confirmed through FTIR analysis. The phase composition was determined using XRD, and elemental analysis revealed a Ca/P ratio ranging from 1.5 to 1.8, confirming the HAp nature of the nano-powder, which ranged in size from 73 to 375 nm. Importantly, preliminary in vitro tests were conducted using mouse preosteoblast cell line MC3T3-E1 to evaluate the cytotoxic effects of the synthesized HAp. The results indicated excellent biocompatibility. Moreover, sample EC exhibited a significantly higher proliferation on days 3, 6, 9, and 12. EC demonstrated promising antimicrobial properties by exhibiting a significantly higher inhibitory effect against the bacteria Streptococcus mutans and Escherichia coli, and the fungi Candida albicans and Aspergillus niger. Additionally, EC displayed notable antioxidant activity, with IC₅₀ values of 271.543 µg/ml and 407.764 µg/ml in DPPH and H₂O₂ assays, respectively. Furthermore, it showed strong anti-inflammatory properties, with a dose-dependent inhibition against protein denaturation. Given these findings, the synthesized HAp holds promise as a potential bone filler and could be beneficial for bone remodeling applications.