OBJECTIVESThis study aims to extract and characterize hydroxyapatite (HAp) nanoparticles from the exoskeleton of Portunus sanguinolentus (blood-spotted swimming crab) shells for potential biomedical applications, particularly in bone tissue engineering and periodontal regenerative dentistry.MATERIALS AND METHODSCrab shells were cleaned, dried at 100 °C, and ground into powder. The powder was sintered at 1000 °C to obtain calcium carbonate (CaCO₃), which was then reacted with diammonium hydrogen phosphate [(NH₄)₂HPO₄]. in double-distilled water using a wet chemical method at pH >9. The precipitate was filtered, dried at 100 °C, and sintered at 800 °C to synthesize HAp nanoparticles. Characterization using FTIR, EDX, XRD, and SEM confirmed the nanoparticles' chemical composition, crystallinity, and nanoscale morphology. Biocompatibility was evaluated through MTT and live/dead cell assays on human gingival fibroblasts (HGF) and periodontal ligament fibroblasts (HPDLF). Osteogenic potential was assessed via real-time qPCR for ALP, BMP2, and RUNX2 gene expression and Alizarin Red S staining for calcium mineralization. Statistical analysis was conducted using ANOVA with Tukey's test.CONCLUSIONResults demonstrated that crab shell-derived HAp nanoparticles exhibited excellent crystallinity, biocompatibility, and osteogenic potential. Enhanced cell viability and significant upregulation of osteogenic markers confirmed their role in periodontal bone regeneration. Increased calcium deposition further validated their extracellular matrix mineralization capability. These findings suggest that Portunus sanguinolentus-derived HAp nanoparticles are a promising, sustainable biomaterial for periodontal regenerative applications.CLINICAL SIGNIFICANCEThe use of Portunus sanguinolentus-derived hydroxyapatite presents a sustainable and cost-effective alternative to synthetic biomaterials in dental applications. With excellent biocompatibility and the ability to promote osteogenic differentiation, these nanoparticles hold promise for bone grafting, implant coatings, and periodontal regeneration, supporting eco-friendly and efficient solutions for clinical bone repair and regenerative dentistry.