Biogenic synthesis of copper oxide nanoparticles using Alpinia calcarata extract promotes osteoblasts...

Copper-based biomaterials are increasingly investigated for bone tissue engineering due to their osteogenic and angiogenic properties. Green synthesis using plant extracts offers an eco-friendly route for producing biocompatible nanoparticles. This study aimed to synthesize copper oxide nanoparticles (CuO NPs) using Alpinia calcarata leaf extract and evaluate their cytocompatibility and osteogenic potential in vitro.

CuO NPs were synthesized by reacting copper nitrate trihydrate with A. calcarata leaf extract under alkaline conditions, followed by calcination. The nanoparticles were characterized using SEM, TEM, XRD, and FTIR to confirm morphology, crystallinity, and functional group interactions. C3H10T1/2 murine mesenchymal stem cells were used to assess cytocompatibility (MTT assay, FDA staining), osteogenic differentiation (Alizarin Red S staining and quantification), and gene expression (qRT-PCR for Runx2, Col-I, and ALP).

SEM and TEM revealed aggregated, spherical nanoparticles of <100 nm, while XRD confirmed crystalline monoclinic CuO and FTIR indicated phytochemical capping. MTT and FDA assays showed CuO NPs were cytocompatible up to 50 μg/mL, with dose-dependent cytotoxicity observed at higher concentrations. Under osteogenic conditions, cells treated with 50 μg/mL CuO NPs displayed significantly enhanced mineral deposition compared to controls. Gene expression analysis demonstrated upregulation of Runx2, Col-I, and ALP, confirming promotion of osteogenic differentiation.

Biogenically synthesized CuO NPs using A. calcarata extract are structurally pure, biocompatible at defined concentrations, and capable of enhancing osteoblast differentiation by stimulating matrix mineralization and osteogenic gene expression. These findings position A. calcarata-mediated CuO NPs as sustainable, multifunctional nanomaterials with promising applications in bone tissue engineering.