Bone healing is a complex process, and if not managed successfully, it can lead to non-union, metal-work failure, bacterial infections, physical and psychological patient impairment. Due to the growing urgency to minimise antibiotic dependency, alternative treatment strategies, including the use of nanoparticles, have attracted significant attention. In the present study, cerium oxide nanoparticles (Ce4+, Ce3+) have been selected due to their unique antibacterial redox capability. We found the processing routes affected the agglomeration tendency, particle size distribution, antibacterial potential, and ratio of Ce3+:Ce4+ valence states of the cerium oxide nanoparticles. The antibacterial efficacy of the nanoparticles in the concentration range of 50–200 µg/ml is demonstrated against Escherichia coli, Staphylococcus epidermis, and Pseudomonas aeruginosa by determining the half-maximal inhibitory concentration (IC50). Cerium oxide nanoparticles containing a more significant amount of Ce3+ ions, i.e., FRNP, exhibited 8.5 ± 1.2%, 10.5 ± 4.4%, and 13.8 ± 5.8% increased antibacterial efficacy compared with nanoparticles consisting mainly of Ce4+ ions, i.e., nanoparticles calcined at 815 °C.

Dicalcium Phosphate Dihydrate (DCPD) mineral scaffolds alone do not possess the mechanical flexibility, ease of physicochemical properties’ tuneability or suitable porosity required for regenerative bone scaffolds. Herein, we fabricated highly porous freeze-dried chitosan scaffolds embedded with different concentrations of Dicalcium Phosphate Dihydrate (DCPD) minerals, i.e., 0, 20, 30, 40 and 50 (wt)%. Increasing DCPD mineral concentration led to increased scaffold crystallinity, where the % crystallinity for CH, 20, 30, 40, and 50-DCPD scaffolds was determined to be 0.1, 20.6, 29.4, 38.8 and 69.9%, respectively. Reduction in scaffold pore size distributions was observed with increasing DCPD concentrations of 0 to 40 (wt)%; coalescence and close-ended pore formation were observed for 50-DCPD scaffolds. 50-DCPD scaffolds presented five times greater mechanical strength than the DCPD mineral-free scaffolds (CH). DCPD mineral enhanced cell proliferation for the 20, 30 and 40-DCPD scaffolds. 50-DCPD scaffolds presented reduced pore interconnectivity due to the coalescence of many pores in addition to the creation of closed-ended pores, which were found to hinder osteoblast cell proliferation.

Bone damage arising from fractures or trauma frequently results in infection, impeding the healing process and leading to complications. To overcome this challenge, we engineered highly porous chitosan scaffolds (S1, S2, and S3) by incorporating 30 (wt)% iron-doped dicalcium phosphate dihydrate (Fe-DCPD) minerals and different concentrations of cerium oxide nanoparticles (CeO2) (10 (wt)%, 20 (wt)%, and 30 (wt)%) using the lyophilisation technique. The scaffolds were specifically designed for the controlled release of antibacterial agents and were systematically characterised by utilising Raman spectroscopy, X-ray diffraction, scanning electron microscopy, and energy-dispersive X-ray spectroscopy methodologies. Alterations in the physicochemical properties, encompassing pore size, swelling behaviour, degradation kinetics, and antibacterial characteristics, were observed with the escalating CeO2 concentrations. Scaffold cytotoxicity and its impact on human bone marrow mesenchymal stem cell (BM-MSCs) proliferation were assessed employing the 2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide (XTT) assay. The synthesised scaffolds represent a promising approach for addressing complications associated with bone damage by fostering tissue regeneration and mitigating infection risks. All scaffold variants exhibited inhibitory effects on bacterial growth against Staphylococcus aureus and Escherichia coli strains. The scaffolds manifested negligible cytotoxic effects while enhancing antibacterial properties, indicating their potential for reducing infection risks in the context of bone injuries.

We have developed two-layered osteoconductive and antibacterial scaffolds, which mimic the natural structure of bone. The synthetic cortical (Type-1) biomaterials were fabricated from porous titanium embedded with 10% iron-doped brushite minerals. The trabecular (Type-2) biomaterials were fabricated from freeze-dried porous chitosan embedded with 10% iron-doped brushite and cerium oxide solution. The Type-1 and Type-2 biomaterials were conjoined during a freeze-drying process in a chamber maintained at -100 ℃ and 40 mTorr pressure; this approach aided the integration of the trabecular and cortical biomaterials, thus forming dual-layered scaffold. The two-layered scaffolds were characterised using Raman, and Fourier Transform Infrared (FTIR) Spectroscopy, Energy-dispersive X-ray (EDX) Spectroscopy and Scanning Electron Microscopy (SEM) for their physicochemical and structural.

An IR femtosecond pulsed laser was used for micropatterning of biomineral containing chitosan membranes, aiming to enhance bone mineralization and angiogenesis. Pre and post irradiation materials have been characterized with XRD, SEM and spectroscopic techniques.