Biomolecule-functionalized exfoliated-graphene and graphene oxide as heteronucleants of nanocrystalline apatites to make hybrid nanocomposites with tailored mechanical, luminescent, and biological properties

Abstract

Nanocrystalline apatite (Ap), known for its exceptional biological properties, faces limitations in hard tissue engineering due to its poor mechanical properties. To overcome this limitation, we investigated the preparation of nanocomposites through heterogeneous nucleation of calcium phosphate on exfoliated graphene (G) and graphene oxide (GO) flakes, selected for their outstanding mechanical properties. The flakes were treated (functionalized) with amino acids of varying isoelectric points—namely L-Arginine (Arg), L-Alanine (Aln) and L-Aspartic acid (Asp)— as well as citrate (Cit) molecules. Furthermore, Tb³⁺ was incorporated into the formulations to introduce luminescence and further enrich the functionality of the composite. The synthesis was conducted using the sitting drop vapor diffusion method. Functionalized GO/Ap nanocomposites significantly improved roughness, adhesion forces and elastic modulus compared to Ap and G-based particles. GO-Asp-Ap-Tb nanocomposites exhibited the highest roughness (163.8 ± 116.2 nm), while G-Cit-Ap had the lowest (6.8 ± 5.6 nm). In terms of adhesion force, GO-Cit-Ap-Tb reached the highest value (31.06 ± 13.3 nN), while G-Arg-Ap had the lowest (3.7 ± 1.8 nN) compared to Ap (13.6 ± 3.2 nN). For the elastic modulus, GO-Aln-Ap-Tb demonstrated the greatest stiffness (3489 ± 101.01 MPa) compared to Ap (30.2 ± 6.5 MPa), while G-Aln-Ap-Tb showed the lowest (17.2 ± 8.4 MPa). Concerning their luminescence, regardless of G/Ap and GO/Ap, the relative luminescence intensities depended on the biomolecule used and decreased in the order Arg > Aln > Asp and Cit. Furthermore, G/Ap and GO/Ap nanocomposites demonstrated good biocompatibility on murine mesenchymal stem cells at low concentrations, showing cell viabilities exceeding 80 % at 0.1 μg/mL. This research offers a novel approach to enhancing the mechanical properties of apatites while preserving their good biocompatibility properties and introducing new functionalities (i.e. luminescence) in the composites, thereby expanding their range of applications in hard tissue engineering.