Preparation and characterization of novel Ag doped hydroxyapatite-Fe<inf>3</inf>O<inf>4</inf>-chitosan hybrid composites and in vitro biological evaluations for orthopaedic applications

Abstract

This investigation represents the fabrication of novel composites having the biopolymer chitosan with Ag doped hydroxyapatite (HAP)-magnetite nanoparticles (Fe3O4 NPs). In this study, Ag doped HAP was synthesized by a sol-gel method using different silver concentrations such as 1%, 3% and 5%. Furthermore, Fe3O4 NPs were prepared by a co-precipitation technique and the composite was developed with Ag doped HAP. The fabricated Ag:HAP-Fe3O4 composites were incorporated into the chitosan matrix by a planetary ball milling technique and were tested for their reliability as a promising biomaterial for orthopaedic applications. The present study aims to evaluate the in vitro hemocompatibility of Ag doped HAP-Fe3O4 incorporated chitosan composites using a hemolytic assay at the concentrations of 200, 400, 600, 800 and 1000 μg ml-1. The results have shown that the developed composites exhibit a hemolytic ratio of less than 5%, which proved them as good blood compatibles in nature. The antibacterial activity of the composites was evaluated in Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) bacteria, which determined that the composites can extensively inhibit the active growth of microorganisms. In addition to this, in vitro bioactivity behavior was performed in SBF solution for 7 days in order to analyze carbonated apatite formation on the surfaces of the composites and they were found to have characteristic bone bonding ability. The in vitro biocompatibility of the fabricated nanocomposite (C-3) was examined by MTT assay using NIH-3T3 fibroblast cells for incubation periods of 24 and 48 h, and the exposed concentrations are similar to the hemolytic assay. This MTT assay indicates that the fibroblast cells were non-toxic up to the concentration of 400 μg ml-1 with prominent cell attachment and proliferation on C-3 composites. These prepared composite materials were further characterized by ATR-FTIR, powder-XRD and SEM-EDAX analysis. The research findings have shown that the hybrid composites of 5%@Ag:HAP-Fe3O4-chitosan can be widely used as a favorable material for orthopaedic and dental applications.