Bone-like growth of hydroxyapatite in the biomimetic coating of Ti-6Al-4V alloy pretreated with protein at 25 °C

Commercial-grade dense Ti-6Al-4V alloy substrate was mechanically roughened, cleaned, and treated with a globular protein [bovine serum albumin (BSA)] for 4 h. Biomimetic calcium phosphate (Ca-P) coating was applied onto the above-treated substrate by immersion into simulated body fluid (SBF) at 25 °C for a period of 4 days, with periodic replacement by freshly prepared SBF at 48-h intervals. After 4 days, branched micron-sized fibers of hydroxyapatite (HAp), resembling the structure of bone, were obtained, connecting the clusters of HAp crystal plates in the coating (thickness ∼200 μm) on the substrate surface. Structural and compositional characterization of the coating was carried out using field emission scanning electron microscopy (FE-SEM) with energy-dispersive x-ray analysis unit (EDX) facility, x-ray diffraction (XRD), and Fourier transform infrared (FTIR) data. In vitro cytotoxicity (ISO 10993-5, 1999), cell adhesion assays, and phase contrast microscopy were performed using NIH 3T3 fibroblast cell lines to ascertain the bioactivity of the coated substrates, with and without protein treatment. Based on our study, we propose a correlation between a specific physical structure of the HAp coating and its biological properties.