Phosphorylation-driven rapid intrafibrillar mineralization: A biomimetic strategy for dentin remineralization

This study aimed to investigate whether collagen phosphorylation using polyphosphate (P45) could accelerate intrafibrillar mineralization (IM) of collagen fibrils and achieve efficient remineralization of demineralized dentin.

The collagen fibrils were first pretreated with P45 at 37°C for 2 h, followed by mineralization in a solution containing 1.67 mM CaCl₂, 9.5 mM KH₂PO₄, 150 mM NaCl, 240 μg/mL pAsp at 37°C for 2 h. The collagen fibrils were characterized by Fourier-transform infrared spectroscopy (FTIR), atomic force microscope-infrared spectroscopy (AFM-IR), inductively coupled plasma-mass spectrometry (ICP-MS) and Zeta potential before and after P45 modification. The mineralized collagen fibrils (MCFs) were observed by using the transmission electron microscopy (TEM), stochastic optical reconstruction microscopy (STORM), and scanning electron microscopy (SEM). In addition, the demineralized dentin (etched with 37% H₃PO₄ for 20 s) was pretreated with P45 and remineralized in the above mineralized medium for 3 days. The remineralized dentin were characterized by SEM to evaluate the microstructure, X-ray diffraction (XRD) to determine the mineral phase, and nanoindentation to assess the mechanical properties. Statistical analysis was performed using one-way ANOVA with a significance level of p < 0.05.

After P45 modification, zeta potential of the collagen fibrils decreased from -17.20 ± 1.94 mV to -31.91 ± 4.05 mV, and calcium ion adsorption increased from 0.18 ± 0.03 μg/g to 0.67 ± 0.05 μg/g. The FTIR results showed that the P45 could bind with collagen fibrils via H-bond. The P45 modification accelerated IM of the collagen fibrils, remarkably reducing the time required to achieve complete mineralization from 6 h to 2 h. Similarly, P45 modification significantly promoted dentin remineralization, resulting in the restoration of both the microstructure and mechanical properties. The hardness (0.73 ± 0.03 GPa) and elastic modulus (24.63 ± 4.43 GPa) of the remineralized dentin were comparable to native dentin (0.73 ± 0.04 GPa and 26.03 ± 2.86 GPa, respectively; p > 0.05).

This study develops a phosphorylation-driven interfacial modulation strategy that enables rapid and efficient IM of collagen fibrils, offering a promising biomimetic approach for functional dentin remineralization and hard tissue repair.