Concentration-Dependent Effect of Phosphate Ester on Apatite Induction In Vitro

Abstract

The effect of immobilized phosphate ester on hydroxyapatite formation was studied in vitro using metastable solutions that do not precipitate spontaneously. Phosphophoryn, an acidic phosphoprotein secreted by odontoblast in dentin, was used in this study. Increasing the solution saturation with respect to hydroxyapatite nucleation decreased the mineral induction time (the lag time until mineral formation). Progressive dephosphorylation (50% and 70%) of phosphophoryn by acid phosphatase prolonged the mineral induction time. 95% dephosphorylated phosphophoryn no longer induced mineral formation after 48 hours. The mineral induced by immobilized phosphophoryn was identified to be an apatite by micro-area X-ray diffraction. Using these data and classical nucleation theory, the interfacial tension between the immobilized phosphophoryn and apatite crystal formed on the surface of phosphophoryn was calculated. The interfacial tension on phosphophoryn containing 0.65 mmol phosphate per ml beads was relatively low (89.0 ± 2.0 ergs/cm2), similar to that on hydroxyapatite (91.0 ergs/cm2). Moreover, dephosphorylation of phosphophoryn increased the interfacial tension for hydroxyapatite nucleation to 91.6 ± 1.6 ergs/cm2 for 50%-dephospphorylated phosphophoryn, and 96.0 ± 2.4 ergs/cm2 for 70%-dephosphorylated phosphophoryn. These results indicate that phosphophoryn possesses a significant potential of apatite nucleation and induces apatite nucleation in a phosphate concentration-dependent manner.