Sequential nano-amorphous calcium phosphate application modulates resin–dentin bonding performance and aging stability on demineralized dentin

OBJECTIVES: This study evaluated the effect of sequentially applying a nano-amorphous calcium phosphate (NACP) solution within a two-step self-etch bonding procedure on resin-dentin bonding performance, comparing a fluoride-free and a fluoride-containing adhesive. METHODS: In total, 72 artificially demineralized and 60 sound bovine dentin samples were prepared. A fluoride-free adhesive (Clearfil SE Bond 2, SE) and a fluoride-containing adhesive (Clearfil SE Protect Bond, PB) were each combined with a 5 wt% NACP solution applied either pre (Np) or post primer (pN), or used without NACP as controls (p), yielding six experimental groups in total. Micro-tensile bond strength (µTBS; n = 6 teeth/group) was tested after 24 h (TC0) and after 10,000 thermocycles (TC10K). Two-way ANOVA, Tukey's post hoc test and Weibull analysis were used. Degree of conversion (DC) was measured by Raman spectroscopy, and failure modes were examined under SEM. RESULTS: Surface treatment and its interaction with time significantly affected µTBS (p < 0.001). At TC0, Np-PB and pN-PB showed higher µTBS than most groups, while Np-SE was the lowest. After TC10K, most between-group differences were no longer significant, except for pN-SE, which maintained significantly higher values than Np-SE. Weibull analysis showed declining α for Np-PB and pN-PB, increasing α for pN-SE, and decreased β for Np-PB. DC was largely unaffected except for increases in pN-SE and Np-SE after aging. SIGNIFICANCE: NACP solution influenced bonding in a sequence- and system-dependent manner. Post-primer application was associated with more stable long-term outcomes, supporting its potential as a proof-of-concept "bio-functional booster" approach for improving adhesion on demineralized dentin.