ObjectivesThe objectives were to investigate the hardness and chemical composition of sound, demineralized and pH-cycled bovine enamel and determine their influence on demineralization and remineralization behavior. MethodsNinety-four, 5 × 5 × 2-mm bovine enamel specimens were demineralized using three different times [(24 h (n = 33), 48 h (n = 30), 96 h (n = 31)]. The specimens were then pH-cycled using either 367 ppm F sodium fluoride or deionized water. Knoop hardness (HK) and energy-dispersive X-ray spectroscopy (measured elements: Ca, P, F, C, Mg, N) were performed at three stages (sound, after demineralization, after pH-cycling) and transverse microradiography was performed after demineralization and pH-cycling. Comparisons were determined by ANOVA. ResultsResults showed that HK, integrated mineral loss and lesion depth were significantly different between stages, demineralization times and treatments. The weight% of F at the surface was significantly affected by treatment, irrespective of demineralization time, while the Ca:P ratio of the enamel remained stable even after de- and remineralization protocols. The F in fluoride groups and the artificial saliva in non-fluoride groups were both able to induce enamel remineralization, indicating the protective effect of salivary pellicle against demineralization even in the absence of fluoride. ConclusionsHarder specimens and those with greater surface F weight% were less susceptible to demineralization and were more likely to remineralize. However, the amount of surface Ca and P did not influence de- or remineralization behavior. Clinical SignificanceThis in vitro study can help clinicians better understand the caries process and the impact of the physical and chemical characteristics of enamel on its behavior during de- and remineralization. The over-the-counter fluoride toothpaste containing 1100 ppm-F was used, and was able to produce a mineralized enamel surface layer.