Bleaching agents have effect on chemical/physical and morphological structure of enamel and dentin that must be taken into account when this therapy is used. The aim of this in vitro study was to evaluate the effects of two bleaching agents containing a high concentration of hydrogen peroxide for professional use on human enamel and dentin surface and to evaluate the potential remineralizing effect of amorphous calcium phosphate gel (ACP). Twenty-five human third molars were divided into two groups and dissected in half and both surfaces were bleached with either ZOOM2 or Opalescence BOOST for 3 x 15 minutes. Vickers microhardness of enamel and dentin was measured before, after the bleaching treatment, and after treatment with artificial saliva and ACP gel or 2-week storage in deionized water. Surface microstructure was evaluated using scanning electron microscopy. The mixed model ANOVA and Wilcoxon Rank Sum test were used. Both bleaching agents showed significant reduction in surface microhardness (p < 0.001 for both BOOST and ZOOM2 application). ZOOM2, which had a lower pH value showed greater decrease in surface microhardness (p = 0.005) compared to BOOST. Post-treatment with artificial saliva and ACP showed significant increase in surface microhardness (p < 0.001). After the bleaching procedure, enamel and dentin surface microstructure showed mild or slight alterations with no loss of superficial structure. In conclusion, both bleaching agents resulted in reduction in surface enamel and dentin microhardness. Treatment with ACP led to increase in surface microhardness, improved surface roughness, and enhanced remineralization of the hard dental tissues.