To investigate the effect of different toothpaste formulations used prior to dental bleaching with 35% hydrogen peroxide (HP) on the mineral content and surface morphology of enamel. Seventy bovine enamel blocks (4×4×2 mm) were submitted to in vitro treatment protocols using a toothbrushing machine prior to dental bleaching or a placebo procedure (n=10) as proposed in the following groups: unbleached control (PLA), bleached control (HP), and brushing with differing toothpastes prior to HP bleaching, including: potassium nitrate toothpaste containing sodium fluoride (PN), sodium monofluorophosphate/MFP toothpaste (FT), arginine-carbonate (8% arginine) (PA) or arginine-carbonate (1.5% arginine) toothpaste (SAN), and toothpaste containing bioactive glass (NM). Phosphorus concentration in gel ([P]) was evaluated (μg of P/mg of gel), and the elemental levels (wt%) of Ca, P, and Na as well as the proportion between Ca and P and spectra graphics were determined using an energy-dispersive X-ray spectrometer (EDS). The surface morphology was assessed using scanning electron microscopy (SEM). The data were subjected to analysis of variance and the Tukey test (α=0.05). HP demonstrated the greatest [P] values in gel, being statistically different from PLA. The [P] of NM was statistically similar to PLA. HP showed a significant decrease in the Ca% and Ca/P values when compared to PLA in EDS analysis. PA showed Ca/P values statistically different from HP. In accordance with SEM analysis, the PA, SAN, and NM groups presented a smooth and uniform enamel surface, while HP and FT demonstrated some alterations in morphology. The toothpastes containing bioactive glass or arginine carbonate used prior to dental bleaching were effective in protecting enamel against mineral loss promoted by the whitening procedure.