Objectives: The action of endodontic instruments on root canal walls leaves dentine shavings, which, mixed with organic residues typical of the canal and with preparation auxiliary substances, form the smear layer. It is clear that the presence of this magma greatly compromises the quality of subsequent phases, hindering suitability of the therapeutic protocol. The purpose of this study was to evaluate the root dentine permeability of different chemical substances used during chemical-surgical preparation. Methods: Twenty human single-rooted teeth were selected, divided into 4 groups: G1: 2% chlorhexidine gel combined with distilled water; G2: 2% chlorhexidine gel combined with distilled water and final irrigation with 17% EDTA-T; G3: Endo PTC gel combined with 0.5% sodium hypochlorite and 17% EDTA-T; and G4: Endo PTC cream combined with 0.5% sodium hypochlorite and 17% EDTA-T. The specimens were made impermeable with ethyl cyanoacrylate and dyed with 1% Rhodamine B. The roots were sectioned longitudinally and the dyed area was analyzed using the ImageLab program. Results: The results showed that the greatest average infiltration (%) was observed for G4 (38.53), followed by G2 (31.95), G3 (30.45) and G1 (20.18). The Kruskall-Wallis statistical test showed significant statistical differences between groups G1 and G2 (p<0.05) and G1 and G4 (p<0.01). Conclusions: According to the methodology used, it could be concluded that 0.5% sodium hypochlorite combined with Endo PTC cream and EDTA-T enabled greater dye infiltration.