This paper aims at assessing the use of nanomaterials in painting conservation and in cleaning practices that could be alternative to the traditional ones to overcome the limits of new green materials. Titanium dioxide nanoparticles have been spread and studied on historical painting surfaces with good results. In particular, the properties of TiO2 nanoparticles have been considered useful for self-cleaning and protective purposes against the accumulation of dirt and dust that represents the first phase in deterioration of historical painting surfaces. TiO2 nanoparticles, prepared in distilled water by Pulsed Laser Ablation in Liquids, were applied on painting mock-ups realized in the laboratory according to old recipes and using historical binders and pigments. The surface characteristics of the painting were investigated by contact angle measurement and by Atomic Force Microscopy. The optical and aesthetical compatibility of the colloidal dispersions with the painting was assessed by spectrophotometry, and then the cleaning efficiency was evaluated by discoloration of a dye under ultraviolet irradiation, at fixed time intervals, using colorimetric technique. Because of the high reactivity of nanoparticles, the possibility of degrading the painting surface, together with the chromatic marker, was examined by colorimetric measurements. The evaluation of the color changes is important for all the materials belonging to cultural heritage, especially painting, for which the color modification induced by protective and/or cleaning interventions could irremediably compromise the work of art.