Understanding the interactions of adsorbed molecules with TiO2 surfaces is important for diverse applications, such as photocatalysis and self-cleaning surfaces. Herein, we investigated the interactions of adsorbed molecules with surface sites on TiO2. The 1H NMR spectra showed that surface healing led to an interesting upfield shift in the chemical shifts of hydroxyls of adsorbed molecules, which is caused by rapid proton exchange between the defect-involved bridging hydroxyls and adsorbed molecules. Infrared spectroscopy confirmed that surface healing decreases the number of surface hydroxyl groups and enhances the number of non-defective surface Ti sites required for methanol adsorption. Therefore, we proposed that undissociative adsorption of methanol is the preferential interaction mode with the non-defective sites of the TiO2 surface but dynamic equilibrium between dissociative and undissociative adsorption exists around surface defect sites. The increase in non-defective surface sites enhanced the surface ζ potential and hindered interfacial electron transfer activity.