In the present work, the surface hydroxyls on rutile TiO2 that evacuated at different temperature were characterized by in suit FTIR. The action of OH groups in photocatalytic reaction was tested by adsorption and photooxidation of toluene in an in situ IR cell. The results show that there are seven OH groups(with bands at 3 724, 3 700, 3 652, 3 648, 3 610, 3 413 and 3 362 cm-1) on rutile TiO2 with different thermal stability. Among the seven hydroxyl groups, the OH groups with bands at 3 724, 3 700 and 3 652 cm-1 are thermal stable and are ascribed to isolate hydroxyl groups, while the OH groups with bands at 3 648, 3 610, 3 413 and 3 362 cm-1 are thermal unstable that almost disappear after evacuation at 250 ℃. During the adsorption of toluene, only the surface hydroxyls with bands 3 648, 3 610 and 3 652 cm-1 act as adsorption sites for adsorption of toluene.