Interfacial chemistry and adhesion between titanium dioxide (TiO<sub>2</sub>) nanotube layers and titanium (Ti) substrates were studied in this Article. TiO<sub>2</sub> nanotube layers were produced on pure Ti by anodization and annealed in air for different time durations. The adhesion of the TiO<sub>2</sub> nanotube layers was then investigated by Rockwell C indentation test. Results show that adhesion of TiO<sub>2</sub> nanotube layers improved with the extension of annealing time. This improvement in adhesion of TiO<sub>2</sub> nanotube layers was analyzed from the viewpoint of interfacial chemistry using energy dispersive X-ray spectrometry (EDS) and X-ray photoelectron spectroscopy (XPS). It suggests that more Ti-O bonds formed in the interface after annealing, and this led to the improved adhesion of the TiO<sub>2</sub> nanoube layers.<br>