The activities of Ru/Al2O3 catalysts for hydrodesulfurization (HDS) of thiophene were studied, using a differential microreactor at atmospheric pressure. The surface properties of the catalysts were determined by NH3-TPD, XPS analysis, and IR spectroscopy. The Ru/Al2O3 catalysts were pretreated by three different methods: oxidized in air and subsequently reduced in H2 (Ru-OR catalyst), directly reduced in H2 (Ru-R catalyst), and sulfided in 10% H2S/H2 (Ru-S catalyst). All of the catalysts, pretreated at a temperature of 300°C, were among the most active, while the activity of Ru-S at 400°C was the least. Mild sulfidation at 300°C in flowing 10% H2S/H2 promoted the activity of the Ru/Al2O3 catalysts for the HDS of thiophene, while severe sulfidation at 400°C decreased the activity. Two NH3 peaks at low and middle temperatures and one peak of N2 and H2 at high temperature were observed for the three kinds of the catalysts in the NH3-TPD study. The XPS analysis showed that the low-, middle-, and high-temperature peaks in NH3-TPD were assigned to RuO2, acid sites on alumina, and Ru0 metal, respectively. The mass spectrometry indicated that the high-temperature peak of NH3-TPD consisted of N2, formed from the decomposition of NH3 on the ruthenium sites of the catalysts. The FTIR study indicated that the low-temperature peak of NH3-TPD was related to Lewis acid sites of the Ru catalysts. The HDS activity of the Ru/Al2O3 catalysts, for the HDS of thiophene, depended on the metallic ruthenium on the surface of the catalysts, rather than surface acidity measured in NH3-TPD.