The deactivation of sulfate-promoted iron oxide in the esterification of acetic acid and n-butanol was studied. The sulfate-promoted iron oxide was used ten runs and 10 h, continually and accumulatively. After ten-run continual use of the catalyst, a considerable deactivation happened to it. The fresh and the deactivated catalysts were compared by means of many characteristic methods including FTIR, XRD, BET, SEM, TG-DSC, and NH3-TPD, to disclose some possible reasons for the deactivation of sulfate-promoted iron oxide in the esterification. Based on the comparative analyses of IR patterns of the fresh catalyst and the deactivated one, a deactivation mechanism is tentatively proposed. Namely, surface sulfate groups, which are originally coordinated to Fe3+ cations and can so induce and generate strong Lewis acidity of Fe3+ cations, may have been gradually turned into free sulfate groups and sulfate esters arisen from strong Lewis-acidic Fe3+ cations' being hydrolyzed by H2O and their being alcoholyzed by n-butanol, which leads to a gradual destruction of the originally strong coordination between Fe3+ cations and surface sulfate groups, so leading to the acidity degradation of the catalyst, and so finally leading to the deactivation of it. Emphatically, in the proposed mechanism, the water produced from the esterification may play a key role on the deactivation of the catalyst, because it can directly hydrolyze some strong Lewis-acidic Fe3+ cations of the catalyst and indirectly promote the alcoholysis of them, to form weak Lewis-acidic Fe-OH species. The deactivated catalyst has a larger crystallinity, a smaller specific surface area, a smaller sulfate groups content, a weaker acidity than the fresh. All these phenomena, accompanying the deactivation of sulfate-promoted iron oxide, can be interpreted by the proposed deactivation mechanism very well.