Water-based enhanced oil recovery (EOR) from carbonates is usually restricted by initial wetting properties, especially in naturally fractured carbonates. Carboxylic material in the crude oil, quantified by the acid number (AN), is regarded as the most important wetting parameter. In this paper, it is shown experimentally that, for a given AN, the temperature and the amount of sulfate present in the formation water will affect the wetting condition significantly. The concentration of sulfate in the formation water is usually low because of a high amount of Ca2+, anhydrite, CaSO4(s), is precipitated at high temperatures. The interaction between small amounts of sulfate dissolved in the formation water and the rock surface was studied using chalk cores. The relative amount of sulfate dissolved in the pore water, SO42-(aq), and sulfate adsorbed onto the chalk surface, SO42-(ad), was quantified at different temperatures of 20, 50, 90, and 130 degrees C. Below 50 degrees C, the relative amount of SO42- (aq) and SO42-(ad) was quite constant, but above 50 degrees C, SO42- (aq) decreased, while SO42-(ad) was not significantly affected by increasing the temperature. Sulfate was precipitated as CaSO4(s) and retained in the core at 130 degrees C. Spontaneous imbibition of formation water, free from sulfate, was also conducted at 50 degrees C into mixed wet chalk cores, which were aged at different temperatures. When the aging temperature increased, the oil recovery by spontaneous imbibition decreased. Separate wettability tests also confirmed the increase in water-wetness as the aging temperature was lowered. The amount of sulfate present in the pore water, SO42- (aq), appeared to be the active sulfate species to increase the water-wetness, which was in line with the previously suggested mechanism for wettability alteration by seawater in carbonates. For the tested aging temperatures of 50, 90, and 130 degrees C, changes in wetting properties appeared to take place at sulfate concentrations in the formation water below 2 mmol/L. At higher concentrations of sulfate, the wetting properties were not significantly affected.