Studies of fluid inclusions in sphalerite and biomarkers from the Upper Mississippi Valley zinc district show homogenization temperatures to be primarily between 90 and 150 ° C, yet show relatively low levels of thermal maturity. We use numerical calculations to simulate fluid and heat flow through fracture-controlled ore zones and heat transfer to the adjacent rocks. Combining a best-fit path through fluid-inclusion data with measured thermal alteration of biomarkers, we calculated the time interval during which mineralizing fluids circulated through the Upper Mississippi Valley district to be on the order of 200 ka. Cambrian and Ordovician aquifers underlying the district, principally the St. Peter and Mt. Simon Sandstones, were the source of the mineralizing fluid. The duration of mineralization thus reflects the fluid-flow history of these regional aquifers.