In the Midwestern US, perennial rhizomatous grasses (PRGs) are considered one of the most promising vegetation types to be used as a cellulosic feedstock for renewable energy production. The potential widespread use of biomass crops for renewable energy production has sparked numerous environmental concerns, including the impacts of land-use change on the hydrologic cycle. We predicted that total seasonal evapotranspiration (ET) would be higher for PRGs relative to maize resulting from higher leaf area and a prolonged growing season. We further predicted that, compared with maize, higher aboveground biomass associated with PRGs would offset the higher ET and increase water-use efficiency (WUE) in the context of biomass harvests for liquid biofuel production. To test these predictions, ET was estimated during the 2007 growing season for replicated plots of Miscanthus×giganteus (miscanthus), Panicum virgatum (switchgrass), and Zea mays (maize) using a residual energy balance approach. The combination of a 25% higher mean latent heat flux (λET) and a longer growing season resulted in miscanthus having ca. 55% higher cumulative ET over the growing season compared with maize. Cumulative ET for switchgrass was also higher than maize despite similar seasonal-mean λET. Based on total harvested aboveground biomass, WUE was ca. 50% higher for maize relative to miscanthus; however, when WUE calculated from only maize grain biomass was compared with WUE calculated from miscanthus harvested aboveground biomass, this difference disappeared. Although WUE between maize and miscanthus differed postsenescence, there were no differences in incremental WUE throughout the growing season. Despite initial predictions, aboveground biomass for switchgrass was less than maize; thus WUE was substantially lower for switchgrass than for either maize scenario. These results indicate that changes in ET due to large-scale implementation of PRGs in the Midwestern US would likely influence local and regional hydrologic cycles differently than traditional row crops.