Prairie vegetation has the potential to serve as a bioenergy feedstock with favorable environmental impacts but generally yields less than corn (Zea mays L.). To more fully assess the potential of prairie vegetation to serve as a biofuel feedstock, more needs to be understood about prairie responses to different management strategies, including fertilization. We hypothesized that (i) N fertilization would increase prairie productivity and nutrient content and (ii) fertilized prairie would produce similar amounts of total biomass as corn but would allocate more biomass and nutrients to roots. Our hypotheses were tested in a field experiment using prairie species representing different functional groups-C-3 grasses, C-4 grasses, legumes, and multifunctional group mixtures grown with and without N fertilizer as well as fertilized corn. Fertilized and unfertilized C-4 prairie grasses produced as much total biomass as corn but allocated up to 65% of their biomass belowground compared to 3% for corn. Corn yields decreased over the 3-yr period of the study whereas yields of fertilized C-4 grasses and multifunctional group mixtures were stable, and yields of unfertilized C-4 grasses and mixtures increased and became equal to their fertilized counterparts. Calculated annual costs for replacing nutrients removed in the harvested portions of corn and fertilized and unfertilized C-4 grasses were US$403, $137, and $40 ha(-1), respectively. Results of this experiment show tradeoffs among corn and prairie systems with respect to harvested biomass, root production, and nutrient exports, emphasizing the need for a multifaceted approach to fully evaluate bioenergy feedstock production systems.