In the Midwestern USA, current biofuel production systems rely on high input monoculture crops that do little to support native biodiversity. The University of Northern Iowa's Tallgrass Prairie Center is investigating the feasibility of cultivating and harvesting diverse mixes of native prairie vegetation for use as a sustainable biofuel in a manner that also conserves biodiversity and protects soil and water resources. In 2009, we established 48 research plots on three soil types at an Iowa site with a uniform history of row crop production. We seeded each plot with one of four treatments of native prairie vegetation: (1) switchgrass monoculture, (2) warm-season grass mix (5 grass species), (3) biomass mix (16 species of grasses, legumes, and forbs), or (4) prairie mix (32 species of grasses, legumes, forbs, and sedges). In 2010, we measured vegetation characteristics and studied butterfly use of the plots to investigate the hypothesis that more diverse plant communities would support a greater abundance and diversity of butterflies. Habitat characteristics varied significantly among the plots by treatment and soil type, and butterflies responded rapidly to variation in floral abundance and richness. Averaged over the entire growing season, butterflies were six times more abundant and twice as species rich in the biomass and prairie mix plots compared to the warm-season grass and switchgrass plots. Our results suggest that implementation of biomass production using diverse mixes of native prairie vegetation on marginal lands could have positive effects on the maintenance of butterfly populations in agricultural landscapes.