The extent to which land use influences soil organic matter processing, nitrogen transformations, and the generation of CO₂ and N₂O in mesic grasslands is unclear. To quantify how land management influences these processes, we examined soil nitrogen availability, N₂O production, denitrification potential, and microbial biomass in grasslands and afforested grasslands in northeastern Kansas. Measurements were obtained every 3-4 weeks from June 2006 to the present, from plots representing five land management techniques. Nitrogen availability was greatest in all fertilized plots (14.3±0.8 μg_N g^-1soil); hayed plots had the lowest available N levels (4.4±0.3 μg_N g^-1soil). Differences between fertilized plots and all other land management practices for inorganic N were the greatest during the growing season; inorganic N was reduced and differences were minimized by December 2006. In fertilized plots, NO₃^- availability explained 87.0% of the variation in N₂O fluxes. Early succession, “old field” grasslands experienced the highest N₂O fluxes, with a maximum flux rate of 3.4 μg_N2O-N m^-2 d^-1. Denitrification potential data suggest that wooded plots possess similar potential for denitrification as fertilized grasslands.  Microbial biomass C:N ranged from 72.1 to 11.8, and fluctuated with season; values peaked in August for early succession and hayed grassland soils, and were relatively low (~30) for fertilized and wooded plots. These data suggest that mesic grasslands subjected to fertilization and afforestation experience changes in microbial structure that can directly influence soil N transformations.  Both fertilization and woody succession, processes occurring widely at the prairie/forest ecotone, can result in relatively high fluxes of N₂O.