Nitrite (NO2−) can accumulate during nitrification in soil following fertilizer application. While the role of NO2− as a substrate regulating nitrous oxide (N2O) production is recognized, kinetic data are not available that allow for estimating N2O production or soil-to-atmosphere fluxes as a function of NO2− levels under aerobic conditions. The current study investigated these kinetics as influenced by soil physical and biochemical factors in soils from cultivated and uncultivated fields in Minnesota, USA. A linear response of N2O production rate () to NO2− was observed at concentrations below 60 μg N g−1 soil in both nonsterile and sterilized soils. Rate coefficients (Kp) relating to NO2− varied over two orders of magnitude and were correlated with pH, total nitrogen, and soluble and total carbon (C). Total C explained 84% of the variance in Kp across all samples. Abiotic processes accounted for 31–75% of total N2O production. Biological reduction of NO2− was enhanced as oxygen (O2) levels were decreased from above ambient to 5%, consistent with nitrifier denitrification. In contrast, nitrate (NO3−)-reduction, and the reduction of N2O itself, were only stimulated at O2 levels below 5%. Greater temperature sensitivity was observed for biological compared with chemical N2O production. Steady-state model simulations predict that NO2− levels often found after fertilizer applications have the potential to generate substantial N2O fluxes even at ambient O2. This potential derives in part from the production of N2O under conditions not favorable for N2O reduction, in contrast to N2O generated from NO3− reduction. These results have implications with regard to improved management to minimize agricultural N2O emissions and improved emissions assessments.