Determination of potential N2O-reductase activity in soil

Abstract

Determination of N2O-reductase activity in soil is important for understanding the microbial regulation of nitrous oxide (N2O) concentrations in soil. Unfortunately, there are no easily applicable and accurate methods for determining N2O-reductase activity, which frustrates the understanding of the mechanisms that control soil management effects on denitrification and N2O emissions. The objectives of the study reported here were (i) to define the optimal experimental conditions for the determination of potential N2O-reductase activity in soil, (ii) to compare the direct-N2 method with the acetylene inhibition technique, and (iii) to investigate the effects of long-term nitrogen (N) fertilization on the potential N2O-reductase activity.Various substrate concentrations, water/soil ratios, incubation times, temperatures and pH values were tested to find the optimal conditions for the potential N2O-reductase activity in soils from two sites. Then, the potential N2O-reductase activity was determined under optimal conditions (10 g soil, 10 ml buffer solution, pH 9, 40 °C, 100 ppmv N2O) in soils from the long-term N fertilization experiment.There were significant differences between soils in potential N2O-reductase activity, but the optimal experimental conditions were similar. The acetylene inhibition technique underestimated N2O-reductase activity in soil relative to the direct-N2 method, especially in the treatment where fertilizer N was withheld for 15 yrs. We recommend that the optimal experimental conditions for the determination of the potential N2O-reductase activity are established also for other soils. More studies are needed to fully understand the interactive effects of long-term N fertilization on nosZ gene expression and N2O-reductase activity in soils.