Measuring denitrification activity in soils under pasture: Optimizing conditions for the short-term denitrification enzyme assay and effects of soil storage on denitrification activity

The short-term denitrification enzyme assay measures the potential denitrification activity of a soil in the field. The technique involves anaerobic incubation of soil samples and the measurement of N2O emission in the presence of C2H2. We examined the effects of incubation conditions and previous soil treatment on the denitrification enzyme activity (DEA) of two soils under permanent pasture. A standardized procedure is recommended. To avoid any effect of growth of denitrifying organisms on the DEA, the incubation period should not exceed 5 h at 20°C. The Km value for substrate NO3− concentrations was in the range 21–38 μg NO3−N g−1 soil, but NO3−N concentrations > 100 μg N g−1 soil inhibited production of N2O. The optimum concentrations for NO3−N and glucose-C were 50 and 300 μg g−1 soil, respectively. Field-moist samples of soil retained their DEA during 5 daysʹ storage at either 2 or 20°C, but thereafter the DEA declined with time. The denitrification activity of air-dried soil increased relative to fresh soil after 1 weekʹs storage, but subsequently declined. The effects were generally consistent with a change in the availability of substrate C in the stored soil (moist or air-dry), but there also appeared to be a decline in the potential activity of the soil denitrifier populations after 5–7 daysʹ storage.