Application of the ¹⁵N gas-flux method for measuring in situ N₂ and N₂O fluxes due to denitrification in natural and semi-natural terrestrial ecosystems and comparison with the acetylene inhibition technique

Abstract. Soil denitrification is considered the most un-constrained process in the global N cycle due to uncertain in situ N₂ flux measurements, particularly in natural and semi-natural terrestrial ecosystems. ¹⁵N tracer approaches can provide in situ measurements of both N₂ and N₂O simultaneously, but their use has been limited to fertilized agro-ecosystems due to the need for large ¹⁵N additions in order to detect ¹⁵N₂ production against the high atmospheric N₂. For ¹⁵N–N₂ analyses, we have used an “in-house” laboratory designed and manufactured N₂ preparation instrument which can be interfaced to any commercial continuous flow isotope ratio mass spectrometer (CF-IRMS). The N₂ prep unit has gas purification steps and a copper-based reduction furnace, and allows the analysis of small gas injection volumes (4 µL) for ¹⁵N–N₂ analysis. For the analysis of N₂O, an automated Tracegas Preconcentrator (Isoprime Ltd) coupled to an IRMS was used to measure the ¹⁵N–N₂O (4 mL gas injection volume). Consequently, the coefficient of variation for the determination of isotope ratios for N₂ in air and in standard N₂O (0.5 ppm) was better than 0.5 %. The ¹⁵N gas-flux method was adapted for application in natural and semi-natural land use types (peatlands, forests, and grasslands) by lowering the ¹⁵N tracer application rate to 0.04–0.5 kg ¹⁵N ha⁻¹. The minimum detectable flux rates were 4 µg N m⁻² h⁻¹ and 0.2 ng N m⁻² h⁻¹ for the N₂ and N₂O fluxes respectively. Total denitrification rates measured by the acetylene inhibition technique in the same land use types correlated (r =  0.58) with the denitrification rates measured under the ¹⁵N gas-flux method, but were underestimated by a factor of 4, and this was partially attributed to the incomplete inhibition of N₂O reduction to N₂, under a relatively high soil moisture content, and/or the catalytic NO decomposition in the presence of acetylene. Even though relatively robust for in situ denitrification measurements, methodological uncertainties still exist in the estimation of N₂ and N₂O fluxes with the ¹⁵N gas-flux method due to issues related to non-homogenous distribution of the added tracer and subsoil gas diffusion using open-bottom chambers, particularly during longer incubation duration. Despite these uncertainties, the ¹⁵N gas-flux method constitutes a more reliable field technique for large-scale quantification of N₂ and N₂O fluxes in natural terrestrial ecosystems, thus significantly improving our ability to constrain ecosystem N budgets.