Continuous measurements of nitrous oxide isotopomers during incubation experiments

Abstract. Nitrous oxide (N₂O) is an important and strong greenhouse gas in the atmosphere and part of a feed-back loop with climate. N₂O is produced by microbes during nitrification and denitrification in terrestrial and aquatic ecosystems. The main sinks for N₂O are turnover by denitrification and photolysis and photo-oxidation in the stratosphere. The position of the isotope ¹⁵N in the linear N = N = O molecule can be distinguished between the central or terminal position (isotopomers of N₂O). It has been demonstrated that nitrifying and denitrifying microbes have a different relative preference for the terminal and central position. Therefore, measurements of the site preference in N₂O can be used to determine the source of N₂O i.e. nitrification or denitrification. Recent instrument development allows for continuous (on the order of days) position dependent δ¹⁵N measurements at N₂O concentrations relevant for studies of atmospheric chemistry. We present results from continuous incubation experiments with denitrifying bacteria, Pseudomonas fluorescens (producing and reducing N₂O) and P. chlororaphis (only producing N₂O). The continuous position dependent measurements reveal the transient pattern (KNO₃ to N₂O and N₂, respectively), which can be compared to previous reported site preference (SP) values. We find bulk isotope effects of −5.5 ‰ ± 0.9 for P. chlororaphis. For P. fluorescens, the bulk isotope effect during production of N₂O is −50.4 ‰ ± 9.3 and 8.5 ‰ ± 3.7 during N₂O reduction. The values for P. fluorescens are in line with earlier findings, whereas the values for P. chlororaphis are larger than previously published δ¹⁵N_bulk measurements from production. The calculations of the SP isotope effect from the measurements of P. chlororaphis result in values of −6.6 ‰ ± 1.8. For P. fluorescens, the calculations results in SP values of −5.7 ‰ ± 5.6 during production of N₂O and 2.3 ‰ ± 3.2 during reduction of N₂O. In summary, we implemented continuous measurements of N₂O isotopomers during incubation of denitrifying bacteria and believe that similar experiments will lead to a better understanding of denitrifying bacteria and N₂O turnover in soils and sediments and ultimately hands-on knowledge on the biotic mechanisms behind greenhouse gas exchange of the Globe.