Continuous measurements of nitrous oxide isotopomers during incubation experiments

Abstract

<p><strong>Abstract.</strong> Nitrous oxide (N<sub>2</sub>O) is an important and strong greenhouse gas in the atmosphere. It is produced by microbes during nitrification and denitrification in terrestrial and aquatic ecosystems. The main sinks for N<sub>2</sub>O are turnover by denitrification and photolysis and photo-oxidation in the stratosphere. In the linear N = N = O molecule <sup>15</sup>N substitution is possible in two distinct positions, central and terminal. The respective molecules, <sup>14</sup>N<sup>15</sup>N<sup>16</sup>O and <sup>15</sup>N<sup>14</sup>N<sup>16</sup>O, are called isotopomers. It has been demonstrated that N<sub>2</sub>O produced by nitrifying or denitrifying microbes exhibits a different relative abundance of the isotopomers. Therefore, measurements of the site preference (difference in the abundance of the two isotopomers) in N<sub>2</sub>O can be used to determine the source of N<sub>2</sub>O i.e. nitrification or denitrification. Recent instrument development allows for continuous position dependent δ<sup>15</sup>N measurements at N<sub>2</sub>O concentrations relevant for studies of atmospheric chemistry. We present results from continuous incubation experiments with denitrifying bacteria, <i>Pseudomonas fluorescens</i> (producing and reducing N<sub>2</sub>O) and <i>Pseudomonas chlororaphis</i> (only producing N<sub>2</sub>O). The continuous analysis of N<sub>2</sub>O isotopomers reveal the transient pattern (KNO<sub>3</sub> to N<sub>2</sub>O and N<sub>2</sub>, respectively). We find bulk isotopic fractionation of −5.01 ‰ ± 1.20 for <i>P. chlororaphis</i>, in line with previous results for production from denitrification. For <i>P. fluorescens</i>, the bulk isotopic fractionation during production of N<sub>2</sub>O is −52.21 ‰ ± 9.28 and 8.77 ‰ ± 4.49 during N<sub>2</sub>O reduction. <br><br> The SP isotopic fractionation for <i>P. chlororaphis</i> is −3.42 ‰ ± 1.69. For <i>P. fluorescens</i>, the calculations result in SP isotopic fractionation values of 5.73 ‰ ± 5.26 during production of N<sub>2</sub>O and 2.41 ‰ ± 3.04 during reduction of N<sub>2</sub>O. We interpret the slightly increased isotopic fractionation during reduction to diffusive isotopic fractionation and a difference in active enzymes during production of N<sub>2</sub>O. In summary, we implemented continuous measurements of N<sub>2</sub>O isotopomers during incubation of denitrifying bacteria and believe that similar experiments will lead to a better understanding of denitrifying bacteria and N<sub>2</sub>O turnover in soils and sediments and ultimately hands-on knowledge on the biotic mechanisms behind greenhouse gas exchange of the globe.</p>