title: Isotope signatures of N₂O in a mixed microbial population system: constraints on N₂O producing pathways in wastewater treatment.
abstract: We present measurements of site preference (SP) and bulk (15)N/(14)N ratios (δ(15)N(bulk)(N2O)) of nitrous oxide (N(2)O) by quantum cascade laser absorption spectroscopy (QCLAS) as a powerful tool to investigate N(2)O production pathways in biological wastewater treatment. QCLAS enables high-precision N(2)O isotopomer analysis in real time. This allowed us to trace short-term fluctuations in SP and δ(15)N(bulk)(N2O) and, hence, microbial transformation pathways during individual batch experiments with activated sludge from a pilot-scale facility treating municipal wastewater. On the basis of previous work with microbial pure cultures, we demonstrate that N(2)O emitted during ammonia (NH(4)(+)) oxidation with a SP of -5.8 to 5.6 ‰ derives mostly from nitrite (NO(2)(-)) reduction (e.g., nitrifier denitrification), with a minor contribution from hydroxylamine (NH(2)OH) oxidation at the beginning of the experiments. SP of N(2)O produced under anoxic conditions was always positive (1.2 to 26.1 ‰), and SP values at the high end of this spectrum (24.9 to 26.1 ‰) are indicative of N(2)O reductase activity. The measured δ(15)N(bulk)(N2O) at the initiation of the NH(4)(+) oxidation experiments ranged between -42.3 and -57.6 ‰ (corresponding to a nitrogen isotope effect Δδ(15)N = δ(15)N(substrate) - δ(15)N(bulk)(N2O) of 43.5 to 58.8 ‰), which is considerably higher than under denitrifying conditions (δ(15)N(bulk)(N2O) 2.4 to -17 ‰; Δδ(15)N = 0.1 to 19.5 ‰). During the course of all NH(4)(+) oxidation and nitrate (NO(3)(-)) reduction experiments, δ(15)N(bulk)(N2O) increased significantly, indicating net (15)N enrichment in the dissolved inorganic nitrogen substrates (NH(4)(+), NO(3)(-)) and transfer into the N(2)O pool. The decrease in δ(15)N(bulk)(N2O) during NO(2)(-) and NH(2)OH oxidation experiments is best explained by inverse fractionation during the oxidation of NO(2)(-) to NO(3)(-).
authors.affiliation: Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, P.O. Box 611, 8600 Duebendorf, Switzerland. pascal.wunderlin@eawag.ch
authors.forename: Pascal
authors.initials: P
authors.lastname: Wunderlin
authors.forename: Moritz F
authors.initials: MF
authors.lastname: Lehmann
authors.forename: Hansruedi
authors.initials: H
authors.lastname: Siegrist
authors.forename: Béla
authors.initials: B
authors.lastname: Tuzson
authors.forename: Adriano
authors.initials: A
authors.lastname: Joss
authors.forename: Lukas
authors.initials: L
authors.lastname: Emmenegger
authors.forename: Joachim
authors.initials: J
authors.lastname: Mohn
mesh_terms: D001419:Bacteria / Q000378:metabolism*; D061249:Batch Cell Culture Techniques; D053898:Biosynthetic Pathways; D058440:Denitrification; D052836:Heterotrophic Processes; D007553:Isotope Labeling; D009573:Nitrites; D009587:Nitrogen Isotopes; D009609:Nitrous Oxide / Q000032:analysis*; D010084:Oxidation-Reduction; D000644:Quaternary Ammonium Compounds / Q000378:metabolism; D062065:Wastewater / Q000382:microbiology*; D018508:Water Purification
chemical_list: D009573:Nitrites; D009587:Nitrogen Isotopes; D009609:Nitrous Oxide; D000644:Quaternary Ammonium Compounds; D062065:Wastewater