Advanced spectroscopic analysis and N-isotopic labelling study of nitrate and nitrite reduction to ammonia and nitrous oxide by .

Nitrate and nitrite reduction to ammonia and nitrous oxide by anaerobic batch cultures is investigated by advanced spectroscopic analytical techniques with N-isotopic labelling. Non-invasive, analysis of the headspace is achieved using White cell FTIR and cavity-enhanced Raman (CERS) spectroscopies alongside liquid-phase Raman spectroscopy. For gas-phase analysis, White cell FTIR measures CO, ethanol and NO while CERS allows H, N and O monitoring. The 6 m pathlength White cell affords trace gas detection of NO with a noise equivalent detection limit of 60 nbar or 60 ppbv in 1 atm. Quantitative analysis is discussed for all four N/N-isotopomers of NO. Monobasic and dibasic phosphates, acetate, formate, glucose and NO concentrations are obtained by liquid-phase Raman spectroscopy, with a noise equivalent detection limit of 0.6 mM for NO at 300 s integration time. Concentrations of the phosphate anions are used to calculate the pH using a modified Henderson-Hasselbalch equation. NO concentrations are determined by sampling for colorimetric analysis and NH by basifying samples to release N/N-isotopomers of NH for measurement in a second FTIR White cell. The reductions of NO, NO, and mixed NO and NO by anaerobic batch cultures are discussed. In a major pathway, NO is reduced to NH NO, with the bulk of NO reduction occurring after NO depletion. Using isotopically labelled NO, NH production is distinguished from background NH in the growth medium. In a minor pathway, NO is reduced to NO the toxic radical NO. With excellent detection sensitivities, NO serves as a monitor for trace NO reduction, even when cells are predominantly reducing NO. The analysis of NO isotopomers reveals that for cultures supplemented with mixed NO and NO enzymatic activity to reduce NO occurs immediately, even before NO reduction begins. Optical density and pH measurements are discussed in the context of acetate, formate and CO production. H production is repressed by NO; but in experiments with NO supplementation only, CERS detects H produced by formate disproportionation after NO depletion.