Nitrite fixation by humic substances: nitrogen-15 nuclear magnetic resonance evidence for potential intermediates in chemodenitrification

Studies have suggested that NO2-, produced during nitrification and denitrification, can become incorporated into soil organic matter and, in one of the processes associated with chemodenitrification, react with organic matter to form trace N gases, including N2O. To gain an understanding of the nitrosation chemistry on a molecular level, soil and aquatic humic substances were reacted with (15)N-labeled NaNO2, and analyzed by liquid phase (15)N and (13)C nuclear magnetic resonance (NMR). The International Humic Substances Society (IHSS) Pahokee peat and peat humic acid were also reacted with (15)NO2 and analyzed by solid-state (15)N NMR. In Suwannee River, Armadale, and Laurentian fulvic acids, phenolic rings and activated methylene groups underwent nitrosation to form nitrosophenols (quinone monoximes) and ketoximes, respectively. The oximes underwent Beckmann rearrangements to 2 degrees amides, and Beckmann fragmentations to nitriles. The nitriles in turn underwent hydrolysis to 1 degree amides. Peaks tentatively identified as imine, indophenol, or azoxybenzene nitrogens were clearly present in spectra of samples nitrosated at pH 6 but diminished at pH 3. The (15)N NMR spectrum of the peat humic acid exhibited peaks corresponding with N-nitroso groups in addition to nitrosophenols, ketoximes, and secondary Beckmann reaction products. Formation of N-nitroso groups was more significant in the whole peat compared with the peat humic acid. Carbon-13 NMR analyses also indicated the occurrence of nitrosative demethoxylation in peat and soil humic acids. Reaction of (15)N-NH3 fixated fulvic acid with unlabeled NO2- resulted in nitrosative deamination of aminohydroquinone N, suggesting a previously unrecognized pathway for production of N2 gas in soils fertilized with NH3.