Carbon and nitrogen isotope effects associated with the dioxygenation of aniline and diphenylamine

Dioxygenation of aromatic rings is frequently the initial step of biodegradation of organic subsurface pollutants. This process can be tracked by compound-specific isotope analysis to assess the extent of contaminant transformation, but the corresponding isotope effects, especially for dioxygenation of N-substituted, aromatic contaminants, are not well understood. We investigated the C and N isotope fractionation associated with the biodegradation of aniline and diphenylamine using pure cultures of Burkholderia sp. strain JS667, which can biodegrade both compounds, each by a distinct dioxygenase enzyme. For diphenylamine, the C and N isotope enrichment was normal with ε_C- and ε_N-values of -0.6 ± 0.1% and -1.0 ± 0.1%, respectively. In contrast, N isotopes of aniline were subject to substantial inverse fractionation (ε_N of +13 ± 0.5%), whereas the ε_C-value was identical to that of diphenylamine. A comparison of the apparent kinetic isotope effects for aniline and diphenylamine dioxygenation with those from abiotic oxidation by manganese oxide (MnO₂) suggest that the oxidation of a diarylamine system leads to distinct C-N bonding changes compared to aniline regardless of reaction mechanism and oxidant involved. Combined evaluation of the C and N isotope signatures of the contaminants reveals characteristic δδ ¹⁵N/δδ¹³C-trends for the identification of diphenylamine and aniline oxidation in contaminated subsurfaces and for the distinction of aniline oxidation from its formation by microbial and/or abiotic reduction of nitrobenzene.