Identification of a hydride-Meisenheimer complex as a metabolite of 2,4,6-trinitrotoluene by a Mycobacterium strain

Abstract

A bacterial strain, Mycobacterium sp. strain HL 4-NT-1, enriched with 4-nitrotoluene as its sole source of nitrogen, was able to metabolize 2,4,6-trinitrotoluene under aerobic conditions. The dark red-brown metabolite, which accumulated in the culture fluid, was identified as a hydride-Meisenheimer complex by comparison with an authentic synthetic sample.In the course of large-scale manufacturing and handling of explosives, both soil and groundwater were contaminated extensively with polynitroaromatic compounds such as 2,4,6-trinitrotoluene (TNT) (5). Large amounts of TNT were synthesized during World War II. The high concentrations still found in the environment indicate the resistance of TNT to microbial degradation. Nitro groups, just like chloro substituents, reduce the electron density of the aromatic IT system, and thus hamper electrophilic attack by oxygenases. Initial reactions in the course of a degradative process of polynitroarenes thus must be reductive rather than oxidative. Several reports in the literature (1, 4, 15) deal with the reductive metabolism of TNT under anaerobic conditions. Little is known, however, about aerobic catabolism of TNT; most reports deal with cometabolic reactions by bacterial species leading to aminodinitrotoluenes (2, 6, 12, 14, 16). The hydroxylamino-or nitrosodinitrotoluenes, intermediates of such coreductions, usually couple with each other, generating biologically inert azoxy compounds (2,6,12,14,16).Recently, a novel reductive degradation mechanism by an aerobic organism has been described for the utilization of picric acid by Rhodococcus erythropolis HL PM-1 (9). An orange-red metabolite, accumulating transiently in the culture fluid, was characterized as the hydride-Meisenheimer complex formed by the nucleophilic addition of a hydride ion at C-3 of picric acid. Subsequently, this Meisenheimer complex was converted to 2,4-dinitrophenol and nitrite. From these data, though without any direct evidence, Duque et al. (3) proposed a corresponding mechanism for the metabolism of TNT yielding nitrite, dinitrotoluenes, nitrotoluenes, toluene, and major amounts of dead-end metabolites.We now present unequivocal evidence that the hydrideMeisenheimer complex of TNT (H --TNT complex) is the product of TNT bioconversion by Mycobacterium sp. strain HL 4-NT-1. 4-Nitrotoluene (4-NT)-grown cells of this organism produced sufficient amounts of the H --TNT complex to allow a spectroscopic characterization, in contrast to another newly isolated strain, CV TNT-8, which utilized TNT as a nitrogen source but accumulated only minor amounts of this rather unstable metabolite (iSa).

Biological remediation of explosives and related nitroaromatic compounds

“…These resist further oxidative microbial degradation and form a very serious problem due to their higher solubility, toxicity and genotoxicity as compared to TNT (see chapter 1. Another reductive degradation mechanism by aerobic organisms has been described: the nucleophilic attack by a hydride ion to form a hydride-Meisenheimer complex, which may be followed by the release of nitrite (Vorbeck et al 1994). Duque et al (1993) reported the isolation and characterisation of Pseudomonas sp.…”

“…clone A. The presence of an FF-TNT complex was successfully demonstrated when Rhodococcus erythropolis HL PM-1 or Mycobacterium strain HL 4-NT-1 were exposed to TNT (Vorbeck et al 1994). Unlike the H--picric acid complex, the corresponding TNT complex undergoes neither nitrite elimination nor re-aromatisation to 2,4-DNT or 2,6-DNT.…”

Initial characterization of a highly contaminated high explosives outfall in preparation for in situ bioremediation

“…Also, aerobic pathways result in formation of azoxy polymers formed through condensation of nitroso intermediates under aerobic conditions. Vorbeck et al (1994) also isolated a transient hydride-Meisenheimer complex product from TNT degradation under aerobic conditions. The other major aerobic degradative mechanism involves direct removal of the nitro group as nitrite (Spain et al, 1979;Zeyer and Kearney, 1984;Bruhn et al, 1987).…”

Gas Phase Reactions of 1,3,5-Triazine: Proton Transfer, Hydride Transfer, and Anionic σ-Adduct Formation

“…Of particular interest to our current work is the presence of a hydride transfer channel with 1,3,5-triazine to form a hydride-Meisenheimer complex. Recently, these complexes have been identified in the biodegradation of nitroaromatics [30][31][32][33][34]. Although fragmentation patterns of the hydrideMeisenheimer complex of trinitrotoluene, [TNT + H] σ-adducts [35].…”