Hydride-Meisenheimer Complex Formation and Protonation as Key Reactions of 2,4,6-Trinitrophenol Biodegradation by Rhodococcus erythropolis

Abstract

Biodegradation of 2,4,6-trinitrophenol (picric acid) byRhodococcus erythropolis HLPM-1 proceeds via initial hydrogenation of the aromatic ring system. Here we present evidence for the formation of a hydride-Meisenheimer complex (anionic ς-complex) of picric acid and its protonated form under physiological conditions. These complexes are key intermediates of denitration and productive microbial degradation of picric acid. For comparative spectroscopic identification of the hydride complex, it was necessary to synthesize this complex for the first time. Spectroscopic data revealed the initial addition of a hydride ion at position 3 of picric acid. This hydride complex readily picks up a proton at position 2, thus forming a reactive species for the elimination of nitrite. Cell extracts ofR. erythropolis HLPM-1 transform the chemically synthesized hydride complex into 2,4-dinitrophenol. Picric acid is used as the sole carbon, nitrogen, and energy source by R. erythropolisHLPM-1.

Function of Coenzyme F 420 in Aerobic Catabolism of 2,4,6-Trinitrophenol and 2,4-Dinitrophenol by Nocardioides simplex FJ2-1A

“…Due to the electron-withdrawing effect of the nitro substituents, the -electron deficiency of the aromatic system favors nucleophilic hydride additions. Such reactions are known for chemical hydride donors like sodium borohydride generating hydride or even dihydride -complexes of nitroaromatic compounds (14,23,27). More recently, hydride complexes were observed in our lab as microbial metabolites of picrate and TNT.…”

“…DOC concentration was analyzed with a Beckmann Industrial 915B total organic carbon (TOC) analyzer. Nitrite concentration was determined by HPLC as described previously (23). Fluorescence of compound C was detected with a common UV lamp at 360 nm.…”

“…Finally, the reaction was stopped with phosphoric acid. Picrate was tested as a substrate for the hydride transferase because the first step in picrate metabolism is the formation of a hydride -complex (21) which subsequently leads to 2,4-DNP (14,23). As shown in Fig.…”

“…Due to the presence of electron-withdrawing nitro groups as substituents, dinitroaromatic compounds and particular trinitroaromatic compounds like 2,4,6-trinitrotoluene (TNT) and 2,4,6-trinitrophenol (picric acid) are readily susceptible to initial reductive rather than oxidative attack (23,27). Consequently, initial oxidations by microbial mono-or dioxygenases of aerobic microorganisms are unknown for this class of xenobiotic compounds.…”

“…Besides specific and unspecific reductions of the nitro groups (26), unusual hydrogenations of the aromatic ring system have been observed for picrate and TNT. Thus, hydride and dihydride complexes have been identified as initial metabolites (23,27). In addition, the identification of 2,4dinitrophenol (2,4-DNP) and 4,6-dinitrohexanoate (4,6-DNH) (13,14,23,22) as metabolites of picrate indicates that extensive hydrogenation of the aromatic system (6 H per mol of picrate and 4 H per mol of 2,4-DNP) gives rise to a nonoxygenolytic ring cleavage.…”