Anaerobic degradation of toluene and o-xylene by a methanogenic consortium

Abstract

Toluene and o-xylene were completely mineralized to stoichiometric amounts of carbon dioxide, methane, and biomass by aquifer-derived microorganisms under strictly anaerobic conditions. The source of the inoculum was creosote-contaminated sediment from Pensacola, Fla. The adaptation periods before the onset of degradation were long (100 to 120 days for toluene degradation and 200 to 255 days for o-xylene). Successive transfers of the toluene-and o-xylene-degrading cultures remained active. Cell density in the cultures progressively increased over 2 to 3 years to stabilize at approximately 109 cells per ml. Degradation of toluene and o-xylene in stable mixed methanogenic cultures followed Monod kinetics, with inhibition noted at substrate concentrations above about 700 FM for o-xylene and 1,800 FM for toluene. The cultures degraded toluene or o-xylene but did not degrade m-xylene, p-xylene, benzene, ethylbenzene, or naphthalene. The degradative activity was retained after pasteurization or after starvation for 1 year. Degradation of toluene and o-xylene was inhibited by the alternate electron acceptors oxygen, nitrate, and sulfate. Degradation was also inhibited by the addition of preferred substrates such as acetate, H2, propionate, methanol, acetone, glucose, amino acids, fatty acids, peptone, and yeast extract. These data suggest that the presence of natural organic substrates or cocontaminants may inhibit anaerobic degradation of pollutants such as toluene and o-xylene at contaminated sites. Soil, sediment, and groundwater are frequently contaminated with petroleum products as a result of leaks in underground storage tanks, improper disposal techniques, and inadvertent spills. Of the many constituents of petroleum, the nonoxygenated, homocyclic aromatic compounds (including benzene, toluene, xylenes, and ethylbenzene) are of particular concern because they are confirmed or suspected carcinogens, even at very low concentrations (6). These compounds are relatively water soluble compared with other components of petroleum and thus frequently migrate through groundwater systems to contaminate drinking water supplies far removed from the actual spill (5, 29). The microbial degradation of compounds such as toluene and benzene under aerobic conditions has been studied in great detail (13, 30); in contrast, the fate of homocyclic aromatic compounds in anaerobic environments is very poorly understood. Until the mid-1980s, it was generally believed that monoaromatic compounds were recalcitrant to degradation under anaerobic conditions. Certain monoaromatic hydrocarbons, most frequently toluene, have since been shown to be degraded by microorganisms under denitrifying (8, 11, 20, 21, 25, 36), iron-reducing (22, 23), sulfate-reducing (2, 10, 17) and methanogenic (16, 32, 33, 34) conditions. We report here the enrichment and maintenance of a mixed culture derived from contaminated aquifer sediments that specifically degrades toluene and o-xylene under methanogenic conditions. The culture has been maintained with toluene or...

Field metabolomics and laboratory assessments of anaerobic intrinsic bioremediation of hydrocarbons at a petroleum‐contaminated site

“…Previous studies have demonstrated a significant lag or inhibition in the degradation of benzene, toluene, ethylbenzene or xylene when one or more of those hydrocarbons were added to laboratory incubations degrading a single BTEX component under a variety of electron-accepting conditions (Evans et al, 1991;Edwards and Grbic-Galic, 1994;Meckenstock et al, 2004b). It has also been shown that once degradation occurred, the microbial enrichments could be inhibited by the presence of other organic acids and alcohols (Edwards and Grbic-Galic, 1994), or co-metabolites as demonstrated with polycyclic aromatic hydrocarbons . Thus, the incomplete biodegradation of various hydrocarbons or the presence of their metabolic breakdown products could potentially interfere with the biodegradation of the COC.…”

Methane Production and Isotopic Fingerprinting in Ethanol Fuel Contaminated Sites

“…The evaluation of the isotopic signature of methane generated from petroleum hydrocarbons was carried out during biodegradation of toluene under methanogenic conditions. The culture was kindly provided by Dr. E. Edwards (Department of Chemical Engineering and Applied Chemistry, University of Toronto) and is a mixed consortium, which was described elsewhere (Edwards and Grbic-Galic 1994;Ficker et al 1999;Ahad et al 2000;Ward et al 2000).…”

“…Two bottles were prepared with 200 mL of culture and 200 mL of medium and are referred to as active 1 and 2. The medium contains no other carbon source and its composition is described in Edwards and Grbic-Galic (1994). Two controls were prepared with 400 mL of medium and 0.4 mL of 10% sodium azide solution.…”

Anaerobic Metabolism of Aromatic Compounds

“…Methyl substituents of the aromatic ring are difficult to attack in the absence of molecular oxygen, which normally initiates degradation. Methylated aromatic compounds, that are readily degraded anaerobically include toluene (Vogel and Grbic-Galic, 1986;Wilson et al, 1986;Zeyer et al, 1986;Dolfing et al, 1990: Lovley andLonergan, 1990;Schocher et al, 1991 ;Rabus et al, 1993;Fries et al, 1994: Rueter et al, 1994 and the ortho, rnetu and para isomers of xylene (Zeyer et al, 1986;Evans et al, 1991Evans et al, , 1992Seyfried et al, 1994;Edwards and Grbic-Galic, 1994;Rueter et al, 1994;Haner et al, 1995) and of cresol (Lovley and Lonergan, 1990;Hopper et al, 1991 ;Rudolphi et al, 1991 ;Flyvbjerg et al, 1993;Bonting et al, 1995). At least two different pathways can be used for the first attack under anaerobic conditions.…”