Toluene cometabolism

Aerobic biodegradation of trichloroethylene occurs by cometabolism with aromatie eompounds (Ensley 1991) and thus requires a cosubstrate such as phenol (Nelson et al. 1987, 1988) or toluene (Fan and Scow 1993). Trichloroethylene degradation by toluene-degrading baeteria has been demonstrated in the presence, but not absence, of toluene (Mu and Scow 1994). Isoprene, a structural analog of trichloroethylene, has also been used as a cosubstrate for triehloroethylene oxidation by some bacteria (Ewers et al. 1990). One source of inhibition of degradation in the absence of cosubstrate may be the toxieity of triehloroethylene itself to indigenous bacteria. 

Hopkins GD, PL McCarty (1995) Field evaluation of in situ aerobic cometabolism of trichloroethylene and three dichloroethylene isomers using phenol and toluene as primary substrates. Environ Sci Technol 29 1628-1637. 

Fries MR, GD Hopkins, PL McCarty, LJ Forney, JM Tiedje (1998a) Microbial succession during a field evaluation of phenol and toluene as the primary substrates for trichloroethene cometabolism. Appl Environ Microbiol 63 1515-1522. 

Fries MR, LJ Forney, JM Tiedje (1998b) Phenol- and toluene-degrading microbial populations from an aquifer in which successful trichloroethene cometabolism occurred. Appl Environ Microbiol 63 1523-1530. 

McCarty PL, MN Goltz, GD Hopkins, ME Dolan, IP Allan, BT Kawakami, TJ Carrothers (1998) Full-scale evaluation of in situ cometabolic degradation of trichloroethylene in groundwater through toluene injection. Environ Sci Technol 32 88-100. 

Incomplete aerobic transformations may involve cometabolic transformations and reactions resulting in recalcitrant dead-end metabolites. Cometabolic o-hydroxylation of MCPs and DCPs by a phenol monooxygenase has been shown, for example, in a Pseudomonas sp. (Knackmuss Hellwig, 1978) and by the toluene dioxygenase reaction in Pseudomonas putida (Spain Gibson, 1988 Spain et al., 1989)- Cometabolic transformation of CPs is also possible in aerobic mixed culture systems. Phenol- and toluene-enriched cultures completely removed 2,4-DCP, and the toluene enrichment also removed 2,4,6-TCP and PCP (Ryding et al., 1994). This PCP attack by the toluene enrichment involved an o-hydroxylation. 

Schafer A, Bouwer EJ. Toluene induced cometabolism of cis-l,2-dichloro-ethylene and vinyl chloride under conditions expected downgradient of a permeable Fe(0) barrier. Water Res 2000 34 3391-3399. 

Chang M-K, Voice TC, Criddle CS. 1993. Kinetics of competitive inhibition and cometabolism in the biodegradation of benzene, toluene, and p-xylene by two Pseudomonas isolates. Biotechnol Bioeng 41(11) 1057-1065. 

Fig. 2. Cometabolic interaction between toluene and o-xylene and corresponding nitrate reduction and nitrite accumulation.

This study proved degradation of toluene, benzene, o-xylene, ethylbenzene and butylbenzene by mixed denitrifying wastewater cultures. The study further revealed that o-xylene could not serve as only carbon and energy source for the culture, since it was not degraded without toluene availabihfy. Previous studies on the denitrifying degradation of alkylbenzenes did not address such interactions, but on the other hand those studies do not exclude cometabolic phenomena (Kuhn et al. 1986 Major et al. 1987 Zeyer et al. 1986). 

When the batches containing ethylbenzene and butylbenzene were inoculated, another type of cosubstrate interactions was revealed, see fig. 3. In this case the culture required either toluene or ethylbenzene as a primary substrate for butylbenzene degradation to occur. But a simultaneous degradation, as is normally observed in cases with cometabolism, was not observed, but in stead degradation of butylbenzene started just after either of the two other compounds was depleted. Thus it seemed as if this compound required a built up of biomass for degradation to occur. In fact etlylbenzene could not initially be degraded without toluene availability, but this dependency was rapidly overcome. 

Our observations did not leave ar r doubt whether cometabolism occurred. Maity theories have been evolved to explain why bacteria would degrade a eompound which cannot serve as eaibon or energy source. One definition is based on the capability of a compound to induce production of metabolic enzymes. Only growth substrates have this capability and with depletion of this substrate a deeay of the enzymes will start. In this case metabolic products need not accumulate. In our stutfy we in fact observed a lag after toluene depletion, which indicated such a relationship. Toluene thus induced the enzymes, which had a slightly slower turnover than toluene itself and therefore remained active in the batches a while after toluene depletion. 

Products were identified as 2,2,2-trichloroethanol and 2,2,2-trichloro-acetaldehyde by GC-MS and GC, respectively. Trichloroethanol (6.7 mu M) was completely degraded in 72 hours. When toluene was added to reaction mixtures, there was a 50% increase in the mineralization of (sup 14 C) TCE. M. vaccae s ability to cometabolize 2,4,6-trinitrotoluene (TNT) was also investigated. Two novel metabolites, as well as known reduction products, have been identified during catabolism of TNT with propane as cosubstrate. When M. vaccae was incubated with propane and (sup 14 C) TNT, 50% of the labelled carbon was found in the lipid fraction of cells. Analysis of this fraction demonstrated metabolism of sup 14 C into known phosphatides and other polar lipids indicating that ring cleavage had occurred. TNT or reduced intermediates were not found in any portion of the lipid fractions. 

The third most common biodegradation pathway for chlorinated hydrocarbons is cometabolism via enzymatic reactions occurring fortuitously with oxidation of compounds such as methane or toluene. These three pathways are, of course, complicated by the fact that 1) each can only occur under specific chemical conditions 2) the dominant pathway changes as the source of electron donor and/or acceptor (both anthropogenic and indigenous) is reduced and 3) There are several different isomers, and other compounds of chlorinated hydrocarbons, that can enter these pathways from biotic or abiotic processes. In other words, it is a complicated process. 

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