The biological dehalogenation of common water pollutants such as trichloromethane (chloroform) and other halogenated aliphatic compounds was the subject of this project. Samples from diverse water environments such as from groundwater contaminated with halogenated compounds and wastewater from regional treatment plants were studied to identify microorganisms in samples from a secondary anaerobic digestor were able to dechlorinate trichloromethane but not dichloromethane (DCM) and samples from a denitrification tank were able to dechlorinate DCM but not trichloromethane. Gene probe analyses of DNA extracted from the dichloromethane-degrading wastewater indicated the presence of the gene coding for dichloromethane dehalogenase, indicating the genetic basis for the dechlorination activity observed. The chloroform dechlorination activity observed in the digestor samples was inhibited by the methanogenesis inhibitor bromoethanesulfonic acid (BES), and this inhibition could be reversed by coenzyme M (CoM), a coenzyme unique to methanogenic bacteria. These studies indicate that methanogenic bacteria are the organisms responsible for the chloroform dechlorination. Interestingly, very low levels of chloroform (ca. 10 ppb) potently inhibited methane production in the samples, yet when the chloroform was converted to dichloromethane, methanogenesis resumed quickly. The chloroform dechlorination thus appears to be a means of detoxification for the methanogens, and we propose that the site where chloroform inhibits methanogenesis is also the site by which it is reduced and detoxified to dichloromethane. Applying the chloroform and DCM activities resulted in the design of a 2-stage anaerobic bioreactor which completely dehalogenated chloroform stepwise to DCM, CO₂ and microbial biomass.

Dechlorination of a common chlorofluorocarbon (CFC-11) was identified in samples taken from a regional aquifer contaminated with halogenated aliphatic compounds. We have shown that sulfate-reducing bacteria were responsible for this activity and that the process depended on both an electron donor (2-4 carbon fatty acids) and an electron acceptor (sulfate, but not thiosulfate or elemental sulfur). Dechlorination by these aquifer bacteria exhibited an unusual kinetic response in that the reaction was inhibited at either very low (ca. 10 ppb) or relatively high (ca. 10 ppm) concentrations of CFC-11 (the aquifer was contaminated with 0.1 – 2 ppm CFC-11). Calculations of thermodynamic parameters such as the change in free energy and the reducing potential associated with the CFC dehalogenation reactions has given us a predictive understanding as to why one reaction is favored over another. For example, dechlorination of CFC-11 is energetically more favorable than defluorination, and CFC dechlorination was the reaction observed to occur. Such correlations between the predicted and the observed pathway of dehalogenation can be used in feasibility decisions concerning bioremediation of waters contaminated with halogenated aliphatic compounds.