Project Abstract

Mechanisms, Chemistry, and Kinetics of Anaerobic Degradation of cDCE and Vinyl Chloride

Principal Investigators

Perry L. McCarty and Alfred Spormann
Stanford University
(Supported by DuPont Chemicals and U.S. Department of Energy)
E-mail: mccarty@cive.stanford.edu

Goal

The objectives of this study are to describe the bacterium or groups of bacteria that are responsible for conversion of tetrachloroethene (PCE) to ethene in aquifer samples taken from a contaminated site in Victoria, Texas, and to examine the factors affecting the rate and extent of transformation.

Rationale

Several species of bacteria have been isolated and identified by others that have the ability to reductively dehalogenate chlorinated aliphatic hydrocarbons (CAHs). However, an individual or group of organisms responsible for the complete dehalogenation process from tetrachloroethylene (PCE) to ethene has not yet been identified. Whether or not the complete dehalogenation or the individual dehalogenation steps are the result of cometabolism or of energy metabolism is also not known. In addition, the various factors that affect the rates of dehalogenation have not been adequately evaluated. Such factors may include the electron donor used, the presence or absence of alternative electron acceptors, pH, and temperature. In order to understand the process better so that it can be more widely applied, its suitability for application at a given location can be better determined, and the economics of process implementation can be improved, definitive studies to better understand the nature of the organisms involved and factors affecting transformation rates are needed.

Approach

Anaerobic aquifer material from a contaminated site in Victoria, Texas, was obtained by DuPont Chemicals for this study. Microcosms were constructed of 125 mL bottles containing aquifer material and groundwater from the site. A small portion of the fluid is removed from the bottle periodically and replaced with groundwater amended with various primary substrates and PCE. Samples are analyzed for a range of CAHs and ethene, as well as for primary substrate, sulfate, and methane. The relationships between time and amount of primary substrate on CAH transformation is being evaluated. In separate studies, enrichment cultures are being developed using various electron donors, PCE, sulfate, and other nutrients, and serial dilutions of the cultures are being made as a first step in organism isolation. If satisfactory transformation of PCE is obtained by highly enriched cultures, then attempts will be made to isolate colonies from agar plates or roll tubes. Once isolated, the organisms will be characterized, and factors affecting PCE transformation will be studied in greater detail.

Status

The original project was completed last year, but additional funding was provided last year to extend the project, but with similar objectives. Funding also was received this year from the U.S. Department of Energy to extend this work. Benzoate, acetate, and formate were all found to stimulate dehalogenation, although benzoate appears to be the better of the three. Enrichment cultures have been developed that retain their ability to reduce PCE to ethene. A facultative pure bacterial strain (MS-1) has been isolated that transfers PCE to cis-1,2-dichloroethylene (c-DCE) while growing on a variety of substrates. This is the first facultative bacterium identified with this dehalogenating ability. A new Center grant has been funded to characterize the physiology and biochemistry of this organism and to evaluate its potential for bioaugmentation for degradation of chlorinated solvents. This overall study clearly demonstrated that there are two separate groups involved in the overall dehalogenation of PCE to ethene, the first is the group that converts PCE to c-DCE as represented by strain MS-1, and the second is the group that converts c-DCE to vinyl chloride and ethene. Rates of transformation were evaluated by a mixed culture growing on yeast extract. The rates of transformation of PCE and TCE to cDCE were about four times faster than for the conversion of cDCE to vinyl chloride and vinyl chloride to ethene. The pH optimum for the culture studied was 6.5 and the temperature optimum was 35 degrees C. Organisms appear to obtain energy from the dehalogenation reaction while using the chlorinated organics as electron acceptors. Monod kinetics of the reactions involved are being evaluated through the extension of the project. Also, enzymatic studies are being conducted in order to better understand the biochemistry of the reactions involved.