Grantee Research Project Results

2003 Progress Report: Developing and Optimizing Biotransformation Kinetics for the Bio- remediation of Trichloroethylene at NAPL Source Zone Concentrations

EPA Grant Number: R828772C001
Subproject: this is subproject number 001 , established and managed by the Center Director under grant R828772
(EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
Center: National Research Program on Design-Based/Model-Assisted Survey Methodology for Aquatic Resources
Center Director: Stevens, Don L.
Title: Developing and Optimizing Biotransformation Kinetics for the Bio- remediation of Trichloroethylene at NAPL Source Zone Concentrations
Investigators: Semprini, Lewis , Dolan, Mark E.
Institution: Oregon State University
EPA Project Officer: Aja, Hayley
Project Period: November 1, 2001 through October 31, 2003
Project Period Covered by this Report: November 1, 2003 through October 31, 2004
Project Amount: Refer to main center abstract for funding details.
RFA: Hazardous Substance Research Centers - HSRC (2001) Recipients Lists
Research Category: Hazardous Waste/Remediation , Land and Waste Management

Objective:

The objectives of this research project are to: (1) develop a culture with the ability to reductively dechlorinate trichloroethylene (TCE) to ethylene at very high concentrations (above 1,000 µM) and in the presence of dense nonaqueous phase liquids (DNAPLs); (2) characterize microbial growth and measure maximum substrate utilization rates and half-velocity coefficients for successive dechlorinations of TCE to ethylene; (3) characterize the microbial consortium by investigating molecular methods to evaluate the diversity of the mixed culture developed in the kinetic studies; and (4) provide kinetic information and cultures in support of the following Center projects "Development of the Push-Pull Test To Monitor the Bioaugmentation of Dehalogenating Cultures," and "Development and Evaluation of Field Sensors for Monitoring Bioaugmentation With Anaerobic Dehalogenating Cultures for In Situ Treatment of TCE."

Rationale. Although TCE reductive dechlorination has been demonstrated under a variety of conditions, most laboratory and field projects have been conducted at TCE concentrations of 100 mg/L or less. Near NAPL sources, however, chlorinated aliphatic hydrocarbon concentrations approach their solubilities (>1,000 mg/L for TCE and >150 mg/L for perchloroethylene [PCE]). Studies with different enrichment cultures isolated from contaminated sites have shown good potential for treatment of high concentrations of PCE and TCE. The cultures have different dehalogenation kinetic properties, which indicate that a more effective enrichment culture might be obtained by combining cultures. Research is needed to optimize the transformation kinetics for the consortium that has the ability to reductively dechlorinate high concentrations of TCE and PCE to stoichiometric quantities of ethylene. This project will prove useful for the remediation of chlorinated aliphatic compounds in the NAPL source zone.

Approach. A culture is being developed that can rapidly degrade high concentrations of PCE and TCE to ethylene by mixing two enrichment cultures. The Point Mugu enrichment (PM) rapidly transforms TCE to vinyl chloride (VC), and slowly transforms VC to ethylene at very high PCE and TCE concentrations. The Evanite enrichment (EV) rapidly transforms PCE to cis-DCE (dichloroethylene) and VC to ethylene. By mixing both cultures, we hope to achieve rapid transformation of PCE and TCE to ethylene. We will use batch reactor studies to determine transformation kinetics for both cultures separately and when both cultures are combined. Inhibition among the chlorinated aliphatic hydrocarbons (CAHs) also will be evaluated. Models will be constructed to simulate the results of the sequential transformations over a broad range of concentrations up to the solubility limits of PCE and TCE.

Progress Summary:

Kinetic studies were conducted with two mixed cultures and a binary culture (a mixture of the two cultures) to describe the reductive dechlorination of chlorinated ethylenes. Inhibition of the CAHs also was studied. The EV and PM cultures obtained from different contaminated sites showed different patterns of reductive dechlorination. The simple batch kinetic method that was developed was easy to implement and produced very reproducible kinetic values. The kmax (based on the total protein content of the culture) for cis-DCE of the EV culture was about two times lower than that of the PM culture, reflecting the slower cis-DCE biotransformation of the EV culture. The kmax and KS values for VC (2.44 ± 0.36 µmol/mg of protein/day and 602 ± 7.06 µM, respectively) of the PM culture were very different from those of the EV culture (8.08 ± 0.94 µmol/mg of protein/day and 62.6 ± 2.37 µM, respectively). Inhibition studies were performed on the CAHs, and these studies show that the more chlorinated ethylenes inhibit reductive dechlorination of the less chlorinated ethylenes. PCE inhibited reductive TCE dechlorination but not cis-DCE dechlorination, while TCE strongly inhibited cis-DCE and VC dechlorinations. cis-DCE strongly inhibited the transformation of VC. Inhibition constants of each chlorinated ethylene, KI (µmol/L), were comparable to their respective half-velocity coefficients when a competitive inhibition model was applied.

Batch tests to study the sequential transformation of PCE to ethylene (ETH) also were performed over a factor of 30 changes in concentration of PCE, up to its solubility limit in water (1,128 µM), with the EV, PM, and a 50/50 mixture of both cultures to yield a binary culture (BM). Additional studies were performed with TCE up to a concentration of 4,173 µM (550 mg/L), which represents 50 percent of its solubility limit in water. Simulations of the successive transformations of PCE to ETH, and TCE to ETH using the independently derived kinetic parameters, matched well with the results of the batch kinetic tests for initial PCE concentrations of up to around 317 µM. The simulations included the growth on the chlorinated solvents and Monod kinetics, including competitive inhibition. Above this concentration simulations deviated from the experimental observations and predicted more rapid transformation of VC than was observed. The results suggest potential toxicity or inhibition at the higher concentrations of PCE and TCE. Simulations have been performed with Halden kinetics incorporated into the transformation models, where high concentrations of a contaminant inhibit its transformation. To explain the experimental observations, Halden kinetics for TCE transformation were required for both the EV and the PM cultures and for cis-DCE and VC transformation by the EV culture. The EV culture appears to be more inhibited at higher CAH concentrations. TCE concentrations of up to 4,173 µM (550 mg/L) were transformed by both the EV and PM culture, with the PM culture more rapidly transforming the TCE to VC and ETH. The results indicate less inhibition of the PM culture at higher concentrations. Batch experimental results indicate that the BM culture, which represents a mixture of both cultures, has better transformation abilities than either of the single cultures. Simulations for the BM culture, using individual transformation abilities of each culture, support the experimental observations of more diverse dechlorination ability than either of the single mixed cultures.

Molecular methods analysis using polymerase chain reactions with Dehalococcoides-specific primers and Desulfuromonas-specific primers, found Dehalococcoides-like microorganisms in both of the cultures, but not Desulfuromonas-like microorganisms. The molecular methods could not distinguish between the Dehalococcoides species of the EV and the PM cultures.

Future Activities:

We will create clone libraries to characterize the cultures. We also plan to perform kinetic measurements at high concentrations of the CAHs to determine whether Halden kinetics are observed and consistent with the results obtained from model simulations.

Journal Articles:

No journal articles submitted with this report: View all 2 publications for this subproject

Supplemental Keywords:

groundwater, reductive dechlorination, microbial activity, nonaqueous phase liquid, NAPL, dense nonaqueous phase liquid, DNAPL, waste, biochemistry, bioremediation, chemical engineering, chemicals, groundwater remediation, hazardous, hazardous waste, remediation, treatment technologies, trichloroethylene, TCE, TCE degradation, dichloroethylene, DCE, cis-DCE, perchloroethylene, PCE, tetrachloroethylene, biotransformation, chemical mixtures, chlorinated organic compounds, chlorinated organics, chlorinated solvents, dechlorination., RFA, Scientific Discipline, TREATMENT/CONTROL, Waste, POLLUTANTS/TOXICS, Hazardous, Remediation, Chemical Engineering, Environmental Chemistry, Groundwater remediation, Hazardous Waste, Treatment Technologies, Bioremediation, Chemicals, Environmental Engineering, chlorinated organics, reductive dechlorination, biotransformation, dechlorination, chlorinated organic compounds, Trichloroethylene, reductive dechlorination rates, NAPL, groundwater, NAPLs, TCE degradation, TCE

Relevant Websites:

http://wrhsrc.oregonstate.edu/ Exit

Progress and Final Reports:

Original Abstract

2002

Final

Main Center Abstract and Reports:

R828772    National Research Program on Design-Based/Model-Assisted Survey Methodology for Aquatic Resources

Subprojects under this Center: (EPA does not fund or establish subprojects; EPA awards and manages the overall grant for this center).
R828772C001 Developing and Optimizing Biotransformation Kinetics for the Bio- remediation of Trichloroethylene at NAPL Source Zone Concentrations
R828772C002 Strategies for Cost-Effective In-situ Mixing of Contaminants and Additives in Bioremediation
R828772C003 Aerobic Cometabolism of Chlorinated Aliphatic Hydrocarbon Compounds with Butane-Grown Microorganisms
R828772C004 Chemical, Physical, and Biological Processes at the Surface of Palladium Catalysts Under Groundwater Treatment Conditions
R828772C006 Development of the Push-Pull Test to Monitor Bioaugmentation with Dehalogenating Cultures
R828772C007 Development and Evaluation of Field Sensors for Monitoring Bioaugmentation with Anaerobic Dehalogenating Cultures for In-Situ Treatment of TCE
R828772C008 Training and Technology Transfer
R828772C009 Technical Outreach Services for Communities (TOSC) and Technical Assistance to Brownfields Communities (TAB) Programs
R828772C010 Aerobic Cometabolism of Chlorinated Ethenes by Microorganisms that Grow on Organic Acids and Alcohols
R828772C011 Development and Evaluation of Field Sensors for Monitoring Anaerobic Dehalogenation after Bioaugmentation for In Situ Treatment of PCE and TCE
R828772C012 Continuous-Flow Column Studies of Reductive Dehalogenation with Two Different Enriched Cultures: Kinetics, Inhibition, and Monitoring of Microbial Activity
R828772C013 Novel Methods for Laboratory Measurement of Transverse Dispersion in Porous Media
R828772C014 The Role of Micropore Structure in Contaminant Sorption and Desorption