Abstract The industrial solvents tetrachloroethylene (PCE) and trichloroethylene (TCE) are among the most ubiquitous chlorinated compounds found in groundwater contamination. One potential method for managing PCE/TCE contaminated sites is the intrinsic bioremediation. Recent regulations adopted by U.S. Environmental Protection Agency allow intrinsic bioremediation to be considered as an alternative during development of corrective action plans. In some remediation cases, enhanced bioremediation are performed to accelerate the contaminant biodegradation rate. The main objective of this study was to evaluate the potential of using intrinsic and enhanced bioremediation technologies to clean up PCE/TCE contaminated aquifers. PCE/TCE bioavailability was evaluated by laboratory microcosms under four reduction/oxidation (redox) conditions including aerobic cometabolism, methanogenesis, iron reduction, and reductive dechlorination. Acclimated bacteria, activated sludge, and aquifer sediments from a pentachlorophenol contaminated site were used as the inocula in this study. Methane, toluene, phenol, sludge cake, and cane molasses were used as the primary substrates (carbon sources) in the cometabolism and reductive dechlorination microcosms. Results from this study show that PCE and TCE can be significantly biodegraded under reductive dechlorination and aerobic cometabolism conditions, respectively. All five carbon sources evaluated in this study can be applied as the primary substrates by microbial consortia to enhance the aerobic cometabolism of TCE. The highest TCE degradation rate [Up to 100% of TCE removal (with an initial concentration of 3.6µM)] was observed in the microcosms with toluene enrichment bacteria as the microbial inocula and toluene as the primary substrate. Under reductive dechlorination conditions, both sludge cake and cane molasses could be used as the primary substrates by microbial consortia (from activated sludge and aquifer sediments) and enhanced the biodegradation of PCE. The highest PCE degradation rate [Up to 100% of PCE removal (with an initial concentration of 17µM)] was observed in the microcosms with anaerobic activated sludge as the microbial inocula and sludge cake as the primary substrate. Except for reductive dechlorination microcosms, no significant PCE removal was observed in the microcosms prepared under iron reduction conditions. Results from this feasibility study would be useful in designing a scale-up in situ (e.g., in situ biobarrier system) or on-site bioremediation system (e.g., bioslurry reactor) for field application. Moreover, the application of non-toxic organic waste to enhance PCE/TCE biodegradation has the potential to become an environmentally and economically acceptable technology for the bioremediation of chlorinated-solvent contaminated groundwater.

A cylindrical electrochemical reactor with a 3 in diameter copper or nickel metal foam cathode and a concentric carbon cloth anode was used to destroy aqueous phase carbon tetrachloride (CT). The results show that a high CT conversion can be achieved in regions of the cathode near the anode, but a low CT conversion is obtained in the region around the center of the cathode. This CT conversion distribution in the radial current-conducting direction suggests that a portion of the cathode worked inefficiently even though the overall CT conversion is still adequate. Further research by changing the solution pH and conductivity suggests that the radial conversion distribution is due to radial variations in cathode surface availability. The inherent difficulties that these results imply with regards to reactor scale up suggested a new approach to the design. An annular reactor, consisting of a thin (3.2 mm) nickel foam cathode wrapped around an inert Plexiglas core and separated for an external concentric anode by a semi-permeable membrane was adopted. Under compatible operating conditions, the annular reactor showed a high overall effluent CT conversion. However, experiments at low pH (2.25) yielded higher conversions than under neutral pH conditions. This result suggests that CT conversion is favored by a relatively high proton concentration. This reactor can be simulated by a one dimensional model. The annular reactor was used to destroy PCE and TCE successfully, which suggests that this technique can be employed to treat groundwater contaminated with complex mixtures of chlorinated hydrocarbons.A multi-layer reactor based on the principle of the annular reactor was developed as an option for the scale up of the system. This reactor exhibited high and uniform radial CT conversion.

Nanosized metal particles have become an important class of materials in the field of catalysis, optical, electronic, magnetic and biological devices due to the unique physical and chemical properties. This research deals with the synthesis of structured bimetallic nanoparticles for the dechlorination of toxic organics. Nanoparticle synthesis in aqueous phase for dechlorination studies has been reported. However, in the absence of polymers or surfactants particles can easily aggregate into large particles with wide size distribution. In this study, we report a novel in-situ synthesis method of bimetallic nanoparticles embedded in polyacrylic acid (PAA) functionalized microfiltration membranes by chemical reduction of metal ions bound to the carboxylic acid groups. Membrane-based nanoparticle synthesis offers many advantages: reduction of particle loss, prevention of particle agglomeration, application of convective flow, and recapture of dissolved metal ions. The objective of this research is to synthesize and characterize nanostructured bimetallic particles in membranes, understand and quantify the catalytic hydrodechlorination mechanism, and develop a membrane reactor model to predict and simulate reactions under various conditions. In this study, the PAA functionalization was achieved by filling the porous PVDF membranes with acrylic acid and subsequent in-situ free radical polymerization. Target metal cations (iron in this case) were then introduced into the membranes by ion exchange process. Subsequent reduction resulted in the formation of metal nanoparticles (around 30 nm). Bimetallic nanoparticles can be formed by post deposition of secondary appropriate metal such as Pd or Ni. The membranes and bimetallic nanoparticles were characterized by: SEM, TEM, TGA, and FTIR. A specimen-drift-free X-ray energy dispersive spectroscopy (EDS) mapping system was used to determine the two-dimensional element distribution inside the membrane matrix at the nano scale. This high resolution mapping allows for the correlation and understanding the nanoparticle structure, second metal composition in terms of nanoparticle reactivity. Chlorinated aliphatics such as trichloroethylene and conjugated aromatics such as polychlorinated biphenyls (PCBs) were chosen as the model compounds to investigate the catalytic properties of bimetallic nanoparticles and the reaction mechanism and kinetics. Effects of second metal coating, particle size and structure and temperature were studied on the performance of bimetallic system. In order to predict reaction at different conditions, a two-dimensional steady state model was developed to correlate and simulate mass transfer and reaction in the membrane pores under convective flow mode. The 2-D equations were solved by COMSOL (Femlab). The influence of changing parameters such as reactor geometry (i.e. membrane pore size) and Pd coating composition were evaluated by the model and compared well with the experimental data.

The WBC-2 consortium was characterized using quantitative PCR and analytical techniques to associate growth of dechlorinating bacteria to each step of the 1,1,2,2-Tetrachloroethane (TeCA) degradation pathway. The consortium was found to degrade TeCA through dichloroelimination to trans-1,2-dichloroethene (tDCE), and reductive dehalogenation to Vinyl Chloride (VC) and ethene. Thus the pathway was hypothesized to provide three distinct niches for three genera of dechlorinating bacteria, Dehalobacter, Dehalogenimonas and Dehalococcoides. Using qPCR to track growth over two time course experiments at different inoculum dilutions, the Dehalobacter species showed significant growth on the first step of TeCA dihaloelimination to tDCE Dehalococcoides and Dehalogenimonas species grew on the dechlorination products. The Dehalogenimonas species, a novel non-Dehalococcoides, was found to grow only on tDCE. The Dehalococcoides species also grew on cDCE, less well on tDCE, and on VC.

KB-1 is an anaerobic Dehalococcoides-containing microbial culture used industrially to bioremediate sites impacted with chlorinated solvents. The culture is typically grown at pH 7. However, lower pH is often encountered and therefore the effect of pH was investigated. Both sudden and stepwise decreases in pH from 7 to 6 and 5.5 were investigated over a period of 450 days. An electron balance was also calculated to look at the flow of electrons for dechlorination. More than 95% of the reducing equivalents went towards methanogenesis and acetogenesis. Select microorganisms were compared by quantitative Polymerase Chain Reaction. It was found that lower rates of dechlorination correspond to low Dehalococcoides numbers and that different methanogens were enriched on different electron donors.

The purpose of this research was to investigate the microorganisms, genetics and biochemistry of anaerobic dechlorination of chlorinated ethanes, which are common groundwater contaminants. Specifically, this project used mixed microbial cultures to study the dechlorination of 1,2-dichloroethane (1,2-DCA), 1,1,2-trichloroethane (1,1,2-TCA) and 1,1,1-trichloroethane (1,1,1-TCA). A mixed microbial culture enriched from a contaminated multilayered aquifer in West Louisiana dechlorinated 1,2-DCA, 1,1,2-TCA, tetrachloroethene, trichloroethene, cis-dichloroethene and vinyl chloride (VC) to non-toxic ethene when amended with ethanol as the electron donor. 16S rRNA gene sequence analysis revealed the presence of the putative dechlorinating organisms Dehalobacter and Dehalococcoides spp. Denaturing gradient gel electrophoresis analysis and quantitative PCR (qPCR) with species-specific primers demonstrated that both organisms grew during the dichloroelimination of 1,2-DCA to ethene. Conversely, during the dichloroelimination of 1,1,2-TCA to VC only Dehalobacter grew, while during the reductive dechlorination of VC to ethene only Dehalococcoides grew. Further enrichment with 1,2-DCA, H2 and acetate yielded a co-culture of Dehalobacter and Acetobacterium spp. that did not dechlorinate other chlorinated ethanes or ethenes. Dehalobacter grew in the presence but not in the absence of 1,2-DCA, while Acetobacterium growth was not affected by 1,2-DCA. A novel putative Dehalobacter-associated 1,2-DCA reductive dehalogenase gene was identified and was shown to be transcribed only in the presence of 1,2-DCA. An enrichment microbial culture derived from a 1,1,1-TCA-contaminated site in the northeastern United States was also studied. This culture, referred to as MS, reductively dechlorinated 1,1,1-TCA to 1,1-dichloroethane (1,1-DCA) and then to monochloroethane (CA) when amended with methanol, ethanol, acetate and lactate. 16S rRNA gene sequence analysis revealed the presence of the putative dechlorinating organism Dehalobacter sp., whose growth during 1,1,1-TCA and 1,1-DCA dechlorination was confirmed by qPCR. In the presence of chlorinated ethenes, dechlorination 1,1,1-TCA by the culture MS was slowed, while dechlorination of 1,1-DCA was completely inhibited. Experiments with cell-free extracts and whole cell suspensions of culture MS suggested that chlorinated ethenes have direct inhibitory effects on 1,1,1-TCA reductive dehalogenase(s), while the inhibition of 1,1-DCA dechlorination may be due to effects on non-dehalogenase components of Dehalobacter sp. cells. Additionally, two novel reductive dehalogenase genes associated with 1,1,1-TCA reductive dechlorination were identified.

The WBC-2 consortium was characterized using quantitative PCR and analytical techniques to associate growth of dechlorinating bacteria to each step of the 1,1,2,2-Tetrachloroethane (TeCA) degradation pathway. The consortium was found to degrade TeCA through dichloroelimination to trans-1,2-dichloroethene (tDCE), and reductive dehalogenation to Vinyl Chloride (VC) and ethene. Thus the pathway was hypothesized to provide three distinct niches for three genera of dechlorinating bacteria, Dehalobacter, Dehalogenimonas and Dehalococcoides. Using qPCR to track growth over two time course experiments at different inoculum dilutions, the Dehalobacter species showed significant growth on the first step of TeCA dihaloelimination to tDCE Dehalococcoides and Dehalogenimonas species grew on the dechlorination products. The Dehalogenimonas species, a novel non-Dehalococcoides, was found to grow only on tDCE. The Dehalococcoides species also grew on cDCE, less well on tDCE, and on VC.

KB-1 is an anaerobic Dehalococcoides-containing microbial culture used industrially to bioremediate sites impacted with chlorinated solvents. The culture is typically grown at pH 7. However, lower pH is often encountered and therefore the effect of pH was investigated. Both sudden and stepwise decreases in pH from 7 to 6 and 5.5 were investigated over a period of 450 days. An electron balance was also calculated to look at the flow of electrons for dechlorination. More than 95% of the reducing equivalents went towards methanogenesis and acetogenesis. Select microorganisms were compared by quantitative Polymerase Chain Reaction. It was found that lower rates of dechlorination correspond to low Dehalococcoides numbers and that different methanogens were enriched on different electron donors.

abstract: Contamination by chlorinated ethenes is widespread in groundwater aquifers, sediment, and soils worldwide. The overarching objectives of my research were to understand how the bacterial genus Dehalococcoides function optimally to carry out reductive dechlorination of chlorinated ethenes in a mixed microbial community and then apply this knowledge to manage dechlorinating communities in the hydrogen-based membrane biofilm reactor (MBfR). The MBfR is used for the biological reduction of oxidized contaminants in water using hydrogen supplied as the electron donor by diffusion through gas-transfer fibers. First, I characterized a new anaerobic dechlorinating community developed in our laboratory, named DehaloR^2, in terms of chlorinated ethene turnover rates and assessed its microbial community composition. I then carried out an experiment to correlate performance and community structure for trichloroethene (TCE)-fed microbial consortia. Fill-and-draw reactors inoculated with DehaloR^2 demonstrated a direct correlation between microbial community function and structure as the TCE-pulsing rate was increased. An electron-balance analysis predicted the community structure based on measured concentrations of products and constant net yields for each microorganism. The predictions corresponded to trends in the community structure based on pyrosequencing and quantitative PCR up to the highest TCE pulsing rate, where deviations to the trend resulted from stress by the chlorinated ethenes. Next, I optimized a method for simultaneous detection of chlorinated ethenes and ethene at or below the Environmental Protection Agency maximum contaminant levels for groundwater using solid phase microextraction in a gas chromatograph with a flame ionization detector. This method is ideal for monitoring biological reductive dechlorination in groundwater, where ethene is the ultimate end product. The major advantage of this method is that it uses a small sample volume of 1 mL, making it ideally suited for bench-scale feasibility studies, such as the MBfR. Last, I developed a reliable start-up and operation strategy for TCE reduction in the MBfR. Successful operation relied on controlling the pH-increase effects of methanogenesis and homoacetogenesis, along with creating hydrogen limitation during start-up to allow dechlorinators to compete against other microorgansims. Methanogens were additionally minimized during continuous flow operation by a limitation in bicarbonate resulting from strong homoacetogenic activity. / Dissertation/Thesis / Ph.D. Civil and Environmental Engineering 2012

M.Sc. (Nanoscience) / Polychlorinated biphenyls (PCBs) are endocrine disrupting compounds (EDCs) and are harmful to humans and the environment. These PCBs are grouped under chlorinated organic compounds (COCs). The PCBs find their way to the environment through human activity such as industrialization and farming. Such activity produces wastes and runoffs that eventually end up in the water we use for drinking, farming and sanitation. It has then become necessary for researchers to find viable methods to remove these compounds from the environment. This is because current water treatment methods are not effective in the removal of the PCBs from water. The stages in the conventional treatment methods may include sand filtration, advanced oxidative processes and coagulation among others. These methods need to be energetically eco-friendly to drive the PCB dechlorination processes. Researchers have used a variety of metallic nanoparticles including bimetallic nanoparticles for the removal of COCs from water. However, nanoparticles tend to agglomerate when not supported - leading to a decrease in their activity. Hence it has become necessary to stabilize or immobilize these nanoparticles on suitable support materials, such as, polymer solutions or solid substrates. Solid substrates including metal oxides, carbon and membranes, are currently being explored. Poly(vinylidene difluoride) microfiltration membranes are especially suitable for this function given the high porosity, chemical inertness and other outstanding physical properties. In this work, the objective was to modify commercially hydrophilized poly(vinylidine)difluoride (PVDF) membranes with poly(ethylene glycol) (PEG). PEG is a bidentate polymer with two –OH groups found on either side of the molecule. The -OH groups allows PEG binding to the PVDF polymer backbone and hence high ability to capture or chelate the metal ions followed by their reduction. Nano-zerovalent metal nanoparticles were formed from these metal ions and chelated into the PEG grafted PVDF membrane to give the composite PVDF-PEG-Fe0. Post addition of the secondary metal was then followed by the introduction of the precomposite to a Pd solution to give the final catalytic membrane (PVDF-PEG-Fe0/Pd0). The use of PEG in this system allows for an even dispersion of the nanoparticles in the composite. The resulting nanocomposite membrane was used for the dechlorination of a polychlorinated biphenyl (PCB 77). Attenuated total reflection- Fourier transform infra red spectroscopy (ATR-FTIR) showed that PEG was successfully grafted onto the PVDF backbone. Optical contact angle measurements (OCA) were taken to determine the change in hydrophilicity of the membrane upon modification. X-ray diffraction spectroscopy (XRD) proved that the Pd and Fe nanoparticles immobilized on the system were indeed zerovalent. Scanning electron microscopy (SEM) images and contact angle measurements suggested a less porous membrane and slightly decreased hydrophilicity after modification. On the SEM micrographs the nanoparticles were observed to be quite evenly distributed in the membrane. Transmission electron microscopy (TEM) showed that the nanoparticles were in the range 20-30 nm in diameter, confirming the particle size values as determined by SEM. For the preliminary dechlorination studies done under ambient conditions, two dimensional column gas chromatography- time of flight- mass spectrometry (GCxGC-TOF-MS) results showed a complete dechlorination of PCB 77. A comparative study of the bare PVDF and catalytic membranes showed a slight difference in adsorption of the total PCB 77 concentrations. The catalytic membrane maintained its activity towards the dechlorination of PCB 77 after multiple regeneration cycles.