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Model Simulations in Support of Field Scale Design and Operation of Bioremediation Based on Cometabolic Degradation
Abstract
This paper addresses questions fundamental to the design and operation of aquifer bioremediation based on cometabolic degradation. A model of a full-scale, in situ system for bioremediation of chlorinated ethenes relying on cometabolic degradation was developed and applied to a hypothetical aquifer being considered for a large-scale field demonstration of in situ bioremediation with recirculation. The model was used to identify feasible substrate (electron donor and electron acceptor) delivery schedules. Trichloroethylene (TCE) was the target contaminant. Methane and phenol were considered as electron donors. The delivery of the electron donors and the electron acceptor, oxygen, was varied to evaluate the rate and extent of bioremediation under different substrate delivery schedules. Maximum removal of TCE was predicted when substrates are delivered at ratios near the stoichiometric requirement of electron donor and acceptor for net microbial growth.
Additionally, the decrease in TCE removal that results from using substrate delivery schedules other than those achieving the maximum removal of TCE was quantified. This decrease was greater for the methane-oxygen system because the two gaseous substrates compete for transfer into the recirculated ground water. If one substrate is introduced in excess of the amount required for net microbial growth, it accumulates, thus limiting the ability to introduce the second substrate. This imbalance both limits the introduction of the second substrate and accelerates the accumulation of the substrate added in excess. The phenol-oxygen system is less sensitive to deviation away from the best observed substrate delivery schedule because phenol is a relatively soluble liquid and its introduction does not compete with the mass transfer of oxygen.
... Although numerous studies focused on in-situ pilot-scale applications of aerobic CAH cometabolism [3,5,6,8,[18][19][20], fullscale applications of aerobic cometabolic bioremediation of CAH-contaminated sites are still rare [21][22][23], as several issues still need to be addressed. Of these, two deserve particular mention: i) there is a risk of completing growth substrate consumption near the injection wells; ii) aquifer clogging near the injection wells can occur due to excessive biomass growth [3,[24][25][26]. The supply of alternated pulses of growth substrate and oxygen is effective both in the creation of a long bioreactive zone and in controlling aquifer clogging: as a result of hydrodynamic dispersion and substrate sorption, the overlapping of substrate and oxygen occurs at low concentrations, over a wide aquifer portion and, in each point, in a discontinuous way. ...
... The kinetic model defined by Eqs. (2), (3), (4) and (6), characterized by competitive inhibition and product toxicity, was selected by several authors as the most appropriate one for aerobic cometabolism both for lab-scale studies [2,4,33,34,36,37] and in-situ applications [5,18,25,38,39], although a smaller number of studies obtained better results with non-competitive or mixed inhibition [40][41][42]. Alternative approaches to simulate transformation product toxicity, such as the one proposed by Ely et al. [43,44] were not considered here, given the higher number of parameters in the kinetic equations. ...
... Alternative approaches to simulate transformation product toxicity, such as the one proposed by Ely et al. [43,44] were not considered here, given the higher number of parameters in the kinetic equations. While several models of in-situ CAH biodegradation do not include a suspended biomass fraction [5,18,25,38,39], we distinguished between cXa and cXs, in order to account for the loss of suspended cells in the reactor outlet, to be able to compare the experimental and simulated values of cXs and to develop a tool capable to predict in-situ microbial transport in bioaugmentation treatments. Our assumption of equilibrium between cXa and cXs can be found in similar studies [e.g., 45] and represents a simplification of more complex models of microbial adhesion and detachment [19,[46][47][48][49], aimed at limiting the number of model parameters. ...
A process of chloroform (CF) aerobic cometabolic biodegradation by a butane-growing consortium was studied for 354days in a 2-m continuous-flow sand-filled reactor. The study was aimed at (a) investigating the oxygen/substrate pulsed injection as a tool to control the clogging of the porous medium, to attain a wide bioreactive zone and to reduce substrate inhibition on CF cometabolism; (b) developing a reliable model of CF cometabolism in porous media. While the continuous supply of butane rapidly led to the clogging of the porous medium due to excessive biomass growth, the testing of six types of oxygen/substrate pulsed feeding led to the identification of a feeding schedule capable to prevent aquifer clogging and to lead to the development of a long bioreactive zone and to satisfactory CF degradation performances. The tested model of aerobic cometabolism allowed a suitable interpretation of the experimental data as long as the ratio of CF degraded to butane consumed was ≤0.27mgCFmgbutane−1. A long-term 1-D simulation of the best-performing schedule of pulsed oxygen/substrate supply extended to a 30-m aquifer length resulted in a 20-m bioreactive zone and in a 96% CF removal.
... Indeed, it allows a reduction of the GS-CAH inhibition and a discontinuous microbial growth at each point, but it is characterized by a more rapid biomass growth and therefore a narrower bioreactive zone. Two modeling studies were specifically dedicated to the optimization of the pulsed supply of GS and oxygen in in situ applications of AC [84,85]. These works showed that the mass ratio of O 2 to GS supplied in each cycle should be significantly higher than the stoichiometric value in order to ensure the complete consumption of GS and to avoid the generation of anaerobic zones in the bioreactive area. ...
... Secondly, the fear of an incomplete consumption of the supplied GS seems unjustified, since all the in situ studies that included a rigorous monitoring of GS concentrations indicated that complete microbial uptake occurred. To ensure complete GS consumption, it is crucial to design the pulsed GS/O 2 supply with a large excess of O 2 [84]. Third, the great advances made during the last 15 years in the simulation of the different approaches for in situ AC allow for a very high level of understanding and control of the process. ...
The possible approaches for in-situ aerobic cometabolism of aquifers and vadose zones contaminated by chlorinated solvents are critically evaluated. Bioaugmentation of resting-cells previously grown in a fermenter and in-well addition of oxygen and growth substrate appear to be the most promising approaches for aquifer bioremediation. Other solutions involving the sparging of air lead to satisfactory pollutant removals, but must be integrated by the extraction and subsequent treatment of vapors to avoid the dispersion of volatile chlorinated solvents in the atmosphere. Cometabolic bioventing is the only possible approach for the aerobic cometabolic bioremediation of the vadose zone. The examined studies indicate that in-situ aerobic cometabolism leads to the biodegradation of a wide range of chlorinated solvents within remediation times that vary between 1 and 17 months. Numerous studies include a simulation of the experimental field data. The modeling of the process attained a high reliability, and represents a crucial tool for the elaboration of field data obtained in pilot tests and for the design of the full-scale systems. Further research is needed to attain higher concentrations of chlorinated solvent degrading microbes and more reliable cost estimates. Lastly, a procedure for the design of full-scale in-situ aerobic cometabolic bioremediation processes is proposed.
... Interestingly , when first-order decay models were applied to experimental data, the first-order rate constant (deactivation rate) for the T4MO-expresing R. pickettii PKO1 bacterium (4.68 d )1 ) was more than an order of magnitude greater than deactivation rates observed for the other oxygenaseexpressing bacteria ( <0.2–0.36 d )1 ) (Costura & Alvarez 2000; Jenkins & Heald 1996; Jones & Morita 1985; Park 2001; Roslev & King 1994). Such a wide range of deactivation rates suggests that deactivation may have a significant influence on the effectiveness of bioremediation processes and the accuracy of in situ biodegradation rate predictions (Lang et al. 1997; Park et al. 2001). However, the dependence of deactivation rates on biological and environmental factors is currently unclear, complicating the ability to incorporate deactivation information in prediction and design methodologies. ...
... Biostimulation processes involving the addition of a primary growth substrate commonly utilize pulse injections to improve mixing and spatial distribution (McCarty et al. 1998). The addition of pulse injections, however, can result in a condition where organisms experience highly fluctuating substrate concentrations and possible periods of carbon and energy limitations (Lang et al. 1997; McCarty et al. 1998; Park et al. 2001). Accounting for starvation effects, therefore, would be useful for designing the length of the substrate pulse such that deactivation of biodegradation activity is minimized. ...
Subsurface bacteria commonly exist in a starvation state with only periodic exposure to utilizable sources of carbon and energy. In this study, the effect of carbon starvation on aerobic toluene degradation was quantitatively evaluated with a selection of bacteria representing all the known toluene oxygenase enzyme pathways. For all the investigated strains, the rate of toluene biodegradation decreased exponentially with starvation time. First-order deactivation rate constants for TMO-expressing bacteria were approximately an order of magnitude greater than those for other oxygenase-expressing bacteria. When growth conditions (the type of growth substrate and the type and concentration of toluene oxygenase inducer) were varied in the cultures prior to the deactivation experiments, the rate of deactivation was not significantly affected, suggesting that the rate of deactivation is independent of previous substrate/inducer conditions. Because TMO-expressing bacteria are known to efficiently detoxify TCE in subsurface environments, these findings have significant implications for in situ TCE bioremediation, specifically for environments experiencing variable growth-substrate exposure conditions.
... To efficiently achieve in situ mixing of contaminants and remedial compounds, these technologies conventionally employ single or multiple injection/extraction well pairs, also known as recirculation wells. In these systems, contaminated water is extracted from a downgradient extraction well, amendments that promote biological or chemical contaminant degradation (i.e., ISB and ISCO, respectively) or enhance the effective aqueous solubility of nonaqueous phase liquid (NAPL) constituents (i.e., SEAR) are mixed with the extracted contaminated water, and then the extracted water containing the amendments is reinjected from an upgradient injection well (McCarty et al., 1998;Lang et al., 1997;Christ et al., 1999;Gandhi et al., 2002a;Cunningham et al., 2004;North et al., 2012). In these recirculation wells, a portion of the reinjected water is recaptured by the extraction well, while the remainder escapes the recirculation zone to flow along the gradient. ...
In general, in situ remediation techniques require that treatment agents come into contact with contaminants to facilitate the treatment process. Greater contact causes more in situ mixing of the two compounds and greater contaminant reduction. In a recirculation well system featuring an injection/extraction well pair, delivery controls the remedial and economic efficiency of decontamination, and is therefore a key consideration for successful in situ remediation. In this study, we numerically evaluated the remedial and economic efficiency of a recirculation well system with sinusoidal temporally varying pumping and injection rates for enhancing remediation; the results were compared with those of a traditional recirculation well system with constant injection/extraction rates. We performed sensitivity analyses to determine the optimal values of four operational parameters associated with the effects of temporally variable pumping or injection rates on the cumulative swept area of injected chemical amendment for a given operation time or cumulative injected volume, which are good measures of remediation and economic efficiency. The findings of this study provide insight into the mechanical process of plume spreading in response to injection/pumping operational strategies, and demonstrate that enhanced plume spreading is a key requirement for achieving sufficient contact between chemical amendments and contaminants.
... As for the kinetic model, for the initial elaboration of each experimental phase the reaction rates ri were based on the AC model with competitive CAH-growth substrate mutual inhibition and transformation product toxicity proposed by Alvarez-Cohen and McCarty [4] and satisfactorily utilized in numerous studies of CAH AC: [33,39,57,58] ...
Chlorinated solvents are toxic and poorly biodegradable pollutants frequently found in contaminated aquifers. Experimental data of chloroform (CF) aerobic cometabolic biodegradation in a sand column with butane as growth substrate were simulated with a system of non‐stationary second‐order partial differential equations with non‐linear kinetic terms. A MATLAB optimization code based on the Gauss‐Newton method and coupled with the Comsol Multiphysics finite elements solver was developed to calibrate the model. For each experimental phase, the best‐fit quality was evaluated by an innovative multi‐variable model adequacy test. The proposed code solved systems of up to 5 partial differential equations and optimized up to 6 unknown parameters, leading to statistically acceptable best‐fits. The optimization of the butane/oxygen pulsed feed led to an 82% CF biodegradation and to a 0.27 gCF /gbutane transformation yield. When the substrate/pollutant ratio was minimized, the standard model of aerobic cometabolism initially tested required additional terms aimed at taking into account the depletion of reducing energy, in order to attain a statistically acceptable best‐fit. This is the first work in which a model of aerobic cometabolism taking into account reducing energy availability was applied to a continuous‐flow process. The proposed optimization code can be used for model calibration in a wide range of physical problems described by non‐stationary, non‐linear partial differential equations, a task that no commercial software can perform. The developed code is made available in the Supplementary Material. This article is protected by copyright. All rights reserved
... (5) Only species in the aqueous phase can be degraded. Other models based on the above assumptions are presented by Lang [1995], Lang et al. [1997], andMacDonald et al. [1999]. Unlike the code developed by Lang [1995] and used by McCarty et al. [1998] to design the Edwards experiment, which was limited to transport and reaction along one-dimensional stream tubes in a two-dimensional flow field, our model is fully threedimensional. ...
... • Sia il VC (figura 24) sia il cDCE erano già stati fortemente degradati durante la fase 3. Sembra che la loro degradazione sia rallentata durante la Fase 4, anche se non è perfettamente chiaro il meccanismo responsabile: si tratta probabilmente del contemporaneo uso dell'ossigeno nella degradazione veloce del propano. accettori di elettroni, substrati primari e, se necessario, di nutrienti nelle aree di interesse e per i tempi necessari ad ottenere la degradazione dei contaminanti ai livelli desiderati (Lang et al., 1997). ...
The TMVOC V. 2.0 reservoir simulator was used to model a pilot test involving the stimulation of direct and cometabolic aerobic degradation reactions of chlorinated organic compounds in a confined aquifer present in the subsurface of a coastal industrial site. Prior to the test, the flow field and the pressure distribution generated under steady state conditions by an extraction/reinjection well doublet were simulated by means of analytical and semi-analytical methods. Then, a numerical model was developed with TMVOC, that adequately reproduces the flow field and the pressure distribution calculated previously.
The model was used to simulate the various phases of the biodegradation pilot test that include first the injection of oxygen, the injection of a primary substrate and then the degradation reactions in the aquifer when the well doublet is operating. The principal parameters that describe the degradation processes were derived from experimental activities conducted in the laboratory, and were partly calibrated to reproduce the experimental results obtained with a slurry microcosm prepared with soil and water samples collected at the site.
The simulations allowed to determine the spatial distribution in the aquifer over time, as well as the concentrations in the fluid extracted from the production well, of injected oxygen and primary substrate, and of the chlorinated solvents dissolved in the groundwater. The numerical simulation underlined the strong coupling of flow and transport phenomena in the aquifer with the degradation reactions of chlorinated solvents. It also confirmed, within the limits of available knowledge and of the simplified assumptions adopted, that the proposed stimulation of identified degradation processes is capable of promoting the degradation of contaminants directly on site.
... Chlorinated compounds generally do not act as primary substrates, but may be degraded by cometabolic degradation. In cometabolic degradation, an enzyme produced by degradation of another carbon compound (the primary substrate) fortuitously catalyzes the reduction of the another compound (Lang et al., 1997). ...
... Taylor and Jaffe (1991) applied a bioremediation model to evaluate in situ bioremediation design for sorbing and nonsorbing contaminants. Lang et al. (1997) designed in situ bioremediation systems relying on cometabolic degradation. These approaches employ only bioremediation models to evaluate the efficiency of alternative system designs. ...
Presented is a simulation/optimization (S/O) model combining optimization with BIOPLUME II for optimizing in situ bioremediation system design. The S/O model uses a new hybrid method combining genetic algorithms and simulated annealing to search for an optimal design and applies the BIOPLUME H model to simulate aquifer hydraulics and bioremediation. This new hybrid method is parallel recombinative simulated annealing, which is a general-purpose optimization approach that has the good convergence of simulated annealing and the efficient parallelization of a genetic algorithm. We propose a two-stage management approach. The first-stage design goal is to minimize total system cost (pumping/treatment, well installation, and facility capital costs). The second-stage design goal is to minimize the cost of a time-varying pumping strategy using the optimal system chosen by the first-stage optimization. Optimization results show that parallel recombinative simulated annealing performs better than simulated annealing and genetic algorithms for optimizing system design when including installation costs. New explicit well installation coding improves algorithm convergence. Threshold accepting reduces computation time 43% by eliminating unnecessary simulation runs. Applying the optimal time-varying pumping strategy in the second stage reduces pumping cost by 31%.
... The simplest configuration, depicted in Fig. 1, has an extraction well drawing contaminated water out of an aquifer. Nutrients and other amendments can be added to the water, Ž which is then reinjected into the aquifer Buchanan et al., 1997;Lang et al., 1997;Spuij . et al., 1997 . ...
Innovative in situ remediation technologies using injection/extraction well pairs which recirculate partially treated groundwater as a method of increasing overall treatment efficiency have recently been demonstrated. An important parameter in determining overall treatment efficiency is the fraction of flow that recycles between injection and extraction points. Numerical and semi-analytical methods are currently available to determine this parameter and an analytical solution has been presented for a single injection/extraction well pair. In this work, the analytical solution for fraction of recycled flow for a single injection/extraction well pair is extended to multiple co-linear well pairs. In addition, a semi-analytical method is presented which permits direct calculation of fraction recycle for a system of arbitrarily placed injection/extraction wells pumping at different rates. The solution is used to investigate the behavior of multiple well pairs, looking at how well placement and pumping rates impact capture zone width and fraction recirculation (and, therefore, overall treatment efficiency). Results of the two-well solution are compared with data obtained at a recent field-scale demonstration of in situ aerobic cometabolic bioremediation, where a pair of dual-screened injection/extraction wells was evaluated. A numerical model is then used to evaluate the impact of hydraulic conductivity anisotropy on the overall treatment efficiency of the dual-screened injection/extraction well pair. The method presented here provides a fast and accurate technique for determining the efficacy of injection/extraction systems, and represents a tool that can be useful when designing such in situ treatment systems.
... [3] Numerical models have been used to both design subsurface delivery strategies [Lang et al., 1997;Hyndman et al., 2000;Scheibe et al., 2001] and evaluate biodegradation processes in laboratory experiments [Chen et al., 1992;Phanikumar et al., 2002;Kim et al., 2003]. Such models can provide significant insight into the nature and rates of the processes that drive bioremediation when adequate laboratory data are available for model development and calibration. ...
1] Bioremediation can be a cost-effective approach to clean up groundwater contaminants, especially in cases where numerical models have been used to evaluate microbial processes and design remediation strategies. This paper describes the three-dimensional reactive transport modeling of carbon tetrachloride (CT) bioremediation at the Schoolcraft site in western Michigan. A ''biocurtain'' was created to remediate this site using a recirculation well gallery installed normal to groundwater flow, which was inoculated with nonnative microbes and fed with weekly additions of electron donor (acetate) and nutrients. The model simulates the transport and reactions of aqueous and sorbed phase CT, acetate, electron acceptor (nitrate), mobile and immobile microbes, and tracer (bromide). Simulated microbial processes include growth, decay, attachment, detachment, and endogenous respiration. This model was used to predict solute concentrations across the site using laboratory-based reaction parameters and to evaluate changes in rates from the laboratory to the field. A reasonable agreement was found between predicted and observed acetate and nitrate concentrations; however, a lower CT degradation rate was needed to describe the CT concentrations observed after the inoculation event.
... Several other processes become important when analyzing multiple interacting microbial populations. Population interactions that have received the most attention are competition, predation, and cometabolism (Bailey and Ollis 1986; Semprini et al. 1991; Mohn and Tiedje 1992; Semprini and McCarty 1992; Harvey et al. 1995; Lang et al. 1997;). Although competition has a much broader definition in population dynamics, in terms of representing this process in Monod kinetics, competition is simply when two or more microbial species compete for the same nutrients. ...
The incorporation of microbial processes into reactive transport models has generally proceeded along two separate lines of investigation: (1) transport of bacteria as inert colloids in porous media, and (2) the biodegradation of dissolved contaminants by a stationary phase of bacteria. Research over the last decade has indicated that these processes are closely linked. This linkage may occur when a change in metabolic activity alters the attachment/detachment rates of bacteria to surfaces, either promoting or retarding bacterial transport in a groundwater-contaminant plume. Changes in metabolic activity, in turn, are controlled by the time of exposure of the microbes to electron acceptors/donor and other components affecting activity. Similarly, metabolic activity can affect the reversibility of attachment, depending on the residence time of active microbes. Thus, improvements in quantitative analysis of active subsurface biota necessitate direct linkages between substrate availability, metabolic activity, growth, and attachment/detachment rates. This linkage requires both a detailed understanding of the biological processes and robust quantitative representations of these processes that can be rested experimentally. This paper presents an overview of current approaches used to represent physicochemical and biological processes in porous media, along with new conceptual approaches that link metabolic activity with partitioning of the microorganism between the aqueous and solid phases.
... A first criterion for the choice between continuous and pulsed substrate supply is given by the necessity to avoid any clogging of the porous medium. Indeed, several in-situ studies of aerobic cometabolism reported that the continuous substrate supply determines a high risk of aquifer clogging as a result of an excessive biomass growth near the injection well [21][22][23][24]44,45]. On the contrary, in this work a 72-day continuous substrate/oxygen supply did not lead to any clogging of the reactor porosity. ...
This work focuses on chloroform (CF) cometabolism by a butane-grown aerobic pure culture (Rhodococcus aetherovorans BCP1) in continuous-flow biofilm reactors. The goals were to obtain preliminary information on the feasibility of CF biodegradation by BCP1 in biofilm reactors and to evaluate the applicability of the pulsed injection of growth substrate and oxygen to biofilm reactors. The attached-cell tests were initially conducted in a 0.165-L bioreactor and, then, scaled-up to a 1.772-L bioreactor. Glass cylinders were utilized as biofilm carriers. The continuous supply of growth substrate (butane), which led to the attainment of the highest CF degradation rate (8.4 mgCF day−1 m
biofilm surface−2), was compared with four schedules of butane and oxygen pulsed feeding. The pulsed injection technique allowed the attainment of a ratio of CF mass degraded per unit mass of butane supplied equal to 0.16 mgCF mg
butane−1, a value 4.4 times higher than that obtained with the continuous substrate supply. A procedure based on the utilization of integral mass balances and of average concentrations along the bioreactors resulted in a satisfactory match between the predicted and the experimental CF degradation performances, and can therefore be utilized to provide a guideline for optimizing the substrate pulsed injection schedule.
Homoacetogenesis and methanogenesis, which usually occur during anaerobic trichloroethene (TCE) dechlorination, affect the removal of TCE and its daughter products. This study develops a one-dimensional, multispecies H2-based biofilm model to simulate the interactions among six solid biomass species [Dehalococcoides, Geobacter, methanogens, homoacetogens, inert biomass (IB), and extracellular polymeric substances (EPS)] and 10 dissolved chemical species [TCE, dichloroethene (DCE), vinyl chloride (VC), ethene, hydrogen (H2), methane, acetate, bicarbonate, utilization associated products (UAP), and biomass associated products (BAP)]. To evaluate and parameterize the model, parameter values from the literature were input into the model to simulate conditions reported for an experiment. The biomass species distribution in the biofilm and the chemical species concentrations in the reactor effluent at a steady state were generally consistent between the experiments and the model. The predicted 15-μm biofilm consisted of three layers, each dominated by a different active biomass type: homoacetogens in the layer next to the membrane, Geobacter in the biofilm surface layer (next to the water), and Dehalococcoides in-between.
In this chapter we discuss how recirculation systems can be engineered to achieve mixing in order to facilitate in situ remediation. As noted in Chapters 1 and 5, recirculation systems have a number of advantages. First, as active systems, they can be designed to maximize the probability of contaminant control and capture, even under changing hydrological conditions. Second, since the systems use wells, the recirculation zone(s) can be established at depths that can’t be impacted by other systems (e.g., permeable reactive barriers). Third, as will be discussed subsequently, mixing of the amendment and the contaminant occurs in an engineered reactor, either in-well or aboveground, and is therefore relatively complete. Fourth, for systems that rely upon biodegradation, it has been shown that recirculation systems can be used to establish in situ biological treatment zones that are effective in biodegrading the target contaminant, even when initially biodegradation activity is sparse (Hoelen et al., 2006). Finally, as net loss of water is minimized when recirculation is used, the systems are very useful in regions where water needs to be conserved. They can be designed to confine a source of contamination and treat it there, or to act similar to a barrier wall by removing contaminants in a passing plume to prevent downgradient contamination. Of course, as active systems that use wells, there are a number of attendant disadvantages to recirculation; the most obvious being the operation, maintenance, and monitoring costs associated with a “pump-and-treat” system (the fact that the treatment happens to occur in situ notwithstanding). Additionally, as systems which rely on pumping, the water which is captured and amended will come preferentially from the most permeable zones of the aquifer. Contaminated water that is resident in low permeability regions may not be treated.
Bioremediation technology is analogous to all other technologies that gradually become accepted by society. Bioremediation is based on fundamental scientific and engineering principles that are translated into applications that reliably serve the public good. All technologies (from airplanes to automobiles to electric lights to personal computers) traverse various developmental stages — each of unpredictable duration, that include conception, proof of concept, prototype development, scale up, design refinements, applications testing, and marketing (among others). Implicit in the title of this chapter is the fact that many applications of bioremediation technology are still in their infancy. Bioremediation still needs quality control — this technology still needs to define its boundaries between promise and reality.
Papers addressing fate and transport processes for analyzing groundwater quality are presented. These papers are separated into water-saturated systems, vanadose zone systems and systems containing nonaqueous phase liquids. Within each of these categories, the articles are further divided based on physical, chemical and biological processes relevant to each system. Papers dealing with groundwater monitoring and groundwater remediation are then treated, with papers grouped technology. Finally, papers describing risk assessment and groundwater protection are presented.
This paper provides a review of literature published in 1995 on the subject of Hazardous waste treatment technologies. Topics covered include: General; Biological treatment - aerobic processes, anaerobic processes, membrane bioreactors, and biosorption/bioleaching, biofiltration and vapor-phase treatment, in situ bioremediation; Chemical and physical treatment - advanced oxidation, membrane processes, vapor sparging/extracting, solution flushing/extracting, and other chemical/physical processes; and, Thermal treatment - combustion/incineration, thermal desorption, and thermal plasma/vitrification.
Phenol, or hydroxybenzene, is both a synthetically and naturally produced aromatic compound. Microorganisms capable of degrading phenol are common and include both aerobes and anaerobes. Many aerobic phenol-degrading microorganisms have been isolated and the pathways for the aerobic degradation of phenol are now firmly established. The first steps include oxygenation of phenol by phenol hydroxylase enzymes to form catechol, followed by ring cleavage adjacent to or in between the two hydroxyl groups of catechol. Phenol hydroxylases ranging from simple flavoprotein monooxygenases to multicomponent hydroxylases, as well as the genes coding for these enzymes, have been described for a number of aerobic phenol-degrading microorganisms. Phenol can also be degraded in the absence of oxygen. Our knowledge of this process is less advanced than that of the aerobic process, and only a few anaerobic phenol-degrading bacteria have been isolated to date. Convincing evidence from both pure culture studies with the denitrifying organism Thauera aromatica K172 and with two Clostridium species, as well as from mixed culture studies, indicates that the first step in anaerobic phenol degradation is carboxylation in the para-position to form 4-hydroxybenzoate. Following para-carboxylation, thioesterification of 4-hydroxybenzoate to co-enzyme A allows subsequent ring reduction, hydration, and fission. Para-carboxylation appears to be involved in the anaerobic degradation of a number of aromatic compounds. Numerous practical applications exist for microbial phenol degradation. These include the exploitation of indigenous anaerobic phenol-degrading bacteria in the in situ bioremediation of creosote-contaminated subsurface environments, and the use of phenol as a co-substrate for indigenous aerobic phenol-degrading bacteria to enhance in situ biodegradation of chlorinated solvents.
Microporous gas-permeable membranes are being investigated as a means to provide in situ delivery of hydrogen to contaminated groundwater to stimulate the biodegradation of chlorinated solvents. Because of the potential for sulfate reducing biofilms to form on the membrane and indirectly produce iron sulfides at the membrane surface, the effects of biological fouling and inorganic fouling on gas transfer performance were evaluated. In general, the gas transfer coefficient decreased as the quantity of foulant accumulated on the membranes increased. Despite the accumulation of a thick (up to 100 μm) foulant layer comprised of micro-organisms and iron sulfide minerals, gas delivery continued under the conditions tested [Reynolds numbers (Re) from 650 to 5460]. More importantly, gas transfer would not have been significantly impeded by these foulants at typical groundwater velocities (Re from 4 × 10-4 to 4 × 10-3) due to the dominance of the liquid film resistance under those conditions. Therefore, gas-permeable membranes should be able to provide sustained gas delivery to groundwater, even in a fouling environment.
Halogenated volatile organic compounds, including chlorinated solvents, are the most frequently-occurring type of soil and groundwater contaminant at Superfund and other hazardous waste sites in the United States. The U.S. Environmental Protection Agency (EPA) estimates that, over the next several decades, site owners will spend billions of dollars to clean up these sites. New technologies that are less costly and more effective are needed to accomplish hazardous waste site remediation. As these new and innovative technologies are being developed and used, site managers require information on how they work, their performance to date, and how to evaluate their application at a particular site. This report provides an overview of the fundamentals and field applications of in situ bioremediation to remediate chlorinated solvents in contaminated soil and groundwater. In situ treatment is increasingly being selected to remediate sites because it is usually less expensive, and does not require waste extraction or excavation. In addition, in situ bioremediation is more publicly acceptable than above-ground technologies because it relies on natural processes to treat contaminants. This document presents information at a level of detail intended to familiarize federal and state project managers, permit writers, technology users, and contractors with in situ bioremediation. The report describes how chlorinated solvents are degraded, how to enhance the process by the addition of various materials and chemicals, design configurations, and the typical steps taken to evaluate technology feasibility at a specific site. It also includes a list of technology vendors and nine case studies of field applications. It is important to note that this report cannot be used as the sole basis for determining this technology's applicability to a specific site.
“Biodegradation” of organic compounds is the partial simplification or complete destruction of their molecular structure by physiological reactions catalyzed by microorganisms. Understanding and proving biodegradation processes under laboratory and field conditions is a science of ongoing discovery. This discovery requires a close dialog among many disciplines. It must be recognized that only under relatively rare circumstances is a proof of field bioremediation unequivocal when a single piece of evidence is relied on. In the majority of cases, the complexities of contaminant mixtures, their hydrogeochemical settings, and accounting for competing abiotic mechanisms of contaminant loss make it a challenge to document biodegradation processes.
1] We present an analysis of an extensively monitored full-scale field demonstration of in situ treatment of trichloroethylene (TCE) contamination by aerobic cometabolic biodegradation. The demonstration was conducted at Edwards Air Force Base in southern California. There are two TCE-contaminated aquifers at the site, separated from one another by a clay aquitard. The treatment system consisted of two recirculating wells located 10 m apart. Each well was screened in both of the contaminated aquifers. Toluene, oxygen, and hydrogen peroxide were added to the water in both wells. At one well, water was pumped from the upper aquifer to the lower aquifer. In the other well, pumping was from the lower to the upper aquifer. This resulted in a ''conveyor belt'' flow system with recirculation between the two aquifers. The treatment system was successfully operated for a 410 day period. We explore how well a finite element reactive transport model can describe the key processes in an engineered field system. Our model simulates TCE, toluene, oxygen, hydrogen peroxide, and microbial growth/death. Simulated processes include advective-dispersive transport, biodegradation, the inhibitory effect of hydrogen peroxide on biomass growth, and oxygen degassing. Several parameter values were fixed to laboratory values or values from previous modeling studies. The remaining six parameter values were obtained by calibrating the model to 7213 TCE concentration data and 6997 dissolved oxygen concentration data collected during the demonstration using a simulation-regression procedure. In this complex flow field involving reactive transport, TCE and dissolved oxygen concentration histories are matched very well by the calibrated model. Both simulated and observed toluene concentrations display similar high-frequency oscillations due to pulsed toluene injection approximately one half hour during each 8 hour period. Simulation results indicate that over the course of the demonstration, 6.9 kg of TCE was degraded and that in the upper aquifer a region 40 m wide extending 25 m down gradient of the treatment system was cleaned up to less than 100 mg L À1 from initial concentrations of approximately 700 mg L À1 . A smaller region was cleaned up to less than 30 mg L À1 . Simulations indicate that the cleaned up area in the upper aquifer would continue to expand for as long as treatment was continued.
Vertical circulation wells can efficiently provide microorganisms with substrates needed for enhanced bioremediation. We present a travel-time based approach for modeling bioreactive transport in a flow field caused by a series of circulation wells. Mixing within the aquifer is due to the differences in sorption behavior of the reactants. Neglecting local dispersion, transport simplifies to a single one-dimensional problem with constant coefficients for each well. Recirculation is characterized by the discharge densities over travel time. We apply the model to the stimulation of cometabolic dechlorination of trichloroethene (TCE) by alternate injection of oxygen and toluene into the circulation wells. Mixing within the wells can be minimized by interposing sufficiently long breaks between the oxygen and toluene pulses. In our simulation, the proposed injection scheme stimulates biomass growth without risking biofouling of the aquifer.
For aerobic co-metabolism of chlorinated solvents to occur, it is necessary that oxygen, a primary substrate, and the chlorinated compound all be available to an appropriate microorganism--that is, a microorganism capable of producing the nonspecific enzyme that will promote degradation of the contaminant while the primary substrate is aerobically metabolized. Thus, the transport processes that serve to mix the reactants are crucial in determining the rate and extent of biodegradation, particularly when considering in situ biodegradation. These transport processes intersect, at a range of scales, with the biochemical reactions. This paper reviews how the important processes contributing to aerobic co-metabolism of chlorinated solvents at different scales can be integrated into mathematical models. The application of these models to field-scale bioremediation is critically examined. It is demonstrated that modeling can be a useful tool in gaining insight into the physical, chemical, and biological processes relevant to aerobic co-metabolism, designing aerobic co-metabolic bioremediation systems, and predicting system performance. Research needs are identified that primarily relate to gaps in our current knowledge of inter-scale interactions.
Groundwater contamination due to releases of petroleum products is a major environmental concern in many urban districts and industrial zones. Over the past years, a few studies were undertaken to address in situ bioremediation processes coupled with contaminant transport in two- or three-dimensional domains. However, they were concentrated on natural attenuation processes for petroleum contaminants or enhanced in situ bioremediation processes in laboratory columns. In this study, an integrated numerical and physical modeling system is developed for simulating an enhanced in situ biodegradation (EISB) process coupled with three-dimensional multiphase multicomponent flow and transport simulation in a multi-dimensional pilot-scale physical model. The designed pilot-scale physical model is effective in tackling natural attenuation and EISB processes for site remediation. The simulation results demonstrate that the developed system is effective in modeling the EISB process, and can thus be used for investigating the effects of various uncertainties.
In-situ methanotrophic bioremediation of ground water contaminated with petroleum hydrocarbons and chlorinated aliphatic hydrocarbons (CAHs) often require the addition of Oâ for compound oxidation. The purpose of this research was to test the feasibility of a down-well gas-transfer apparatus for adding Oâ directly into recirculating ground water. At times it may be desirable to add other gases as well, such as methane for CAH co-metabolism A high downward velocity of water through the inlet tube of an inverted cone prevented upward movement of injected Oâ bubbles. The addition of gas-liquid mixing cones with in the transfer section of the inverted cone increased the turbulence below the inlet. Complete dissolution of gases is possible. The relationships between Oâ transfer rate and system variables such a pressure loss, water flow rate, and Nâ content in the influent water were evaluated. A significant factor affecting Oâ mass transfer was the total dissolved gas content in the water entering the transfer device. From the results of this study, a design for practical conditions of operation was developed.
A series of percolation experiments was carried out to determine the extent to which an obligately aerobic bacterial strain, Arthrobacter sp., is able to clog permeameters filled with fine sand. The experimental results indicate that strictly aerobic bacteria are able to reduce saturated hydraulic conductivity (K) by up to four orders of magnitude. This decrease in K within the sand does not appear on scanning electron micrographs to be caused by exopolymers, which are entirely absent, but rather seeems to be due to the presence of large aggregates of cells that form local plugs within the pores. Under conditions of severe N limitation, the same mechanism seems to be largely responsible for the observed clogging. In all cases, the coverage of the solid surfaces by the bacterial cells is sparse and heterogeneous. -from Authors
Tetrachloroethylene (PCE) and trichloroethylene (TCE), common industrial solvents, are among the most frequent contaminants found in groundwater supplies. Due to the potential toxicity and carcinogenicity of chlorinated ethylenes, knowledge about their transformation potential is important in evaluating their environmental fate. The results of this study confirm that PCE can be transformed by reductive dehalogenation to TCE, dichloroethylene, and vinyl chloride (VC) under anaerobic conditions. In addition, [14C]PCE was at least partially mineralized to CO2. Mineralization of 24% of the PCE occurred in a continuous-flow fixed-film methanogenic column with a liquid detention time of 4 days. TCE was the major intermediate formed, but traces of dichloroethylene isomers and VC were also found. In other column studies under a different set of methanogenic conditions, nearly quantitative conversion of PCE to VC was found. These studies clearly demonstrate that TCE and VC are major intermediates in PCE biotransformation under anaerobic conditions and suggest that potential exists for the complete mineralization of PCE to CO2 in soil and aquifer systems and in biological treatment processes.
The kinetics of the degradation of trichloroethylene (TCE) and seven other chlorinated aliphatic hydrocarbons by Methylosinus trichosporium OB3b were studied. All experiments were performed with cells grown under copper stress and thus expressing soluble methane monooxygenase. Compounds that were readily degraded included chloroform, trans-1,2-dichloroethylene, and TCE, with Vmax values of 550, 330, and 290 nmol min-1 mg of cells-1, respectively. 1,1-Dichloroethylene was a very poor substrate. TCE was found to be toxic for the cells, and this phenomenon was studied in detail. Addition of activated carbon decreased the acute toxicity of high levels of TCE by adsorption, and slow desorption enabled the cells to partially degrade TCE. TCE was also toxic by inactivating the cells during its conversion. The degree of inactivation was proportional to the amount of TCE degraded; maximum degradation occurred at a concentration of 2 mumol of TCE mg of cells-1. During conversion of [14C]TCE, various proteins became radiolabeled, including the alpha-subunit of the hydroxylase component of soluble methane monooxygenase. This indicated that TCE-mediated inactivation of cells was caused by nonspecific covalent binding of degradation products to cellular proteins.
The trichloroethylene (TCE) transformation rate and capacity of a mixed methanotrophic culture at room temperature were measured to determine the effects of time without methane (resting), use of an alternative energy source (formate), aeration, and toxicity of TCE and its transformation products. The initial specific TCE transformation rate of resting cells was 0.6 mg of TCE per mg of cells per day, and they had a finite TCE transformation capacity of 0.036 mg of TCE per mg of cells. Formate addition resulted in increased initial specific TCE transformation rates (2.1 mg/mg of cells per day) and elevated transformation capacity (0.073 mg of TCE per mg of cells). Significant declines in methane conversion rates following exposure to TCE were observed for both resting and formate-fed cells, suggesting toxic effects caused by TCE or its transformation products. TCE transformation and methane consumption rates of resting cells decreased with time much more rapidly when cells were shaken and aerated than when they remained dormant, suggesting that the transformation ability of methanotrophs is best preserved by storage under anoxic conditions.
Microorganisms that biosynthesize broad-specificity oxygenases to initiate metabolism of linear and branched-chain alkanes, nitroalkanes, cyclic ketones, alkenoic acids, and chromenes were surveyed for the ability to biodegrade trichloroethylene (TCE). The results indicated that TCE oxidation is not a common property of broad-specificity microbial oxygenases. Bacteria that contained nitropropane dioxygenase, cyclohexanone monooxygenase, cytochrome P-450 monooxygenases, 4-methoxybenzoate monooxygenase, and hexane monooxygenase did not degrade TCE. However, one new unique class of microorganisms removed TCE from incubation mixtures. Five Mycobacterium strains that were grown on propane as the sole source of carbon and energy degraded TCE. Mycobacterium vaccae JOB5 degraded TCE more rapidly and to a greater extent than the four other propane-oxidizing bacteria. At a starting concentration of 20 microM, it removed up to 99% of the TCE in 24 h. M. vaccae JOB5 also biodegraded 1,1-dichloroethylene, trans-1,2-dichloroethylene, cis-1,2-dichloroethylene, and vinyl chloride.
Trichloroethylene was shown to degrade aerobically to carbon dioxide in an unsaturated soil column exposed to a mixture of natural gas in air (0.6%).
The biodegradation of trichloroethylene (TCE) and toluene, incubated separately and in combination, by indigenous microbial populations was measured in three unsaturated soils incubated under aerobic conditions. Sorption and desorption of TCE (0.1 to 10 micrograms ml-1) and toluene (1.0 to 20 micrograms ml-1) were measured in two soils and followed a reversible linear isotherm. At a concentration of 1 micrograms ml-1, TCE was not degraded in the absence of toluene in any of the soils. In combination, both 1 microgram of TCE ml-1 and 20 micrograms of toluene ml-1 were degraded simultaneously after a lag period of approximately 60 to 80 h, and the period of degradation lasted from 70 to 90 h. Usually 60 to 75% of the initial 1 microgram of TCE ml-1 was degraded, whereas 100% of the toluene disappeared. A second addition of 20 micrograms of toluene ml-1 to a flask with residual TCE resulted in another 10 to 20% removal of the chemical. Initial rates of degradation of toluene and TCE were similar at 32, 25, and 18 degrees C; however, the lag period increased with decreasing temperature. There was little difference in degradation of toluene and TCE at soil moisture contents of 16, 25, and 30%, whereas there was no detectable degradation at 5 and 2.5% moisture. The addition of phenol, but not benzoate, stimulated the degradation of TCE in Rindge and Yolo silt loam soils, methanol and ethylene slightly stimulated TCE degradation in Rindge soil, glucose had no effect in either soil, and dissolved organic carbon extracted from soil strongly sorbed TCE but did not affect its rate of biodegradation.
Trichloroethylene was metabolically activated by toluene dioxygenase to produce toxic effects in Pseudomonas putida F1. Cytotoxicity was indicated by growth inhibition and by the covalent modification of cellular molecules in P. putida F1 exposed to [C]trichloroethylene. With a toluene dioxygenase mutant, neither growth inhibition nor alkylation of intracellular molecules was observed.
Trichloroethylene (TCE), a common groundwater contaminant, is a suspected carcinogen that is highly resistant to aerobic biodegradation. An aerobic, methane-oxidizing bacterium was isolated that degrades TCE in pure culture at concentrations commonly observed in contaminated groundwater. Strain 46-1, a type I methanotrophic bacterium, degraded TCE if grown on methane or methanol, producing CO(2) and water-soluble products. Gas chromatography and C radiotracer techniques were used to determine the rate, methane dependence, and mechanism of TCE biodegradation. TCE biodegradation by strain 46-1 appears to be a cometabolic process that occurs when the organism is actively metabolizing a suitable growth substrate such as methane or methanol. It is proposed that TCE biodegradation by methanotrophs occurs by formation of TCE epoxide, which breaks down spontaneously in water to form dichloroacetic and glyoxylic acids and one-carbon products.
The effects of carbon deprivation on survival of methanotrophic bacteria were compared in cultures incubated in the presence and absence of oxygen in the starvation medium. Survival and recovery of the examined methanotrophs were generally highest for cultures starved under anoxic conditions as indicated by poststarvation measurements of methane oxidation, tetrazolium salt reduction, plate counts, and protein synthesis. Methylosinus trichosporium OB3b survived up to 6 weeks of carbon deprivation under anoxic conditions while maintaining a physiological state that allowed relatively rapid (hours) methane oxidation after substrate addition. A small fraction of cells starved under oxic and anoxic conditions (4 and 10%, respectively) survived more than 10 weeks but required several days for recovery on plates and in liquid medium. A non-spore-forming methanotroph, strain WP 12, displayed 36 to 118% of its initial methane oxidation capacity after 5 days of carbon deprivation. Oxidation rates varied with growth history prior to the experiments as well as with starvation conditions. Strain WP 12 starved under anoxic conditions showed up to 90% higher methane oxidation activity and 46% higher protein production after starvation than did cultures starved under oxic conditions. Only minor changes in biomass and morphology were seen for methanotrophic bacteria starved under anoxic conditions. In contrast, starvation under oxic conditions resulted in morphology changes and an initial 28 to 35% loss of cell protein. These data suggest that methanotrophic bacteria can survive carbon deprivation under anoxic conditions by using maintenance energy derived solely from an anaerobic endogenous metabolism. This capability could partly explain a significant potential for methane oxidation in environments not continuously supporting aerobic methanotrophic growth.
Trichloroethylene was shown to degrade aerobically to carbon dioxide in an unsaturated soil column exposed to a mixture of natural gas in air (0.6%).
Studies were conducted using sand-packed column reactors for which variations in piezometric head, substrate concentration, and biomass measured as organic carbon were monitored in space and time. Methanol was used as a growth substrate. Permeability reductions by factors of order 10 -3 were observed. The data show that a limit on permeability reduction exists, having a magnitude of 5 × 10 -4 in the present study. The limit on permeability reduction and the existence of high densities of bacteria in substrate depleted zones are explained with an open pore model. Permeability reduction was observed to correlate well with biomass density for values less than about 0.4 mg/cm 3, and exhibited independence at higher densities. -from Authors
An experimental investigation was conducted to quantify the permeability reduction caused by enhanced biological growth in a porous medium. Studies were conducted using sand-packed column reactors for which variations in piezometric head, substrate concentration, and biomass measured as organic carbon were monitored in space and time. Methanol was used as a growth substrate. Permeability reductions by factors of order 10−3 were observed. The data show that a limit on permeability reduction exists, having a magnitude of 5 × 10−4 in the present study. The limit on permeability reduction and the existence of high densities of bacteria in substrate depleted zones are explained with an open pore model. Permeability reduction was observed to correlate well with biomass density for values less than about 0.4 mg/cm3, and exhibited independence at higher densities.
The enhanced biotransformation was accomplished by stimulating the growth of indigenous methane-oxidizing bacteria (methanotrophs), which transform chlorinated aliphatic compounds by a cometabolic process to stable, nontoxic end products. Experiments were performed in the presence and absence of biostimulation by means of controlled chemical addition, frequent sampling, and quantitative analysis. The degree of biotransformation was assessed using mass balances and comparisons with bromide as a conservative tracer. Biostimulation of the test zone was successfully achieved by injecting methane- and oxygen-containing ground water in alternating pulses under induced gradient conditions. After a few weeks of stimulation, methane concentrations gradually decreased below the detection limit within two meters of travel. Under active biostimulation conditions, 20 to 30% of the trichloroethylene was biotransformed during the first season of testing. -from Authors
The results of the US Environmental Protection Agency, Office of Drinking Water, sampling and analysis of volatile organic compounds (VOCs) in finished water supplies that use groundwater sources are discussed. Concentrations of 29 VOCs in addition to five trihalomethanes and total organic carbon from 945 water supplies were measured. The five most frequently found compounds other than trihalomethanes were trichloroethylene, 1,1,1-trichloroethane, tetrachloroethylene, cis- and/or trans-1,2-dichloroethylene, and 1,1-dichloroethane. Approximately half of the samples were taken from a random list of water systems. The nonrandom samples were taken from systems selected by the states, using groundwater sources that were likely to show VOCs in drinking water. Large systems in the random sample had a significantly higher frequency of occurrence of VOC contamination than small systems and were also more likely to have higher levels of contamination.
Trichloroethylene (TCE) and cis-1,2-dichloroethylene (DCE) cometabolic degradation by a filamentous, phenol-oxidizing enrichment from a surface-water source were investigated in batch tests. No intermediate toxicity effects were evident during TCE or DCE degradation for loadings up to 0.5 mg TCE/mg VSS or 0.26 mg DCE/mg VSS. Phenol addition up to 40 mg/L did not inhibit TCE or DCE degradation. TCE specific degradation rates ranged from 0.28 to 0.51 g TCE/g VSS-d with phenol present, versus an average endogenous rate of 0.18 g TCE/g VSS-d. DCE specific degradation rates ranged from 0.79 to 2.92 g DCE/g VSS-d with phenol present, versus 0.27 to 1.5 g DCE/g VSS-d for endogenous conditions. There was no inhibition of DCE degradation rates at concentrations as high as 83 mg/L. TCE degradation rates declined between 40 and 130 mg/L TCE. Perchloroethylene, 1,1,1-trichloroethane, and chloroform were not degraded.
A pilot study of in-situ aerobic cometabolic degradation of trichloroethylene (TCE) through the injection of phenol and oxygen into a confined aquifer was conducted at the Moffett Field test site together with a related laboratory study. With injected phenol and dissolved oxygen concentrations of 12.5 and 35 mg/L, respectively, first-order TCE removal of 88% was obtained over a concentration range of 62-500 [mu]g/L. With 1000 [mu]g/L TCE, removal was lower (77%), but increased to 90% when the phenol concentration was raised to 25 mg/L. The maximum field transformation yield of 0.062 g of TCE/g of phenol compared favorably with the highest measured resting-cell laboratory yield of 0.11 g of TCE/g of phenol. These results demonstrate high promise for in-situ aerobic cometabolic biodegradation of TCE with phenol-induced enzymes. 30 refs., 6 figs., 2 tabs.
Results are presented that demonstrate the in-situ biotransformation of vinyl chloride (VC), trans-l,2-dichloroethylene (t-DCE), cis-l,2-dichloroethylene (c-DCE), and trichloroethylene (TCE) by an enhanced population of methane-utilizing (methanotrophic) bacteria. Biostimulation was accomplished by introducing dissolved methane and oxygen into a shallow, confined aquifer, to encourage the growth of the native methanotrophic bacteria. Biotransformation of the target compounds ensued immediately after the commencement of methane utilization, and reached steady-state values within three weeks. The approximate extents of transformation achieved in the two meter biostimulated zone were as follows: VC, 95%; t-DCE, 90%; c-DCE, 50%; and TCE, 20%. The biotransformation of VC and t-DCE was observed to be competitively inhibited by methane. Cyclic variations in methane concentration caused by the alternate pulse injection of dissolved methane into the test zone caused oscillations of the aqueous concentrations of VC and t-DCE. When formate and methanol were substituted for methane as alternative electron donors, inhibition ceased (no oscillations), and concentrations were reduced to levels achieved during periods when no methane was present, confirming the inhibition by methane. Higher transformation rates were achieved temporarily, i.e., for several days, through the addition of formate or methanol. When electron donor addition was terminated, the concentration of target compounds rapidly increased, indicating that the transformation promptly ceased. Although these experiments indicated that methane competitively inhibits transformation rates, this competition is a second-order effect: methane as substrate for growth was also required for transformation of VC, t-DCE, c-DCE, and TCE by methanotrophs.
A nonsteady-state model is presented for the in-situ biostimulation of a microbial population in saturated porous media. The model includes basic processes of microbial growth, utilization of electron donor and acceptor, advective transport, dispersion, and sorption in porous media. Model simulations are compared with results from a series of controlled field experiments at the Moffett Naval Air Station, where the growth of an indigenous population of methane-utilizing bacteria (methanotrophs) was stimulated by the controlled addition of dissolved methane and oxygen (DO) into a semiconfined aquifer. Simulations provide good matches to the observed transient uptake of methane and DO, demonstrating that the observed response resulted from the growth of methanotrophs in the test zone. Simulations duplicate results from alternate pulsed addition of methane (electron donor) and oxygen (electron acceptor), used as a means for distributing microbial growth throughout the test zone. The model permits estimation of changes in microbial population distribution at various stages of the two-year experiment. Temporal changes in model-fitted kinetic parameters indicate that the microbial population evolved to one that more effectively utilized the methane at lower concentrations. These analyses demonstrate that a relatively simple model, which includes basic microbial and transport processes, can be of use in the design and evaluation of in-situ biotreatment processes. The model user, however, must provide judgment in the selection of appropriate input parameters, as well as being aware of model limitations.
A nonsteady‐state model is presented and used in simulating the results of field experiments where chlorinated ethenes were cometabolically degraded by methanotrophic bacteria. In cometabolism, contaminants are degraded fortuitously by microbes growing on a primary substrate, which in this case is methane. The model includes microbial processes of microbial growth, utilization of the electron donor (methane) and the electron acceptor (oxygen), and the cometabolic transformation of the chlorinated ethenes coupled with the transport processes of advection, dispersion, and sorption onto the aquifer solids.
Model simulations of the chlorinated ethenes biotransformation resulting from the biostimulation of methanotrophic bacteria agreed well with the field observations. A kinetic model for the cometabolic transformation that included the competitive inhibition by methane was required to match the field observations. The simulations duplicated the cyclic oscillations in concentration of the chlorinated ethenes that resulted from the pulse feeding of methane. Rate‐limited sorption and desorption were also required in order to match the extended tailing in the concentration response that was observed along with the pulsed oscillations in the concentration due to competitive inhibition.
The kinetic parameters for the cometabolic transformation, derived from model fits to the field observations, differed for the different chlorinated ethenes tested. Vinyl chloride (VC) and trans‐dichloroethylene (t‐DCE) had the highest biotransformation rate coefficients (k/Ks), which were in the range of methane itself, the primary substrate for methanotrophic growth. Cis‐dichloroethylene (c‐DCE) and trichloroethylene (TCE) were degraded at rates one and two orders of magnitude lower than methane, respectively. The transformation rate for TCE was consistent with laboratory‐derived rates from microbes enriched and isolated from the test zone and grown under conditions that most represented those in the field. The kinetic model also permitted the activation and the deactivation of microbial mass towards the cometabolic transformation, which was based on cell growth and decay, and first‐order deactivation.
Careful site characterization and implementation of quantitative monitoring methods are prerequisites for a convincing evaluation of enhanced biostimulation for aquifer restoration. This paper describes the characterization of a site at Moffett Naval Air Station, Mountain View, California, and the implementation of a data acquisition system suitable for real-time monitoring of subsequent aquifer restoration experiments. A shallow, confined aquifer was chosen for the enhanced biodegradation demonstration, and was shown to have suitable hydraulic and geochemical characteristics. Injection and extraction wells were installed at a distance of 6 m, with intermediate monitoring wells at distances of 1, 2.2, and 4 meters from the injection well. Bromide tracer tests revealed travel times of S to 27 hours from the injection well to the various monitoring wells, and 20 to 42 hours from the injection well to the extraction well. Complete breakthrough of the tracer at the monitoring wells was facilitated by choosing a line of wells aligned with the regional flow, and selecting injection and extraction flow rates of approximately 1.5 and 10 liters/min. Transport studies were conducted with selected halogenated organic compounds. The retardation factors were found to range from approximately 2 to 12. The breakthrough responses for the more strongly sorbing compounds, e.g. TCE, exhibited pronounced tailing, such that a minimum period of several weeks was required to achieve complete saturation of the aquifer. The microcomputer-driven sampling, analysis and data management system provided automated data acquisition at sample intervals of 40 minutes, with coefficients of variation smaller than 20%, and allowed for real-time surveillance of the dynamic responses. Overall, the conditions were favorable for a quantitative evaluation of in-situ aquifer restoration by enhanced biodegradation.
The Moffett field site was used for further evaluation of in situ biotransformation of chlorinated aliphatic hydrocarbons with phenol and toluene as primary substrates. Within the 4 m test zone, representing a groundwater travel time of less than 2 days, removal efficiencies for 250 mu g/L TCE and 125 mu g/L cis-1,2-dichloroethylene were greater than 90%, and that of 125 mu g/L trans-1,2-dichloroethylene was similar to 74%, when either 9 mg/L toluene or 12.5 mg/L phenol was used. Phenol and toluene were removed to below 1 mu g/L. Vinyl chloride removals greater than 90% were also noted. However, only 50% of the 65 mu g/L 1,1-dichloroethylene was transformed with phenol addition, and significant product toxicity was evident as concomitant TCE transformation was here reduced to similar to 50%. Hydrogen peroxide addition performed as well as pure oxygen addition to serve as a required electron acceptor.
For a mixed methanotrophic culture grown under copper deficiency, the relative transformation capacities of the chloroethenes with 10 mM formate present was in the order from highest to lowest: 1,2-trensdichloroethylene (t-OCE); 1,2-cis-dichloroethylene (c-DCE); vinyl chloride (VC); trichloroethylene (TCE); and 1,1-dichloroethylene (1,1-DCE). Respective values were 4.8, 3.6, 2.3, 0.85, and 0.13 μmol transformed per mg of cells. Chloroethenes with asymmetric chlorine distributions had lower transformation capacities, probably due to higher transformation product toxicity. While 1,1-DCE itself was not toxic at the concentrations evaluated (up to 1 mg/L), its transformation products were highly toxic. Aquifer microcosms transformed up to 4.8 mg/L VC with no apparent toxic effects, but when both VC and 1,1- DCE were present, about 75% less transformation of VC and a marked decrease in methane oxidation rate resulted because of 1,1-DCE transformation product toxicity. About 25 times more VC was transformed in the soil microcosms per unit of methane consumed than in aqueous batch tests.
The soluble, three-protein component methane monooxygenase purified from Methylosinus trichosporium OB3b is capable of oxidizing chlorinated, fluorinated, and brominated alkenes, including the widely distributed ground-water contaminant trichloroethylene (TCE). The oxidation rates for the chloroalkenes were observed to be comparable to that for methane, the natural substrate, and up to 7000-fold higher than those reported for other well-defined biological systems. The competitive inhibitor tetrachloroethylene was found to be the only chlorinated ethylene not turned over. However, this appears to be due to steric effects rather than electronic effects or the lack of an abstractable proton because chlorotrifluoroethylene was efficiently oxidized. As evidenced by the formation of diagnostic adducts with 4-(p-nitrobenzyl)pyridine, the halogenated alkenes were oxidized predominantly by epoxidation. Stable acidic products resulting from subsequent hydrolysis were identified as the major products. However, additional aldehydic products resulting from intramolecular halide or hydride migration were observed in 3-10% yield during the oxidation of TCE, vinylidene chloride, trifluorethylene, and tribromoethylene. Product analysis of the hydrolysis reaction of authentic TCE epoxide showed little or no 2,2,2-trichloroacetaldehyde (chloral) formation, indicating that atomic migration occurred prior to product dissociation from the enzyme. The occurrence of atomic migration products shows that an intermediate in the substrate to product conversion carries significant cationic character. Such a species could be generated through interaction with a highly electron-deficient activated oxygen in the active site.(ABSTRACT TRUNCATED AT 250 WORDS)
Intact cells of Pseudomonas cepacia G4 completely degraded trichloroethylene (TCE) following growth with phenol. Degradation kinetics were determined for both phenol, used to induce requisite enzymes, and TCE, the target substrate. Apparent Ks and Vmax values for degradation of phenol by cells were 8.5 microM and 466 nmol/min per mg of protein, respectively. At phenol concentrations greater than 50 microM, phenol degradation was inhibited, yielding an apparent second-order inhibitory value, KSI, of 0.45 mM as modeled by the Haldane expression. A partition coefficient for TCE was determined to be 0.40 +/- 0.02, [TCEair]/[TCEwater], consistent with Henry's law. To eliminate experimental problems associated with TCE volatility and partitioning, a no-headspace bottle assay was developed, allowing for direct and accurate determinations of aqueous TCE concentration. By this assay procedure, apparent Ks and Vmax values determined for TCE degradation by intact cells were 3 microM and 8 nmol/min per mg of protein, respectively. Following a transient lag period, P. cepacia G4 degraded TCE at concentrations of at least 300 microM with no apparent retardation in rate. Consistent with Ks values determined for degradation, TCE significantly inhibited phenol degradation.
Chlorinated ethenes are toxic substances which are widely distributed groundwater contaminants and are persistent in the subsurface environment. Reports on the biodegradation of these compounds under anaerobic conditions which might occur naturally in groundwater show that these substances degrade very slowly, if at all. Previous attempts to degrade chlorinated ethenes aerobically have produced conflicting results. A mixed culture containing methane-utilizing bacteria was obtained by methane enrichment of a sediment sample. Biodegradation experiments carried out in sealed culture bottles with radioactively labeled trichloroethylene (TCE) showed that approximately half of the radioactive carbon had been converted to 14CO2 and bacterial biomass. In addition to TCE, vinyl chloride and vinylidene chloride could be degraded to products which are not volatile chlorinated substances and are therefore likely to be further degraded to CO2. Two other chlorinated ethenes, cis and trans-1,2-dichloroethylene, were shown to degrade to chlorinated products, which appeared to degrade further. A sixth chlorinated ethene, tetrachloroethylene, was not degraded by the methane-utilizing culture under these conditions. The biodegradation of TCE was inhibited by acetylene, a specific inhibitor of methane oxidation by methanotrophs. This observation supported the hypothesis that a methanotroph is responsible for the observed biodegradations.
Trichloroethylene (TCE) is also known as 1,1–2-trichloroethylene, 1, 1-dichloro-2-chloroethylene, acetylene trichloride, and ethylene trichloride (Mycroft and Fan 1985). It has the following chemical and physical properties.
Trichloroethylene (TCE) was metabolized by the natural microflora of three different environmental water samples when stimulated by the addition of either toluene or phenol. Two different strains of Pseudomonas putida that degrade toluene by a pathway containing a toluene dioxygenase also metabolized TCE. A mutant of one of these strains lacking an active toluene dioxygenase could not degrade TCE, but spontaneous revertants for toluene degradation also regained TCE-degradative ability. The results implicate toluene dioxygenase in TCE metabolism.
A number of soil and water samples were screened for the biological capacity to metabolize trichloroethylene. One water sample was found to contain this capacity, and a gram-negative, rod-shaped bacterium which appeared to be responsible for the metabolic activity was isolated from this sample. The isolate degraded trichloroethylene to CO(2) and unidentified, nonvolatile products. Oxygen and water from the original site of isolation were required for degradation.
Design and optimization of in situ bioremediation relying on cometabolic degradation
- M M Lang