Project no.

(GOCE) 003985 Acronym: EuroDemo Project title: European Platform for Demonstration of Efficient Soil and Groundwater Remediation

Instrument: Coordination Action Thematic Priority: Global change and ecosystems

D6-2 Status report on technological reliability for demonstrated soil and groundwater management technologies with special focus on the situation in Europe (update on the bioremediation part only)
Due date of deliverable: November 2005 Actual submission date: November 2005, revision February 2007

Start date of project: 01.01.2005

Duration: 3 years

Organisation name of lead contractor for this deliverable: Universität Lüneburg

Revision [draft, 1, 2, …]
Project co-funded by the European Commission within the Sixth Framework Programme (2002-2006)
PU PP RE CO Dissemination Level Public Restricted to other programme participants (including the Commission Services) Restricted to a group specified by the consortium (including the Commission Services) Confidential, only for members of the consortium (including the Commission Services) PU

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1 Enhanced In Situ Bioremediation Technologies (EISB)
1.1 Introduction
Bioremediation relies on microorganisms to biologically degrade groundwater contaminants through a process called biodegradation. It may be engineered and accomplished in two general ways: (1) stimulating native microorganisms by adding nutrients, oxygen, or other electron acceptors (a process called biostimulation); or (2) providing supplementary pregrown microorganisms to the contaminated site to augment naturally occurring microorganisms (a process called bioaugmentation) (AFCEE, 2004; ITRC, 2002; U.S.EPA, 2000a; U.S.EPA, 2004b). This technology mainly focuses on remediating organic chemicals such as fuels and chlorinated solvents. One approach, aerobic bioremediation, involves the delivery of oxygen (and potentially other nutrients) to the aquifer to help native microorganisms reproduce and degrade the contaminant (U.S.EPA, 2004b). Another approach, anaerobic bioremediation, circulates electron donor materials—for example, food-grade carbohydrates such as edible oils, molasses or lactic acid whey—in the absence of oxygen throughout the contaminated zone to stimulate anaerobic microorganisms to consume the contaminant (AFCEE, 2004; DoD, 2002). In some cases, pregrown microbes may be injected into the contaminated area to help supplement existing microorganisms and enhance the degradation of the contaminant, a process known as bioaugmentation. A potential advantage of bioremediation is its ability to treat the contaminated groundwater in place with naturally occurring microorganisms, rather than bringing contaminants to the surface. By using native microorganisms, rather than injecting additional ones, cleanup can be more cost-effective at some sites. However, heterogeneous subsurfaces can make delivering nutrient/oxygen solutions to the contaminated zone difficult by trapping or affecting movement of both contaminants and groundwater. Also, nutrients for stimulating the microorganisms can be consumed rapidly near the injection well, thereby limiting the microorganisms’ contact with the contaminants, or stimulating biological growth at the injection site. In summary, this technology avoids the costs associated with bringing water to the surface for treatment; instead, the main costs associated with bioremediation include: delivery of the amendments to the subsurface (which varies depending on the depth of contamination), the cost of the amendments themselves, and monitoring of the treatment. This report focuses on the approach of enhanced in situ anaerobic bioremediation of chlorinated solvents, because chlorinated solvents are the most common groundwater contaminants and therefore numerous laboratory studies as well as pilot and field applications have been or currently are being conducted. To date, enhanced anaerobic bioremediation has been applied at over 600 sites (AFCEE, 2004).

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1.2 Enhanced In Situ Anaerobic Bioremediation of chlorinated solvents
Enhanced in situ anaerobic bioremediation can be an effective method of degrading various chlorinated solvents dissolved in groundwater, including chloroethenes, chloroethanes, and chloromethanes. Collectively, these compounds (some of which are degradation products of chlorinated solvents) are referred to as chlorinated aliphatic hydrocarbons (CAHs) (AFCEE, 2004; Cope and Hughes, 2001; ITRC, 2004; U.S.EPA, 2000a). Bioremediation of CAHs can occur through natural mechanism (natural attenuation or intrinsic bioremediation or by enhancing the natural mechanisms (enhanced bioremediation). The addition of an organic substrate to an aquifer has the potential to further stimulate microbial growth and development, creating an anaerobic environment in which rates of anaerobic degradation of CAHs may be enhanced. Therefore, a variety of organic substrates have been applied to the subsurface to promote anaerobic degradation of CAHs to innocuous end products. In some cases, microorganisms also may be added (bioaugmentation), but only if the natural microbial population is incapable of performing the required transformations. The most common chlorinated solvents released to the environment include tetrachloroethene (PCE, or perchloroethene), trichloroethene (TCE), trichloroethane (TCA), and carbon tetrachloride (CT). These chlorinated solvents are problematic because of their health hazards and their resistance to natural degradation processes. Because these compounds exist in an oxidized state, they are generally not susceptible to aerobic oxidation processes (with the possible exception of cometabolism). However, oxidized compounds are susceptible to reduction under anaerobic conditions by either biotic (biological) or abiotic (chemical) processes. Enhanced anaerobic bioremediation is intended to exploit primarily biotic anaerobic processes to degrade CAHs in groundwater (AFCEE, 2004; Suthersan, 2001). Other common groundwater contaminants that are subject to reduction reactions are also susceptible to enhanced anaerobic bioremediation. While not addressed in this report, constituents that can also potentially be treated with this approach include the following: • • • • • Chlorobenzenes; Chlorinated pesticides (e.g., chlordane), polychlorinated biphenyls (PCBs), and chlorinated cyclic hydrocarbons (e.g., pentachlorophenol); Oxidizers such as perchlorate and chlorate; Explosive and ordnance compounds; Dissolved metals (e.g., hexavalent chromium); and

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Nitrate and sulphate

Detailed informations for the design and implementation of enhanced anaerobic bioremediation systems for theses groundwater contaminants listed above are described e.g. by (ITRC, 2002; Liles et al., 2004; Mikszewski, 2004; Suthersan, 2001).

1.2.1 Degradation Mechanisms
There are several potential reactions that may degrade CAHs in the subsurface, but not all CAHs are amenable to degradation by each of these processes (Table 1). For example, PCE is not amenable to any process of aerobic degradation, while TCE may only be degraded by aerobic cometabolism that typically requires addition of a substrate in the presence of oxygen (McCarty et al., 1998). However, anaerobic biodegradation processes may potentially degrade not only PCE and TCE, but all of the common chloroethenes, chloroethanes, and chloromethanes. Altogether, the degradation pathways and microbiology of aerobic and anaerobic dechlorination of chlorinated ethenes are better studied than for chloroethanes and chloromethanes (AFCEE, 2004; U.S.EPA, 2000a). This is primarily because they occur more frequently as contaminants in groundwater. Anaerobic reductive dechlorination is the degradation process targeted by enhanced anaerobic bioremediation. Through addition of organic substrates to the subsurface, enhanced anaerobic bioremediation converts naturally aerobic or mildly anoxic aquifer zones to anaerobic and microbiologically diverse reactive zones, making them conducive to anaerobic degradation of CAHs. Biodegradation of an organic substrate depletes the aquifer of dissolved oxygen (DO) and other terminal electron acceptors (e.g., nitrate or sulfate), and lowers the oxidationreduction potential (ORP) of groundwater, thereby stimulating conditions conducive to anaerobic degradation processes. After DO is consumed, anaerobic microorganisms typically use native electron acceptors (as available) in the following order of preference: nitrate, manganese and ferric iron oxyhydroxides, sulfate, and finally carbon dioxide. Figure 1 illustrates a CAH plume where substrate has been injected into the source area. An anaerobic treatment area is created with the development of progressively more anaerobic zones closer to the source of organic carbon as electron acceptors are depleted. Anaerobic dechlorination has been demonstrated under nitrate, iron, and sulfate reducing conditions, but the most rapid biodegradation rates, affecting the widest range of CAHs, occur under methanogenic conditions (Bouwer, 1994).

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Table 1: Potential Degradation Prozesses for CAHs (AFCEE, 2004).
Chloroethenes Degradation Process Aerobic Oxidation Aerobic Cometabolism Anaerobic Oxidation Direct Anaerobic Reductive Dechlorination Cometabolic Anaerobic Reduction Abiotic Transformation PCE N N N Y Y Y TCE N Y N Y Y Y DCE P Y P Y Y Y VC Y Y Y Y Y Y PCA N P N Y P Y Chloroethanes TCA N Y N Y Y Y DCA Y Y Y Y Y Y CA Y Y P Y P Y CT N N N Y Y Y Chloromethanes CF N Y N Y Y Y MC Y Y Y Y Y Y CM P Y P Y P Y

PCE = tetrachloroethene, TCE = trichloroethene, DCE = dichloroethene, VC = vinyl chloride, PCA = tetrachloroethane, TCA = trichloroethane, DCA = dichloroethane, CA = chloroethane, CT = carbon tetrachloride, CF = chloroform, MC = methylene chloride, CM = chloromethane. N = Not documented in the literature. Y = Documented in the literature. P = Potential for reaction to occur but not well documented in the literature.

Figure 1: Reducing zone established downgradient of substrate Injection (AFCEE, 2004).

Under anaerobic conditions, reductive dechlorination mechanisms can effectively biodegrade CAHs. Reductive dechlorination generally involves the sequential replacement of a chlorine atom on a CAH with a hydrogen atom (that is, converting PCE to TCE to DCE, and so on) and has been observed to occur both directly and cometabolically. In anaerobic reductive dechlorination (direct), the mediating bacteria use the CAH directly as an electron acceptor in energy-producing redox reactions. Anaerobic reductive dechlorination (cometabolic) occurs when bacteria incidentally dechlorinate a CAH in

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the process of using another electron acceptor to generate energy. The three general reactions that may degrade CAHs by anaerobic reductive dechlorination are Direct Anaerobic Reductive Dechlorination, Cometabolic Anaerobic Reductive Dechlorination and Abiotic Reductive Dechlorination. More detailed information about the different pathways by which microorganisms transform contaminants is given by (AFCEE, 2004; DoD, 2002; U.S.EPA, 2000a).

1.3 APPLICATION OF ENHANCED ANAEROBIC BIOREMEDIATION
Application of enhanced anaerobic bioremediation starts with a review of site-specific conditions and evaluation of remedial objectives to determine if this remedial approach is appropriate for a site. Once enhanced bioremediation is selected as a remedial alternative, design criteria for implementation are developed including selection of a substrate and system configuration (AFCEE, 2004). The following subsections describe some common technology screening criteria, substrate alternatives, and system configurations used for enhanced anaerobic bioremediation.

1.3.1 Technology Screening
The addition of an organic substrate to the subsurface to stimulate and enhance the anaerobic dechlorination process in situ has been explored at many sites (AFCEE, 2004). Enhanced anaerobic bioremediation has been applied under a broad range of site conditions, including the following: Hydrogeologic Settings. Enhanced anaerobic bioremediation has been applied in a variety of hydrogeologic settings, from low permeability silts and clays to high permeability alluvial sand and gravel deposits to fractured bedrock. Enhanced bioremediation has been applied at depths up to 400 feet below ground surface (bgs) and with groundwater velocities ranging from a few feet per year to several feet per day. However, there are limits to applying the technology in settings with the extremes of very high and very low rates of groundwater flow. It may be impractical to maintain reducing conditions in high flow settings, due to the magnitude of groundwater and native electron acceptor flux. On the other hand, it may be difficult to inject substrates into tight formations, and under low flow settings mixing of substrate with groundwater due to advection and dispersion may be limited (AFCEE, 2004). Contaminant Levels and Distribution. The technology has typically been applied to groundwater plumes with concentrations of CAHs ranging from 0.01 to 100 mg/L. Sites

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with indications of residual or sorbed DNAPL (dissolved CAH concentrations in excess of 100 mg/L) also have been successfully treated. However, it may not be realistic to expect rapid remediation of source areas with DNAPL pools (AFCEE, 2004). Geochemical Conditions. During anaerobic dechlorination, CAHs function as electron acceptors in competition with naturally occurring (inorganic) electron acceptors. For example, a high rate of groundwater flow coupled with high concentrations of DO may create an oxygen electron acceptor demand that cannot practically be overcome with substrate addition. In some cases, adverse site conditions can be mitigated with proper system design. For example, recirculation systems may be used to impose a hydraulic gradient and enhance groundwater flow at sites with very low natural hydraulic gradients. However, when pumping of significant quantities of groundwater is required, the technology may not be cost competitive with pump and treat; this becomes a sitespecific issue. Once enhanced bioremediation has been selected as an appropriate technology, there are several substrate alternatives and system configurations to consider (AFCEE, 2004).

1.3.2 Substrate
There are many organic substrates which can be naturally degraded and fermented in the subsurface that result in the generation of hydrogen. Examples of easily fermentable organic substrates include alcohols, low-molecular-weight fatty acids (e.g., lactate), carbohydrates (e.g., sugars), vegetable oils, and plant debris (e.g., mulch). The substrates most commonly added for enhanced anaerobic bioremediation include lactate, molasses, Hydrogen Releasing Compound (HRC®), and vegetable oils. Substrates used less frequently include ethanol, methanol, benzoate, butyrate, high-fructose corn syrup (HFCS), whey, bark mulch and compost, chitin, and gaseous hydrogen. Table 2 summarizes the attributes of several substrate types. These substrates are classified here as soluble substrates, viscous fluids and low viscosity fluids, solid substrates, and experimental substrates. The physical nature of the substrate dictates the frequency of addition, the addition technique, and potential system configurations.

Table 2: Substrates used for enhanced anaerobic bioremediation (AFCEE, 2004).

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1.3.3 System configuration
Enhanced in situ anaerobic bioremediation can be implemented to provide source area or dissolved plume treatment or containment, or a combination of source area and dissolved plume remediation can be used. Enhanced bioremediation and conventional source treatment or containment approaches (e.g., chemical oxidation or groundwater extraction) will be subject to the same difficulties associated with mass transfer limitations of a continuing source and preferential flow paths in heterogeneous formations (AFCEE, 2004). The single largest difference between conventional remedial technolo-

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gies and enhanced bioremediation may be that enhanced bioremediation, if properly implemented, can maintain effectiveness over a longer period of time at a lower overall cost. This may make enhanced bioremediation an effective remedial approach due to the substantial challenges associated with significant CAH source mass removal. Typical system configurations and associated remedial action objectives that engineered anaerobic bioremediation may be used to address include the following: Source Zone Treatment: Remediation strate/contaminant contact is possible.
•Plume

of

source

zones

where

good

sub-

Containment using a Biologically Reactive Barrier: Reduction of mass flux from a source zone or across a specified boundary.
•Plume-Wide

Restoration: Total treatment of an entire dissolved plume.

In some cases, several approaches may be combined. For example, a source area may be targeted for remediation using a grid configuration, combined with a linear barrier configuration upgradient from a downgradient point of compliance (Figure 2).

Figure 2: Schematic of Source Area and Biobarrier Injection Configurations (AFCEE, 2004).

The appropriate application of enhanced anaerobic bioremediation will be site-specific and based on a strategy that takes into account final remedial objectives, feasibility of the application, and regulatory issues. Ultimately however, there will be an economic limit to the size of a plume that can be treated with a complete plume-wide application of

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enhanced bioremediation. For plume sizes greater than 10 to 20 acres, use of containment strategies combined with other remedial approaches may be more feasible. Source Zone Treatment Enhanced anaerobic bioremediation has been used to address source zones either to limit mass flux from the source zone or to accelerate source mass removal. Mass flux reduction is achieved by stimulating biodegradation in the dissolved phase, while reducing contaminant massis available to migrate downgradient. Source mass removal is achieved by accelerating DNAPL dissolution and then stimulating biodegradation of the dissolved contaminants. Anaerobic dechlorination is a process that takes place in the aqueous phase and does not directly attack DNAPL mass. Therefore, enhanced bioremediation may be limited in its ability to rapidly treat DNAPL source zone areas (AFCEE, 2004; DoD, 2002). On the other hand, treatment to reduce mass flux and to perhaps increase the rate of dissolution and treatment as compared to natural attenuation or groundwater extraction may be more achievable. Enhanced bioremediation of DNAPL sources is being researched and may someday be a proven and feasible long-term remedial alternative (AFCEE, 2004). Alternatively, injection of a low solubility, persistent carbon source such as vegetable oil into a source area may serve to reduce mass flux and to effectively sequester the source due to partitioning and lowering of hydraulic conductivity. However, while degradation of dissolved constituents may be stimulated, this may not accelerate destruction of DNAPL or sorbed source mass. Plume Containment using Biologically Enhanced Barrier Systems For large plumes having poorly defined, widely distributed, or inaccessible source areas, enhanced bioremediation systems may be configured as permeable reactive barriers (biobarriers) to intercept and treat a contaminant plume. For example, biobarriers may be employed at a property boundary or upgradient from a point of regulatory compliance to prevent plume migration to potential receptors. Biobarriers typically consist of either rows of substrate injection wells or a solid-substrate trench located perpendicular to the direction of groundwater flow. Passive biobarriers typically use slow-release, long-lasting substrates (e.g., HRC®, vegetable oils, or mulch) that can be either injected or otherwise placed in a trench, and that are designed to remain in place for long periods to maintain the reaction zone. Contaminant mass is delivered to the treatment zone via natural groundwater flow. Capital and operating costs for a passive biobarrier configuration are typically lower than for plume-wide configurations because of a limited treatment area. However, life-cycle costs could be significant if the source of the CAHs upgradient of the biobarrier is not addressed. Semi-passive or active biobarriers are

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similar to passive biobarriers except that a soluble substrate is typically injected periodically (semi-passive) or via a recirculation system (active). Soluble substrates migrate with groundwater flow, are depleted more rapidly, and require frequent addition. However, these systems offer the advantage of being able to adjust the rate or type of substrate loading over time, and soluble substrates may be easier to distribute throughout larger volumes of the contaminant plume. Recirculation can improve substrate distribution, contaminant/substrate mixing, and retention time for treatment; but the overall groundwater flux downgradient of the system does not change (AFCEE, 2004). Plume-Wide Restoration Enhanced bioremediation systems may be configured to treat dissolved CAHs across an entire contaminant plume. Creating an anaerobic reaction zone across broad areas of a plume is an aggressive approach that may reduce the overall timeframe for remediation. Plume-wide delivery systems will typically be configured as a large injection grid, or a recirculation well field may be employed to increase the effective area of substrate distribution. Higher initial capital and operating costs of recirculation systems may be offset by shorter remedial timeframes with lower monitoring and total long-term operating costs. However, plume-wide applications where substrate is delivered to the entire plume may be cost prohibitive for very large plumes or cost inefficient for low-level contaminant plumes. At sites where larger plumes are present (greater than several acres), or the depth of the plume makes installing injection wells difficult and expensive, multiple treatment lines can be established perpendicular to the direction of groundwater flow, typically separated by 6 to 12 months of groundwater travel time. A recirculation approach may not be practical or cost effective at a large scale due to the large volumes of groundwater to be processed and ineffective in situ mixing in heterogeneous environments. There is some controversy as to the cost effectiveness of using enhanced anaerobic bioremediation for plume-wide restoration. For any kind of recirculating system, groundwater pumping rates may have to be similar to pump and treat methods; the cost of enhanced bioremediation must be carefully compared to pump and treat. If substrate addition is done by some kind of multiple point injection relying on natural groundwater flow for dispersion, this may require very close spacing of injection points and or it may not result in good mixing of substrate and CAHs in situ (AFCEE, 2004).

1.3.4 Delivery Options Common substrate delivery options include direct injection or recirculation of fluid substrates, or emplacement of solid substrates in biowall trenches. Where direct-push methods can be used, substrate may be injected directly through the probe rods. This is

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a common approach for both slow-release and soluble substrates. Otherwise, injection wells are used. Soluble substrates may be injected in batch mode, or in the case of frequent injections, the use of automatic injection systems may be warranted. Recirculation systems may also be employed for distribution of soluble substrates. Figure 3 is an example of a horizontal circulation system. Recirculation may be continuous or in a pulsed mode. Substrates are added to the groundwater as it is reinjected into the treatment zone. Recirculation systems may be effective for difficult hydrogeological conditions. For example, recirculation may be used to effectively mix substrate and contaminated groundwater at sites with very low hydraulic gradients and low rates of groundwater flow (AFCEE, 2004).

Figure 3: Schematic of a horizontal recirculation system (AFCEE, 2004).

1.4 Case Studies
Dry Cleaning Facility, Arlington, Texas
TECHNOLOGY: Hydrogen Release Compounds (HRC®) SCALE: Full GEOLOGY: Clayey silt underlain by medium to dark grey shale CONTAMINANTS: PCE, TCE, cDCE, VC

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The site is located in Arlington, Texas, covering approximately seven acres. A former dry cleaner was situated in a small suite and operated at the site between 1982 and 1992. Environmental site assessments were conducted in 1996 and the site was admitted into the TCEQ (Texas Commission on Environmental Quality) Voluntary Cleanup Program. An unknown amount of chlorinated solvents was released at the site and had contaminated soil and groundwater. The concentrations of contaminants of concern (COC) in subsurface soils at a depth of approximately eight feet below the building were in excess of site target concentrations. Site investigations revealed that the groundwater with COC concentrations above the site target levels occupied approximately 3,500 square feet directly beneath and downgradient of the source area (U.S.EPA, 2004). Immediately prior to injection of HRC®, the groundwater concentrations of COCs were 4,500 µg/L for PCE, 1,000 µg/L for TCE, 7,300 µg/L for cis-1,2-DCE, and 870 µg/L for VC. The underlying rock at the site is predominantly medium to dark grey shale, which readily weathers to a thick, clayey soil. Site geology consists of a very low permeability light to dark brown, soft, moist clay from the ground surface to approximately 22 feet bgs. groundwater is encountered at 7 feet bgs. Table 3 shows the target cleanup concentrations at the site, established by Conceptual Environmental Assessment Model (CEAM), a health-based risk assessment for groundwater. The cleanup approach included soil excavation from the source area, performed in 1998, followed by in situ bioremediation using HRC® in 2002. In May 2000, HRC® was injected into 45 borings that are located within the shallow aquifer area to a depth of approximately 22 feet bgs. Of the 45 injection borings, 29 borings were advanced perpendicular to the ground surface, and 16 borings were advanced at angles of 15 to 30 degrees from vertical to extend beneath the building’s foundation. A total of approximately 7,000 pounds of HRC® were injected. Quarterly sampling of groundwater was conducted as a part of the response action.

Table 3: Cleanup criteria Contaminant PCE TCE cDCE VC CEAM Cleanup Criteria for Groundwater [µg/L] 500 500 7,000 200

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Approximately 18 months after HRC injection, PCE, TCE, cDCE, and VC in the monitoring well located nearest to the contaminant source area had decreased to 408 µg/L, 87.4 µg/L, 438 µg/L, and 132 µg/L, respectively. Based on these results, the site received a Conditional Certificate of Completion from the TCEQ Voluntary Cleanup Program on October 26, 2001.

INEEL, Test Area North, Idaho Falls, Idaho
TECHNOLOGY: Sodium lactate SCALE: Full GEOLOGY: Deep-fractured basalt aquifer CONTAMINANTS: PCE, TCE, radionuclides

The historical injection of liquid wastes and concentrated sludges into the Snake River Plain Aquifer using Well TSF-05 has resulted in a TCE, PCE, and tritium plume in groundwater at the TAN facility of the Idaho National Engineering and Environmental Laboratory (INEEL). The depth to water at TAN is approximately 200 ft. The aquifer and most of the unsaturated zone are comprised primarily of layered basalt flows, intercalated with sedimentary interbeds deposited during periods of volcanic quiescence. Groundwater flow in the aquifer is controlled by the highly transmissive zones that occur at the contacts between individual basalt flows, and to a lesser extent, by fractured zones within flow interiors. The scale of the basalt geology dictates that preferential flow can be very important at spatial scales less than approximately 100 m (330 ft), after which a transition to continuum behaviour occurs and the aquifer can be thought of essentially as a macro-porous medium (ITRC, 2004). Figure 4 shows the TCE plume at the TAN facility. A very large, low-concentration fringe surrounds and emanates from a much smaller, high concentration core. Within the core is a very small residual source area that continues to contaminate fresh groundwater flowing through from up-gradient. The transition from the scale of the residual source, where preferential flow is significant, to the scale of the fringe, where sufficient vertical communication has been present along the flow path to create a relatively well-mixed, predictable groundwater plume. The residual source of contamination in the aquifer near TSF-05 is believed to be composed primarily of the sludge that was injected into the well over 15–20 years. The pore water of the sludge probably contains large amounts of TCE, with PCE and tritium also present in significant amounts. Given the organic content of the sludge, sorbed PCE and TCE are also likely to be present. TCE concentrations as high as 300,000 µg/L have been measured in groundwater from well TSF-05. Some of the sludge has been

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shown to have TCE concentrations as high as 3%. The sludge therefore represents a long-term source of contamination to the aquifer.

Figure 4: Medial zone of TCE plume at TAN (TCE-concentrations in µg/L).

The 1995 Record of Decision (ROD) identifies the primary Remedial Action Objectives (RAO) for TAN, which is to restore the contaminated aquifer groundwater by 2095 (100 years from the date of the ROD) by reducing all contaminants of concern to below MCLs and a 1 × 10-4 total cumulative carcinogenic risk-based level for future residential groundwater use and for noncarcinogens, until the cumulative hazard index is less than 1. The ROD selected pump and treat as the default remedy for the residual DNAPL source area, but because this approach was unlikely to restore the site in a meaningful time frame, it also identified five alternative technologies to be evaluated for their potential to enhance or replace the default remedy. One of the technologies identified was enhanced in situ bioremediation (ISB). Field activities for enhanced ISB began in November 1998 with the shutdown of the interim pump and -treat facility that was operating in the hotspot and the initiation of the first phase of ISB field operations. The primary performance objective of the field operation was to determine whether anaerobic reductive dechlorination of TCE in the residual DNAPL source area could be enhanced through the addition of an electron donor (sodium lactate). Initially, baseline sampling and a conservative tracer test were performed,

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followed by initiation of weekly lactate injections into the original disposal well and biweekly groundwater monitoring for the parameter sets described above. Seven months after sodium lactate injections began, significant dechlorination activity was stimulated to distances of >40 m from the injection well. For example, TCE concentrations in well TSF-05 decreased to <10 µg/L, with increases in ethene of up to 2,500 µg/L. Dechlorination was strongly correlated with elevated electron donor concentrations and the establishment of strongly reducing conditions. In particular, dechlorination of TCE to cis-DCE coincided with sulphate reduction, while dechlorination of cis-DCE to vinyl chloride and ethene was observed only in the presence of methanogenesis. Electron donor concentrations [as chemical oxygen demand (COD)] of >4,000 mg/L and >5,800 mg/L were observed in wells TAN-25 and TAN-26 (refer to Figure 4 for well locations). By the end of the field evaluation, methane concentrations in these wells were 20,000 µg/L and 17,000 µg/L, while TCE concentrations had dropped to 68 µg/L and nondetect. A significant outcome of the field evaluation work was the strong evidence indicating that the injected sodium lactate solution significantly enhanced the bioavailability of TCE in the residual DNAPL source area, thereby accelerating degradation of the source. This effect was evidenced by the near 21-fold increase in TCE concentrations that corresponded to arrival of lactate in well TAN-26, followed by complete dechlorination of this newly bioavailable TCE to ethane. ISB has been demonstrated to be a much more effective technology for treatment of residual source areas than previously believed. Figure 5 shows the pre-lactate TCE concentrations in the residual source area, and the TCE concentrations after 21 months of lactate injections. Based on these substantial concentration reductions within and immediately downgradient of the DNAPL residual source area, coupled with the evidence for accelerated degradation of the residual DNAPL source, an amended ROD was signed by the State of Idaho and EPA Region 10 to replace pump and treat with bioremediation for restoration of the TCE source area at TAN. Operations conducted since the conclusion of the field evaluation have been focused on optimization (the current phase of operations) and long-term implementation of ISB as defined by the objectives described below.

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Figure 5: TCE concentrations before lactate injection (a) and TCE concentrations after 21 months of lactate injection (ITRC, 2004).

Two stages of operations remain for the TAN ISB hotspot remedy-optimization activities and long-term operations. The primary performance objective for ISB optimization activities, which is the current phase of ISB operations, is to distribute electron donor throughout the entire residual source area to eliminate VOC migration beyond the ISB treatment cell. The performance metric for this objective, as it was for the ISB field evaluation, is changes in groundwater concentration. Once down- and cross-gradient migration has been eliminated, then the ISB remedy will move into its final phase, longterm operations. The primary performance objective of ISB long-term operations is to maintain the sufficiently large biologically active zone for a period of time such that the residual source material is degraded to a point where natural attenuation processes will address the residual levels of groundwater contamination. The performance objective for ISB optimization activities is to distribute electron donor throughout the entire residual DNAPL source area to completely eliminate contaminant migration to down-gradient wells. During the field evaluation, a relatively constant supply of lactate was provided through weekly injections. During initial optimization activities, however, the presence of propionate and acetate combined with the absence of lactate resulted in increased dechlorination efficiency as evidenced by conversion of all accumulated cis-DCE to ethene throughout the residual source area. Therefore, the primary operational change during optimization activities was to inject larger pulses of lactate less frequently, approximately every six to eight weeks, to increase periods when propionate was the primary electron donor. This has resulted in significant progress towards meeting the performance objective.

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Although substantial progress toward the performance objective for optimization activities has been made, the objective has not yet been achieved. To achieve this objective, a second injection well, was incorporated into ISB operations in December 2003. Although the ISB remedy has not yet moved into long-term operations, progress toward this objective can still be measured by monitoring chloroethene and ethene groundwater concentrations in ISB wells. During optimization activities, the same pattern of enhanced TCE dissolution in response to sodium lactate injections, followed by complete dechlorination of the newly bioavailable TCE to ethene that was observed in the field evaluation still persisted. During optimization activities, TCE concentrations in well TSF05 have routinely increased from nondetect to 500–1000 µg/L in response to lactate injections, but then have been subsequently reduced below detection by 4–5 weeks after injections. Overall, significant progress towards the performance objective for both optimization activities and long-term operations has been made at TAN. Additional optimization activities are needed to distribute electron donor throughout the entire residual DNAPL source area to completely eliminate flux to down-gradient wells, evaluate alternative electron donors that may decrease long-term costs, and study microbial competition to determine whether the system can be optimized to favour the organisms of interest. Lessons Learned Enhanced in situ bioremediation can be very effective even in complex source areas, toxicity of chlorinated solvents to bacterial communities, once thought to be a problem for ISB of DNAPLS, may not be an issue. In fact, dechlorinating bacteria may have an ecological niche in these high concentration areas, reductive dechlorination is strongly correlated both to electron donor distribution and redox conditions, and, dechlorination efficiency can be controlled in the field by the electron donor addition strategy and type.

NASA’s Launch Complex 34 (LC34) at Cape Canaveral
TECHNOLOGY: Biostimulation and Bioaugmentation through ethanol and a consortium of naturally occurring microorganism (KB-1TM) SCALE: Demonstration GEOLOGY: Surficial fill and silty sands CONTAMINANTS: TCE, DCE

The purpose of the project was to evaluate the technical and cost performance of the

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biostimulation and bioaugmentation technology when applied to dense, nonaqueousphase liquid (DNAPL) contaminants in the saturated zone. This demonstration was conducted at Launch Complex 34, Cape Canaveral Air Force Station, Florida, where chlorinated volatile organic compounds (CVOCs), mainly trichloroethylene (TCE), are present in the subsurface as DNAPL. Smaller amounts of cis-1,2-dichloroethylene (DCE) and vinyl chloride (VC) also are present as a result of the natural degradation of TCE. The part of the source zone used as a test plot for the demonstration is entirely underneath the Engineering Support Building (Figure 6).

Figure 6: Location map of Launch Complex 34 site (Battelle, 2004).

The biostimulation and bioaugmentation project was conducted under the National Aeronautics and Space Administration (NASA) Small Business Technology Transfer Research (STTR) Program. For this project, the Small Business Concern vendor was GeoSyntec Consultants (GeoSyntec). This demonstration was independently evaluated by Battelle under the United States Environmental Protection Agency’s (U.S. EPA’s) Superfund Innovative Technology Evaluation (SITE) Program (Battelle, 2004). A sequential process of biostimulation and bioaugmentation is a promising remediation technology for enhancing the extent and rate of degradation of CVOCs. Biostimulation involves stimulating indigenous microbial cultures by adding nutrients (i.e., biostimulation), whereas bioaugmentation involves introducing microbial cultures that are particularly adept at degrading these contaminants into the target aquifer. The premise is that

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although many aquifers contain native microorganisms that can degrade CVOCs, the native microorganisms can be supplemented by specific cultures that enhance the degradation of chlorinated solvents. Natural microorganisms, such as Dehalococcoides ethenogenes, can be separately cultured and introduced into the aquifer to enhance the degradation rates and extent of degradation that would normally be achievable by natural attenuation or by biostimulation (addition of nutrients) alone. Bioaugmentation using specific cultures is claimed to be particularly effective in (1) degrading byproducts of reductive dehalogenation, such as cis-1,2- DCE and VC, which would otherwise accumulate in the aquifer; and (2) completing dechlorination processes to non-chlorinated products such as acetylene, ethene, ethane, and methane. This demonstration involved biostimulation followed by bioaugmentation in the same test plot. During the biostimulation phase of treatment, an electron donor (ethanol) was added to provide nutrients for indigenous microorganisms and stimulate CVOC degradation. During the bioaugmentation phase, KB-1™, a consortium of naturally occurring microorganisms known to completely dechlorinate high concentrations of TCE to ethene, was added to the test plot. At Launch Complex 34, the DNAPL source zone was not large enough to conduct a control demonstration using biostimulation alone for comparison. Therefore, the sequential treatment of biostimulation and bioaugmentation was evaluated at Launch Complex 34 in the same test plot. Bioaugmentation was chosen as a second treatment phase to determine if complete dechlorination of a TCEDNAPL source zone was possible. Based on pre-demonstration groundwater and soil sampling by Battelle, a test plot was identified for biostimulation and bioaugmentation that was 20 ft long × 20 ft wide × 20 ft deep (saturated thickness). The Upper Sand Unit, where the treatment was targeted, is the shallowest part of the surficial aquifer, and extends down to a depth of 26 ft. The water table at the site occurs at about 5 to 6 ft below ground surface (bgs), thus providing about a 20-ft-thick zone of aquifer for treatment. The Upper Sand Unit is underlain by the Middle Fine-Grained Unit, which is made up of finer sand and silt, and constitutes somewhat of a hydraulic barrier to the Lower Sand Unit below. These three stratigraphic units constitute the surficial aquifer. The Lower Clay Unit forms a thin aquitard under the surficial aquifer. The bioaugmentation treatment was particularly targeted at depths of 16 to 24 ft bgs in the Upper Sand Unit, where most of the DNAPL appeared to be present. The predemonstration soil and groundwater characterization was done in January 2002, before the vendor began installing the treatment system. Prior to beginning the demonstration, the vendor installed a recirculating groundwater system to establish a controlled hydraulic flow field. This was done to facilitate the distribution of electron donor, simplify the placement of monitoring points, and accelerate the degradation process to a point where it could be monitored in the reasonable time-

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frame allotted to this demonstration. The groundwater was recirculated from the extraction wells to the injection wells for several weeks to establish hydraulic control. During this testing and modification period (May 23 to September 12, 2002), the recirculated groundwater was passed through carbon canisters and treated prior to reinjection. CVOCs were removed from groundwater in the treatment plot during this time. Prior to beginning the biostimulation phase of the treatment, the carbon canisters were removed from the recirculating system. The electron donor (ethanol) was injected inside the plot to begin the biostimulation phase of the demonstration (October 23, 2002). Approximately 14 weeks later (February 6, 2003), the KB-1™ culture was injected in the aquifer to begin the bioaugmentation phase. Groundwater sampling was conducted in December 2002 (one month after electron donor injection) and March 2003 (one month after KB-1™ culture injection).

Figure 7: KB-1™ Dechlorinator Culture Containers (Battelle, 2004).

Post-demonstration soil and groundwater characterization was done in June 2003. Performance assessment activities for the biostimulation and bioaugmentation demonstration included pre-demonstration investigations, installation of wells, operation, monitoring, and post-treatment evaluation. Battelle conducted detailed soil and groundwater characterization activities to establish the DNAPL distribution and mass inside the test cell. The vendor conducted additional operational measurements. The objectives of the performance assessment were to:

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Determine changes in total TCE (dissolved and free-phase) and DNAPL mass in the test plot due to the biostimulation and bioaugmentation treatment; Determine changes in aquifer quality due to the treatment; Determine the fate of TCE, the primary DNAPL contaminant; and, Determine operating requirements and cost of the technology. Changes in Total TCE and DNAPL Mass Detailed pre-demonstration and post-demonstration soil sampling was the main tool for estimating changes in total TCE and DNAPL mass in the plot due to the treatment technology. In general, the eastern portion of the plot had the highest predemonstration TCE concentrations. TCE concentrations were higher at approximatelyi 26 ft bgs, which is at the interface between the Upper Sand Unit and Middle Fine- Grained Unit. The rest of the plot appeared to contain mostly dissolved-phase TCE. The soil sampling results were evaluated using both linear interpolation and kriging to obtain mass estimates for the entire treatment zone (i.e., Upper Sand Unit). Linear interpolation indicated that, under pre-demonstration conditions, 25.5 kg of total TCE (dissolved and free phase) was present in the Upper Sand Unit. Approximately 2.6 kg of the total TCE was estimated to be DNAPL. Following the demonstration, soil sampling indicated that 0.4 kg of total TCE remained in the Upper Sand Unit; the post-demonstration mass of TCE-DNAPL was estimated as 0.0 kg because no post-demonstration TCE concentrations were observed above the threshold of 300 mg/kg. Therefore, the overall decrease in TCE mass due to the treatment, as indicated by linear interpolation, was 98.5% for total TCE and >99% for DNAPL in the Upper Sand Unit. Kriging of the soil data indicated that the total TCE mass in the target zone before the biostimulation and bioaugmentation treatment ranged from 17.6 to 46.6 kg, with an average of 32.1 kg. After treatment, the total TCE mass in the plot ranged from 0.1 to 0.3 kg, with an average of 0.2 kg. The decline in TCE mass due to the biostimulation and bioaugmentation treatment ranged from 98.6 to 99.7%, with an estimated average decline of 99%. Because few data points were available for DNAPL estimation, only the total TCE data were subjected to kriging. These estimated TCE mass ranges are based on an 80% confidence level and incorporate the uncertainty and spatial variability in the data. The linear interpolation estimates are within the range of the kriging estimates. These results indicate that the biostimulation and bioaugmentation treatment caused a significant decrease in total TCE and DNAPL mass in the target treatment zone. Changes in Aquifer Quality Dissolved TCE concentrations, as measured in the monitoring wells, declined substantially in the Upper Sand Unit of the demonstration area following the bioaugmentation treatment. DCE levels increased following biostimulation, and then decreased after bio-

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augmentation. Vinyl chloride levels increased immediately after biostimulation and bioaugmentation, and then decreased during subsequent post-demonstration monitoring. Ethene concentrations increased substantially toward the end of the demonstration. These changes indicate sequential degradation of TCE to DCE, and ultimately to vinyl chloride and ethane during the demonstration. In order to verify that the DNAPL source had been substantially reduced and that the CVOC reductions observed during the demonstration could be sustained (without encountering rebound), one further round of groundwater monitoring was conducted in January 2004, almost one year after injection of the KB-1™ culture. This long-term monitoring showed further substantial reductions in TCE (to below detection), cis-1,2- DCE, and vinyl chloride. These results show that DNAPL mass was substantially removed by the treatment and that the reduced CVOC levels were sustainable. Oxidation-reduction potential (ORP) and dissolved oxygen (DO) levels decreased in the demonstration area after biostimulation began. The decreases continued through the bioaugmentation phase of the demonstration and post-demonstration sampling. These data indicate that strongly reducing anaerobic conditions were created in the Upper Sand Unit during the demonstration. Groundwater pH in the shallow wells remained relatively steady. Dissolved iron concentrations in the center well of the test plot generally decreased after the bioaugmentation treatment. The secondary drinking water limit for iron is 0.3 mg/L, which was exceeded in the majority of wells before, during, and after the demonstration. Chloride levels in the monitoring wells, which were already high partly due to saltwater intrusion in the aquifer, showed a slight increase over the course of the demonstration. Anaerobic reductive dechlorination of TCE, cis- 1,2-DCE, and VC, which was observed in this demonstration, releases chloride from contaminant molecules and leads to increases in chloride levels in groundwater. Increases in dissolved methane, as well as decreases in sulfate concentrations, indicate that an increase in biological activity occurred as a result of the biostimulation and bioaugmentation treatment. Biological oxygen demand (BOD) levels in the groundwater increased, indicating an increase in the bioavailable organic matter in the aquifer, most likely due to the addition of a carbon electron donor to the recirculating groundwater. Total organic carbon (TOC) levels also increased, probably as a result of the carbon electron donor addition. The hydraulic conductivity of the Upper Sand Unit does not appear to have been affected by the treatment, suggesting that the addition of electron donor and KB-1™ culture did not noticeably affect the aquifer. There were no substantial changes in permeability in the test plot according to slug tests conducted in the center well before and after the demonstration.

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Fate of TCE/DNAPL in the Aquifer The performance assessment indicates that biodegradation was a substantial pathway accounting for the decrease in TCE, cis-1,2-DCE, and vinyl chloride measured in the test plot. An increasing trend in dissolved ethene and chloride levels is evidence of dechlorination reactions in the aquifer. The combination of biostimulation and bioaugmentation treatments accounted for the enhanced biodegradation seen in the plot. In addition, some TCE and other VOCs were likely extracted by the recirculation system and captured by adsorption in the aboveground carbon canisters. However, an analysis of the amounts of water and TCE potentially extracted from the test plot by the recirculation system showed that biostimulation and bioaugmentation contributed substantially to the TCE removal observed in the test plot, even after adjusting for any dilution due to the water recirculation system and carbon. Operating Requirements and Cost In general, the treatment system operated smoothly through the recirculation, biostimulation, and bioaugmentation phases. Relatively good hydraulic control appeared to have been maintained in the test plot, and the electron donor and KB-1™ culture were welldistributed in the target zone. The vendor reported that biofouling in the injection wells became apparent after amending the recirculating groundwater with electron donor. To mitigate the biofouling, the duration of ethanol was decreased to one concentrated dose administered daily; the injection wells were scrubbed, surged, and purged on a weekly basis to removed biofilm from the screen; and the reinjected groundwater was amended with sodium hypochlorite to inhibit microbial growth in the injection wells. It is unclear what the long-term effect of the change in electron donor dose/timing and the addition of sodium hypochlorite into the aquifer had on the microorganisms throughout the demonstration plot. Future applications of the biostimulation and bioaugmentation technology may benefit from a study of optimizing electron donor dosing schedules, and establishing procedures to monitor for biofouling and treat occurrences of biofouling. A present value (PV) analysis was conducted to compare the cost of DNAPL source treatment with biostimulation and bioaugmentation to the cost of installing and operating an equivalent pump-and-treat system for a long period of time (30 years). It was assumed that the biostimulation and bioaugmentation treatment would reduce the DNAPL presence in the aquifer sufficiently for the rest of the contamination to attenuate naturally. This analysis showed that the cost of source treatment with biostimulation and bioaugmentation was lower than the PV of the costs of long-term treatment with a pump-and-treat system at this site.

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1.5 Status and Applicability
Enhanced in situ anaerobic bioremediation has emerged in recent years as a remediation strategy for CAHs in groundwater. Advantages include complete mineralization of the contaminants in situ with little impact on infrastructure and relatively low cost compared to more active engineered remedial systems (e.g., groundwater extraction, permeable reactive iron barriers, or chemical oxidation). There are many considerations to take into account when selecting and designing an enhanced bioremediation system. Enhanced anaerobic bioremediation as a remediation technology may not be appropriate at all sites due to the complexity of chlorinated solvent contaminant plumes (e.g., DNAPL source areas) and potential site-specific limitations (e.g., difficult hydrogeologic conditions). At some sites, it may have utility only when coupled with other remedial technologies. Enhanced anaerobic bioremediation may be appropriate at sites where: • Site-specific data indicate that the contaminants present (including any toxic degradation products) can be readily degraded by native microbial populations under anaerobic conditions. Subsurface conditions (e.g., aquifer permeability) are conducive to adequate emplacement and distribution of a substrate, and creation of an in situ reactive zone conducive to anaerobic degradation of the targeted contaminants. A cost/benefit analysis indicates that the technology is cost-effective relative to other remedial measures (e.g., monitored natural attenuation [MNA], air sparging, groundwater extraction, permeable iron reactive barriers, or chemical oxidation).

Conditions that may preclude the use of enhanced anaerobic bioremediation are: • • • • Sites with impacted receptors, or with short travel time or distance to potential discharge and/or exposure points Sites with inaccessible DNAPL sources Difficult hydrogeologic conditions that may preclude cost-effective delivery of amendments, such as low permeability or a high degree of aquifer heterogeneity Geochemical conditions (e.g., unusually low or high pH) that inhibit the growth and development of dechlorinating bacteria

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Contaminant Distribution Enhanced anaerobic bioremediation takes advantage of natural processes that may already be contributing to the degradation of CAHs. The presence of degradation products that indicate that anaerobic dechlorination of CAHs is occurring, or has occurred, naturally is a favourable indicator. Conversely, the lack of any dechlorination products is a “red flag” that either enhanced bioremediation may not be a suitable approach or that further evaluation is required (AFCEE, 2004). The release of CAHs is often associated with release of other potential electron donors such as fuels or landfill leachate. A review of historical records may indicate that anaerobic dechlorination occurred in the past, but that the system has stalled (e.g., at cisDCE) once the initial electron donor supply was depleted. In this case, complete and rapid degradation can often be restored by substrate addition. Enhanced anaerobic bioremediation has been successfully applied to a few sites with residual or sorbed DNAPL. Application to sites with large quantities of free-phase DNAPL has yet to be proven effective, and in these instances enhanced anaerobic bioremediation may be more suitable to reduce source mass or as a polishing step following application of more aggressive source removal technologies (Stroo et al., 2003). Highly elevated concentrations of solvents may act as toxic inhibitors to biodegradation, especially for sites where the release is relatively recent (e.g., within 1 to 3 years). However, dechlorinating bacteria (at least for chloroethenes) are known to be tolerant of concentrations nearing solubility limits (Yang and McCarty, 2000). Successful site closures to date (involving enhanced anaerobic bioremediation) typically have involved relatively small- to moderate-size plumes associated with small commercial operations such as dry cleaners (AFCEE, 2004). An area-wide treatment using enhanced anaerobic bioremediation may simply not be economical where treatment areas exceed tens to hundreds of acres. Microbiology Enhanced anaerobic bioremediation of CAHs is targeted at stimulating microbially mediated anaerobic reductive dechlorination. The success of the technology largely depends on the presence of appropriate dechlorinating bacteria and the ability to stimulate sufficient growth and activity to degrade contaminants to the extent (and at a rate) that meets the intended remedial objectives. Incomplete dechlorination (e.g., cis-DCE or VC stall) due to insufficiently reducing conditions or lack of appropriate dechlorinating populations are common microbial issues when applying enhanced anaerobic bioremediation (AFCEE, 2004). Initially, a site can fall into one of three microbiological categories:

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1. Sites where appropriate dechlorinating microorganisms are present, geochemical conditions are appropriate for their growth, and sequential dechlorination products (e.g., VC and ethene) are observed. 2. Sites where appropriate dechlorinating microorganisms are present, but at insufficient quantity or level of activity for complete sequential dechlorination to innocuous end products. 3. Sites where appropriate dechlorinating microorganisms are completely absent (rare). In the first case listed above, biostimulation alone can be applied with a high degree of confidence. In the second case listed above, biostimulation alone may or may not be successful. It may be difficult to distinguish the second case from the third case, because detection and identification of appropriate microbial species in these systems is problematic (AFCEE, 2004). Hydrogeology The uncertainty in characterizing subsurface hydrogeology complicates all in situ treatment technologies, and must be considered during the site selection and design process. Inadequate characterization of the site hydrogeology can lead to remedial system failure. However, in many cases, the system can be designed to mitigate difficult hydrogeologic conditions. Difficult hydrogeologic conditions that may preclude cost-effective delivery of amendments include excessive groundwater flow velocity, low permeability, high levels of aquifer heterogeneity, or excessive depth to groundwater (i.e., high drilling costs) (AFCEE, 2004). Depth to Groundwater. Depth to water and the vertical thickness of the plume primarily impact the capital cost of drilling and delivering the substrate to the intended treatment zone. The capital expense of installing multiple injection wells in deep settings (e.g., greater than 100 feet bgs), or across thick formations needs to be compared to the costs associated with competing technologies. There are practical limits (perhaps 15 to 20 feet) to the maximum length of well screen across which a substrate can be uniformly injected; therefore, large saturated thicknesses may require multiple vertical injection points (AFCEE, 2004). Hydraulic Conductivity. Hydraulic conductivity is a primary factor in effective distribution of substrate in the subsurface. In general, hydraulic conductivities greater than 1 foot per day (ft/day), or approximately 3 x 10-4 centimeters per second (cm/sec), are suitable for injection of dissolved substrates. It is generally infeasible to effectively distribute substrates in zones having a hydraulic conductivity less than 0.01 ft/day (3 x 10-6

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cm/sec). Alternate injection techniques such as hydraulic fracturing may be used in some cases, but the timeframe for remediation may still be many years as remediation of the entire aquifer volume will likely be diffusion-limited (AFCEE, 2004). Groundwater Flow. Groundwater velocity, flow direction, and horizontal and vertical gradients will impact the effectiveness of substrate addition. Most applications rely to some extent on advective groundwater flow or recirculation to distribute substrate uniformly throughout the intended treatment zone. Excessively high rates of groundwater flow (greater than 5 to 10 ft/day) may require large amounts of substrate to overcome a large influx of native electron acceptors migrating into the reactive zone (AFCEE, 2004). It may be impractical to maintain sufficiently reducing conditions in high-flow aquifers. Cross-gradient distribution of soluble substrates in high-flow regimes also may be limited by lower transverse dispersion. Where rates of groundwater flow are very low (less than 10 to 30 feet per year [ft/yr]), closer injection well spacing will be required and the timeframe for remediation may be extended due to reduced mixing of substrate and contaminant mass. Groundwater Geochemistry Redox processes in natural systems are rarely in equilibrium, and the predominant electron acceptor being utilized by microbial populations to derive energy often varies in zones across the site. Addition of an organic substrate is intended to consume native electron acceptors and to maintain optimal conditions for high rates of anaerobic dechlorination. Excessive levels of competing electron acceptors (e.g., DO, bioavailable iron, and sulfate) may limit the effectiveness of substrate addition. Groundwater geochemical characteristics across the site should be reviewed to identify any undesirable conditions (AFCEE, 2004). Dissolved Oxygen and Oxidation-Reduction Potential. Background levels of DO and values of ORP are an indicator of the pre-injection redox conditions that must be lowered to achieve efficient dechlorination. In general, elevated levels of DO and nitrate in most aquifer systems can be overcome by providing adequate organic substrate. However, the problem may be compounded by other factors such as high rates of groundwater flow (AFCEE, 2004). Bioavailable Iron. High levels of bioavailable ferric iron (as iron oxide or iron hydroxide minerals) may inhibit microbial anaerobic dechlorination in a manner similar to other competing electron acceptors. In particular, it has been theorized that the free energy associated with electron transfer during reduction of bioavailable iron by iron-reducing bacteria is greater than that associated with the reduction of cis-DCE. Therefore, an-

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aerobic dechlorination of cis-DCE to VC may potentially be inhibited in the presence of relatively high levels of bioavailable iron because iron-reduction is more energetically favorable (Evans and Koenigsberg, 2001; Wilson et al., 2003). This may be a temporal phenomenon until the bioavailable iron is depleted; the concentrations or levels of bioavailable iron that may inhibit anaerobic dechlorination have not been well documented or defined. Because bioavailable iron cannot be determined from groundwater sampling alone, this parameter is frequently underestimated. Sulfate/Sulfides. Existing guidance documents tend to suggest that, while CAH dechlorination under sulfate reducing conditions is feasible, high sulfate levels are problematic for CAH bioremediation. High sulfate levels may lower the efficiency at which substrate is used for anaerobic dechlorination (AFCEE, 2004). However, there is ample evidence in the literature for dechlorination of a variety of CAHs at sites containing elevated dissolved sulfate levels (ITRC, 1998; Devlin and Muller, 1999). ARCADIS (Suthersan et al., 2002) reports successful application of enhanced anaerobic bioremediation at sites containing up to 500 to 700 mg/L of sulfate. Complete anaerobic dechlorination has been stimulated at several high-sulfate Air Force sites including Altus AFB, Oklahoma (sulfate up to 2,600 mg/L) and Travis AFB, California (sulfate up to 5,400 mg/L). Therefore, the presence of high sulfate concentrations does not necessarily preclude effective application of this technology (AFCEE, 2004). Excessive levels of sulfides produced by reduction of sulfate may potentially inhibit anaerobic dechlorination. Elevated levels of dissolved sulfides or hydrogen sulfide have been shown to inhibit sulfate reducing bacteria and methanogens, as well as some fermentation reactions that produce hydrogen. The levels of sulphide that may potentially inhibit dechlorinating microorganisms (and whether these levels are commonly encountered in the field) are not well documented. In general, dissolved sulphide and hydrogen sulfide are rapidly co-precipitated with ferrous iron (a byproduct of ferric iron reduction), but this may not be sufficient to reduce sulfide levels at high sulfate/low iron sites, where there is insufficient iron to react with the sulfides (AFCEE, 2004). pH and Alkalinity. A pH close to neutral (i.e., 6 to 8) is the most conductive to the proliferation of healthy, diverse microbial populations. Low pH conditions (<5) are detrimental to sulfate-reducing, methanogenic, and dechlorinating bacteria. Fermentative organisms favour lower pH conditions, and therefore will out-compete sulfate-reducing and methanogenic bacteria in more acidic environments; this can result in the formation of undesirable byproducts of fermentation, such as ketones, alcohols, and aldehydes. In such cases, pH buffering, typically using common basic salts such as sodium bicarbonate, may be used during implementation to raise and/or neutralize pH against further

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decreases (AFCEE, 2004). Sites with pH outside of the 5 to 9 range may require more thorough biological screening (e.g., using microcosm studies) to evaluate the effect of pH manipulation on the existing dechlorinating microbial populations. In practice, care must be taken in evaluating site-specific behaviour. For example, if groundwater pH is below 5 but complete dechlorination is observed in the field, then it may be clear that the local microbial population has adapted to low pH conditions. Aquifer systems with lower buffering capacities are more susceptible to decreases in pH. Alkalinity is a general indicator of the buffering capacity of an aquifer system. However, because of the importance of the aquifer solids in establishing buffering capacity, groundwater alkalinity may underestimate the true buffering capacity. From a practical standpoint, alkalinities greater than 300 mg/L are generally sufficient to buffer against adverse pH changes. Alkalinity less than 100 mg/L is cause for concern, and pH should be monitored carefully. Lowering of pH and problems with adequate buffering are more likely to occur where organic acids (e.g., lactic acid), organic acid salts (e.g., sodium lactate), or soluble sugars (e.g., HFCS or molasses) are used. Substrate selection, substrate loading rate, and the addition of buffering reagents should be carefully evaluated at sites with low alkalinity or in response to field observations of excessive drop in pH (AFCEE, 2004). The Air Force Center for Environmental Excellence and the Naval Facilities Engineering Service Center (AFCEE, 2004) has reviewed 17 case studies to evaluate common attributes of either successful or unsuccessful applications of enhanced anaerobic bioremediation. Site conditions and system design were evaluated to identify factors that contributed to a successful application of enhanced bioremediation, or that limited the effectiveness of the application. The case studies reviewed include the application of different substrate types and bioaugmentation. Overall six field applications of soluble substrates were reviewed, including applications of lactate and molasses, and a site with an acetate/fructose combination. Eight applications of HRC® and vegetable oil were reviewed to evaluate the performance of slow-release (viscous fluid) substrates. The solid substrate case studies reviewed include two applications of bark mulch biowalls for remediation of chlorinated ethenes and furthermore the study included two bioaugmentation cases studies. Table 4 (attached) contains a summary of these sites. In the following the main statements of this evaluation are summarized.

Soluble Substrate Applications The soluble substrates used in the preceding cases studies were readily injected and distributed in the subsurface, and may be particularly effective in difficult hydrogeologic settings. The applications at Test Area North and Hanscom AFB used only a single well

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to distribute substrate throughout the treatment zone. Recirculation systems have the potential to impact even larger treatment zones, which may be necessary at sites with complex hydrogeology or low rates of groundwater flow. The primary disadvantage of using these substrates is the requirement for frequent injection and resulting higher operations and maintenance (O&M) costs relative to slow release substrates. The cost of O&M alone is a large percentage of the life-cycle costs of soluble substrate systems. Factors that resulted in incomplete dechlorination observed in some cases was either due to 1) an inadequate supply (low concentration) of substrate, resulting in insufficient reducing conditions, or 2) a lack of microorganisms capable of complete anaerobic dechlorination. Limiting the substrate loading rate in order to avoid methanogenesis may result in large areas of the treatment zone not becoming sufficiently reducing. Although soluble substrates require frequent injection, the substrate concentration and volume can be modified to increase the substrate loading rate, as needed. However, this also may result in extra cost and time to monitor and adjust the substrate volume, concentration, and frequency of injection until an optimal scenario is obtained. Furthermore, difficult hydrogeologic conditions are the cause of failure at some sites, including a high flux of aerobic groundwater, unexpected groundwater flow direction, and inadequate characterization of complex stratigraphy. For example, the demonstration project conducted at Hanscom AFB required modifications to the injection scenario over a period of approximately 1 year to fully deplete native electron acceptors and to induce methanogenic conditions under which degradation of DCE and VC was observed. Temporal fluctuations in the direction of groundwater flow also complicated substrate distribution at this site. The substrate limitation was overcome by adjustments to the system, and system performance objectives were achieved when sufficient substrate loading was accomplished. Advantages of soluble substrate systems include the following: Soluble substrates are readily mixed and distributed in the subsurface relative to other substrate types. This may be beneficial for deep treatment zones where direct-push techniques cannot be used. In this case, the high cost of well installation may be offset by the use of soluble substrates that require fewer injection wells for distribution of substrate over larger volumes of the aquifer. The ability to vary the concentration, volume, and frequency of injection allows for optimizing substrate delivery and manipulating the geochemical conditions in the reaction zone over time. Recirculation systems may be used for hydraulic containment and/or for a greater residence time of contaminants in the reaction zone. Recirculation systems also may treat larger aquifer volumes with fewer wells.

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Disadvantages or limitations of soluble substrate systems include the following: O&M costs are high relative to long-lasting substrates as a result of the need for frequent injection. Biofouling of injection wells may require additional maintenance. The use of potable water to make up large volumes of substrate mixture may result in dilution and displacement of the contaminant plume. Low-molecular-weight substrates (e.g., lactate or butyrate) can be more expensive than bulk food-grade products, while the presence of impurities may require the use of higher grades of molasses or the use of high fructose corn syrup. Slow –release viscous fluid substrate systems HRC® Applications HRC® has been demonstrated to be an effective substrate for enhanced anaerobic bioremediation, with over 474 field applications since 1999 (Willett et al., 2004). Complete degradation of PCE and TCE parent compounds to VC and ethene was observed at both the Fisherville Mill Site and the Springdale Cleaners Site. Although the ability to reach federal MCLs has yet to be demonstrated at these sites, significant contaminant reductions were achieved. The results obtained for the Springdale Cleaners Site suggests the potential for remediating DNAPL source areas using HRC® products. Conversely, application of HRC® at the Atlas 10 Site in Nebraska ((USACE), 2003) did not stimulate significant reduction of TCE, largely due to a high flux of native electron acceptors (DO, nitrate, and sulfate). Limited substrate distribution and a consequent inability to induce highly reducing conditions was observed at the Naval Air Station Dallas Site (CH2M Hill Constructors, 2001) due to low aquifer permeability and lack of groundwater flow. These limitations are not unique to the application of HRC®; rather, they can be problematic with enhanced anaerobic bioremediation applications in general. Vegetable Oil Applications Vegetable oil is currently being developed as a low-cost alternative substrate, designed to induce or enhance anaerobic dechlorination of CAHs for several years with a single injection. Vegetable oil applications to date have mostly been performed to achieve source reduction using grid configurations, or to construct biobarriers using rows of injection points. Applications at Altus AFB, Travis AFB, and Cape Canaveral Air Force Station have demonstrated that concentrations of chlorinated ethenes can be reduced by several orders of magnitude to below federal drinking water MCLs. Applications of vegetable oil at Travis AFB and Cape Canaveral Air Station indicate anaerobic dechlorination can be stimulated for periods of at least 3 to 4 years with a single application. Some difficulty in achieving effective substrate distribution has been encountered, and

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uniform distribution of the substrate may be complicated by low permeability soils or by a high degree of aquifer heterogeneity. Distribution problems may be expected with injection of neat vegetable oil, and generally this is no longer recommended. Lowviscosity microemulsions are currently the state-of-the-art for distribution of vegetable oils, which allows more uniform distribution. Stable emulsions with very fine droplet sizes may be difficult to prepare in the field. Alternately, more expensive commercial emulsion products are available. Advantages of using slow-release viscous fluids include the following: Application of HRC® or vegetable oil emulsions can be very cost-effective for treating shallow groundwater plumes using inexpensive direct-push techniques, where close injection point spacing (5- to 15-foot centers) can be utilized. The use of fast-acting HRC®-primer and long-lasting HRC-XTM products provides design alternatives for varied site conditions. Vegetable oil emulsions are easily modified to fit site-specific conditions. Oil-inwater emulsions can be modified to include fast-acting soluble substrates or to modify the effective oil saturation. The effective lifespan of a single application of these substrates may last from 1 to 5 years. Slow-release substrates also may be used in trenches and excavations to supplement solid substrates (e.g., spraying vegetable oil onto sand or mulch, or backfilling with a layer of HRC®). Disadvantages of using slow-release viscous fluids include the following: Slow-release substrates typically require more injection points and may not be as cost-effective as soluble substrate applications under very deep or difficult hydrogeologic conditions where injection costs are high. High rates of groundwater flow or high rates of native electron acceptor flux may require higher loading rates and additional injections. In some cases (e.g., the Atlas 10 site), the native electron acceptor flux may be too high to overcome. HRC® products are relatively expensive compared to other substrate types. However, a cost-benefit analysis should be conducted that considers other cost factors and the overall life-cycle costs. Creating stable emulsions in the field with an appropriate droplet size may be difficult in practice. Use of pre-mixed commercial microemulsion products will increase cost. Bark Mulch Solid Substrate Systems

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Two applications of bark mulch in a permeable biowall configuration were reviewed to evaluate the performance of solid substrate systems. The Offutt AFB and Altus AFB biowalls were both capable of effectively reducing concentrations of TCE. Reduction of TCE concentrations to below federal drinking water MCLs occurs in the immediate vicinity of the Altus AFB biowall. Both sites exhibited substantial degradation of cis-DCE without an accumulation of VC or ethene. Degradation processes other than biotic anaerobic reductive dechlorination of TCE, cis-DCE, and VC are likely occurring, including abiotic degradation by reactive iron-monosulfides. Humic acids in the mulch and compost mixtures may also serve as electron acceptors in energy yielding reactions that result in the oxidation of cis-DCE and VC under anaerobic conditions (Bradley et al., 1998). Although exhibiting decreasing trends, cis-DCE has persisted at concentrations above initial conditions and above its MCL at both biowall sites. Nonetheless, overall mass destruction rates for total CAHs are impressive, ranging from 64 percent (Offutt AFB pilot-scale biowall) to 86 percent (Altus AFB biowall). Both of the biowalls appear to be effective at treating shallow groundwater plumes in highly heterogeneous formations having a low to moderate permeability. It is yet to be determined whether retention time and substrate loading using mulch biowalls is sufficient for degrading concentrations of CAHs in excess of 10 to 100 mg/L, or for flow rates greater than 1.0 ft/day. The use of wider trenches (greater than 2 feet in width) or multiple parallel trenches may be necessary to treat higher CAH concentrations at sites with high rates of groundwater flow or high rates of native electron acceptor flux. In summary, the advantages and disadvantages of solid substrate mulch and compost biowalls are listed below. Advantages related to the use of mulch and compost biowalls include the following: Effective for shallow groundwater plumes in low to moderate permeability or highly heterogeneous formations. The continuity of the trench eliminates the potential for groundwater bypass due to preferential flow paths, or non-uniform distribution of substrate that may occur with delivery of liquid substrate via injection wells. Mulch, compost, and sand are relatively inexpensive when purchased in bulk quantities. Tree mulch can often be obtained for the cost of shipping and handling alone. Mulch biowalls require no O&M other than periodic performance monitoring. However, it has yet to be determined how many years biowall systems will be able to sustain anaerobic reductive dechlorination of CAHs. Trenches can be modified to include wells or perforated pipe for addition of liquid substrates to supplement carbon loading, if necessary. In addition, the relatively

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small treatment volume of the trench (relative to other substrate configurations) makes biowall systems ideal candidates for relatively low-cost inoculation with bioaugmentation cultures. Disadvantages or limitations of mulch biowalls include the following: The depth that can be trenched in a practical and cost-effective manner is limited to approximately 35 feet bgs. Excavation of a bench for the trenching equipment may provide for an additional 5 to 10 feet of depth. Trenching may interfere with site infrastructure and utilities. The contaminant retention time in the trench and substrate loading capacity (i.e., rate at which organic carbon is added to the groundwater passing through the trench) may be insufficient to treat concentrations of CAHs in excess of 10 to 100 mg/L. Use of wider trenches or multiple parallel trenches may be necessary to treat higher CAH or to deplete high concentrations of native electron acceptors. The effective life-span of mulch biowalls has yet to be determined. Bioaugmentation Systems The bioaugmentation applications at the Aerojet Facility in California and the Bachman Road site in Michigan demonstrate that bioaugmentation can be highly effective with small-scale recirculation systems. It has yet to be shown whether bioaugmentation can be used successfully with large-scale recirculation or passive enhanced bioremediation systems. The ability to increase the scale of bioaugmentation systems will be dependent to a large extent on the ability to uniformly distribute the culture over large volumes of the treatment zone. Most bioaugmentation demonstrations performed to date have used low-molecular-weight soluble substrates (e.g., lactate or ethanol) in recirculation configurations to carefully control aquifer redox conditions. The fermentation reactions for low-molecular weight substrates are generally better understood than for more complex substrates (e.g., molasses or vegetable oils). However, there is no reason to believe that bioaugmentation cannot be effective with all substrate types. Commercially available bioaugmentation cultures consist predominantly of the microbial strain Dehalococciodes ethenogenes. Dehalococciodes ethenogenes is the only microbial strain that has been shown to be capable of complete sequential dechlorination of PCE and TCE to cis-DCE, VC, and ethene. The ability of these bioaugmentation cultures to completely dechlorinate chloroethanes and chloromethanes is less well understood. The advantages and disadvantages of bioaugmentation are listed below. Advantages of bioaugmentation include the following:

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Bioaugmentation can be used to reduce acclimation periods and/or to provide populations of known dechlorinating microorganisms where dechlorination is incomplete. Bioaugmentation may be used to increase the confidence of using an enhanced anaerobic bioremediation approach. In some cases, bioaugmentation may be cost-effective in that the overall time for remediation can be decreased, thereby reducing costs for O&M and performance monitoring. Bioaugmentation in a carefully controlled reaction zone may result in complete anaerobic dechlorination without inducing strongly reducing (i.e., methanogenic) conditions. Avoidance of strongly reducing conditions may not be necessary in most cases, but this capability can be used to advantage where strict adherence to drinking water standards is being enforced. Limitations of bioaugmentation include the following: Enriched cultures may quickly attach to the aquifer matrix and do not migrate as readily as soluble substrates. Therefore, it may be difficult to distribute the cultures throughout large volumes of an aquifer. Although the cost of bioaugmentation cultures is decreasing as more commercial vendors enter the market, the cost of the recirculation systems commonly used to carefully control geochemical conditions is high. The ability to inoculate passive treatment systems in a barrier configuration may represent a more cost-effective use of the technology. The following subsections describe some common limitations and reasons for failure of enhanced anaerobic bioremediation applications that have been encountered in this survey of case studies. Hydrogeology and Substrate Delivery High rates of groundwater flow, low aquifer permeability, and low hydraulic gradient have all resulted in ineffective applications of enhanced anaerobic bioremediation. In general, a high flow rate results in a high native electron acceptor flux and difficulty in establishing a highly reducing environment. For example, a high flux of DO, nitrate, and sulfate at the Atlas 10 Missile site in Nebraska is thought to have resulted in native electron acceptor demand that could not be overcome by two applications of HRC®. High groundwater flow rates may also limit the effective residence time of CAHs in the reactive zone. This may be a critical design factor for systems such as mulch biowalls, where the reaction zone and residence time may be relatively short. Low permeability limited the effectiveness of an application of HRC® at the Naval Air

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Station Dallas site. HRC® and vegetable oil have been applied successfully in low permeability settings at other sites (e.g., the Travis AFB), but it is typical for very long lag times to occur and for distribution of the soluble constituents of these substrates to be diffusion limited (a slow process). A low hydraulic gradient and slow rates of groundwater flow (less than 10 to 30 ft/yr) may similarly inhibit effective distribution of substrate in passive systems. While it is possible that slow migration of soluble substrate via diffusion may occur over a period of several years, for most applications this may not be an acceptable timeframe. The use of recirculation systems that enhance hydraulic gradients and the rate of groundwater flow should be considered for treatment of dissolved plumes in low gradient or low permeability groundwater systems. Geochemistry, Redox Conditions, and Substrate Loading Insufficient substrate loading and high native electron acceptor flux can result in redox conditions that are not sufficiently reducing to stimulate effective rates of anaerobic dechlorination. Although anaerobic dechlorination has been demonstrated to occur under sulfate-reducing conditions without methanogenesis (e.g., methane less than 1 mg/L), the number of sites that exhibit complete anaerobic dechlorination under methanogenic conditions would suggest that an attempt to limit substrate loading, in order to prevent methanogenesis and increase substrate utilization, may be counter productive. The cost of additional substrate necessitated by diminished utilization of CAHs for anaerobic dechlorination is small compared to the cost incurred for longer O&M and monitoring due to slower rates of anaerobic dechlorination. Due to aquifer and microbial heterogeneity, controlling subsurface redox conditions with precision is difficult. Such control typically requires frequent substrate injection and/or recirculation, and possibly also temporal variation in loading rates, both of which are labour and cost intensive. Furthermore, the soluble (aqueous) substrate applications at several sites show that, anaerobic dechlorination of cis-DCE or VC to ethene did not occur until methanogenic conditions were observed. Methanogenic conditions are often an indication that bioavailable iron and sulfate have been adequately depleted, and that high rates of fermentation are occurring. These conditions favour high rates of anaerobic dechlorination, and should not be considered detrimental. At Naval Support Activity Mid-South, production of VC and ethene were only observed after a high concentration slug of acetate was injected at the end of a recirculation test that had utilized a lower substrate loading rate. Furthermore, it has been suggested that rapid stimulation of strongly reducing (i.e., methanogenic) conditions is one way to expedite an evaluation of microbial sufficiency. If a complete dechlorination pathway is not observed after inducing strongly methano-

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genic conditions for several months, then it is likely that the requisite microbial species are not present. In contrast, the Aerojet Facility application illustrated in Appendix E.9 is an example of where bioaugmentation in a carefully controlled reaction zone was successful in stimulating complete anaerobic dechlorination without inducing methanogenic conditions or creating undesirable levels of dissolved metals or other fermentation products. The bioaugmentation culture was capable of complete dechlorination of TCE to ethene without the need to cultivate methanogenic conditions. Microbial Sufficiency Microbial sufficiency is likely the most difficult condition to assess during the site selection process. Multiple conditions may cause an accumulation of intermediate dechlorination products (e.g., cis-DCE, VC, or 1,2-dichloroethane [DCA]). Examples of incomplete or slow dechlorination of cis-DCE or VC include the Point Mugu IRP Site 24and Seal Beach IRP Site 40. No singular analysis, including determining the presence or absence of Dehalococcoides, can be used to confirm the lack of a sufficient microbial population. For example, Dehalococcoides was detected at the Atlas 10 site in a well that had elevated levels of TOC and metabolic acids. Yet the presence of this species, combined with the presence of reducing conditions and elevated organic carbon levels, did not result in significant degradation of TCE or evidence of complete dechlorination. Quantitative, real-time polymerase chain-reaction (PCR) analysis may provide estimates of the quantity or concentration of Dehalococcoides in a sample. However, it is not well known what concentrations are required to effect efficient and complete anaerobic dechlorination. Furthermore, molecular screening methods cannot currently identify the strains of Dehalococcoides that may be present. Isolation of different strains in the laboratory indicates they have different dechlorinating capacities. In addition to the absence of appropriate dechlorinating microorganisms, insufficient substrate loading, failure to achieve sufficient reducing conditions, inhibition due to high levels of native electron acceptors (e.g., sulfate), inhibition due to preferential degradation of more highly-chlorinated compounds in the contaminant mixture, and kinetic disparity all may contribute to accumulation of intermediate dechlorination products. At many sites, a relatively high percentage of the CAH mass may be present in the form of DNAPL or sorbed to the aquifer matrix. Constant dissolution of CAH mass into the groundwater reaction zone may occur. In these cases, cis-DCE and VC may be constantly produced by dechlorination of the parent compounds (PCE and TCE). However, unless the rate of dechlorination of cis-DCE and VC is greater than the rate of dechlorination of PCE and TCE, cis-DCE and VC will accumulate or persist until the mass of PCE and TCE is depleted. Because cis-DCE and

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VC are typically dechlorinated more slowly than PCE and TCE, this rate disparity should be anticipated. Given the current state-of-the-practice, it is important to evaluate all potential causes of the accumulation of intermediate dechlorination products and of incomplete degradation pathways before considering the need for bioaugmentation. While bioaugmentation should not be a default remedy for poorly designed or poorly implemented biostimulation applications, there are sites that will benefit from its application. Bioaugmentation has been successfully applied at several sites, and is a potential option for sites with an absence, or insufficient quantities or activity, of appropriate dechlorinating microorganisms. Application of enhanced anaerobic bioremediation using bioaugmentation at the Aerojet Facility in California and the Bachman Road site in Michigan benefited from bioaugmentation. It should be noted that these projects were specifically designed to demonstrate bioaugmentation using small-scale recirculation systems. Application of a biostimulation-only test typically uses a much different design and allows for a much longer acclimation period. Nonetheless, bioaugmentation was required to meet the project-specific goals of achieving complete dechlorination within a specified time period at these sites.

1.6 References
AFCEE, N., 2004. Principles and Practices of Enhanced Anaerobic Bioremediation of chlorinated solvents. 022/738863/28.doc, Air Force Center for Environmental Excellence, Brooks City-Base, Texas Naval Facilities Engineering Service Center, Port Hueneme, California Environmental Security Technology Certification Program, Arlington, Virginia. (AFRL), A.F.R.L., 2001. Reductive Anaerobic Biological In-Situ Treatment Technology (RABITT), Battelle Memorial Institute, Cornell University, USEPA, NFESC. ARCADIS Geraghty and Miller, I.A., 2003. Final Report: In-situ Substrate Addition to Create Reactive Zones for Treatment of Chlorinated Aliphatic Hydrocarbons, Hanscom Air Force Base., Prepared for the Air Force Center for Environmental Excellence and the Environmental Security Technology Certification Program. Battelle, 2004. Demonstration of Biodegradation of Dense, Nonaqueous-Phase Liquids (DNAPL) through Biostimulation and Bioaugmentation at Launch Complex 34 in Cape Canaveral Air Force Station, Florida, Battelle Press, Columbus, OH. Bouwer, E.J., 1994. Bioremediation of Chlorinated Solvents Using Alternate Electron Acceptors. Publishers. In: R.D. Norris, R.E. Hinchee, R. Brown, P.L McCarty, L. Semprini, J.T. Wilson, D.H. Kampbell, M. Reinhard, E.J. Bouwer, R.C. Borden, T.M. Vogel, J.M. Thomas, and C.H.Ward (Editor), Handbook of Bioremediation. Lewis Publishers, pp. 149-175. Bradley, P.M., Chapelle, F.H. and Lovely, D.R., 1998. Humic Acids as Electron Acceptors for Anaerobic Microbial Oxidation of Vinyl Chloride and Dichloroethene. Applied Environmental Microbiology, 64: 3102-3105. CH2M Hill Constructors, I.C.M.H., 2001. Technical Memorandum, HRC Pilot Study Evaluation at SWMU 138 - Boat Ramp Naval Air Station Dallas, TX.

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Cope, N. and Hughes, J.B., 2001. Biologically-enhanced removal of PCE from NAPL source zones. Environ. Sci. Technol., 35: 2014-2021. Dean, S., Wiseman, L., Accashian, J., Edgar, M. and Callender, J., 2001. Design of an Enhanced Bioremediation System for a Low Permeability Aquifer, Proceedings of the Sixth In-Situ and On-Site Bioremediation Symposium, San Diego, California. DoD, 2002. Evaluation of performance and costs associated with anareobic dechlorination techniquesRevision 02 (Phase 1 site survey), Prepared for: Environmental security technology certification programm, Arlington, Virginia. DoE, 1999. In Situ Bioremediation for the Hanford Carbon Tetrachloride Plume. French, J. et al., 2003. Phased In situ Biostimulation/Bioaugmentation Pilot Testing in a Coastal Aquifer, Proceedings of the Seventh International Symposium of In Situ and On-Site Bioremediation. Battelle Press, Columbus, Ohio, Orlando, Florida, June 2003. GeoSyntec, 2002. Pilot Test for In Situ Bioremediation of Perchlorate & Trichloroethene in Groundwater Using an Active Biobarrier. Granade, S., D.P. Leigh, and C.D. Johnson., 2003. Chlorinated Solvent Bioremediation: 3 Case Studies, Proceedings of the Seventh International Symposium of In Situ and On Site Bioremediation. Battelle Press, Columbus, Ohio, Orlando, Florida, June 2003, pp. Paper A-13. Haas, P., Gonzales, J., Cork, P. and Henry, B., 2003. Remedial Performance of Organic Mulch Biowalls at Two Geochemically Distinct Sites, Proceedings of the 2003 AFCEE Technology Transfer Workshop, San Antonio, Texas, Air Force Center for Environmental Excellence, Brooks CityBase, Texas. February. Henry, B.M. et al., 2003. Permeable Mulch Biowall for Bioremediation of Chlorinated Solvents, Proceedings of the Seventh International Symposium of In Situ and On-Site Bioremediation. Battelle Press, Columbus, Ohio, Orlando, Florida, June 2003, pp. Paper K-03. Horst, J.F., Beil, K.A., Burdick, J.S. and Suthersan, S.S., 2000. Comparison of Natural and Enhanced Rates Through Substrate Amendments, Proceedings of the Second International Conference on Remediation of Chlorinated and Recalcitrant Compounds, Monterey, California. ITRC, 2002. Technical/Regulatory Guidelines: A Systematic Approach to In Situ Bioremediation in Groundwater Including Decision Trees on In Situ Bioremediation for Nitrates, Carbon Tetrachloride, and Perchlorate, Interstate Technology and Regulatory Council In Situ Bioremediation Team. ITRC, 2004. Strategies for Monitoring the Performance of DNAPL Source Zone Remediation. Koch, S.A., Rice, J.M., 2001. In Situ Enhanced Reductive Dechlorination of PCE, Proceedings of the Sixth International In-Situ and On Site Bioremediation Symposium, San Diego, California, pp. 181. Lee, M.D. and Lieberman, M.T., 2002. Low Cost Emplacement of Insoluble Organic Substrate for Enhanced In Situ Reductive Dechlorination, Altus Air Force Base. Lee, M.D. et al., 2003. Pilots to Enhance Trichloroethene Reductive Dechlorination and Ferrous Sulfide Abiotic Transformation, Proceedings of the Seventh International Symposium of In Situ and OnSite Bioremediation. Battelle Press, Columbus, Ohio, Orlando, Florida, June 2003, pp. Paper K14. Leigh, D.P. et al., 2000. Enhanced Anaerobic In Situ Bioremediation of Chloroethenes at NAS Point Mugu, Proceedings of the Second International Conference on Remediation of Chlorinated and Recalcitrant Compounds, Monterey, California, pp. 229-235. Lendvay, J.M. et al., 2001. Plume control using bioaugmentation with halorespiring microorganisms, Groundwater Quality 3rd International Conference, Sheffield, UK. Liles, D.S., Lut, C.C., Page, G.B. and Suthersan, S.S., 2004. In Situ Chemical Stabilization of Metals and Radionuclides Through Enhanced Anaerobic Reductive Precipitation: Application of a Commercial Technology to DOE Needs. http://www.arcadisus.com/resources/insituchemicalstabilization.pdf. Maierle, M.S. and Cota, J.L., 2001. Complete PCE Degradation and Site Closure Using Enhanced Reductive Dechlorination, Proceedings of the Sixth International In-Situ and On-Site Bioremediation Symposium, pp. 149-156. Martin, J.P., Sorenson, K.S. and Peterson., L.N., 2001. Favoring Efficient In Situ Dechlorination through Amendment Injection Strategy, Proceedings of the Sixth International In-Situ and On Site Bioremediation Symposium, San Diego, California, pp. 265-272. McCarty, P.L. et al., 1998. Full-Scale Evaluation of In Situ Cometabolic Degradation of Trichloroethylene in Groundwater Through Toluene Injection. Environ. Sci. Technol., 32(1): 88-100. Mikszewski, A., 2004. Emerging Technologies for the In Situ Remediation of PCB-Contaminated Soils and Sediments: Bioremediation and Nanoscale Zero-Valent Iron, National Network for Environ-

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mental Management Studies Fellow for U.S. Environmental Protection AgencyOffice of Solid Waste and Emergency Response Office of Superfund Remediation and Technology Innovation Technology Innovation Program Washington, DC www.clu-in.org. Murray, W., Dooley, M. and Koenigsberg, S., 2001. Enhanced Bioremediation of Chlorinated Solvents, Proceedings of the Sixth International In-Situ and On-Site Bioremediation Symposium, San Diego, California, pp. 197-204. Murt, V., Huscher, T., Easley, D., 2000. A Reductive Dechlorination Treatability Study of a Shallow Alluvial Aquifer, Proceedings of the Second International Conference on Remediation of Chlorinated and Recalcitrant Compounds, Monterey, California. Parsons, 2002. Final Phase II Field Feasibility Test for In Situ Bioremediation of Chlorinated Solvents Via Vegetable Oil Injection at Hanger K Area, Cape Canaveral Air Force Station, Florida. Parsons, 2004. Final Project Completion Report for a Field Feasibility Test for In Situ Bioremediation of Chlorinated Solvents Via Vegetable Oil Injection at Site SS015, Travis Air Force Base, California. Peeples, J.A., Warburton, J.M., Al-Fayyomi, I. and Haff, J., 2000. Enhanced Reductive Dechlorination of Ethenes Large-Scale Pilot Testing, Proceedings of the Third International Conference on Remediation of Chlorinated and Recalcitrant Compounds, Monterey, California. Sandefur, C.A., Parrett, K. and Lapus, K.A., 2002. Bioremediation of a PCE Plume at a Dry Cleaner Site. In: A.R.G.a.A.S.C. Chen (Editor), Proceedings of the Third International Conference on Remediation of Chlorinated and Recalcitrant Compounds. Battelle Press, Columbus, Ohio, Monterey, California, May 2002, pp. Paper 2B-52. Sorenson, K.S., 2000. Biodegradation of TCE Improved with Lactate Injection in Deep, Fractured Rock. Groundwater Currents, 38. Sorenson, K.S., 2003. Enhanced Bioremediation for Treatment of Chlorinated Solvent Residual Source Areas. In: S.M.H.a.S.D. Warner (Editor), Chlorinated Solvent and DNAPL Remediation: Innovative Strategies for Cleanup. ACS Symposium Series. Vol. 837, pp. 119-131. Stroo, H.F. et al., 2003. Remediating Chlorinated Solvent Source Zones. Environ. Sci. Technol., 37(11): 224A-230A. Suthersan, S.S., 2001. Natural and Enhanced Remediation Systems. ARCADIS. Lewis Publishers, Boca Raton, Florida. U.S.EPA, 2000a. Engineered Approaches to In Situ Bioremediation of Chlorinated Solvents: Fundamentals and Field Applications. EPA 542-R-00-008 http://www.epa.gov/clu-in.org. U.S.EPA, 2000b. Cost and Performance Report: Bioaugmentation (Accelerated Anaerobic Bioremediation) at Area 6 of the Dover Air Force Base, Dover, Delaware. U.S.EPA, 2004. DNAPL Remediation: Selected projects approaching regulatory closure. EPA 542-R-04016. U.S.EPA, 2004b. How to Evaluate Alternative Cleanup Technologies for Underground Storage Tank Sites: A Guide for Corrective Action Plan Reviewers. EPA 510-R-04-002. (USACE), U.S.A.C.o.E., 2003. Project Management Plan for Former Lincoln AFB, Atlas Site 10, York, NE. Willett, A., Tseng, J., Gillespie, R. and Koenigsberg, S., 2004. Hydrogen Release Compound (HRC®): A Review of Published Papers and Case Histories 1999-2003. Yang, Y. and McCarty, P.L., 2000. Biologically enhanced dissolution of tetrachloroethene DNAPL. Environ. Sci. Technol., 34(14): 2979-2984.

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Table 4: Summary of case studies results (AFCEE, 2004)
Observed Dechlorinaton Pathway Soluble Substrates – Lactate Test Area North, TCE to cis-DCE to Idaho National VC to ethene. Energy and Environmental Laboratory Idaho Site Name and Location Effectiveness of Substrate Distribution Substrate distribution was effectively attained with complete dechlorination observed up to 150 feet downgradient of the injection well. Site Status Criteria for Success Factors Contributing to Success or Failure References MCLs were achieved for TCE and DCE at the base of the aquifer. Concentrations of VC are generally less than 15 µg/L (October 2003) and VC has not accumulated. The pilot-scale application resulted in a ROD amendment (approved September 2001) to switch from groundwater pump and treat to enhanced in situ bioremediation. Dechlorination of VC was deemed too slow to meet remedial objectives, and the system converted to aerobic cometabolism. VC was effectively oxidized by cometabolic processes. It is unknown whether injection of additional substrate after 57 days would have stimulated faster dechlorination of VC. Strongly reducing conditions were induced (sulfate reduction and methanogenesis), but after 8 months of lactate injection, dechlorination of PCE and TCE stalled at cis- DCE. Dehalococcoides species were not detected by molecular screening. A bioaugmentation pilot test is planned. References

System is being expanded with larger volume injections and a second injection well.

Demonstrate that enhanced bioremediation of a DNAPL source area can reduce CAHs to below MCLs. Demonstrate that enhanced bioremediation can replace groundwater extraction as a final remedy for the source area. Pilot test to demonstrate that lactate addition can remediate TCE and DCE to ethene. Sequential dechlorination of TCE to DCE to VC and ethene were observed over approximately 36 months. Pilot test to demonstrate that lactate addition can stimulate complete dechlorination of PCE and TCE to ethene.

(AFCEE, 2004; Martin et al., 2001; Sorenson, 2003)

IRP Site 24, Naval Base Ventura County, Point Mugu California

TCE to cis-DCE to VC to ethene (degradation of VC to ethane relatively slow).

Substrate distribution was effective within the recirculation cell. Substrate was allowed to be depleted after last injection at day 57. Substrate was added by direct injection into a single well, and substrate distribution was effective within the treatment zone.

Pilot test monitored for 3 years, then converted to aerobic cometabolism. Full-scale sequential anaerobic/aerobic treatment is planned. Biostimulation pilot test conducted for 8 months. A bioaugmentation pilot test is planned.

(AFCEE, 2004; Granade, 2003; Leigh et al., 2000)

IRP Site 40, Naval Weapons Station Seal Beach, California

PCE and TCE to cis-DCE. After 8 months of biostimulation, transformation of DCE to VC and ethene was not observed.

(AFCEE, 2004; French et al., 2003)

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Observed Dechlorinaton Pathway Soluble Substrates – Molasses Washington PCE to TCE to cisSquare Mall DCE to VC to ethene/ethane. Wisconsin

Site Name and Location

Effectiveness of Substrate Distribution Substrate was effectively distributed using an initial injection into 182 direct-push points and during follow-on injections using a limited array of 12 injection points. Substrate distribution was limited initially; frequent injections were required to extend the zone of influence. Locations without adequate substrate showed only a relatively small decrease in TCE concentration, and incomplete dechlorination.

Site Status

Criteria for Success

Factors Contributing to Success or Failure References Closure with MNA was granted by WDNR 30 months after the initial application. Closure was based on complete dechlorination of PCE to ethene and ethane. Residual levels of DCE and VC were sufficiently low to allow for closure with MNA. Concentrations of total CAHs above 200 µg/L were reduced by over 80% at most locations. Concentrations of total CAHs between 50 and 200 µg/L were reduced by at least 75% at only a few locations. Complete dechlorination to ethene was observed at ocations that received a continuous and adequate supply of substrate.

References

Conditional closure with MNA.

Achieve closure under Wisconsin Department of Natural Resources (WDNR) flexible closure rules (negotiated preventative action limits).

(AFCEE, 2004; Maierle and Cota, 2001)

Hanscom AFB Massachusetts

TCE to cis-DCE to VC to ethene.

Pilot test was completed in September 2002. Future remedial actions are under evaluation.

Performance objectives included 1) for concentrations of total CAHs >200 µg/L – reduce concentration by 80% within 1 year.; 2) for concentrations of total CAHs from 50-200 µg/L – reduce concentration by 75% within 1 year; 3) for concentrations of total CAHs <50 ppb – reduce concentration by 50% within 1 year. Show that TCE can be completely dechlorinated to ethane. Demonstrate complete dechlorination of PCE and TCE to DCE and VC in an anaerobic reactive zone, with complete oxidation of residual DCE and VC in a natural downgradient aerobic treatment zone.

(ARCADIS Geraghty and Miller, 2003)

Other Soluble Substrates (Fructose/Acetate) Anaerobic/Aerobic TCE to DCE during The system was effecPilot Study, Naval pilot test. tive in recirculating Support Activity Further dechlorinagroundwater, although Mid-South tion of DCE to VC distribution of substrate observed 6 months appeared to be limited Tennessee after a final pulsed as levels of TOC averinjection of acetate. aged less than 20 mg/L throughout the treatment zone.

System is inactive pending regulatory approval of a final remedy for the site. MNA with hotspot reduction using direct injection of sodium acetate is proposed.

Concentrations of PCE and TCE were reduced by only 40 to 60% during active recirculation, with accumulation of cis-DCE. The rate and efficiency of substrate loading may not have been adequate to completely degrade TCE, or for dechlorination of DCE to VC and ethane.

(AFCEE, 2004)

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability

43

Observed Dechlorinaton Pathway Slow-Release Substrates – HRC® Fisherville Mill TCE to cis-DCE to Site VC to ethene. Massachusetts

Site Name and Location

Effectiveness of Substrate Distribution Substrate was effectively distributed throughout the treatment zone. Levels of TOC and metabolic acids remained elevated for a period of at least 27 months. Substrate was effectively distributed throughout the treatment zone. Levels of TOC and metabolic acids remained elevated for a period of approximately 18 months (HRC®) to 40 months (HRC-XTM).

Site Status

Criteria for Success

Factors Contributing to Success or Failure References Concentrations of TCE declined by 88 to 98%, coupled with dechlorination of cis-DCE and VC to ethene (completed dechlorination pathway). Elevated levels of TOC and metabolic acids persisted for at least 27 months. However, concentrations of TCE, cis-DCE, and VC within and downgradient of the treatment zone remain above MCLs. Sequential increases and decreases of TCE, cis-DCE, VC, and ethane were observed over approximately 18 months, indicating that PCE was co pletely dechlorinated to ethene. Co centrations of dechlorination products were observed to rebound after approximately 18 months, indicating that the HRC® substrate had been depleted. Regulatory approval for a fullscale application was granted based upon results of the pilot scale application. Significant reductions in TCE concentration did not occur. Only limited production of cis-DCE was observed in one location towards the end of the pilot test. Development of a stable and highly reducing reaction zone was not achieved. It is believed that high levels of DO (> 6 mg/L), nitrate (> 10 mg/L), and sulfate (>20 mg/L), combined with a high rate of groundwater flow, inhibited formation of adequate reducing conditions.

References

Results of the pilot test are being evaluated for consideration of a full-scale application.

Demonstrate that application of HRC® can remediate PCE, TCE, and dechlorination products to ethene. Demonstrate that application of HRC® can control the migration of CAHs to potential downgradient receptors. Demonstrate that application of HRC® can remediate PCE, TCE, and daughter products to ethene. Obtained regulatory approval for full-scale application.

(AFCEE, 2004; Murray et al., 2001)

Springdale Cleaners, Portland Oregon

TCE to cis-DCE to VC to ethene.

Pilot testing complete. Full-scale application has been approved and is pending funding for the project.

(AFCEE, 2004; Sandefur et al., 2002)

Atlas 10 Site Nebraska

Limited dechlorination of TCE to cisDCE only.

Elevated levels of TOC and organic acids were only observed in the immediate vicinity of the injection array during the 11-month test.

Site conditions were determined to be unsuitable for enhanced in situ bioremediation. Groundwater extraction and ex situ treatment using aeration are being considered.

Remediate chloroethenes in groundwater to federal drinking water MCLs.

((USACE), 2003)

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability

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Observed Dechlorinaton Pathway Slow-Release Substrates – HRC® SWMU 138, Limited dechlorinaNaval Air Station tion of PCE and TCE to cis-DCE. Dallas, Texas No significant concentration trends could be discerned in 9 months of monitoring.

Site Name and Location

Effectiveness of Substrate Distribution Elevated levels of substrate were not observed in any monitoring locations during the pilot test. Distribution appears to have been limited by low permeability soils and a lack of advective groundwater flow.

Site Status

Criteria for Success

Factors Contributing to Success or Failure References A lack of groundwater flow and mixing is thought to have resulted in insufficient distribution of substrate. Hydrogeologic conditions at this site do not appear to be suitable for use of slowrelease substrates. The contractor determined that HRC® did not offer an effective means to provide short term (less than 1 year) degradation of chloroethenes. Complete dechlorination of TCE to VC and ethene was observed. Concentrations of TCE were reduced to below detection along the centerline of the treatment zone. Although a complete dechlorination pathway has been observed, concentrations of cis-DCE and VC remain elevated above MCLs after 13 months of monitoring. Substrate distribution appears to be controlled by aquifer heterogeneity and secondary permeability. Complete dechlorination of PCE and TCE to ethene was observed, with over 80% reduction in total CAH mass site wide. Although MCLs for PCE, TCE, cis-DCE and VC were attained at multiple locations, this site may require several years to attain uniform reduction in CAHs to MCLs due to low permeability and slow desorption of CAH mass. Sequential increases and decreases of TCE, cis-DCE, VC, and ethane were observed over approximately 18 months. Approximately 40 months after injection, MCLs were attained for PCE, TCE, and cis-DCE at all locations within the treatment zone. MCLs for VC were also attained at multiple monitoring locations.

References

Pilot test completed, full-scale expansion was not recommended.

Reduce concentrations of PCE and TCE to below MCLs within the pilot test treatment zone.

(CH2M Hill Constructors, 2001)

Slow-Release Substrates – Vegetable Oil Site SS-17, Altus TCE to cis-DCE Injection of the vegetaAFB to VC to ethene. ble oil emulsion was readily accomplished, Oklahoma but uniform distribution was not observed at all locations downgradient of the treatment zone. Distribution appears to be controlled by secondary permeability.

Pilot test results continue to be evaluated.

Determine if a vegetable oil inwater emulsion can be effectively distributed and a reaction zone maintained in a barrier configuration. Determine whether complete dechlorination of TCE to ethene can be stimulated.

(AFCEE, 2004; Lee and Lieberman, 2002; Lee et al., 2003)

SS-015, Travis AFB, California

TCE to cis-DCE to VC to ethene.

Elevated levels of TOC were observed through out the entire treatment area, indicating that substrate was distributed effectively.

The site is approved for MNA and is being redeveloped by the Air Force. Monitoring will continue after construction.

Demonstration project to determine if vegetable oil is capable of complete dechlorination of TCE to ethene in a high sulfate, low permeability environment, and to evaluate different vegetable oil injection scenarios.

(Parsons, 2004)

Hangar K, Cape Canaveral Air Force Station Florida

PCE to TCE to cisDCE to VC to ethene.

Based on monitoring data for TOC and metabolic acids, substrate was effectively distributed throughout the source zone area.

Field test complete. Continue to monitor source area for longterm depletion of substrate.

Demonstration project to determine if vegetable oil is capable of complete dechlorination of PCE and TCE to ethene in a source area application. Results were evaluated to determine if federal drinking water MCLs could be achieved.

(Parsons, 2002)

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability

45

Observed Dechlorinaton Pathway Solid Substrates – Mulch Biowalls Building 301, TCE to cis-DCE, Offutt AFB with limited evidence of cis- DCE Nebraska to VC and ethene/ethane.

Site Name and Location

Effectiveness of Substrate Distribution Bark mulch was uniformly distributed within the biowall using a continuous one-pass trencher.

Site Status

Criteria for Success

Factors Contributing to Success or Failure References TCE was dechlorinated to cis-DCE, with limited evidence of dechlorination to VC to ethene. VC did not accumulate. A full-scale biowall was installed in 2001 based on results of the pilot test. The extent of dechlorination may be limited by the residence time of CAHs in the reactive zone. Concentrations of TCE within and immediately downgradient of the biowall have been reduced to below the MCL for TCE. TCE was dechlorinated to cisDCE, with no accumulation of VC. The average reduction in concentrations of total CAHs is 86 percent, although elevated concentrations of cis-DCE persist. Substrate addition alone was incapable of completely dechlorinating TCE to ethene within the 90-day test period. PCE was successfully dechlorinated to ethane with the addition of the bioaugmentation culture. The pilot test was also successful in degrading perchlorate to innocuous end products.

References

Full-scale biowall system was installed in 2001, with continued long-term monitoring.

Determine effectiveness of a mulch biowall to completely dechlorinate TCE to ethene. Develop operations and cost data to support a full-scale application.

(AFCEE, 2004; Haas et al., 2003)

OU1, Altus AFB Oklahoma

Primarily TCE to cis-DCE. Low levels (< 10 ug/L) of VC were observed. Abiotic degradation of TCE and cis-DCE is also occurring.

Bark mulch was uniformly distributed within the biowall using a continuous trencher. Elevated levels of TOC (greater than 20 mg/L) were observed up to 30 feet downgradient of the biowall. The recirculation system was effective at distributing substrate within the pilot test area. Distribution of the bioaugmentation culture was limited to the immediate area of injection.

Long-term process Monitoring continues. Fullscale biowalls are currently being installed for other CAH plumes at Altus AFB.

Intercept and contain a shallow TCE and DCE groundwater plume in order to prevent surface water discharge and off-base migration. Evaluate the technology for application at other Air Force sites.

(Haas et al., 2003; Henry et al., 2003)

Bioaugmentation Aerojet Facility California

PCE to TCE to cisDCE to VC to ethene. Also perchlorate to chlorate to chlorite to chloride.

Additional pilot testing was recommended. System is being evaluated for fullscale application.

Bachman Road Residential Wells Site Michigan

PCE to TCE to cis-DCE to VC to ethene.

Recirculation systems were effective at distributing substrate and bioaugmentation culture.

Full-scale application under consideration.

Determine if TCE and perchlorate can be completely degraded through substrate addition. Because microcosm studies indicated that a suitable microbial population may not be present, demonstrate the effectiveness of bioaugmentation to increase dechlorination rates. Assess feasibility of full-scale application. Determine relative effectiveness of bioaugmentation (lactate plus bioaugmentation culture) versus biostimulation alone (lactate only) to increase dechlorination rates of PCE and TCE to ethene.

(AFCEE, 2004; GeoSyntec, 2002)

A 92% conversion of chloroethenes to ethene was observed in the bioaugmentation test plot within 43 days of inoculation. Dechlorination of PCE and TCE to ethene was observed in the biostimulation test plot after appr. 3 months from the start of lactate injection. The lag time for dechlorination of cis-DCE to VC and ethene, and overall dechlorination rates were enhanced by addition of the bioaugmentation culture.

(AFCEE, 2004)

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability

46

Field Applications of In Situ Enhanced Bioremediation
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Removal Efficiency/Post treatment concentrations TCE: ND DCE: ND VC: ND Ethene: 0.36 mg/L Substrate Cost Comments Contacts/ References

Soluble Substrates – Lactate Bachman Road Sept. 2000 Residential Well 140 days Site injection Oscoda, MI

TCE, DCE, VC TCE: 0.13 mg/L DCE: 0.19 mg/L VC: 0.25 mg/L

Relatively uniform glacial outwash sands and gravels.

Groundwater remediation Pilot-scale 2 Surface area: 270 ft , depth: 8 ft Two injection wells; one extraction well; network of monitoring points around the plot. Lactate was injected as a 6% solution in water. Bioaugmentation after 20 days Amendments: nitrogen, phosphorous Technology demonstration; groundwater remediation Pilot-scale (Sodium lactate: 200 mg/L) 2 Surface area: 2,400ft , depth: 5 ft 3 injection wells, screen 3848' bgs. Closed loop recirculation cell w/ 3 recovery wells. Cyclic injection of nutrients and unamended GW. Bioaugmentation with a microbial culture from the DOE Pinellas Site Amendments: ammonia, and phosphate Source reduction by 50% Full-scale Lactate: 200 mg/L, amended with 2 mg/L yeast extract, sodium sulfite Injection wells

NA

Contaminant concentration reduction, as measured in one monitoring well, was nearly 100% for TCE, DCE, and VC.

John Lenvay, Univ of San Francisco Erik Petrovskis, HSI Geotrans

(DoD, 2002; Lendvay et al., 2001)

Area 6, Dover AFB Dover, DE

April 1998 to June 1999

TCE, DCE PCE: 0.046 mg/L TCE: 7.5 mg/L DCE: 2 mg/L VC: 0.034 mg/L

Sand with varying amounts of clay, silt and gravel.

PCE: ND TCE: 75 µg/L DCE: 45 µg/L VC: 20 µg/L

NA

As of March 1998, 98.5% of TCE and DCE in groundwater were converted to ethene and 75% to 80% of the TCE and DCE mass had been recovered as ethene. Biofouling had been reported as a problem, affecting the consistency of injections, the injection concentration, and the consistency of injection rates.

David Ellis, DuPont 302-892-7445 (U.S.EPA, 2000b)

Former Drycleaning Facility WI

Aug.1997 to April 2000

PCE PCE: 520 µg/L TCE: ND DCE: ND

20-feet of glacially deposited sand and gravel overlying sandstone bedrock

PCE: 220 µg/L TCE: 3 µg/L DCE: 28 µg/L

NA

PCE concentrations were reduced by approximately 58% while TCE and DCE concentrations increased form below method detection limits to concentrations greater than the Federal MCL for each contaminant.

Stacey Koch, RMT Inc. (Koch, 2001)

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design The four injections after the first were made under pressure to reduce biofouling. Groundwater remediation Full-scale 2 Surface area: 700 ft , depth:200 ft 300 gallons of sodium Lactate was injected weekly for 8 months. Approximately 26 to 27 feet of fine to medium grained sand. Technology assessment Pilot-scale 2 Surface area: 420 ft , depth:10 ft Sodium lactate injected at about 60% lactate in water in 3 to 4 week intervals over 10 months. One extraction well, two injection wells, 6 monitoring wells installed. Substrate injection was performed concurrently with groundwater extraction from the downgradient extraction well to form a closed recirculation system. Compare enhanced and intrinsic biodegradation rates Pilot-scale Recirculation with nutrient addition (Yeast extract, vitamins) PCE: 60 µg/L TCE: 1 µg/L DCE: 140 µg/L VC: NA Na Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

47

Test Area North (TAN), INEEL ID

Pilot Nov. 1998

TCE TCE: 3.2 mg/L

Fractured basalt bedrock.

TCE: <5 µg/L

$54,000 total

Within 6 weeks from the start of lactate injection nearly 100% of TCE concentrations in groundwater had been dechlorinated to DCE and complete dechlorination was occurring within 4 months. Contaminant reduction calculations could not be performed due to the limited availability of analytical data.

Kent Sorenson, Northwind Environmental (ITRC, 2004; Sorenson, 2000)

Ogallala Superfund Site Ogallala, NE

April 1998

PCE, TCE PCE: 30 µg/L TCE: ND DCE: ND VC: ND

Vicki Murt, Nebraska Department of Environmental Quality (DoD, 2002; Murt, 2000)

NAS Fallon Fallon, NV

1998 500 days

PCE, TCE, DCE PCE: 680 µg/L TCE: 915 µg/L DCE: 866 µg/L VC: 4.3 µg/L

Seal Beach Site 40

PCE: 415 µg/L TCE: ND

Approximately 4-feet of sandy soil underlain by approximately 2-feet of clayrich silts and sands. Bottom is sandy silt, and clay layer greater 20 feet thick. Surficial soils , sands and silty

PCE: <25 µg/L TCE: 100 µg/L DCE: 250 µg/L VC: 1.5 µg/L

NA

Contaminant concentration reduction in groundwater ranged from a minimum 65% in the case of VC to a maximum of 99.7% in the case of PCE.

Victor Magar, Battelle (DoD, 2002)

Pilot-scale

PCE: ND TCE: ND

NA

Strongly reducing conditions were induced (sulfate

Kent Sorenson, Northwind

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level DCE: ND VC: ND Description of Geology Remedial Objective Treatment System Design sands intercalated with low permeable intervals of clay, silt and silty clay. Area: 1,300 ft , depth: 15 ft Sodium lactate: 3 g/L One injection well, with 6 monitoring wells interspersed within a 20 ft radius. Weekly injections for 2 months, then none for 2 months, lastly every 3 weeks.
2

48
Removal Efficiency/Post treatment concentrations DCE: 485 µg/L VC: ND Substrate Cost Comments Contacts/ References

CA

Tennessee Air national Guard TN

2002

Groundwater remediation to reduce total VOC to < 500 ppb Pilot-scale Sodium lactate Multiple manifold wells, bimonthly injections Technology demonstration Pilot-scale Sodium lactate Single well injection, one time for demo. Pilot-scale Sodium Benzoate, sodium lactate and methanol Groundwater remediation Pilot-scale Sodium lactate Amendments: Ammonia and ortho-phosphate 2 Area: 12,000 ft , depth: 10 ft Each treatment cell consists of 4-5 extraction wells on cell perimeter around central injection well. Treatability study Pilot-scale 2 Area: 340 ft , depth: 10 ft

reduction and methanogenesis), but after 8 months of lactate injection, dechlorination of PCE and TCE stalled at cis- DCE. Dehalococcoides species were not detected by molecular screening. A bioaugmentation pilot test is planned. Analytical data was not provided for this site.

Kent Sorenson, Northwind

Black’s Drycleaner OR

2001

PCE, TCE

Analytical data was not provided for this site.

Kent Sorenson, Northwind

Pinellas Northeast Site Largo, FL Former PEC Industries Site Orlando, FL

Analytical data was not provided for this site.

(DoE, 1999)

2001

PCE, TCE, DCE, VC PCE: 43 µg/L TCE: 17 µg/L DCE: 8.1 mg/L VC: 112 µg/L

The surficial aquifer consists of three sand units separated by sandy clay and clayey sand aquitards.

NA

The well installation and baseline monitoring phases have been completed as of May 2001. The injection phase was scheduled for 2001 while monitoring was to be extended through 2002.

(Dean et al., 2001)

Cape Canaveral Facility 1381 FL

March 1999 149 days

TCE, DCE, VC TCE: 1.97 mg/L DCE: 11.1 mg/L VC: 1.25 mg/L

Poorly sorted coarse to fine sands and shell material to 35 feet.

TCE: ND DCE: 970 µg/L VC: 625 µg/L

NA

Rapid dechlorination of TCE and cDCE to VC, followed by slower subsequent dechlorination of VC to ethene under sulfate

Jeff Morse, Battelle

((AFRL), 2001)

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Lactic Acid Two communicating (recirculation) wells, 13 tri-level groundwater monitoring probes, and upgradient and downgradient monitoring wells. Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

49

Watertown Industrial Area Watertown, MA

July 1997

TCE; DCE PCE: 1.5 mg/L TCE: 12 mg/L DCE: 3.5 mg/L VC: 100 µg/L

Industrial site adjacent to river, 20 ft of silty sand on top of thin till and bedrock.

Feasibility study Pilot-scale Lactic Acid Amended with ammonia chloride, tripolyphosphate, yeast extract, and sodium hydroxide. System of injection and recirculation cells

PCE: 100 µg/L TCE: 1 mg/L DCE: 3 mg/L VC: 1 mg/L

NA

and methanogenic conditions. RABITT protocol slightly modified because Florida's UIC regulations do not allow for reinjection of contaminated groundwater. Reduction of TCE, DCE, and VC by 88.7%, 90.6%, and 66.3%, respectively Contaminant concentration reduction ranged from 7% in the case of PCE to 92% in the case of TCE. Contaminant mass reductions were measured in a single well (IN-2). After 8 months of anaerobic operation, the system was converted to an aerobic circulating cell for 3 months.- ORC,methane addition => 70% reduction of total ethenes in 11 months. TCE concentrations were reduced by nearly 100% 125 days of injection.

Willard Murray, Harding Lawson Associates (DoD, 2002)

Aerojet Superfund Site CA

Sept. 2000

TCE TCE: NA DCE: NA VC: NA

Technology demonstration Pilot-scale Lactic acid Bioaugmentation with a commercially produced microbial augment (KB-1) Technology demonstration Pilot-scale Lactic acid One extraction well, one injection well and 5 monitoring wells. Groundwater was circulated in a closed loop between the

TCE: < 5 µg/L DCE: NA VC: NA

Evan Cox or David Major, Geosyntec Consultants

(DoD, 2002) TCE: < 5 µg/L DCE: <5 µg/L VC: 15 µg/L Within approximately 180 days TCE and DCE concentrations had been reduced by nearly 100% at one well. However VC concentrations increased by a factor of 15 within the same time period. Pilot test monitored for 3 years, then converted to Daniel Leigh, IT (Leigh et al., 2000)

Naval Air Station Point Mugu IRP Site 24 CA

Dec. 1998 1 year

TCE, DCE, VC TCE: 1.7 mg/L DCE: 750 µg/L VC: 1 µg/L

Upper unconfined aquifer: sand and gravel overlying approximately 4 feet of clay. Lower confined aquifer: massive sand unit.

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design extraction well and the injection well. Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

50

Weyerhauser Sycan Maintenance Shop Beatty, OR

Pilot test complete

PCE, TCE PCE: 1.7 mg/L TCE: 450 µg/L DCE: 3.6 mg/L VC: 90 µg/L

Sandy silts and silty sands

Feasibility study Pilot-scale Lactic acid Injection and monitoring system was placed to inject into and monitor changes within the upper aquitard. One well was used for groundwater extraction, substrate injection, and process monitoring. Treatability study Pilot-scale Lactic acid Amendments: Vitamin B12, yeast extract

PCE: 200 µg/L TCE: 160 µg/L DCE: 1400 µg/L VC: 90 µg/L

aerobic cometabolism for faster VC degradation Full-scale sequential anaerobic/aerobic treatment is planned. The PCE, TCE, and DCE concentrations were reduced by approximately 88%, 64%, and 61% respectively while the VC concentration stayed about the same.

Sanjay Vancheeswaran, CH2M Hill (DoD, 2002)

Lauderick Creek Area of Concern, Main Plume, Aberdeen Proving Ground

Pilot test complete

PCA, TCA PCA: 20 mg/L TCE: 1 mg/L DCE: ND VC: ND

PCA: 3.5 mg/L TCE: 1.8 mg/L DCE: 3.9 mg/L VC: ND

Within 14 weeks the PCA concentration at one well was reduced by 83% while the TCE and DCE concentrations increased.

Andrew Jackson, Texas Tech University (DoD, 2002)

MD Soluble Substrates – Molasses Avco Lycoming Pilot: NovemSuperfund Site ber 1995 Full: January Williamsport, PA 1997 to July 1998

TCE TCE: 48 µg/L DCE: 10 µg/L VC: <1 µg/L

Glacial sandy silt overburden overlying fractured limestone bedrock

Groundwater remediation Pilot, Full Area: 5,000 ft , depth: 11 20 injection wells Soil and groundwater remediation Full-scale Area: 11,500 ft , depth 25-28’ Geoprobe, infiltration gallery
2 2

TCE: 1.4 µg/L DCE: 2 µg/L VC: <1 µg/L

Daniel Jacobs, ARCADIS 215-752-6840 (Horst et al., 2000)

Cedarburgh Drycleaners Cedarburgh, WI

PCE, TCE, DCE PCE: 6.8 mg/L TCE: 810 µg/L DCE: 1 mg/L

Poorly drained silty clay topsoil with little sand and gravel (14'). Silty clay and clayey silts (4-16'). Silty clay with trace sand and gravel (15-39')

NA

Jim Drought, ARCADIS 414-276-7742 (DoD, 2002)

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design overlying silty find sands. Sand and gravel Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

51

Industrial Site Central Ohio

Phase I: November 1996 Phase II: September 1998 Phase III: June 1999

DCE, VC DCE: 265 µg/L VC: 15 µg/L

Contaminant mass reduction in a downgradient groundwater plume Pilot-scale Area: 2 Phase 1: 500 ft 2 Phase II: 4,500 ft 2 Phase III: 32,000 ft Depth: 25 ft Phase I consisted of 1 injection well, 1 injection well, and 1 monitoring well. Phase II consisted of 2 injection wells and 9 monitoring wells. Phase III used the same injection wells installed for Phase II but added 20 more monitoring wells. Source reduction with residual natural attenuation Full-scale 2 Area: 30,000 ft , depth: 5 ft Molasses (47% carbohydrate during pilot; 66% during full) Geoprobe, infiltration gallery. Site closure Pilot-scale, Full-scale 2 Area: 170000 ft , depth: 2573 ft Fully automated Mass reduction, natural attenuation for low-level residuals

DCE: 25 µg/L VC: 4 µg/L

Phase I was designed to confirm the capability of enhancing reductive dechlorination in-situ. Phase II was designed to evaluate amendment distribution strategies. Phase II was designed to the maximum volume of aquifer that could be impacted by a single well injection. Contaminant concentration data were collected during Phase I only.

Jim Peeples, formerly Parsons (Peeples et al., 2000)

Washington Square Mall Germantown, WI

August '98; 6 months Pilot March '99; 6 months Full

PCE PCE: 1.5 mg/L

Manufacturing Facility Greenville, SC

Pilot - March 1998 Full - October 2001 2 years

CT CT: 0.19-1.2 mg/L

Approximately 14 feet of clay and silt overlying a 2 to 5 foot thick saturated silt and sand bed, clay till aquitard below the saturated silt/sand bed Silty sand and sandy silt overlying a highly weathered transition zone. Approximately 4 to 12 feet of fill material overlying 4 to 8'

PCE: ND TCE: ND DCE: 400 µg/L VC: 100 µg/L

Site closure from WDNR, less than 2.5 yrs after initiating the in situ reaction zone.

Michael Maierle, ARCADIS 414-276-7742 (Maierle and Cota, 2001)

CT: 5 µg/L

$30,000/yr.

Crestwood Site Glendale, WI

1998

PCE: 44 mg/L

NA

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design of sand; clay till aquitard underlies the sand bed. Interbedded sand and clay to a depth of 24 feet underlain by impermeable clay Full-scale Geoprobe, infiltration gallery Reduce average concentration of total chromium and TCE in groundwater by more than 90% Pilot-scale, Full-scale 91 injection points with Geoprobe Manufacturing Facility Northeastern USA Pilot - October 1998 Full - April 2000 2 to 5 years of injection PCE PCE: 80 mg/L TCE: 1 mg/L DCE: 1 mg/L VC: ND 10-20' thick overburden of glacial sands, silts and clays; water table in underlying bedrock. Full-scale 2 Area: 450000 ft , depth: 50190 ft Molasses, 200 gal; 10%carbohydrates in each well Batch delivery under pressure to 11 injection wells. Reduce VOC concentrations, contain plume migration Pilot-scale PCE: 1 µg/L TCE: 1 µg/L DCE: 1 µg/L VC: 2 µg/L $5,000/year Zero valent iron proposed for source area. Full-scale containment in place in mid-plume 2000 to 2002. Full scale source area injection program began in October 2001. TCE: <5 µg/L DCE: <5 µg/L VC: 21 µg/L $3 per gallon Closure, all contaminant concentrations were below MCLs. Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

52

Emeryville Manufacturing Facility Emeryville, CA

Pilot - August 1995 Full - April 1997

TCE, DCE TCE: 6.5 mg/L DCE: 590 µg/L VC: 10 µg/L

Nuclear Fuel Services Site Erwin, TN

August 2000 8 months of injection

PCE, TCE PCE: 12.4 mg/L TCE: 1.14 mg/L DCE: 660 µg/L VC: 130 µg/L

0-30' Alluvial silts, clays, clayey sand, sand w/ gravel and cobbles underlain by weathered, fractured, steeply.

PCE: 850 µg/L TCE : 320 µg/L DCE: 67.8 mg/L VC: 13 mg/L

Within 6 months following substrate injection PCE, and DCE contaminant mass had been reduced by 93% and 82% respectively. Within the same time period DCE, VC, and ethene concentrations increased by approximately 2 orders of magnitude. The pilot scale application was considered a success and full scale applications are scheduled for "designated source areas" in the 2nd quarter of 2002.

TRW Automotive Rogersville, TN

4 months of injection

PCE PCE: 22 mg/L

0-30' Alluvial silts, clays, clayey sand,

Reduce VOC concentrations Pilot-scale

PCE: 2 µg/L TCE: <2 µg/L DCE: 8.6 µg/L

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level TCE: 5 µg/L DCE: 42 µg/L VC: 1 µg/L Description of Geology Remedial Objective Treatment System Design sand w/ gravel and cobbles underlain by weathered, fractured bedrock. Upper and lower aquifers consisting of glacial till overburden overlying bedrock and separated by a stiff. Alluvial marine sands (with gravel, silt, and clay) over siltstone Removal Efficiency/Post treatment concentrations VC: 22.6 µg/L Substrate Cost Comments Contacts/ References

53

Hanscom AFB Bedford, MA

October 2000

TCE, DCE, VC TCE: 1.9 mg/L DCE: 5.3 mg/L VC. 1.3 mg/L

Technology demonstration Pilot-scale 2 Area: 5000 ft , depth: 50 ft One injection well. Manual, pressurized injections. Postinjection water pushes. Technology demonstration Pilot-scale 2 Area: 1000 ft , depth: 30 ft Molasses: variable frequency and amount based on injection well process monitoring data. 3 injection wells. Manual, pressurized injections. Plume restoration by reactive barrier Full-scale 5 to 10% molasses solution biweekly for 6 months, intermittent thereafter Reactive barrier configuration

TCE: 1.9 mg/L DCE: 5.3 mg/L VC. 1.3 mg/L

Post-treatment concentrations are those from existing monitoring well after 19 months of demonstration testing.

Vandenberg AFB Lompoc, CA

February 2001

TCE TCE: 4 mg/L

TCE: 19 µg/L

Concentrations from existing monitoring well showing good treatment.

Ohio Industrial Site Southwestern Ohio

1999

PCE, TCE PCE: 500 µg/L TCE: 700 µg/L VC: 3 µg/L

Porous, high carbonate aquifer

PCE: < 1 µg/L TCE: < 1 µg/L VC: < 1 µg/L

Manufacturing Facility Southeast England

1999

TCE TCE: 22 mg/L DCE: 12 mg/L VC: 45 µg/L

Sandy gravel alluvium overlying a thick regional aquitard known as the London Clay

Plume restoration, partially underneath the facility Full-scale A total of 53 injection wells were installed under the future site of a new building.

TCE: 14 µg/L DCE: 20.8 mg/L VC: 4.5 mg/L

Within approximately 440 days following substrate injection PCE, TCE, DCE, VC concentrations in groundwater had been reduced by nearly 100%. At day 180 DCE concentrations peaked and began to decline while ethane concentrations peaked at day 240. Since substrate injection in 1999 the average TCE concentration has decreased by nearly 100% while VC and ethane concentrations have increased by 1 and three orders of magnitude re-

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Each injection well was piped separately to the injection system installed in a separate building so that substrate injection could be controlled by well. 7 monitoring wells were installed up and down gradient from the injection area. Technology Demonstration Full-scale Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

54

spectively and the average DCE concentration has increased by approximately 73%..

Former Manufacturing Facility North Carolina

2001 1 year

PCE, TCE PCE: 11 mg/L TCE: 180 µg/L

Silty clay

NA

PCE and TCE concentration decreases of 89% and 94% respectively, were reported although analytical data was not provided.

Soluble Substrates – Butyrate Naval Air Station June 1999 Alameda Building 360 (Site 194 days #4) Alameda, CA

TCE, DCE, VC TCE: 24 mg/L DCE: 8.6 mg/L VC: 2.2 µg/L

Treatability study Pilot-scale 2 Area: 45 ft , depth: 3 ft Butyric acid (3 mM) at 236 gal/day (0.62 L/min) Amendments: Yeast extract, 20 mg/L Single injection well, single extraction well, and nine monitoring points at depths of 24 to 27 feet bgs

TCE: 700 µg/L DCE: 2.4 mg/L VC: 600 µg/L

Fort Lewis East Gate Disposal Yard Fort Lewis, WA

August 2000 179 days

TCE, DCE TCE: 1.5-6.3 mg/L

Treatability study Pilot-scale 2 Area: 120 ft , depth: 3 Butyric acid, 1.5 L/min Amendments: Yeast extract, 20 mg/L; Sodium bicarbonate, 279 mg/L Three injection wells spaced 2 ft apart; system of monitoring wells; existing well in

TCE: 69 µg/L DCE: NA VC: 217 µg/L

Water injected with amendments contained average TCE, cDCE, and VC concentrations of 81.7 mM, 7.0 mM, and 3.4 mM, respectively. Average TCE concentrations were reduced by 94% by the end of the demonstration. On average, 87% of injected chloroethenes could be accounted for, with ethane levels accounting for approximately 50% of the total chloroethene concentration. Injected TCE concentrations spiked as high as 169 mg/L, but this did not prove toxic to the microorganisms. Calculated TCE half lives of 4.4 to 4.7 hours. cDCE accumulated with VC comprising a small percentage of overall mass.

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design zone of contamination used to supply injection water for test plot. Treatability study Pilot-scale 2 Area: 120 ft , depth: 3 Butyric acid, 0.65 L/min Amendments: Yeast extract, 20 mg/L Three injection wells, nine monitoring wells at depths of 45 to 48 feet bgs. Technology demonstration Pilot-scale 2 Area: 400 ft , depth: 10 Amendments: Nitrate One injection well and one extraction well were installed and run continuously as a recirculation system; two monitoring wells were installed between the injection and extraction wells for process monitoring. Technology demonstration Pilot-scale Eastern US One injection well and one extraction well were installed. Acetate was injected to reduce sulfate mass and to push geochemistry to more reducing conditions. 3 months after acetate injection, bioaugmentation with methanol and KB-1 was performed. Manufacturing 1998 PCE, TCE Thin saturated Technology demonstration NA Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

55

Ethene and ethane remained at or below detection. Average instead of max: PCE: 5.5 mg7l TCE: 400 µg/L DCE: 80 µg/L VC:<6 µg/L

Marine Corps Camp Lejeune Site 88 Camp Lejeune, NC

June 2001 180 days

PCE, TCE NA

Sand and clay sediments to 75 ft bgs. A series of sand and limestone beds occur between 50 and 300 ft bgs.

Soluble Substrates – Acetate Hanford 200 Area January 1995 Site to March 1996 Richland, WA

CT, Nitrate NA

Unsaturated zone extends to a depth of 250 feet. Two permeable zones are located at 250 to 256 feet bgs and 285 to 302 feet bgs with a competant aquitard between.

NA

Approximately 2 kg of CT was reported to have been destroyed during the 14 month pilot test. Within the same time period approximately 30 kilograms (dry weight) of bacterial biomass was produced. Acetate and nitrate were injected and recirculated through 2 aquifer zones to treat carbon tetrachloride and nitrate contamination. This site had high sulfate (>1000 mg/L), high salinity, and tidal impact due to close proximity of the coast. The target zone was also deep >100 ft. TCE degradation to DCE was noted early on but DCE degradation and VC/ethene accumulation was not observed until 4 months after bioaugmentation with the KB-1 commercial bioaugmentation product. Acetate injection into a

Gillette Company Site

TCE

NA

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design soil over fractured basaltic bedrock. Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

56

Facility NJ Soluble Substrates –Methanol/Acetate Building 360, Kelly December, PCE, TCE, DCE Air Force 1999 Base PCE: 8.1 mg/L 320 days TCE: 190 µg/L South Central TX DCE: 2.1 mg/L VC: ND

Pilot-scale

surficial overburden aquifer/fractured bedrock aquifer.

20 to 40 feet of alluvial gravel, sand, and silt overlying impermeable clay.

Technology demonstration/ source reduction Bench, Pilot-scale 2 Area: 90 ft , depth: 25 ft KB-1 ("A naturally occurring microbial consortia isolated by GeoSyntec and Univ. of Toronto") Closed loop circulation system consisting of 3 extraction wells, 1 injection well and 5 monitoring wells. Technology Demonstration Pilot-scale 2 Area: 5000 ft , depth 10 ft 0.35% methanol, 1.44% compost leachate, 0.9 g/L ammonium chloride 10 Injection wells were installed upgradient of 5 extraction wells. Groundwater was extracted, amended, and reinjected.

PCE : 810 µg/L TCE : 19 µg/L DCE : 315 µg/L VC : NA

Reported a 90% reduction in PCE. Cis- 1,2-DCE degradation was observed only after the addition of the KB-1 microbial consortium amendment.

Rademarkt Site Groningen, Netherlands

35 weeks

PCE, TCE, DCE, VC PCE : 1.28 mg/L TCE : 1.03 mg/LPCE DCE : 2.6 mg/L VC : 940 µg/L

PCE : 600 µg/L TCE : 10 µg/L DCE: 290 µg/L VC: 2.26 mg/L

Contaminant concentration reductions represent site average concentrations before injection and 25 weeks after injection. This application used methanol and compost leachate as a substrate to augment reductive dechlorination already occuring at the site.

Soluble Substrates –Lactose Former Municipal 1997-2000 Waste Water pilot Treatment Facility 2000-2003 (Lactose) full-scale 2003-pursue NH closure with

PCE, TCE, DCE PCE: 20 µg/L TCE: 300 µg/L DCE: 8.4 mg/L VC: 1.5 mg/L

Upper fine sand and silt unit (5-20'); Lower sandy and silt unit (30-50' bgs)

Groundwater remediation to potable water standards Full-scale Lactose (approx. 70%) and

PCE: <1 µg/L TCE: <1 µg/L DCE: 2.7 mg/L VC: 300 µg/L

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status MNA Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Yeast Extract (approx. 30%) (proprietary blend) Injection of blend pumped into 3 locations: 5,050 lbs biostimulant per 65,950 gallons injected during pilot; 22,100 lbs per 375 gallons injected during full-scale TCE Fluvial deposits aquifer Feasibility test for groundwater restoration Pilot-scale Amendment: Acetate Recirculation system with one downgradient extraction well. NA Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

57

Soluble Substrates –Fructose Naval Support March 2000 Activity MidSouth (Fructose, Acetate) Millington, TN

Soluble Substrates –Sodium Benzoate New England January 1998 DCE, Super Fund Site Tetrahydrofuran, Sages Drycleaner 2 years Fuel Hydrocarbons New England

Technology demonstration for enhanced bioremediation as a cost effective alternative to pump and treat Pilot-scale Sodium Benzoate, injected as a4 mg/L solution at a rate of 5 liters per point per day. 30 electron donor injection points installed in sets of three (three injection intervals) in a barrier configuration. 1 upgradient and 3 downgradient rows of monitoring wells are used for process monitoring.

NA

Degradation rates were calculated for cis- 1,2-DCE and VC based on production rates of ethene and ethane for the first two years of operation. The average half life calculated for cis-1,2-DCE was 255 days. The calculated average half life for VC was 48 days.

Soluble Substrates –Ethanol June 1998 Jacksonville, FL 1 year

PCE DNAPL PCE: 150 mg/L

Technology Demonstration to remove PCE DNAPL

PCE: 44 mg/L

Approximately 43 liters of PCE were recovered from the extracted water/ etha-

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Pilot-scale Ethanol 8,980 gallons injected as a 95% ethanol in water mixture. Amendments: methanol, n hexanol 3 injection wells surrounded by 6 recovery wells. Ethanol was injected at a total of 15 liters per minute for three days. Groundwater was extracted at a rate of 30.4 liters per minute to ensure containment of injected fluids. Slow-Release Substrates – HRC® Contemporary Cleaners 1 year Orlando, FL PCE, TCE, DCE, VC PCE: 19.2 mg/L TCE: 2.5 mg/l DCE: 2.4 mg/L Fine grained quartz sand (25-30'), clay 112'; fine grained sand and sandy clay 20-25 Groundwater remediation via accelerated anaerobic biodegradation Full-scale 2 Area: 14600 ft , depth: 15 ft HRC; 6,800-lbs was injected into the shallow aquifer 18 months later 6,810 pounds were injected into the deep aquifer 144 injection points installed within the upper aquifer on a 10 foot grid spacing. Second injection consisted of 145 injection points on a 10 foot grid. PCE: 820 µg/L TCE: 1.3 mg/L DCE: 4.0 mg/L VC: 1.0 mg/L $40,800 1st application $40,860 2nd application Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

58

nol waste stream. This PCE mass represents approximately 63% of the total PCE mass present in the flushed area indicating that the extraction efficiency is 63%.

Within 152 days following injection PCE, TCE, DCE, and VC concentrations were reduced by 96%, 48%, 38%, and 58%, respectively, over 5 wells screened in the shallow aquifer. Two HRC applications were performed on this site. The first application was performed in the shallow aquifer. The second application was performed 18 months later in the deep aquifer because the first application did not appear to be affecting contaminant mass present in the deep aquifer.

Decorah Shopping Center Drycleaners Decorah, WI

1999

PCE PCE: 18 µg/L TCE: 4 µg/L DCE: <1 µg/L

Sandy silt and silty sand with varying amounts of clay (5'), med grained sand (5'-11'bgs) silty sand and

Contaminant source removal, plume stabilization, groundwater restoration Full-scale

NA

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design sandy silt with silty clay (1124'bgs), silty clay (24-28'bgs) Silty , finegrained sand (0-18' bgs); clayey finegrained sand (18-30' bgs); limestone (3032' bgs) Shallow sand (15-22'), dense silty clay (30') Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

59

Dixie Cleaners Jacksonville, FL

June 2000 1 year

PCE, TCE, DCE, VC PCE: 5.2 mg/L TCE: 12.7 mg/L DCE: 7.5 mg/L VC: 1.1 mg/L

Source zone remediation Pilot-scale 2 Area: 18400 ft , depth: 10 ft 175 injection points installed on a 10 foot grid spacing. 30 monitoring wells, 10 installed within each aquifer zone. Source zone and plume remediation Pilot-, Full-scale 2 Full: 3,900 ft , depth: 5 ft Pilot - 8 direct injection wells installed in a semicircle 2.5 feet from one monitoring well. A second monitoring well was installed 3 feet downgradient from the outside edge of the injection point circle. Full - 159 injection points installed in 3 grids and 1 barrier. Polish source area after removal of 2-Phase extraction system Full-scale 2 Area: 900 ft , depth: 15 ft 21 injection points at 5-ft centers using a Geoprobe rig

PCE: 42 µg/L TCE: 1.3 mg/L DCE: <5 µg/L VC: NA

Average PCE, TCE, and DCE concentrations were reduced by 99%, 90%, and 99% 7 months after injection.

Dover Park Plaza Drycleaning Facility Yardville, NJ

PCE, TCE PCE: 1.4 mg/L TCE: 20 µg/L DCE: 100 µg/L VC:<5 µg/L

PCE: <5 µg/L TCE: <5 µg/L DCE: 220 µg/L VC: <5 µg/L

After approximately 400 days following the full scale application PCE and TCE concentrations were reduced by approximately 99% and 75% respectively.

Former Industrial Filter Manufacturer Rochester, NY

Aug. 1998

TCE TCE: 26 mg/L DCE: 340 µg/L VC: ND

Relatively low permeability clayey silts

TCE: 4.7 mg/L DCE: 18 mg/L VC: 190 µg/L

Former Landfill

Aug. 2000

TCE

Determine potential for

TCE: ND

Contaminant concentration data used to calculate mass reduction represents maximum values detected in 5 monitoring wells. Within 15 months of injection TCE concentr tions were reduced by approximately 64% while DCE and VC concentrations increased by approximately 288% and 3700% respectively. DCE and VC peaked

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design groundwater remediation via anaerobic biodegradation Pilot-scale 9 injection points using a Geoprobe rig Watertown Industrial Area Watertown, MA Feb. 1998 PCE, TCE PCE: 1 mg/L TCE: 13 mg/L DCE: 3 mg/L VC: 200 µg/L Appr. 13 feet of sand and gravel overlying approximately 7 feet of silty sand. The surficial material is underlain by an aquitard consisting of glacial till. Silty sand Feasibility study for enhanced anaerobic biodegradation Pilot-scale HRC-containing canisters suspended in injection well HRC canisters in wells used with a recirculation system PCE: 8.5 µg/L TCE: 95 µg/L DCE: 163 µg/L VC: < 200 µg/L Average PCE, TCE, and DCE concentrations were reduced by 99%, 99%, and 95% at 206 days after injection. Removal Efficiency/Post treatment concentrations DCE: 700 µg/L VC: 20 µg/L Substrate Cost Comments Contacts/ References

60

Site 7, Duluth International Airport Duluth, MN

TCE: 354 µg/L DCE: 50 µg/L VC: 10 µg/L

before starting to decline at end of pilot test.

Former Manufacturing Site Walled Lake, MI

Jan. 1999

DCE, VC DCE: 9.9 mg/L VC: 1.9 mg/L

Feasibility study for enhanced anaerobic biodegradation Pilot-scale Geoprobe injection

DCE: 5.6 mg/L VC: 3.2 mg/L

Groundwater results showed large amounts of lactic acid fermentation.

Unocal Wichita Wichita, KS

Sept. 1999

PCE PCE: 7 mg/L TCE: 840 µg/L DCE: 560 µg/L VC: ND

Silt and clay

Feasibility study for enhanced anaerobic biodegradation Pilot-scale 15 injection points

PCE: 40 µg/L TCE: 540 µg/L DCE: 15 mg/L VC: 1 mg/L

PCE was degraded from 6 mg/L to 0.2 mg/L within 30 days.

FMC Corporation Site San Jose, CA

June 1999 to /May 2000 Full-scale planned

TCE TCE: 3.62 mg/L DCE: 210 µg/L VC: 20 µg/L

Silty clays (4550'); gravelly sand (3035')

Accelerate groundwater restoration Pilot-, Full-scale 2 Area: 150 ft , depth: 20 ft 6 direct-push points and 5 monitoring wells.

TCE: 3.15 mg/L DCE: 550 µg/L VC: 570 µg/L

Contaminant concentrations used to calculate mass reductions represent averages of 6 site wells used to monitor this pilot test. Within 6 months of injection the average TCE concentration had been reduced by 13%. DCE, VC, and ethene concentrations

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

61

Hayden Island Cleaners Portland, OR

May 1999

PCE PCE: 800 µg/L TCE: 1 µg/L DCE: 3.4 µg/L

20-40 feet of silty sand

Accelerate groundwater restoration Full-scale 2 Area: 200 ft , depth: 15 ft 34 injection points in 2 rows

PCE: 200 µg/L TCE: 35 µg/L DCE: 70 µg/L

increased by 161%, 2750%, and 733% respectively during the same time period. PCE concentrations were reduced by approximately 75% within 20 months of injection, while TCE and DCE concentrations increased within the same time period. Site closed

Naval Air Station Dallas Dallas, TX Naval Air Warfare Center Indianapolis, IN

July 2000 – May 2001

PCE, TCE PCE: 99 µg/L TCE: 44 µg/L DCE: 11 µg/L TCE, DCE TCE: 1 mg/L DCE: 202 µg/L VC: 0.3 µg/L

Fine-grained fill, alluvial sediments, weathered shale

Groundwater remediation Pilot-scale 2 Area: 1500 ft , depth: 10 ft Groundwater remediation Full-scale Injection per Regenesis design Accelerate natural attenuation of groundwater to less than 5 years Pilot-scale 2 Area: 2500 ft , depth: 30 ft 24-25 direct injection points installed in a grid with 5 foot spacing. A total of 7 monitoring wells were installed for process monitoring

PCE: 79 µg/L TCE: 51 µg/L DCE: 23 µg/L

TCE: 540 µg/L DCE: ND VC: 0.5 µg/L

Hurlburt Field Tallahasse, FL

Jan. 1999

TCE, DCE TCE: 9.5 mg/L DCE: 2 mg/L VC: <100 µg/L

The intermediate aquifer consists of sand, silty sand, and discontinuous lenses of clay and is located from 40 to 50 feet below ground surface.

TCE: 1 mg/L DCE: 8.5 mg/L VC: 1.5 mg/L

Within 1 year of injection TCE concentrations had decreased by approximately 89% while DCE, VC, and ethene concentrations increased by approximately 325%, 1,400%, and 400% respectively. These concentration changes are representative of one well installed approximately 5 feet downgradient of the injection area. The highest concentrations detected during the baseline event were detected at this well.

Industrial Site NJ

PCE Pilot-scale PCE: 5.28 mg/L TCE: 227 µg/L 23 injection points using

PCE: 775 µg/L TCE: 866 µg/L DCE: 3.35 mg/L VC: ND

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level DCE: 10 µg/L PCE, TCE PCE: 9 mg/L TCE: 1.4 mg/L DCE: 560 µg/L Description of Geology Remedial Objective Treatment System Design Geoprobe rig Feasibility study Pilot-scale Polymer injected at nine locations using Strataprobe rig Removal Efficiency/Post treatment concentrations PCE: 500 µg/L TCE: 680 µg/L DCE: 17 mg/L Substrate Cost Comments Contacts/ References

62

Closed Industrial Facility CO

Sept. 1999 1 year

Site 1, Santa Clara County Santa Clara County, CA

Dec. 1999/Sept. 2001

PCE, TCE PCE: 27 µg/L TCE: 11 mg/L DCE: 595 µg/L VC: <50 µg/L

Site 2, Santa Clara County Santa Clara County, CA

Oct. 2000/June 2001

PCE, TCE, DCE PCE: 220 µg/L TCE: 820 µg/L DCE: 3 mg/L VC: 300 µg/L

20-25' Unconsolidated silt and clay with occasional lenses of fine sand and sandy clay; severely weathered claystone Interbedded layers of clay, silty sand, and sand (clay is predominant soil type above water-bearing zone) Interbedded layers of clay, silty sand, and sand (clay is predominant soil type above water-bearing zone)

Within first month of HRC injection, PCE concentrations decreased by up to 97% in the source zone.

Accelerate groundwater restoration Pilot-, Full-scale Injection by Geoprobe rig

PCE: <20 µg/L TCE: 71 µg/L DCE: 5.23 mg/L VC: 14 µg/L

Accelerate groundwater restoration in source area Pilot-, Full-scale Injection using Geoprobe rig. First, HRC-primer (fast-release) applied, and then the standard polymer (slowrelease) is injected few inches away. Accelerate groundwater restoration at hotspots Full-scale 2 Area: 4100 ft , depth: 15 ft 33 direct injection points. HRC was injected at a rate of 5 pounds per foot from 5 to 20 feet below ground surface. Groundwater remediation Full-scale 2 Area: 2000 ft

PCE: 23 µg/L TCE: 940 µg/L DCE: 1.5 mg/L VC: 1.4 mg/L

Additional injections to source area after 9 months. Both fast and slow release HRC products were shown to stimulate anaerobic biodegradation of CAHs

Tosco Manufacturing Facility Burien, WA

Aug. 2000

PCE, TCE, DCE PCE: 11.4 mg/L TCE: 1.7 mg/L

30 feet of dense clay overlying approximately 20 feet of sand

PCE: 3.1 mg/L TCE: 2.5 mg/L

Within 159 days following injection the PCE concentration in one well was reduced by 73% while the TCE concentration in the same well increased by 44%. DCE and VC data for this site were not provided.

Strip Mall Dry Cleaner Kent, WA

Dec. 1998

PCE, TCE PCE: 67.4 mg/L TCE: 11.7 mg/L

PCE: 34 µg/L TCE: 17 µg/L DCE: 20 µg/L VC: 70 µg/L

PCE and TCE concentrations had been reduced by nearly 100% within 14 months of injection at one

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level DCE: 360 µg/L VC: 60 µg/L Description of Geology Remedial Objective Treatment System Design Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

63

55 direct injection points and three monitoring wells.

Dayco Manufacturing Facility Eldora, IA

Feb. 1998 35 weeks

TCE TCE: 2.23 mg/L DCE: 5.03 mg/L VC: 134 µg/L

Feasibility study for HRC in a barrier configuration Pilot-scale 2 Area: 250 ft , 15 ft HRC was emplaced using 4 foot long perforated poly carbonate canisters. A total of 4 canisters were installed successively in four treatment event spanning 1 week. A total of 8 monitoring wells were used for process monitoring activities. Feasibility study to compare anaerobic and aerobic degradation of DCE and VC Pilot-scale 2 Area: 450 ft , depth 30 ft ORC and HRC tested in two identical side by side field applications Two pilot test arrays consisting of 4 injection wells and 3 monitoring wells per array DNAPL and source area remediation Pilot-scale 2 Area: 45000 ft , depth: 3 ft A residual source area was treated with 5 injection points

TCE: 560 µg/L DCE: 1.71 mg/L VC: <1 µg/L

source area well. DCE and VC concentrations had been reduced by 44% and 88% respectively in one downgradient well within 2 years of injection. Within 35 weeks of injection TCE, DCE, and VC concentrations were reduced by approximately 75%, 66%, and nearly 100% respectively. These reductions were observed in one well approximately 5 feet downgradient from the injection well. Contaminant concentration data from wells further downgradient from the injection area were not provided. Original COC was TCE but nearly all TCE mass has been reduced naturally to DCE and VC. TCE has not been detected onsite at concentrations exceeding the MCL for several years. Within 180 days of injection DCE and VC concentrations in one well 25 feet downgradient from the injection area were reduced by approximately 97% and 94% respectively. Within 18 months after the pilot scale injection PCE and TCE concentrations had decreased by 99% while DCE and VC concentrations had increased significantly in one well.

Moen Manufacturing Facility Elyria, OH

July 1999 180 days pilot tests with design of full scale application to follow

DCE, VC DCE: 590 µg/L VC: 210 µg/L

DCE: 20 µg/L VC: 10 µg/L

Springdale Drycleaners Portland, OR

Dec. 1999 2-3 years

PCE, TCE PCE: 98 mg/L TCE: 35.9 mg/L DCE: <5 µg/L VC: <5 µg/L

PCE: <250 µg/L TCE: 300 µg/L DCE: 43.9 mg/L VC: 9.5 mg/L

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design (80 lb HRC per point) and the downgradient dissolved phase plume area was treated with 22 injection points (120 lb HRC per point). Groundwater remediation Pilot-scale Area: 4300 ft , depth: 5 ft A single line of 43 injection points were installed at 10 foot intervals. Two monitoring wells (10 feet upgradient and 10 feet downgradient) were used to monitor performance. Groundwater remediation of perchlorate Full-scale 2 Area: 1200 ft . depth: 4 ft 25 injection points installed in a grid on 5 foot centers. 6 monitoring wells installed for process monitoring.
2

64
Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

Reichhold Chemicals Tacoma, WA

Chlorinated Phenols PCP: 1.6 mg/L TetraCP: 200 µg/L TriCP: 50 µg/L DiCP: 50 µg/L

Approximately 400 feet of fine grained marine sediments

PCP: 540 µg/L TetraCP: 100 µg/L TriCP: 50 µg/L DiCP: 50 µg/L

Within 280 days after injection PCP and 2,3,4,6-TCP concentrations had been reduced by approximately 66% and 50% respectively at one monitoring well installed approximately 10 downgradient from the line of injection wells.

Acton Mickelson Ordinance Facility Hollister, CA

Dec. 2000

Perchlorate, Hexavalent Chromium, Freon 113 Perchlorate: 7.5 mg/L CrVI: 160 µg/L Freon 113: 250 µg/L

Fine to medium silty sand

Perchlorate: 1 mg/L CrVI: <10 µg/L Freon 113: 20 µg/L

Pueblo Chemical Depot SWMU-14 Pueblo, CO

Feb. 2000

2,4,6-TNT 1,3,5-TNB 2,4-DNT RDX 2,4-DNT 0.0037 mg/L RDX - 0.0093 mg/L 1,3,5-TNB 0.231 mg/L Hydrazine 0.001 mg/L

Sand and clayey sand overlain by a clay to silty sand confining or semiconfining unit.

Groundwater remediation, plume containment Pilot-scale 2 Area: 350 ft , depth HRC was injected into 30 locations installed in a line. A total of 15 monitoring wells were installed for process monitoring sampling.

2,4-DNT 0.0003 mg/l RDX - < 0.0001 mg/L 1,3,5-TNB 0.005 mg/L Hydrazine – NA

Within 80 days after injection perchlorate, hexavalent chromium, and freon1113 concentrations had been reduced by approximately 87%, nearly 100%, and 92% respectively at one well installed within the injection grid. No analytical data was provided for wells installed downgradient from the injection area. Within approximately 105 days following injection 2,4-DNT, RDX, and 1,3,5TNB concentrations had decreased by approximately 89%, 99%, and 98% respectively. These reduction calculations represent average concentrations calculated from 4 downgradient monitoring wells. Concentration data was provided for only the 4 wells closest to the injection area (approximately 30

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

65

Pueblo Chemical Depot SWMU-14 Pueblo, CO

Feb. 2001

TCE NA

Alluvial aquifer

Groundwater restoration Pilot-scale Barrier configuration

NA

Manufacturing Facility Crozet, VA

Dec. 1999

TCE TCE: 225 µg/L DCE: 125 µg/L VC: <5 µg/L

Fractured crystalline bedrock

Feasibility study for a bedrock site Pilot-scale 2 Area: 4900 ft , depth: 15 ft 3 injection wells spaced approximately 5 feet apart with one monitoring well installed approximately 10 feet downgradient from the injection wells. Feasibility study Pilot-scale 2 Area: 4900 ft , depth: 15 ft 15 direct push points installed in a barrier configuration approximately 5 feet apart. Feasibility study Pilot-scale Three 3.5-inch by 4-foot long PVC canisters filled with gelled HRC were installed in a single 4-inch well. An unknown number of monitoring wells were installed for process monitoring.

TCE: <5 µg/L DCE: 8 µg/L VC: 50 µg/L

Manufacturing Facility NJ

TCE NA

NA

feet downgradient) out of a total of 15 monitoring wells used for process monitoring. Contaminanat data was minimal during first 3 months of pilot test. However, and order of magnitude reduction in TCE concentration was observed 50 feet downgradient of treatment zone after 11 months. Within 13 months of injection TCE and DCE concentrations had decreased by approximately 98% and 94% respectively while VC and ethene concentrations had increased by approximately 900% in one monitoring well installed approximately 10 feet downgradient from the injection wells. Contaminant concentration data was not provided.

Flemington Manufacturing Facility Flemington, NJ

March 2001

PCE, TCE, DCE PCE: 110 µg/L TCE: 100 µg/L DCE: 50 µg/L VC: NA

Very dense till consisting of clayey silts and some sand

PCE: 4 µg/L TCE: 9 µg/L DCE: 22 µg/L VC: NA

Within 80 days of injection PCE, TCE, and DCE concentrations had been reduced by approximately 64%, 10%, and 56% respectively in one monitoring well installed an unknown distance downgradient from the emplacement area. This application involved

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

66

Japanese Electrical Plant Japan

2000 1 year

PCE, TCE, DCE NA

Groundwater remediation Pilot-scale 2 Area: 500 ft , depth: 7 ft 7 injection points in two rows spaced approximately 13 feet apart. Row spacing was also approximately 13 feet. Two monitoring wells were installed, 1 upgradient and 1 downgradient. Groundwater remediation Pilot-scale

NA

installing canisters of gelled HRC in a well as a slowly soluble and therefore longer term HRC source. This site is very similar to the Central US Manufacturing Facility application. HRC accelerated the natural attenuation process.

Coopervision Manufacturing Facility Scottsville, NY Invensys Control Old Saybrook CT

July 2001

1,1,1-TCA NA

NA

Efforts to inject using direct push were unsuccessful. Re-injection using mud rotary is planned

Pilot - Dec. 1999 Full – Sept. 2000

PCE PCE: 13.9 mg/L

Groundwater restoration to facilitate property transfer Pilot-, Full-scale Full - 55 injection points in an unknown configuration, unknown number of monitoring points.

PCE: 10 µg/L

Manufacturing Facility Brighton, NY

TCE NA

Groundwater Restoration Pilot-scale 2 Area: 560 ft

NA

Within 9 months after the pilot scale injection PCE concentrations had decreased by 99% in one well installed an unknown distance from the injection area. No other data has been provided for this site. Electron donor injection was a viable remedial approach for chlorinated VOCs in this low permeability application. 80% of PCE mass was removed. Conditional Closure has been granted.

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

67

Arlington Cleaners Arlington, TX

May 2000

PCE, TCE, DCE, VC PCE: 4.5 mg/L TCE: 1 mg/L DCE: 7.3 mg/L VC: 870 µg/L

24 injection points installed in a grid with 5 foot spacing. 4 monitoring wells installed for process monitoring. Groundwater remediation under voluntary cleanup program Full-scale 2 Area; 3000 ft , depth: 15 ft 45 direct injection points. 16 of the points were installed at a 15 to 30 degree angle to inject beneath a preexisting structure. An unknown number of monitoring wells were installed for process monitoring.

PCE: 410 µg/L TCE: 90 µg/L DCE: 440 µg/L VC: 130 µg/L

Within 18 months after the pilot scale injection PCE, TCE, DCE, and VC concentrations had decreased by 98%, 91%, 94%, and 85% respectively at one well installed within or downgradient from the injection area. During this application 1/3 of the injection wells were installed at a 15 to 30 degree angle to inject HRC under an existing building. Closure under the TNRCC VCP Program.

Slow-Release Substrates – Vegetable Oil Former Zipper April 2000 PCE, TCE, DCE Manufacturing Facility PCE: 120 mg/L TCE: 8 mg/L Newport News, DCE: 14 mg/L VA VC: 1.3 mg/L Hangar K, Cape Canaveral AFS Cape Canaveral, FL Site N-6, Former NSA Mid-South Millington, TN Pilot – June 1999 Full – July 2000 PCE, TCE, DCE PCE: 1.8 mg/L TCE: 260 mg/L DCE: 78 mg/L VC: 950 µg/L TCE, DCE, CT TCE: 1.8 mg/L DCE: 34 µg/L VC: ND Pilot – April 1999 Full – Dec. PCE, TCE PCE: 1 mg/L Sand and silty sand with

Risk-based closure Pilot-scale

PCE: 34 mg/L TCE: 11 mg/L DCE: 27 mg//L VC: 5.3 mg/L

Technology demonstration Pilot-, Full-scale

PCE: 48 µg/L TCE: 310 mg/L DCE: 190 mg/L VC: 13 mg/L

Aug. 2000

Site SS-015, Travis AFB

Fluvial deposits from 40 to 90 feet bgs consisting of sand, silty sand, and clay stringers Silty clay with thin layers of silt and silty

Field feasibility study Pilot-scale

TCE: 1.2 mg/L DCE: 250 µg/L VC: ND

Technology demonstration Pilot-, Full-scale

PCE: 430 µg/L TCE: 2.2 mg/L DCE: 4 mg/L

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status 2000 Contaminants Concentrations level TCE: 4.2 mg/L DCE: 13 mg/L VC: 17 mg/L PCE PCE: 340 µg/L TCE: 13 µg/L DCE: 46 µg/L VC: ND PCE, TCE, VC TCE: 6.7 mg/L DCE: 220 µg/L VC: ND Description of Geology Remedial Objective Treatment System Design sand Removal Efficiency/Post treatment concentrations VC: 550 µg/L Substrate Cost Comments Contacts/ References

68

Travis AFB, CA

Site FF-87, Former Newark AFB Newark, OH

Sept. 2001

Naval Industrial Reserve Ordnance Plant, Fridley Fridley, MN

Nov. 2001

Alluvial medium to coarse sand underlain by Pleistocene glacial silty clay Glacial-fluvial medium to coarse grained sand

Accelerate groundwater restoration Full-scale

PCE: 740 µg/L TCE: 8 µh/L DCE: 36 µg/L VC: 20 µg/L

Field feasibility study Pilot-scale

TCE: 6.2 mg/L DCE: 1.8 mg/L VC: 2 µg/L

At approximately 5 months after injection TCE concentrations were approximately the same as baseline conditions. However, cis-1,2-DCE and ethane concentrations detected at 5 months post injection were substantially higher than the baseline event.

Former Radiator Facility IL Site SS-17 Altus AFB, OK

Sept. 2000

TCE TCE: 1.3 mg/L DCE: 7.4 mg/L VC: 220 µg/L TCE, DCE TCE: 1.66 mg/L DCE: 900 µg/L VC: 440 µg/L

Clayey sand

Field feasibility study Pilot-scale

TCE:< 1mg/L DCE: 41 mg/L VC: 8.3 mg/L

Nov. 2001

Reddish brown, moderately plastic, sandy clay to 15 feet below ground surface underlain by clayey shale with occasional gypsum layers.

Field feasibility study Bench-, Pilot-scale

TCE: 650 µg/L DCE: 1.07 mg/L VC: 790 µg/L

OU-1 Landfill 3 Altus AFB, OK

Nov. 2001

TCE, DCE TCE: 10.4 mg/L DCE: 500 µg/L

Reddish brown, moderately plastic, sandy clay to 15 feet

Field feasibility study Pilot-scale

TCE: 6.44 mg/L DCE: 800 µg/L VC: 240 µg/L

Within approximately 5 months the TCE maximum concentration had been reduced by approximately 61%. DCE, VC, and ethene maximum concentrations increased by approximately 19%, 80%, and 1,470% respectively. TCE concentration reductions were observed in all of the injection wells and monitoring wells with per well reductions ranging from 13 to 99%. Within approximately 5 months the TCE maximum concentration had been reduced by approximately

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level VC:< 120 µg/L Description of Geology Remedial Objective Treatment System Design below ground surface underlain by clayey shale with occasional gypsum layers. Removal Efficiency/Post treatment concentrations Substrate Cost Comments Contacts/ References

69

Whiteman Air Force Base LF-08 Whiteman AFB, MO

July 2002 2 years

TCE, DCE, VC TCE: 2.1 mg/L DCE: 33 µg/L VC: ND

Approximately 24 feet of dense clay overlying a clayey gravel zone which extends to the top of weathered claystone bedrock at 30 feet below ground surface. Stiff to very stiff, reddish brown, low plasticity, silty clay Reddish brown silty clay

Field feasibility study Pilot-scale

NA

37%. DCE, VC, and ethene maximum concentrations increased by approximately 60%, 80% and an order of magnitude, respectively. TCE concentration reductions were observed in all wells except injection well 6, with per well reductions ranging from 23 to 92%. The first process monitoring round is scheduled for October 2002.

Solid Substrates – Bark Mulch Building 301, Pilot – Jan. Offutt AFB 1999 Full – July Omaha, NE 2001 OU1 Landfill 3 Altus AFB, OK July 2000

TCE TCE: 1.9 mg/L DCE: 270 µg/L VC: 2 µg/L TCE, DCE Upgradient well TCE: 6.2 mg/L DCE: 850 µg/L VC: ND Perchlorate

Field feasibility study to accelerate groundwater restoration Pilot-, Full-scale Full-scale application to prevent surface water discharge and prevent potential off-base migration Full-scale Groundwater restoration for offsite migration via a permeable reactive biobarrier Bench-, Pilot-, Full-scale

TCE: 1.22 mg/L DCE: 98 µ/L VC: 4 µg/L

61 percent reduction in mean TCE concentration downgradient of biowall

Biowall well MP01 TCE: 48 µg/L DCE: 640 µg/L VC: ND

Concentration data after 4 weeks of installation. Limited evidence of degradation of TCE to DCE.

Naval Weapons Industrial Reserve Plant McGregor, Texas

2000

Full-scale System Installed

Solid Substrates – Chitin Distler Brickyard Oct. 2001

TCE, cDCE, VC

Relatively low

Groundwater remediation to

TCE: ND

Intermittent wetting of

State-Of-The-Art-Report: Enhanced In Situ Bioremediation Technologies CG 6 Technical Reliability
Site Name/Location Application Date/Project Status Contaminants Concentrations level Description of Geology Remedial Objective Treatment System Design permeability silts and clays potable water standards Pilot-scale Removal Efficiency/Post treatment concentrations DCE: 58 µ/L VC: 9 µ/L Substrate Cost Comments Contacts/ References

70

Superfund Site Louisville, KY Gaseous Hydrogen Injection Launch Complex Feb. 1999 15 18 months Cape Canaveral, FL TCE: 7 µg/L DCE: 90 µ/L VC: 16 µg/L PCE, TCE PCE: 87 mg/L DCE: 370 mg/L VC: 52 mg/L

contaminant zone by infiltration

Offutt AFB Omaha, NE Twin Cities Army Ammunition Plant New Brighton, MN

Nov. 1998

DCE DCE: 430 µg/l

Silica sand with some shell fragments containing clay and organic matter to 70 feet bgs Sand

Technology demonstration Pilot-scale

TCE: 66 mg/L DCE: 270 mg/L VC: 67 mg/L

Technology demonstration Pilot-scale Technology demonstration for use of hollow fiber membranes to deliver hydrogen in situ Bench-, Pilot-scale

DCE: < 5 µg/L

50% ( at 15 feet from injection point ) to 90% (at 3 to 6 feet from injection point) reduction in total chlorinated ethenes in treatment zone over 18 months. Small scale pilot test, complete cis-1,2-DCE (over 99%) removal. A decline in concentrations of TCE and DCE was observed, with an increase in VC and ethene after a lag period.

Aug. 2000 20 months

TCE NA

NA