United States Office of Research and EPA/625/K-96/001

Environmental Protection Development May 1996

Agency Washington, DC 20460

1EPA Seminars

Bioremediation of

Hazardous Waste Sites:

Practical Approaches to

Implementation

May 29--30, 1996----Chicago, IL

June 4--5, 1996----Kansas City, MO

June 6--7, 1996----Atlanta, GA

June 18--19, 1996----San Francisco, CA
EPA/625/K-96/001

May 1996

Seminars on

Bioremediation of Hazardous Waste Sites:

Practical Approaches to Implementation

Office of Research and Development

U.S. Environmental Protection Agency

Washington, DC
Notice

Mention of trade names or commercial products does not constitute endorsement or recommendation for use.

ii
Contents

Background Information for Bioremediation Applications ............................... 1-1

Bioventing ................................................................ 2-1

Bioremediation of Sediments ................................................... 3-1

Aerated Lagoons: A Case Study ................................................ 4-1

Oil-Contaminated Shorelines .................................................. 5-1

Land Treatment ............................................................ 6-1

Land Treatment Unit Case Study:

Champion International Superfund Site ........................................... 7-1

Phytoremediation ........................................................... 8-1

Development and Application of Composting Techniques for Treatment of

Soils Contaminated With Hazardous Waste ........................................ 9-1

Biopile Treatment of Soils Contaminated With Hazardous Waste ........................ 10-1

Effective Treatment of Hazardous Waste Constituents in Soil by

Lignin-Degrading Fungi ..................................................... 11-1

Slurry Bioreactors for Treatment of Contaminated Soils,

Sludges, and Sediments ..................................................... 12-1

Fixed Film Bioreactors ...................................................... 13-1

Suspended Growth Bioreactors ................................................ 14-1

Natural Attenuation of Ground Water ........................................... 15-1

Natural Attenuation of Soils .................................................. 16-1

Natural Attenuation of Landfills ................................................ 17-1

Natural Attenuation of Sediments .............................................. 18-1

Source Control: Free Product Recovery and Hydraulic Containment ...................... 19-1

Air Sparging/Air Injection .............................................. 19-5

iii
State Review: Natural Attenuation of Ground Water and Soils .......................... 20-1

Monitoring .............................................................. 21-1

Modeling ............................................................... 22-1

iv
Sources of Information

Recent EPA Bioremediation Publications

http://www.epa.gov/docs/ORD

Bioremediation in the Field Bulletin

Latest edition EPA/540/N-96/500

Bioremediation in the Field Search System: Database on national and some international field

applications 

Version 2.0 EPA/540/R-95/508b

Also on the Internet

Request to be on EPA's bioremediation mailing list or to request specific bioremediation documents

513-569-7562

NRMRL/SPRD Home Page

http://www.epa.gov/ada/kerrlab.html

v
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

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Background Information for Bioremediation Applications

Ronald C. Sims

Utah State University, Logan, UT

Introduction

This technology transfer seminar series is sponsored by the U.S. Environmental Protection Agency's

(EPA's) Biosystems Program. The Biosystems Program coordinates research, development, and

evaluation of full-scale bioremediation activities. The seminar series provides participants with

state-of-the-art information on the practical aspects of implementing bioremediation. The series is

divided into the following sections:

# Background for Bioremediation Applications

# In Situ Treatment of Soils, Sediments, and Shorelines

# Ex Situ Treatment With and Without a Reactor

# Natural Attenuation

# Treatment of the Subsurface

Each section includes discussion of advantages and limitations, materials handling, types of waste

amenable to the treatment process, pre- and posttreatment requirements, and capital and operation

and maintenance costs. The overall focus is on field applications in use today, with some

information on processes that are nearing readiness for field use.

This section has been organized to address the following topics:

# Biodegradation and metabolism

# Environmental factors affecting biodegradation

# Site characterization

# General concept of treatability studies

Biodegradation and Metabolism

Biodegradation involves chemical transformations mediated by microorganisms that satisfy

nutritional requirements, satisfy energy requirements, detoxify the immediate environment, or occur

fortuitously such that the organism receives no nutritional or energy benefit (1). Mineralization is the

complete biodegradation of organic materials to inorganic products, and often occurs through the

combined activities of microbial consortia rather than through a single microorganism (2). Cometabolism is the partial biodegradation of organic compounds that occurs fortuitously and that

does not provide energy or cell biomass to the microorganism(s). Co-metabolism can result in

partial transformation to an intermediate that can serve as a carbon and energy substrate for

microorganisms, as with some hydrocarbons, or can result in an intermediate that is toxic to the

transforming microbial cell, as with trichloroethylene (TCE) and methanotrophs.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

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Two classes of biodegradation reactions are aerobic and anaerobic. Aerobic biodegradation

involves the use of molecular oxygen (O ), where O (the "terminal electron acceptor") receives 2 2

electrons transferred from an organic contaminant:

organic substrate + O 6 biomass + CO + H O + other inorganics 2 2 2

Thus, the organic substrate is oxidized (addition of oxygen), and the O is reduced (addition of 2

electrons and hydrogen) to water (H O). In this case, the organic substrate serves as the sources 2

of energy (electrons) and the source of cell carbon used to build microbial cells (biomass). Some

microorganisms (chemoautotrophic aerobes or lithotrophic aerobes) oxidize reduced inorganic

compounds (NH , Fe , or H S) to gain energy and fix CO to build cell carbon: 3 2 2

+2

NH3 (or Fe+2 or H2S) + CO2 + H2 + O2 6 biomass + NO3 (or Fe+3 or SO4) + H2O

At some contaminated sites, as a result of consumption of O2 by aerobic microorganisms and slow

recharge of O , the environment becomes anaerobic (lacking O ), and mineralization, 2 2

transformation, and co-metabolism depend upon microbial utilization of electron acceptors other

than O (anaerobic biodegradation). Nitrate (NO ), iron (Fe ), manganese (Mn ), sulfate (SO ), 2 3 4

+3 +4

and carbon dioxide (CO2) can act as electron acceptors if the organisms present have the

appropriate enzymes (3). JP-4 jet fuel constituents were observed to be biodegraded in the presence

of NO3 as the electron acceptor (4). Iron and manganese are important microbial electron

acceptors, with background concentrations in soils ranging from 20 to 3,000 mg/kg for Mn and

3.8 to 5.2 percent for iron. An evaluation of the degradation of polycyclic aromatic hydrocarbons

(PAHs) in aerobic and anaerobic environments was conducted based on thermodynamic principles

(5). Biodegradation of pentachlorophenol (PCP) has been observed to increase the presence of

added Mn (6).

Halogenated compounds can be used as growth substrates or co-metabolized by aerobic and

anaerobic microorganisms. Dehalogenation can be spontaneous, as in the loss of halogens during

ring cleavage, or enzymatically catalyzed through hydrolytic cleavage or reductive dehalogenation

(1). Halogenated compounds can often serve as the electron acceptor and become reduced in

environments where there is a source of electrons; for example, under methanogenic conditions

(production of methane in reduced environments) reductive dehalogenation of perchloroethylene

(PCE) to TCE, trans-1, 2-dichloroethylene (DCE), vinyl chloride, and ethylene occurs (1). In such

situations, alternative electron acceptors such as NO3 and SO4 may compete with the halogenated

compounds for electrons. TCE can also be biodegraded co-metabolically in an aerobic

environment by methanotrophs when methane is added to cause the formation of TCE-epoxide,

which will abiotically transform to dichloroacetic acid, TCE-diol, formic acid, and glyoxylic acid.

Reduced dehalogenated intermediates often undergo rapid biodegradation by aerobic

microorganisms in the presence of O (7). 2
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

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Environmental Factors Affecting Biodegradation

Microbial ecologists have identified ranges of critical environmental conditions that affect the activity

of soil microorganisms (Table 1). Many of these conditions are controllable and can be changed

to enhance the biodegradation of organic constituents. A discussion of the factors identified below,

including principles, status of the technology, secondary impacts, equipment, advantages and

disadvantages, and references is provided in the document Handbook on In Situ Treatment of

Hazardous Waste-Contaminated Soils (7).

Table 1. Critical Environmental Factors for Soil Microbial Activity (8).

Environmental Factor Optimum Levels

Oxygen Aerobic metabolism: greater than 0.2 mg/L

dissolved oxygen, minimum air-filled pore

space of 10%

Anaerobic metabolism: less than 0.2 mg/L

dissolved oxygen, O2 concentration less than

1% air-filled pore space

Nutrients Sufficient nitrogen, phosphorus, and other

nutrients so not limiting microbial growth

(suggested C:N:P ratio of 120:10:1)

Moisture Unsaturated soil: 25-85% of water holding

capacity, -0.01 MPa; will affect oxygen

transfer into soil (aerobic status); 

in saturated zone, water will affect transport

rate of oxygen and therefore will affect rate

of aerobic remediation

Environment (pH) 5.5-8.5

Environment (redox) Aerobes and facultative anaerobes: greater

than 50 millivolts; Anaerobes: less than 50

millivolts

Environment (temperature) 15-45EC (mesophilic)

Oxygen diffuses into the soil from the air above it, and gases in the soil atmosphere diffuse into the

air. Oxygen concentration in a soil may be much less than in air, however, while CO2

concentrations in soil may be orders of magnitude higher than in air. A large fraction of the

microbial population within the soil depends on oxygen as the terminal electron acceptor in

metabolism. When soil pores become filled with water, the diffusion of gases through the soil is

restricted since oxygen diffuses through air 10,000 times faster than through water. Oxygen may

be consumed faster than it can be replaced by diffusion from the atmosphere, and the soil may

become anaerobic. Facultative anaerobic organisms, which can use oxygen when it is present or

switch to alternative electron acceptors such as nitrate in the absence of oxygen (e.g., denitrifying
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

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bacteria), and obligate anaerobic organisms become the dominant populations. Additional

information concerning in situ anaerobic bioremediation can be found elsewhere (7).

Oxygen concentrations in soil systems may be increased by tilling and draining unsaturated soil, for

example, in prepared-bed land treatment systems, in ex situ treatment (e.g., composting, biopiles,

and fungal treatment) and in situ treatment systems, and through the application of bioventing

systems, where air is forced through a soil system and carries oxygen to soil microorganisms to

accomplish aerobic degradation. Hinchee (9) and Hinchee and Downey (10) successfully applied

bioventing for enhancement of biodegradation of petroleum hydrocarbons in JP-4 jet fuel

contaminated soil at Hill Air Force Base, Ogden, Utah, by increasing subsurface oxygen

concentrations. Oxygen and CO2 concentrations were monitored and correlated well with

hydrocarbon biodegradation. A minimum criterion for aerobic biodegradation of PAH in creosotecontaminated soil was established at 2 percent O in air (11). 2

Within saturated environments, oxygen transport is considered to be the rate-limiting step in aerobic

bioremediation of contaminated hydrocarbons when adequate nutrients are present. At the Traverse

City, Michigan, site contaminated with jet fuel (12), an increase in the oxygen concentration in water

through addition of hydrogen peroxide and was observed to positively affect the rate of

biodegradation of the jet fuel components benzene, xylene, and toluene.

Microbial metabolism and growth depend on adequate supplies of essential macro- and

micronutrients. If the wastes present at a site are high in carbonaceous materials and low in

nitrogen (N) and phosphorus (P), the subsurface may become depleted of available N and P

required for biodegradation of the organic contaminants. Addition of nutrients may be required as

a management technique to enhance microbial degradation, and can be used to treat water from

a pump-and-treat system and applied through reinfiltration or irrigation (13). Recommended ratios

for subsurface systems of carbon (C), N, and P are 120:10:1 on a weight basis. Nutrients have

been added to enhance microbial degradation of hydrocarbon contaminants at many sites (14). At

the Champion International Superfund Site in Libby, Montana (15), nutrients are added to enhance

bioremediation in a prepared-bed land treatment system, in an aboveground reactor for treating

extracted ground water, and in injection wells for in situ bioremediation of PAH and PCP.

Moisture content and the soil water matrix potential against which microorganisms must extract

water from the soil regulate their activity. The soil matrix potential is the energy required to extract

water from the soil pores to overcome capillary and adsorptive forces. Soil water also serves as the

transport medium through which many nutrients and organic constituents diffuse to the microbial

cell, and through which metabolic waste products are removed. Soil water also affects soil aeration

status, nature, and amount of soluble materials; soil water osmotic pressure; and the pH of the soil

solution (8). Generally, microbial activity measured as biodegradation rates and rates of

detoxification of contaminants in soil have been found to be highest at soil moisture contents of 60

to 80 percent of field capacity (8). Field capacity is the amount of water held against the force of

gravity, generally equal to 0.1 to 0.3 atmospheres of force.

Soil moisture can be increased using standard agricultural irrigation practices such as overhead

sprinklers or subirrigation. To remove excess water or lower the water table to prevent

water-logging, drainage or well point systems can be used. Also, the addition of vegetation to a site

will increase evapotranspiration (ET) of water and will also retard the downward migration of water

(i.e., leaching) (7, 16). 
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

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Other environmental factors, including pH, redox potential, and temperature, are important

parameters that will affect the rate and extent of bioremediation in unsaturated and saturated

subsurface systems. Outside the pH range of 5.5 to 8.5, microbial activity is generally decreased.

Maintaining soils near neutral pH is most often recommended for enhanced bioremediation (7);

however, acidic soils are known to become colonized by fungi over time. Conventional agricultural

practices for increasing soil pH include adding lime periodically and mixing the lime with the acidic

soil (7).

Redox potential of a subsurface environment has an influence on microbial metabolism and activity

(5). For aerobic metabolism the redox potential should be greater than 50 millivolts, for anaerobic

conditions less than 50 millivolts. At low redox potentials, alternative electron acceptors to oxygen

(e.g., nitrate, iron, manganese, and sulfate) act as electron acceptors. A redox potential higher than

50 millivolts is conducive to biodegradation of hydrocarbons. A redox potential of less than 50 is

condusive to degradation of chlorinated hydrocarbons (7).

Soil temperature has an important effect on microbial activity and has been correlated with

biodegradation rates of specific organic compounds (12). Prepared-bed land treatment and in situ

bioremediation should be planned to take advantage of the warm season in cooler climates.

Vegetation can act as an insulator against heat loss and limit frost penetration. Application of

mulches can help control heat loss at night and heat gain during the day (7, 12).

Site Characterization

A contaminated site is a system generally consisting of four phases: 1) solid, which has an organic

matter component and an inorganic mineral component composed of sand, silt, and clay, 2) oil

(commonly referred to as nonaqueous phase liquid, or NAPL), 3) gas, and 4) aqueous (leachate or

ground water). These phases and compartments need to be characterized with regard to extent and

distribution of contamination as well as potential exposure to human and environmental receptors.

Each phase affects bioavailability, i.e., interactions with microorganisms and exposure to human

health and environmental receptors. Each phase can be a site for biological reactions that results

in the transformation of a parent chemical to CO2, H2O, and other inorganic species through the

process of mineralization, or transformation to intermediates that persist or that react with soil

components to chemically bind to soil and therefore alter the bioavailability of the chemicals. 

Evaluating the extent and distribution of contamination at a site will provide important information

that can be used as a basis to select specific bioremediation technologies that are addressed in this

seminar series, or to select a treatment train that represents a combination of physical/chemical and

biological technologies. If contamination is widespread and low in concentration, then in situ

treatment or natural attenuation may be feasible. Conversely, with high concentrations of

contaminants, soil excavation and placement in a confined treatment facility (CTF) or a land

treatment prepared-bed reactor may be advisable. 

Distribution of contaminants at a site is determined by the physical and chemical properties of the

contaminants and the properties of the site. Contaminant properties will affect whether contaminants

are leachable, volatile, and/or adsorbable, and therefore will indicate which subsurface phases

contain the contaminant(s). Physical phases containing the contaminants require evaluation of
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

1-6

bioremediation potential. When the physical and chemical properties are evaluated within the

context of site characteristics, a site-based waste characterization can be used to identify the

phases/compartments at the site and the chemicals associated with each phase. Additional

information concerning practical aspects of site characterization for bioremediation of contaminated

ground water is available in the document In Situ Bioremediation of Contaminated Ground Water

(17).

General Concept of Treatability Studies

Treatability studies are conducted in laboratory microcosms, at pilot scale, or in the field. EPA,

through the Biosystems Field Initiative, and the Departments of Defense and Energy indicate an

increased emphasis on field-scale evaluation of bioremediation, with a supportive role for

laboratory-scale treatability testing. Parent compounds, intermediates, and electron acceptor

utilization are evaluated. A mass balance conceptual framework for treatability studies, at any scale,

refers to the characterization of the physcial phases in the soil and the determination of the influence

of the phases on the bioavailability and bioremediation of associated target chemicals (18), as

described in the "Site Characterization" section above. 

While in the past the goal for bioremediation implied complete mineralization of chemicals to CO ,2

H2O, and inorganic chemicals, alternative endpoints that are protective of human health and the

environment are currently being evaluated by the Department of Energy, EPA, the National Science

Foundation, and the Office of Naval Research. Treatability studies that examine the bioavailability

of contaminants in waste matrices, potential for toxic effects of intermediate metabolites during the

degradation process, and interactions between waste chemicals and organisms are desired. The

overall goal of treatability studies is to develop a better understanding of factors that threaten

ecosystems and human health and of chemicals and their degradation products during

bioremediation so that the regulatory community can take into consideration the possibility of

alternatives to complete mineralization (19, 20).

References

1. Stoner, D.L. 1994. Biotechnology for the treatment of hazardous waste. Boca Raton, FL:

CRC Press.

2. Shelton, D.R., and J.M. Tiedje. 1984. Isolation and partial characterization of bacteria in

an anerobic consortium that mineralizes 3-chlorobenzoic acid. Appl. Environ. Microbiol.

48:840-848.

3. Sims, R.C. 1990. Soil remediation techniques at uncontrolled hazardous waste sites. J. Air

Waste Mgmt. Assoc. 40(5):703-732.

4. Hutchins, S.R., G.W. Sewell, D.A. Kovacs, and G.A. Smith. 1991. Biodegradation of

aromatic hydrocarbons by aquifer microorganisms under denitrifying conditions. Environ.

Sci. Technol. 25:68-76.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

1-7

5. McFarland, M.J., and R.C. Sims. 1991. Thermodynamic framework for evaluating PAH

degradation in the subsurface. Ground Water 29(6):885-896.

6. Petrie, R.A., J.E. McLean, and R.C. Sims. 1995. Treatment of pentachlorophenol with

manganese oxide addition to biotic and abiotic sediments. Haz. Waste Haz. Mat.

12(3):271-282.

7. U.S. EPA. 1989. Bioremediation of contaminated surface soils. Robert S. Kerr

Environmental Research Laboratory. EPA/600/9-89/073. Ada, OK.

8. U.S. EPA. 1990. Handbook on in situ treatment of hazardous waste-contaminated soils.

EPA/540/2-90/002.

9. Hinchee, R. 1989. Enhanced biodegradation through soil venting. In: Proceedings of the

Workshop on Soil Vacuum Extraction, Robert S. Kerr Environmental Research Laboratory,

Ada, OK (April 27-28).

10. Hinchee, R., and D. Downey. 1990. In situ enhanced biodegradation of petroleum

distillates in the vadose zone. In: Proceedings of the International Symposium on Hazardous

Waste Treatment. Air and Waste Management Association and U.S. EPA Risk Reduction

Engineering Laboratory (February 5-8).

11. Hurst, J., R.C. Sims, J.L. Sims, D.L. Sorensen, and J.E. McLean. 1990. Polycyclic aromatic

hydrocarbon biodegradation as a function of oxygen tension in contaminated soil. J. Haz.

Mat. In press.

12. U.S. EPA. 1991. Site characterization for subsurface remediation. Seminar publication.

EPA/625/4-91/026. Office of Research and Development, Washington, DC.

13. U.S. EPA. 1991. Handbook: Stabilization technologies for RCRA corrective actions.

EPA/625/6-91/026. Office of Research and Development, Washington, DC.

14. U.S. EPA. Bioremediation in the Field Search System (BFSS) database, user documentation.

EPA/540/R-95/508a. Office of Research and Development.

15. U.S. EPA. 1995. Champion International Superfund site, Libby, Montana: Bioremediation

field performance evaluation of prepared bed system, Vols. 1 and 2.

EPA/600/R-95/156a,b.

16. Aprill, W., and R.C. Sims. 1990. Evaluation of the use of prairie grasses for stimulating

polycyclic aromatic hydrocarbon treatment in soil. Chemosphere 20(1-2):253-265.

17. U.S. EPA. 1992. In situ bioremediation of contaminated ground water.

EPA/540/S-92/003. Office of Solid Waste and Emergency Response.

18. Sims, R.C., and J.L. Sims. 1995. Chemical mass balance approach to field evaluation of

bioremediation. Environ. Prog. 14(1):F2-F3.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

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19. Environmental Biotechnology. 1995. In: Biotechnology for the 21st century: New horizons.

National Science and Technology Council. 

20. DOE/EPA/NSF/ONR. 1996. Joint program on bioremediation. Interagency Announcement

of Opportunity. National Center for Environmental Research and Quality Assurance, U.S.

EPA.

21. Hurst, J. 1996. Prepared bed bioremediation as affected by oxygen concentration in soil

gas: Libby, Montana, Superfund site. M.S. Thesis, Department of Civil and Environmental

Engineering, Utah State University, Logan, UT.
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Background

Information for

Bioremediation

Applications

Ronald C. Sims

Utah State University

Logan, UT

Background Information for

Bioremediation Applications

● National Status on Applications

● Biodegradation and Metabolism

● Environmental Factors Affecting

Biodegradation

● Site Characterization

● General Concept of Treatability

Studies

National Status

on Applications

Superfund Remedial Actions

Technologies Selected in FY94

Bioremediation

27%

Flushing

6%

Solvent Extraction

2%

Thermal Desorption

11%

SVE

18%

Incineration

15%

S/S

21%

Superfund Remedial Actions

Technologies Selected in FY89

Bioremediation

13%

Flushing 3%

Solvent Extraction 3%

Thermal Desorption 2%

SVE

24%

Incineration

29%

S/S

24%

Soil Washing 2%

Legislative Authority for Sites

Using Bioremediation

UST

33%

CERCLA

23%

Federal

Facility

17%

Other

13%

RCRA

9% International Government

4%

TSCA 1%

International Private

Sector 0%
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Petroleum

65%

Solvents

12%

Wood Preserving

Wastes 10%

Other

10%

Pesticides/Herbicides

3%

Breakdown of Sites by Type of

Contamination

I

3%

II

5%

III

5%

IV

10%

V

41%

VI

5%

VII

3%

VIII

6%

IX

15%

X

3%

Canada

4%

Distribution of Bioremediation

Projects by Region

Percentage of Sites Treating

Each Medium

Soil

58%

Ground Water

32%

Sediments

6%

Sludge 2%

Air 1%

Surface Water 1%

Breakdown of Processes by

Treatment Technology

(Includes Laboratory-, Pilot-, and Full-Scale)

Ex Situ (with 

reactor)

15%

Ex Situ (without

 reactor)

17%

In Situ

68%

Top 9 Bioremediation Methods

All Other Methods

11%

Fixed Film

4%

Solid Phase, Pile 

Treatment 4%

Attached Growth

5%

Air Sparging 6%

Solid Phase, Prepared Bed Natural Attenuation 6%

11%

Soil Bioremediation

14%

Ground Water

Bioremediation

14%

Bioventing

25%

In Situ Biotreatment Processes

0

20

40

60

80

100

120

140

Air Sparging

Bioventing

Confined

Treatment

Facility forSediments

Ground Water

Bioremediation

Unknown

Treatability

Design

In the Field

Completed
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

In Situ Biotreatment Processes

(continued)

0

5

10

15

20

25

30

35

40

45

In Situ Other

Natural

Attenuation

Sediment

Bioremediation

Soil

Bioremediation

Unknown

Treatability

Design

In the Field

Completed

Biodegradation and

Metabolism

Biodegradation and

Metabolism

● Nutrition

● Energy

● Detoxification

● Fortuitous (co-metabolism)

Chemical transformations

mediated by microorganisms:

Biodegradation

● Biological transformation of an organic compound

to another form without regard to extent

OH

Cl

OH

Cl

OH

m-chlorophenol

3-chlorophenol

m-chlorocatechol

3-chlorocatechol

Mineralization

● Conversion of an organic compound to carbon

dioxide, water, methane, and other inorganic

forms (e.g., Cl–, NH4

+)

■ Aerobic

conditions

■ Anaerobic

(methanogenic)

conditions

OH

Cl

+ O2 CO2 + H2O + Cl– + ATP + Biomass

OH

Cl

 CH4 + CO2 + Cl– + ATP + Biomass

Co-metabolism

CH4 + O2 CH3OH + H2O

TCE + O2 TCE-EPOXIDE + H2O

Methane Methanotrophs Methanol

Methanotrophs

MMO

MMO
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Aerobic Biodegradation

Benzene

+ O2 Biomass + CO2 + H2O

Anaerobic Biodegradation

Toluene Nitrate

+ NO3 CO2 + N2 + H2O

Denitrification

CH3

Anaerobic

Biodegradation

(Reductive

Dechlorination)

of Chlorinated

Alkenes

Cl Cl

Cl Cl

Cl

Cl Cl

Cl

Cl

Cl

Cl

H

H

H

H

H

H

H

H

H H

Cl

H

PCE

TCE

cDCE tDCE

VC

E

H

Environmental

Factors Affecting

Biodegradation

Soil Particle

Nutrients Mass

Transport

Electron

Acceptor

Nutrients Mass

Transport

Electron

Acceptor

Mass transport and toxicity limitations to bioremediation 

as a function of NAPL concentration

Nonaqueous Phase Liquid (NAPL)

(Resistance to mass transport)

Toxicity to Microorganisms

Critical Environmental Factors for

Soil Microbial Activity

Environmental Factor Effects

Oxygen Metabolism: Aerobic/Anaerobic

Degradation Pathways

Nutrients Nitrogen, Phosphorus Activity

Moisture Unsaturated/Saturated Soil

Oxygen Transfer

Environment (pH) 5.5–8.5

Activity

Environment (Redox) Aerobes/Facultative Anaerobes: > 50 mV

Anaerobes: < 50 mV

Degradation Pathways

Environment (Temperature) 15–45°C (Mesophilic)

Activity

Reference: (9)
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Oxygen Supply

Oxygen diffuses through

water at a rate that is 10,000

times less than oxygen

diffuses through air

Mineralization of 14C-pyrene in non-poisoned soil microcosms as a

function of time and oxygen concentration. Error bars represent the least

significant difference of 7.94. Values are the means for triplicate reactors.

Reference: (12)

0 20 40 60 80

0 

10

20

30

40

50

60

0%

2%

5%

10% 

21% 

Time (days) 

% 14C-Pyrene Mineralized

% Oxygen

Mineralization of 14C-pyrene in poisoned soil microcosms as a function of

time and oxygen concentration. Values are the means for triplicate reactors.

Reference: (12)

0 7 14 21 28 35 42 49 56 63 70

0.0 

0.1 

0.2 

0.3 

0.4 

0.5 

0.6 

0.7 

0.8 

0.9 

1.0 

0 % 

2 % 

5 % 

10 %

21 %

Time (days)

% 14C-Pyrene Mineralized

% Oxygen

Mineralization of 14C-PCP in non-poisoned soil microcosms as a function of time and

oxygen concentration. Error bars represent the least significant difference of 4.67%.

0 20 40 60 80

0 

20

40

60

80

0%

2%

5%

10% 

21% 

Time (days) 

% 14C-PCP Mineralization

% Oxygen

Mineralization of 14C-PCP in poisoned soil microcosms as a function of time

and oxygen concentration. Values are the means for triplicate reactors.

0 20 40 60 80

0.0 

0.1 

0.2 

0.3 

0.4 

0.5 

0.6 

0.7 

0.8 

0.9 

1.0 

0%

2%

5%

10% 

21% 

Time (days) 

% 14C-PCP Mineralization

% Oxygen

Environmental Factors

Nutrients: 100:10:1 Weight ratio

Moisture: 60–80% Field capacity

pH: 5.5–8.5

Redox Potential: >50 mV — Aerobic

<35 mV — Dechlorination

Temperature: Adaptation
1-14

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Site Characterization

Solid Phase 

Organic

Matter

Texture 

 Sand

 Silt

 Clay

Gas

 Carbon Dioxide

 Oxygen 

Water

Leachate 

Fluid Phase 

Non-Aqueous Phase Liquids

(NAPLs)

Physical Phases at a Site To Be Considered

for Bioremediation Technologies

General Concept of

Treatability Studies

Treatability Studies

● Field-scale — more emphasis

● Parent compounds

● Intermediates

● Electron acceptors

Mass Balance

Framework

Physical Phases at a Site To Be Considered

For Bioremediation Technologies

Solid Phase 

Organic

Matter

Texture 

 Sand

 Silt

 Clay

Gas

 Carbon Dioxide

 Oxygen 

Water

Leachate 

Fluid Phase 

Non-Aqueous Phase Liquids

(NAPLs)

Treatability Studies

● Alternative endpoints

■ DOE/EPA/NSF/ONR

■ Bioavailability

■ Intermediate metabolites

■ Interactions or chemicals and

organisms

■ Risk impact
1-15

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Intermediate Metabolites

● 1-Hydroxy-2-Naphthoic acid

● 2,3-Dihydroxynaphthalene

Reference: Ginn, J., W.J. Doucette, and R.C. Sims. 1994. Chemical mass balance

approach for estimating fate and transport of polycyclic aromatic metabolites in

the subsurface environment. Polycyclic Aromatic Compounds 5:225-234.

Experimental Design

● Controls: sterile, no treatment,

field background, number?

● Replicates: duplicate or triplicate?

all time points? all controls?

● Treatments: what are the questions

you want answered?

● How are you going to optimize the

degradation process?

Experimental Design (continued)

● Treatment time: how long should

the study be performed?

● Types of analysis: bulk

measurements? waste specific?

● Data reduction: raw data?

massaged data? QC/QA?

● Cost considerations: how will it

limit scope of test? Time Concentration ( )

Relative Toxicity ( )

Distribution of 14C in Non-poisoned

Microcosms Spiked With 14C-Pyrene

Oxygen % 14C % 14C Soil % 14C Mass

Conc. Mineralized Bound Recovered

 0% 13 8 91

 2% 54 15 91

 5% 52 16 88

10% 51 14 86

21% 46 15 86

Reference: (12)

Distribution of 14C in Poisoned

Microcosms Spiked With 14C-Pyrene

Oxygen % 14C % 14C Soil % 14C Mass

Conc. Mineralized Bound Recovered

 0% <0.2 9 95

 2% <0.2 9 91

 5% <0.2 11 89

10% <0.2 12 90

21% <0.2 8 97

Reference: (12)
1-16

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Bioremediation Applications

Contaminated Site Characterization

Contaminants

Bioavailability

Technologies

Capabilities

Limitations

Phases

● Solid

● Liquid

● Gas

● NAPL

Site
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

2-1

Bioventing

Gregory D. Sayles

Office of Research and Development, National Risk Management Research Laboratory,

U.S. Environmental Protection Agency, Cincinnati, OH

Research conducted in the mid to late 1980s by the U.S. Air Force (1, 2), researchers in the

Netherlands (3-6), the Texas Research Institute (7, 8), Battelle Memorial Institute (2, 9-11), Utah

State University (11), and the U.S. Environmental Protection Agency (EPA) (12), among others,

suggests that delivering air to the vadose zone to promote biodegradation could be a low-cost

means of cleaning fuel-contaminated vadose zone soils. This approach was motivated by attempting

to solve two different remediation development problems: 1) soil vacuum extraction for treatment

of contaminated vadose zones involved costly off-gas treatment and only removed the volatile

fraction of the contamination, and 2) oxygen delivery to the vadose zone to promote aerobic

biodegradation by using the approaches attempted in promoting biodegradation in ground water,

namely delivering oxygen-saturated water or aqueous solutions of hydrogen peroxide or nitrate to

the contaminated area, was not efficient or cost-effective. 

A process was needed that could deliver oxygen by introducing air into the vadose at a rate that

minimized volatilization of the contamination. Several groups simultaneously developed what is now

known as bioventing. 

EPA and the Air Force recognized the potential cost savings of such a technology over traditional

remediation approaches and began an aggressive bioventing development program in 1990. To

date, this program has demonstrated or is currently developing the use of bioventing for the

following situations:

# With air injection (10-17)

# In cold climates (18-20)

# With soil warming (18-20)

# For jet fuel and other aviation fuels (10-20)

# For nonfuel contaminants such as acetone, toluene, polycyclic aromatic

hydrocarbons (PAHs) (21), and trichloroethylene (TCE)

The cumulative knowledge of EPA, the Air Force, and Battelle Memorial Institute regarding

bioventing of fuel contaminated sites was distilled in Principles and Practices Manual for Bioventing,

released in 1996 (22). The manual outlines the physical, chemical, and biological principles used

in bioventing, and accepted approaches to determining site-specific treatability using onsite tests,

design and monitoring of bioventing systems, and site closure.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

2-2

Many documents exist that provide valuable information on bioventing. The Army Corps of

Engineers has also released a helpful manual (23). The most current collection of papers on

bioventing research and development is available in the book In Situ Aeration: Bioventing and

Related Remediation Processes (24). The next frontier for aerobic bioventing is the application of

the process to sites contaminated with chlorinated solvents and PAHs. EPA is currently involved in

two laboratory and field projects to develop co-metabolic bioventing. Co-metabolic bioventing is

the promotion of the aerobic biodegradation of chlorinated solvents, such as TCE, in the vadose

zone by delivering oxygen and, if necessary, a volatile co-metabolite to the contaminated site. The

Air Force has developed cost estimates for bioventing of fuels (25). Calculations show that

bioventing can range from $50 to $5 per cubic yard for soil volumes ranging from 2,000 to

20,000 cubic yards, respectively. These costs for bioventing are cheaper than costs estimated for

other onsite remediation methods such as soil vapor extraction, land farming, and excavation

followed by low-temperature thermal desorption. 

The available information on bioventing (experimental, performance, cost) easily convince the reader

that bioventing of fuels is probably the most successful in situ bioremediation technology developed

to date. There are an estimated 1,000 sites in the United States that have used or are currently

using bioventing, mostly for fuel-contamination remediation. In the future, expect the bioventing

approach to be shown useful for the cleanup of almost any aerobically biodegradable contaminant.

References

1. Miller, R.N. 1990. A field-scale investigation of enhanced petroleum hydrocarbon

biodegradation in the vadose zone combining soil venting as an oxygen source with moisture

and nutrient additions. Ph.D. dissertation. Utah State University, Logan, UT.

2. Miller, R.N., C.C. Vogel, and R.E. Hinchee. 1991. A field-scale investigation of petroleum

hydrocarbon biodegradation in the vadose zone enhanced by soil venting at Tyndall AFB,

Florida. In: Hinchee, R.E., and R.F. Olfenbuttel, eds. In situ bioreclamation. Stoneham, MA:

Butterworth-Heinemann. pp. 283-302.

3. Staatsuitgeverij. 1986. Proceedings of a Workshop, 20-21 March, 1986.

Bodembeschermingsreeeks No. 9; Biotechnologische Bodemsanering, pp. 31-33. Rapportnr.

851105002, ISBN 90-12-054133, Ordernr. 250-154-59; Staatsuitgeverij Den Haag: The

Netherlands. 

4. van Eyk, J. and C. Vreeken. 1988. Venting-mediated removal of petrol from subsurface soil

strata as a result of stimulated evaporation and enhanced biodegradation. Med. Fac.

Landbouww. Riiksuniv. Gent, 53(4b):1,873-1,884.

5. van Eyk, J., and C. Vreeken. 1989. Model of petroleum mineralization response to soil

aeration to aid in site-specific, in situ biological remediation. In: Jousma et al., eds. Groundwater contamination: Use of models in decision-making. Proceedings of an International

Conference on Groundwater Contamination. Boston/London: Kluwer. pp. 365-371. 
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

2-3

6. van Eyk, J., and C. Vreeken. 1989. Venting-mediated removal of diesel oil from subsurface

soil strata as a result of stimulated evaporation and enhanced biodegradation. In: Hazardous

waste and contaminated sites, Envirotech Vienna, Vol. 2, Session 3. ISBN 389432-009-5.

Essen, Germany: Westarp Wiss. pp. 475-485.

7. Texas Research Institute. 1980. Laboratory-scale gasoline spill and venting experiment.

American Petroleum Institute, Interim Report No. 7743-5:JST.

8. Texas Research Institute. 1984. Forced venting to remove gasoline vapor from a large-scale

model aquifer. American Petroleum Institute, Final Report No. 8210l-F:TAV.

9. Hinchee, R.E., and M. Arthur. 1991. Bench-scale studies of the soil aeration process for

bioremediation of petroleum hydrocarbons. J. Appl. Biochem. Biotech. 28/29:901-906.

10. Hinchee, R.E., and S.K. Ong. 1992. A rapid in situ respiration test for measuring aerobic

biodegradation rates of hydrocarbons in soil. Air & Waste Mgmt. Assoc. 42(10):1,305-1,312.

11. Dupont, R.R., W.J. Doucette, and R.E. Hinchee. 1991. Assessment of in situ bioremediation

potential and the application of bioventing at a fuel-contaminated site. In: Hinchee, R.E., and

R.F. Olfenbuttel, eds. In situ bioreclamation: Applications and investigations for hydrocarbon

and contaminated site remediation. Stoneham, MA: Butterworth-Heinemann. pp. 262-282.

12. Wilson, J.T., and C.H. Ward. 1986. Opportunities for bioremediation of aquifers

contaminated with petroleum hydrocarbons. J. Ind. Microbiol. 27:109-116.

13. Ostendorf, D.W, and D.H. Kampbell. 1990. Bioremediated soil venting of light hydrocarbons.

Haz. Waste Haz. Mat. 1(4):319-334.

14. Kampbell, D.H., and J.T. Wilson. 1991. Bioventing to treat fuel spills from underground

storage tanks. J. Haz. Mat. 28:75-80.

15. Kampbell, D.H., J.T. Wilson, and C.J. Griffin. 1992. Performance of bioventing at Traverse

City, Michigan. In: Bioremediation of hazardous wastes. EPA/600/R-92/126. pp. 61-64. 

16. Kampbell, D.H., C.J. Griffin, and F.A. Blaha. 1993. Comparison of bioventing and air

sparging for in situ bioremediation of fuels. In: Symposium on Bioremediation of Hazardous

Wastes: Research, Development, and Field Evaluations. EPA/600/R-93/054. pp. 61-67. 

17. Sayles, G.D., R.C. Brenner, R.E. Hinchee, and R. Elliott. 1994. Bioventing of jet fuel spills II:

Bioventing in a deep vadose zone at Hill AFB, Utah. In: Symposium on Bioremediation of

Hazardous Wastes: Research, Development and Field Applications. EPA/600/R-94/075. pp.

22-28. 

18. Sayles, G.D., R.C. Brenner, R.E. Hinchee, A. Leeson, C.M. Vogel, and R.N. Miller. 1994.

Bioventing of jet fuel spills I: Bioventing in a cold climate with soil warming at Eielson AFB,

Alaska. In: Symposium on Bioremediation of Hazardous Wastes: Research, Development and

Field Applications. EPA/600/R-94/075. pp. 15-21. 
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

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19. Leeson, A., R.E. Hinchee, J. Kittel, G. Sayles, C. Vogel, and R. Miller. 1993. Optimizing

bioventing in shallow vadose zones in cold climates. Hydrological Sciences J. 38(4).

20. Sayles G.D., A. Leeson, R.E. Hinchee, C.M. Vogel, R.C. Brenner, and R.N. Miller. 1995. Cold

climate bioventing with soil warming in Alaska. In: Hinchee, R.E., R.N. Miller, and P.C.

Johnson, eds. In situ aeration: Bioventing and related remediation processes. Columbus, OH:

Battelle Press. pp. 297-306.

21. McCauley, P.T., R.C. Brenner, F.V. Kremer, B.C. Alleman, and D.C. Beckwith. 1994.

Bioventing soils contaminated with wood preservatives. In: Symposium on Bioremediation of

Hazardous Wastes: Research, Development and Field Applications. EPA/600/R-94/075. pp.

40-45.

22. U.S. EPA. 1995. Bioventing: Prinicples and practice. EPA/540/R-95/543.

23. U.S. Army Corps of Engineers. 1995. Soil vapor extraction and bioventing, engineering and

design. EM 1110-1-4001. November.

24. Hinchee, R.E., R.N. Miller, and P.C. Johnson, eds. 1995. In situ aeration: Bioventing, and

related remediation processes. Columbus, OH: Battelle Press.

25. U.S. Air Force Center for Environmental Excellence. 1994. Bioventing performance and cost

summary. July.
2-5

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Bioventing

An Aerobic Bioprocess To Treat

Vadose Zone Contaminated Soils

Presented by

Gregory Sayles or Dolloff F. Bishop

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, Ohio

Outline

● What is bioventing?

● Site characterization for

bioventing

● Treatability for bioventing

● Full-scale design

Outline (continued)

● Operation/Monitoring

● Field examples

● Costs

● Bioventing manual

Hydrocarbon Distribution at a

Contaminated Site

Source Vapor 

Phase

Capillary 

Fringe

Dissolved 

Contaminants

Water

Table

Residual

Saturation Ground Water

Distribution of a

148,000 kg Spill (200 m3)

Contaminate % of Mass %

Phase Concentration Volume (m3) Volume (kg) of Mass

Recoverable 100% 63 0.2 47,000 32

NAPL

Soil Gas 1,000 ppm 5,600 17.0 1.7 .000011

Ground Water 100mg/L 20,000 62.0 2.0 .000014

Residual Soil 10,000 mg/kg 6,500 21.0 97,000 66

Sorbed

Courtesy of Rob Hinchee, Parsons Engineering Science Inc.

Natural Oxygen Delivery

Not Adequate

O2 O2 O2

Source

Residual

Saturation

Vapor

Phase
2-6

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Aerobic Biodegradation —

Respiration

C6 H6 + 71/2 O2 6 CO2 + 3 H2O

3.1 lb O2/lb C6 H6

C6 H14 + 91/2 O2 6 CO2 + 7 H2O

3.5 lb O2/lb C6 H14

Oxygen Carrier

Mass Requirements

Carrier/Hydrocarbon

 Oxygen Carrier (lb/lb)

Aqueous Solutions

Air Saturated 400,000

Nitrate (50 mg/L) 90,000

H202 (100 mg/L) 65,000

Air 13

Conceptual Layout of Bioventing

Process With Air Injection Only

Low Rate Air

Injection

Cutoff Well To

Prevent

Migration to

Basement

(if necessary)

Contaminated

Soil

Soil Gas

Monitoring

Biodegradation

of Vapors

Monitoring in

Basement

(as required)

What Is Bioventing?

Definition

Forced air movement through

contaminated vadose zone soils to

supply the oxygen necessary for

otherwise oxygen-limited in situ

bioremediation

Bioventing vs. SVE

Removal Rate

0 Air Flow Rate

Biodegradation

Volatilization and

Biodegradation

Aerobically Biodegradable

Rates vary from fast to slow:

BTEX Ketones (acetone)

Jet fuel PAHs (naphthalene)

Gasoline Alcohols

Diesel Fuel oil

Mono- or di-chlorinated benzenes, phenols

Mono- or di-chlorinated ethanes, ethylenes
2-7

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Site Characterization

● Historical data

● Soil gas survey

● Soil sampling

Historical Data

● Known spills, overfills, leaks

● Soil and GW data

● Location and levels

Purpose: Initial evaluation of feasibility, 

help plan soil gas survey

Soil Gas Survey

● Sample soil gas at various:

■ locations

■ depths

● Analyze gas for O2, CO2, TVH

Purpose: To locate areas where 

oxygen levels are low, 

minimize soil sampling

Schematic of a Soil Gas

Sampling System

Tubing

Pressure Relief 

Port

Vacuum 

Desiccator Vacuum 

Gauge

Tubing

Sampling 

Pump

Outlet

1/8” Flexible Tubing

Soil Probe Drive Tip

Soil Probe Extensions

Tedler Sample Bag

(Inside Desiccator)

Land Surface

Soil Gas Survey Results

● Low O2, high CO2

■ Bioactivity present, but needs O2

■ Candidate location for bioventing

● High O2, low CO2

■ Bioactivity low, something else is

retarding biodegradation

■ Not a candidate site for bioventing

Soil Sampling

● In region of low O2, sample soil at

various:

■ locations

■ depths

● Analyze for contaminants of

regulatory concern (e.g., TPH, BTEX)

Purpose: To confirm type and extent

of contamination, estimate

of cleanup time
2-8

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Site Characterization-Aerial View

G = Gas samples

Low O2

S = Soil samples

G

G

G

G

G

G

G

G

G

G

G

G

G

G

G

G

S S

S

S

S

Field Treatability Tests

Want to know the required:

● Air flow rate 

● Well spacing

● Cleanup time estimate

● Cost estimate

Treatability Test

● In situ respirometry test

● Soil gas permeability test

In Situ Respiration Test

Purpose:

● To measure O2 use rate for

feasibility

● To calculate air flow rate for

design

● To estimate cleanup time

In Situ Respiration Test

Protocol:

1. Install:

■ air injection tube

■ soil gas monitoring points

into contaminated area and 

background 

In Situ Respiration Test (continued)

2. Aerate (air + helium) for 1-2 days,

until soil gas levels steady

3. Shut off aeration

4. Monitor O2, CO2, and He with time
2-9

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

In Situ Respiration Test Apparatus

Inert Gas

Gas

Sampling

Port

Rotometer

Ground

Surface

Small Diameter

Probe

Screen

Pressure Gauge

Air Source

Reference: Hinchee, R.E. and S.K.

Ong. 1992. J. Air Waste Management

Association (42)10:1035-1312.

In Situ Respiration Test Results for

Tinker AFB, Oklahoma

Time (hours)

Oxygen

and

Carbon

Dioxide

(%)

He

k=0.17%/hr

O2

CO2

30

20

10

0

Helium

(%)

5.0

4.0

3.0

2.0

1.0

0.0

0 20 40 60 80 100 120

Background

Reference: U.S. Air Force Center for Environmental

Excellence. Test Plan and Technical Protocol for a

Field Treatability Test for Bioventing.

In Situ Respiration Test Results

for Kenai, Alaska

Time (hours)

Oxygen

and

Carbon

Dioxide

(%)

He

O2

CO2

25

20

15

10

5

0

Helium

(%)

4

3

2

1

0

0 10 20 30 40 50

Reference: U.S. Air Force Center for Environmental Excellence. Test Plan and

Technical Protocol for a Field Treatability Test for Bioventing.

Soil Gas Permeability Test

Purpose:

● Radius of influence of air

injection

● Well-spacing

● Cost

Radius of Influence Test

P P P P

Injection

Pressure

Monitoring

Radius of Influence Data, Saddle

Tank Farm, Galena AFS, Alaska

10

1

0.1

 0 20 40 60 80 100

Distance From Vent Well (feet)

Log Pressures (inches of H2O)
2-10

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Bioventing Decision Tree

O2 High

Soil

Sampling

O2 Low Soil Gas

Survey

Identify Limiting

Factors

Bioventing Decision Tree (continued)

Low Rate

Low Radius

Moderate to 

High Radius

High

Rate

Respiration

Test

Soil Gas

Permeability

Test

Consider

Alternative

Technology

Full-Scale

Design

Full-Scale Design

● Air flow rate

● Wells/Area

● Air injection vs. withdrawal

● Other well configurations

Flow Rate and Wells

● O2 use rate

● Radius of influence

● Total air flow rate

● Number of wells/area

Using

Calculate

Design Approach

Required Injection/ Withdrawal Rate

Oxygen Use Rate Soil Gas

Permeability

Wells/Area

Injection vs. Withdrawal

Injection usually preferred:

● Minimizes off-gas production

● Lowers water table—treats

capillary fringe

● Vapor residence time greater

But, be careful of subsurface structures!
2-11

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Conceptual Layout for Bioventing

Process with Air Injection Only

Low Rate Air

Injection

Cutoff Well To

Prevent

Migration to

Basement

(if necessary)

Contaminated

Soil

Soil Gas

Monitoring

Biodegradation

of Vapors

Monitoring in

Basement

(as required)

Other Configurations

Use injection and withdrawal

well combinations to meet

special site requirements

Air Injection System With

Reinjection of Extracted Soil Gas

Optional 

Makup Air

Contaminated 

Soil

Blower

Soil Gas 

Monitoring

Basement Monitoring Point

Negative Pressure

Blower

Optional

To

Injection

Schematic of Bioventing

Under Buildings

Operation/Monitoring

● Soil sampling at selected

time intervals

● O2 gas measurements

● Soil temperature

Operation/Monitoring (continued)

● Respiration tests at least

semi-annually

● Operate year round

● t = end determined by rate 0
2-12

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Results From the Field

● Hill AFB Field Research Study

■ Arid soil, deep air injection

■ Jet fuel

● Greenwood Chemical Superfund

site

■ Tight soil

■ Toluene, acetone, naphthalene

Hill AFB, Utah, Bioventing Study

● Jet fuel contamination

● From overfills of old USTs

● Contamination to 95 ft deep

● Low moisture, high permeability

soil

● Air injection operated for 31⁄2 yrs

Mean Oxygen Utilization Rate vs. Time Within the IW

25-ft Zone at Hill Air Force Base 280 Site

●

● ●

●

0.060

0.050

0.040

0.030

0.020

0.010

0.000

Nov-90 Jun-91 Dec-91 Jul-92 Jan-93 Aug-93 Mar-94 Sep-94 Apr-95

Date

O2 Utilization Rate

(% O2/hr)

●
2-13

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Greenwood Chemical Superfund

Site, Virginia, Pilot Test

● Specialty chemical company

● Toluene, acetone, naphthalene,

contamination

● Tight silty clay soils

● Air injection operated for 15

months

Costs

Example calculation*

● 5,000 yd3 jet-fuel contaminated soil

● 3,000 mg/kg TPH

● 4 injection wells

● Contamination, wells to 15 ft deep

* “Bioventing Performance and Cost Summary,” 

AFCEE, July 1994.
2-14

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Example (continued)

Item Cost

Project planning $11,000

Pilot testing $27,000

Regulatory approval $3,000

Example (continued)

Item Cost

Full-scale construction $27,000

Monitoring, 2 yrs $6,500

Power, 2 yrs $2,800

Final soil sampling $13,500

Total $90,800

Cost/yd3 $18

Bioventing Manual

 Available on the Internet

The Address is:

http://www.epa.gov/docs/ORD

Summary

If your site:

● Has soil contamination

● Low O2

● The contamination is

aerobically biodegradable

Seriously consider bioventing
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

3-1

Bioremediation of Sediments

Dolloff F. Bishop

Office of Research and Development, National Risk Management Research Laboratory, 

U.S. Environmental Protection Agency, Cincinnati, OH

Introduction

Contaminated sediments in rivers, lakes, and harbors in the United States pose a potential risk to

human health and the environment. Bioremediation (1-3), both through natural attenuation (intrinsic

bioremediation) and through enhanced bioremediation, promises possible approaches for

destruction of contaminants in sediments. Using natural processes involving microbial growth and

enzymatic production, bioremediation can convert target contaminants ultimately to nontoxic end

products. High molecular weight contaminants, however, such as polychlorinated biphenyls (PCBs)

and polynuclear aromatic hydrocarbons (PAHs), persist in sediments, biodegrading only slowly while

strongly partitioning to the sediments and bioaccumulating up the food chain (4), ultimately reaching

humans.

Both PCBs and PAHs are biodegradable under appropriate conditions in laboratory studies (1, 3).

PAHs (5) are typically degraded under aerobic conditions. PCBs (1) are typically degraded under

sequential anaerobic and aerobic conditions. Appropriate anaerobic conditions dehalogenate more

highly chlorinated PCBs, usually the meta- and para-chlorines on the biphenyl structure. Aerobic

conditions usually degrade the resulting lightly chlorinated PCBs with the chlorine atoms at the ortho

position.

Reasons why the persistent contaminants in sediments (6) are resistant to microbial degradation

include: 

# Contaminant toxicity to the microorganisms

# Preferential feeding of microorganisms on other substrates

# Microorganisms' inability to use a compound as a source of carbon and energy

# Unfavorable environmental conditions in sediments for propagation of appropriate

microorganisms 

# Poor contaminant bioavailability to microorganisms

Indeed, while the intrinsic biodegradation of such recalcitrant compounds is not uncommon in

nature, the degradation process can take many years.

The challenge for successful bioremediation of sediments involves combining appropriate microbial

pathways, biochemistry, and the function of natural microbial communities with innovative

engineering methods to overcome the recalcitrance of the compounds in sediments, thus increasing
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

3-2

bioremediation effectiveness. Successful acceleration of degradation rates in situ without a

bioreactor would provide a method for preferred sediment remediation, but such approaches have

exhibited limited effectiveness. Sediment dredging, usually to maintain open channels for shipping,

however, also offers the opportunity for alternative ex situ treatment (6), such as biotreatment in

confined treatment facilities (CTFs), slurry reactors, and composting land treatment applications.

Slurry reactor technology has also been applied in situ to contaminated sediments in water bodies

(5).

Field Bioremediation of Sediments

This review examines two pilot field studies on contaminated sediments: one an ex situ CTF

treatment of PCBs in sediments from the Sheboygan River in Wisconsin, the other an in situ slurry

reactor treatment of PCBs in sediments in the upper Hudson River. The CTF study (6) was

conducted for the U.S. Environmental Protection Agency's (EPA's) Region 5 and included a parallel

laboratory study on the Sheboygan River sediments by EPA's Athens Laboratory. The in situ slurry

reactor study (7) was conducted by the General Electric Company using caisson slurry bioreactors

placed in PCB-contaminated sediments in the river. 

The 14,000-square-foot aboveground CTF (Figure 1) used in the Sheboygan study was constructed

of steel sheet piling with a containment capacity of approximately 2,500 cubic yards of sediment

in four separate cells: two treatment and two control cells. Each cell (Table 1), lined with highdensity polyethylene, was hydraulically independent. Water accumulating in each cell discharged

through a permeable wall. The cells contained an underdrain system to add nutrients, oxygen, and

other amendments which could also be used for leachate control. The cells were filled with dredged

PCB-contaminated sediments (original source: Arochlor 1248 and 1254) obtained from the river

in late 1989 and from March to August 1990. The study attempted to evaluate remediation under

both anaerobic and aerobic conditions in the CTF. Two approaches for oxygenating the contained

sediments in Cell 4 were use of oxygenated (saturated) water from a compressed air saturator (July

1992) and use of dilute hydrogen peroxide solutions (November 1993). Mineral nutrient were also

added to the two treatment cells. Finally, laboratory studies were conducted to evaluate enhancing

anaerobic dehalogenation in the Sheboygan sediments.

In the second field evaluation, six steel caisson slurry reactors (Figure 2) were driven into

contaminated sediments in the upper Hudson River to isolate the natural bacteria and sediment from

the river environment. The experimental design in the study (Table 2) featured a low-mix caisson

and a high-mix caisson as unamended controls; two duplicate low-mix caissons with indigenous

organisms amended with ammonium and phosphate nutrients, biphenyl, and hydrogen peroxide;

and one high-mix and one low-mix caisson with indigenous organisms, both amended with

ammonium phosphate nutrients, biphenyl, hydrogen peroxide, and a culture of PCB degraders, A.

eutrophus H850. 

The sediments were mixed using high-mix turbines turning at 40 revolutions per minute (rpm) and

low-mix rakes turning at 3 rpm. The target dissolved oxygen level, automatically supported by

addition of hydrogen peroxide solution, was maintained between 6.0 and 6.5 mg/L in four caissons.

Other amendments were added to the four caissons as appropriate. The unamended high-mix
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

3-3

control became aerobic but was held to less than 2 mg/L liter by nitrogen purging while the low-mix

control remained anaerobic.

Sediment Remediation Performance

In the CTF study (Tables 3 and 4) at Sheboygan (8), the PCBs in the dredged sediments in the

various cells had an average chlorine per molecule of biphenyl ranging from 2.79 to 3.12,

indicating that only limited amounts of highly chlorinated congeners remained in the sediment.

Heavy oxygen demand in the sediment on Cell 4 minimized the oxygen (less than 0.1 mg/L)

available for degradation of lightly chlorinated PCBs. Attempts to aerobically degrade PCBs in the

sediments in Cell 4 thus produced no increased PCB remediation in the sediments. The oxygenation

attempts were unable to supply enough oxygen to overcome the oxygen demand in the sediment

and the sediment in Cell 4 remained anaerobic. The sediments, loaded into the cells over an

extended period, were dredged from various places in the river and were highly heterogenous with

wide variability in PCB concentrations from sampling location to sampling location in each cell. The

heterogeneity produced high variability in each cell's average concentration over the three sampling

events, as shown in Table 5. Under anaerobic conditions in the other CTF cells, statistically valid

increases in dehalogenation of the PCBs also did not occur.

Parallel laboratory studies at the Athens Laboratory (8) revealed (Figure 3) that addition of

octachlorobiphenyl (octa-CB) substantially increased dehalogenation of the PCBs in the historical

Sheboygan sediment. Sterile and live controls revealed no significant change in the PCBs in the

sediment. Increased dechlorination in historical PCB mixtures in the sediment, induced by the added

octa-CB, delayed the onset of transformation of the added octa-CB by 1 to 2 months.

The PCB homologs (Figure 4) revealed essentially no monohomolog and only modest dihomologs

in the initial sediment. The largest homolog was the trihomolog, which accounted for approximately

50 percent of the PCBs. The control test after 30 weeks revealed insignificant changes in PCB

homolog distribution. The amended system with 20 mg/L of octachlorobiphenyl exhibited significant

dechlorination with major increases of mono- and dihomologs (Figure 5).

Three methods were used to examine PCB concentration changes within the slurry reactors in the

Hudson River field study: direct concentration measurement and concentrations normalized to a

recalcitrant reference congener (peak 61, 34-34-/236-34 chlorobiphenyl) and to sediment total

organic carbon (9). The alternative methods were considered because of sampling variability in the

caissons, reflecting the heterogeneity in PCB distribution and sampling in the field. The two

normalizing methods were the most significant in quantifying PCB changes after 73 days of

treatment in the caissons (Table 5).

The normalized analyses revealed statistically significant PCB losses of 38 to 55 percent in all

amended caissons. The addition of the H850 culture produced no impact on the PCB changes, and

the H850 cultures were not competitive. Congener homolog group analysis (Figure 6) revealed

significant biodegradation of the mono- and dicongeners.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

3-4

Conclusions

The results of the Sheboygan River and the Hudson River studies reveal that partial bioremediation

of PCBs in sediments is possible, even without active biotreatment. The remediation, however, is

incomplete, even with active biotreatment. While sequential anaerobic/aerobic approaches may

completely degrade PCBs in aqueous dispersions, portions of the PCBs in sediments are not

available or only slowly available for biotreatment. Additional research is clearly needed to develop

and evaluate improved approaches for sediment bioremediation. Alternative measurements

(endpoints), based on toxicity, need to be evaluated on bioremediated sediments to assess the

potential environmental and health impacts of the residual PCBs after intrinsic bioremediation

(natural attenuation) and after active biotreatment.

References

1. Abramowicz, D.A. 1995. Aerobic and anaerobic PCB degradation in the environment.

Environ. Health Perspective 103, Supplement 5: 97-99.

2. Liu, S.M., and W.J. Jones. 1995. Biotransformation of dichloromatic compounds in nonadapted and adapted freshwater sediment slurries. Appl. Microbiol. Biotechnol. 43:725-

732.

3. Wilson, S.C., and K.C. Jones. 1993. Bioremediation of soil contaminated with aromatic

hydrocarbons (PAHs): A review. Environ. Pollut. 80:229-249.

4. Safe, S. 1980. Metabolism uptake, storage and bioaccumulation. In: Kimbrough, R., ed.

Halogenated biphenyls, naphthalenes, dibenzodioxins, and related products. Elsevier, North

Holland. pp. 81-107.

5. Seech, A., B. O'Neil, and L.A. Comacchio. 1993. Bioremediation of sediments

contaminated with polynuclear aromatic hydrocarbons (PAHs). In: Proceedings of the

Workshop on the Removal and Treatment of Contaminated Sediments. Environment

Canada's Great Lakes Cleanup Fund, Wastewater Technology Centre, Burlington, Ontario.

6. U.S. EPA. 1994. Assessment and remediation of Contaminant Sediments Program,

remediation guidance document. EPA/905/R-94/003. Great Lakes National Program

Office. October.

7. Flathman, P.E. 1992. Bioremediation technology advances via broad research

applications. Genetic Engineering News. October 15.

8. Jones, W.J. 1996. Personal communication.

9. Harkness, M.R. et al. 1993. In situ stimulation of aerobic PCB biodegradation in Hudson

River sediments. Science 159: 503-507.
3-5

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Bioremediation of

Sediments

Dolloff F. Bishop

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, OH

Acknowledgements

W. J. Jones

Environmental Research Laboratory, Athens

National Environmental Risk Laboratory

U.S. Environmental Protection Agency

and

F. J. Mondello

General Electric Company

Bioremediaton of

Contaminants in Sediments

● Natural attenuation (intrinsic bioremediation)

● Enhanced bioremediation using amendments

● Microbial growth and enzymatic production

often limited by conditions in sediments

● PCBs and PAHs as common high molecular

weight contaminants

Conditions Limiting

Bioremediation of Sediments

● Contaminant toxicity to microorganisms

● Preferential feeding of microorganisms on

other substrates

● Inability of microorganisms to use

contaminant as source of carbon and energy

● Sediment conditions unfavorable for

appropriate microbial propagation

● Contaminants not bioavailable to

microorganisms

Challenge for Sediment

Bioremediation

● Combining appropriate microbial pathways,

biochemistry, and function of natural microbial

communities

● Developing innovative engineering methods in sediments

to overcome contaminant recalcitrance to biodegradation

● Developing in situ biotreatment without reactors

(preferred but has exhibited limited effectiveness)

● Developing in situ treatment of dredged sediments for

enhanced bioremediation

● Developing in situ biotreatment with slurry reactors in

water bodies

Field Bioremediation of

Sediments

● Ex situ treatment of PCBs in

CTFs with supporting

laboratory studies

● In situ aerobic slurry treatment

of PCB in steel caissons

Figure 1. Confined Treatment Facility

for Sheboygan River Sediments

CTF

Discharge 

Piping
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Table 1. CTF Bioreactor Cells

In Situ 

Cell No. PCB mg/kg Treatment Condition

1 225 Anaerobic with nutrients

2 185 Anaerobic control

3 100 Anaerobic control

4* 125 Anaerobic with nutrients

*Cell 4 was intended to be aerobic but D.O. never >0.1 mg/L

Figure 2. In Situ Slurry Biodegradation

of Hudson River Sediments

Table 2. In Situ Slurry Reactor

Experimental Design

Caisson Treatment Initial PCB Conc.

(mg/kg)

R101 High-mix, control 6.0 + 1.9

R102 High-mix, amended H850 20.0 + 11.0

R103 Low-mix, amended H850 30.2 + 10.6

R104 Low-mix, control 39.9 + 15.6

R105 Low-mix, amended indig. 49.7 + 27.8

R106 Low-mix, amended indig. 39.1 + 17.5

In Situ Slurry Reactor Design

● High-mix turbines turning at 40 rpm

● Low-mix rakes turning at 3 rpm

● Amended with ammonium and

phosphate nutrients biphenyl,

hydrogen peroxide (D.O. 6–6.5 mg/L)

● Indigenous organism or indigenous and

H850 organisms

● Low-mix control-anaerobic; high-mix,

<2 mg/L D.O.

Table 3. Average CL Per

Biphenyl*

Sample date Cell 1 Cell 2 Cell 3 Cell 4

6-1-92 3.14 2.78 2.87 3.22

8-20-92 3.11 2.80 2.82 3.12

11-4-92 3.11 2.79 2.75 2.95

Averages 3.12 2.79 2.81 3.10

*Sheboygan River sediments in CTF

Table 4. Average PCB

Concentrations*, mg/kg

Sample date Cell 1 Cell 2 Cell 3 Cell 4**

6-1-92 200 115 91 134

8-20-92 273 132 109 230

11-4-92 323 165 180 236

Averages 265 137 127 200

*Sheboygan River sediments in CTF

**Cell 4 was intended to be aerobic but D.O. never >0.1 mg/L

Anaerobic 

with nutrients

Anaerobic control Anaerobic control Anaerobic

with nutrients
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Figure 3. Induced Dechlorination of

Sheboygan Sediments

Figure 4. Congener Homologs in

Sheboygan River Sediments

Figure 5. Congener Transformation

by Octachlorobiphenyl Amendment

Table 5. PCB Transformations in

Hudson River Sediments

 Percent Changed

Treatment Direct Measure Peak 61* TOC**

High-mix control +8.7 –14.4 –30.7

High-mix, H850 –41.0 –42.4 –44.7

Low-mix, H850 –36.8 –37.8 –55.5

Low-mix, control –41.8 –4.3 +8.4

Low-mix, indig. –72.6 –40.5 –53.1

Low-mix, indig. –68.5 –38.7 –46.0

*Normalized to congener 34-34/236-34 chlorobiphenyl

**Normalized to total TOC

Figure 6. Transformation of PCB

Homologs in Hudson River Sediments

To = Time zero.

Tf = Final time after 73 days.

Conclusions

● Partial bioremediation of PCBs in sediments

occurs even without active biotreatment

● Remediation is incomplete even with active

biotreatment

● Portions of PCBs in sediment are not or only

slowly available for biotreatment

● Alternative measurements (endpoints) based

on toxicity need to be conducted on

bioremediation sediments

● Research is needed to develop improved

methods of sediment bioremediation
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

4-1

Aerated Lagoons: A Case Study

Dolloff F. Bishop

Office of Research and Development, National Risk Management Research Laboratory, 

U.S. Environmental Protection Agency, Cincinnati, OH

Introduction

In the mid-1960s to the early 1970s, the French Limited Superfund site (Figure 1) was a statelicensed waste disposal site near Crosby, Texas. About 90 companies contributed petroleum and

petrochemical wastes that were hauled to the site for disposal. At closure of the disposal site in

1971, about 70 million gallons of wastes were in the main waste lagoon. In late 1983, the

potentially responsible parties (PRPs) formed the French Limited Task Group (FLTG) to consider site

cleanup (1). In early 1987, the U.S. Environmental Protection Agency (EPA) issued a record of

decision (ROD) for the site (2) calling for remediation by incineration, at estimated costs of $75 to

$125 million.

Beginning in late 1985 and continuing through 1986, bench-scale bioremediation had already

been successfully conducted on the contaminated sludges and soils in the lagoon. When the ROD

selecting incineration was issued, FLTG began to explore, at field pilot scale, environmentally

protective and less costly in situ bioremediation for French Limited cleanup. After the successful field

pilot study, EPA in late 1987 modified the ROD to allow in situ bioremediation (2) as the preferred

cleanup technology for the site. Full-scale site remediation, first in one biotreatment cell (one half

of the lagoon) and then in a second cell, was initiated at the site in early 1992 and was completed

by 1994.

Cleanup Approach

Most contaminants were biodegradable and in a water matrix at a site with a warm climate. Practical

bioremediation at the site needed to manage ambient air quality; mechanically mix microorganisms,

nutrients, oxygen, sludge, soil, and mixed liquor to produce acceptable biodegradation rates in the

12-acre lagoon; and accurately measure cleanup effectiveness over time. The major design

challenges that had to be met included providing oxygenation with minimum air emissions, effective

mixing during reintroduction of lagoon sludges and soils into a suspended mixed liquor, and

effective circulation (mixing) to distribute nutrients and dissolved oxygen throughout the biotreatment

cell.

Several technologies (3) were considered for oxygenation, including fine bubble aeration and pure

oxygen contacting. Dissolved pure oxygen (Table 1) provided the lowest air emissions. The Mixflo

system (Figure 2), designed by Proxair Inc., was selected for the site by EPA, the FLTG, and ENSR

Consulting and Engineering. Mixflo uses pure oxygen in a two-stage process. The system, with a

maximum capacity of 25 tons of oxygen per day, is the largest oxygenation and sludge and soil

mixing system in the world.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

4-2

In the first stage, slurry pumped from the lagoon and pressurized in a pipeline was fed high-purity

oxygen. The two-phase mixture flowed turbulently through the pipeline, substantially increasing

oxygen solubility in the slurry under elevated pressure. In the second stage, the oxygen/slurry

dispersion was reinjected into the lagoon using a liquid/liquid eductor (Figure 3) that mixed

unoxygenated slurry with the oxygenated slurry and produced a fine bubble oxygen dispersion before

dispersing the mixture throughout the lagoon.

The mixing of unoxygenated slurry with oxygenated slurry in the eductor before discharging the

mixture reduced the dissolved oxygen concentration below atmospheric pressure saturation. Thus,

dissolved oxygen did not come out of solution in the lagoon. The oxygen not dissolved in the

pipeline contactor also was well distributed as fine bubbles with a low frequency of bubble

coalescence in the lagoon. Further oxygen dissolution then occurred in the lagoon, minimizing air

emissions and providing excellent (90 percent) oxygen dissolution efficiency. To ensure an effective

circulation pattern in the lagoon biotreatment cell, nine 50,000-gallon-per-minute FLYGT banana

mixers were placed on three rafts. The Mixflo system and the FLYGT mixers provided effective

solutions to the engineering challenges. After completion of bioremediation, each biotreatment cell

was subsequently filled with clean soil and planted in cover vegetation.

Bioremediation Performance 

In situ aerobic bioremediation met all sludge soil cleanup requirements (4, 5) for the lagoon. Using

indicator contaminants (Table 2) as examples, residual arsenic had to be at or below 7 parts per

million (ppm); benzene at or below 14 ppm; benzo(a)pyrene at or below 9 ppm; total

polychlorinated biphenyls (PCBs) at or below 23 ppm; and vinyl chloride at or below 43 ppm. Actual

concentrations of the indicator contaminants after bioremediation typically were 1 to 2 ppm arsenic,

0.5 to 10 ppm benzene, 1.8 to 10 ppm benzo(a)pyrene, 1 to 10 ppm PCBs, and 3 to 17 ppm vinyl

chloride.

Ambient air monitoring during remediation (Table 3) revealed that air criteria concentrations to

quantify maximum cumulative concentrations for each of 35 compounds of concern were also fully

achieved. Finally, the direct costs (3) of the lagoon bioremediation (Table 4), including the field pilot

demonstration, were $39 million. Total costs for bioremediation were $59 million, compared with

the estimated $75 to $125 million, for incineration.

Site Closure

A second bioremediation process (6), not presented here, was conducted at the site. The lagoon had

contaminated the surrounding ground water. The ground-water bioremediation process was recently

completed (January 1996). Full site closure with continued ground-water monitoring is nearly

complete.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

4-3

References

1. Biotreatment News. 1991-1992. French Limited: A successful approach to bioremediation.

A three-part series.

2. U.S. EPA. 1992. Superfund at work. EPA/520/P-93/004.

3. Bergman, T.J., et al. 1992. An in situ slurry-phase bioremediation case with emphasis on

selection and design of a pure oxygen dissolution system. Union Carbide Industrial Gases

Technology Corporation, Tarrytown, NY, and ENSR Consulting and Engineering, Houston,

TX.

4. CH2M Hill. 1995. Site remediation report, Part A: Lagoon remediation verification. EPA

Contract No. 68-W8-0112.

5. U.S. EPA. 1994. Hazardous Waste Management Division first 5-year review: French Limited

site, Crosby, TX. CERCLIS TXD-980514814.

6. Biotreatment News. 1993-1994. In situ bioremediation of ground water and subsoils at

French Limited site, TX. A three-part series. 
4-4

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Aerated Lagoons

Acknowledgements

Judith Black

 Region VI

U.S. Environmental Protection Agency

Richard Sloan

ARCO Chemical Company

A Case Study of the French Limited

Superfund Site

Presented by

Dolloff F. Bishop

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, Ohio

French Limited Waste

Disposal Site

● Mid 1960 to 1971

● Petroleum and petrochemicals

● Incineration ROD in 1987 at

estimated costs of $75–125 million

● ROD in late 1987 modified to

permit in situ bioremediation

Figure 1. French Limited

Site Location

Engineering Challenges in

Lagoon Bioremediation

● Minimize air emissions

● Provide efficient shearing and

introduction of sludge and soil into

the lagoon’s suspended mixed liquor

● Maintain mixing of suspended mixed

liquor

● Provide efficient distribution of

nutrients and oxygen

Solutions to Engineering

Challenges

● Pure oxygen dissolution using

Mixflo

● Liquid/liquid eductor

● FLYGT banana mixers on rafts

Figure 2. Mixflo
4-5

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Table 1. Comparison of Mixflow

and Fine Bubble Aeration

Mixflo Fine Bubble

Oxygen transfer efficiency % 90 14

Gas volume, scfm 112 3,418

Off gas volume, scfm 12 3,318

Figure 3. Liquid/Liquid Eductor

Table 2. Performance of

Indicator Compounds

Cleanup Typical

Required Residuals

PPM PPM

Arsenic 7 1–2

Benzene 14 0.5–10

Benzo(a)pyrene 8 1.8–10

Total PCBs 23 1–10

Vinyl Chloride 43 3–17

Table 3. Benzene Ambient Air

Management ACC Ratios

Subdivision ACC* Ratios**

Cell E Cell D/F

Riverdale 0.2393 0.1872

Rogge 0.0597 0.0402

Dreamland 0.0368 0.0277

* Air Criteria Concentrations

** Requirement: ACC ratio must be less than 1.0 at end of 2 years.

Table 4. Incineration and

Bioremediation Costs

Incineration* Bioremediation

$ Millions $ Millions

General 5 13**

Site Preparation 7 7

Remediation 68 19

Indirect Costs 15 10

Contingency 30 5

TOTALS 125 54

* On site incineration

** Includes 10 million dollar cost for field pilot demonstration.

Site Revegetation
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

5-1

Oil-Contaminated Shorelines

Albert D. Venosa

Office of Research and Development, National Risk Management Research Laboratory,

U.S. Environmental Protection Agency, Cincinnati, OH

Introduction

This case study is based on a field study conducted during the summer of 1994 by researchers from

the U.S. Environmental Protection Agency's (EPA's) National Risk Management Research Laboratory

and the University of Cincinnati, in cooperation with the Delaware Department of Natural Resources

and Environmental Control (1).

Light crude oil was intentionally released onto plots to evaluate bioremediation. Past field studies

involving bioremediation of oil-contaminated shores have concluded that bioremediation enhances

the removal of crude oil several times more effectively than the intrinsic rate (2-9). Much skepticism

remains in the field, however, because data from all of these investigations have been equivocal to

some extent. The goals of this project were to quantify the effectiveness of natural attenuation due

to levels of background nutrients already present in the Fowler Beach area of Delaware Bay; to

demonstrate the effectiveness of biostimulation and/or bioaugmentation; to determine the extent of

any resulting rate enhancement; and to provide guidelines that can be used by spill responders and

on-scene coordinators for the effective bioremediation of oil-contaminated sandy shores.

Biodegradation was tracked by gas chromatography/mass spectroscopy (GC/MS) analysis of

selected components, and the measured concentrations were corrected for abiotic removal by

hopane normalization. (Hopane is a nonbiodegradable compound that exists in all crude oils.) Five

replicates of three treatments were evaluated: an oiled no-nutrient control, addition of water soluble

nutrients, and addition of water soluble nutrients supplemented with a natural microbial inoculum

from the site. 

Approach

Without full replication and random interspersion of treatments, it is impossible to ascribe statistically

significant differences in the response variable(s) to the treatments. A randomized complete block

design was used to assess treatment effects. Five areas (blocks) of beach were selected, each large

enough to accommodate four experimental units or test plots. The blocks were positioned on the

beach parallel to the shoreline. Three treatments were tested on oiled plots: a no-nutrient addition

control, addition of water soluble nutrients (biostimulation), and addition of water soluble nutrients

supplemented with a natural microbial inoculum from the site (bioaugmentation). A fourth treatment,

an unoiled and untreated plot, served as a control for background biological measurements. The

four treatments were randomized in each of the five blocks. 

Previously weathered light crude oil from Nigeria (Bonny Light) was the source of crude oil. It was

applied to the plots uniformly by spray nozzles connected to drums. Each plot received 36 gallons
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

5-2

of oil. Laboratory microcosms indicated that a concentration of 0.5 mg N/L and limited oxygen

uptake and CO2 production, whereas at concentrations greater than 2.5 mg N/L, maximum uptake

was observed. Thus, the target nitrate-N was set at about 1.5 mg/L.

A lithium tracer experiment to determine how frequently fertilizer should be added to maintain the

target nutrient level found that tracer diluted quickly as the plots became submerged by the incoming

tides and waves. In fact, there was a direct correlation between plot submergence and the amount

of tracer remaining in the bioremediation zone. Because the plots for the field study were positioned

within the intertidal zone, nutrients had to be applied every day to maintain the desired 1.5 mg/L in

the interstitial pore water.

The bioaugmentation treatment consisted of an inoculum of oil degraders isolated from the site,

grown in batches on the same crude oil, and added back every week. The indigenous inoculum was

grown for 2 weeks in two 55-gallon stainless steel drums. To allow weekly inoculation with fresh 2-

week cultures, each drum was offset in time from the other by 1 week. The drums contained 40

gallons of seawater from Delaware Bay, the weathered Bonny Light crude oil (600 mL) as the sole

carbon source, and the same nutrients used on the beach. 

Results

Nutrient Persistence. The control plots receiving only seawater with no nutrients had measurable

concentrations of nitrate (mean of 0.82 mg/L), which were approximately half the 1.5 mg/L target

level desired for maximum biodegradation. The concentrations in the nutrient and inoculum treated

plots were substantially higher. The Fowler Beach area of Delaware Bay was close to farm land,

where runoff could easily account for the high background levels found. 

Physical Loss of Oil. To distinguish physical loss from biodegradative loss of oil, the concentration

of hopane, a known nonbiodegradable biomarker in all crude oils, was quantified in each sand

sample. Data from the three oiled treatments revealed a hopane half-life of 28 days. This was

interpreted to represent physical loss of crude oil due to wave action and tidal inundation. A similar

study of the temporal loss of total extractable organic material (EOM) from the plots revealed an

EOM half-life of 21 days. The EOM first-order rate coefficient was significantly higher than the

hopane disappearance rate. The difference in loss rates (and half-lives) between hopane and EOM

was attributed to biodegradation because EOM includes both biodegradable and nonbiodegradable

components. EOM, however, was not a sensitive enough indicator to discern treatment differences.

Results of Bioremediation. The bioremediation study revealed that, although substantial

hydrocarbon biodegradation occurred in the untreated plots, statistically significant differences

between treated and untreated plots were observed in the biodegradation rates of the hopanenormalized total alkane and total aromatic hydrocarbons. The rate enhancement was approximately

two-fold for the alkanes and 50 percent for the aromatics. First-order rate constants for

disappearance of individual hopane-normalized alkanes and polycyclic aromatic hydrocarbons

(PAHs) were computed, and the patterns of loss were typical of biodegradation. As the number of

alkyl-substituted groups increased on the aromatic ring structure, the rate of PAH disappearance

decreased. This is known to be typical of biodegradation. In the field, the ratio of biodegradation

rates of unsubstituted parent compounds and lower substituted compounds to the highest substituted
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

5-3

compound in a homologous series revealed strikingly close agreement with the same ratios

computed from laboratory experiments (except for naphthalene and C1-naphthalene, which are

highly volatile). This signifies that the loss of hydrocarbons due to factors other than biodegradation

(i.e., dissolution and volatilization) was negligible.

Significant differences were not observed between plots treated with nutrients alone and plots treated

with nutrients and the indigenous inoculum. The high rate of oil biodegradation observed in the

untreated plots was attributed to the relatively high background nitrogen concentrations that were

measured at the site. 

Conclusions

Significant intrinsic biodegradation of petroleum hydrocarbons occurred naturally when sufficient

nutrients already existed in the affected area. Statistically significant rate enhancement was

demonstrated, even in the presence of an already high rate of natural attenuation, by supplementing

natural nutrient levels with inorganic mineral nutrients; however, bioaugmentation did not

significantly contribute to any further enhancement. Maintenance of a threshold concentration of

about 2 mg nitrate-N/L interstitial pore water permits close to maximum hydrocarbon

bioremediation. The incremental increase in biodegradation rate over the intrinsic rate (i.e., slightly

greater than two-fold for the alkanes and 50 percent for the PAHs) might not have been high

enough to warrant a recommendation to actively initiate a major, perhaps costly, bioremediation

action in the event of a large crude oil spill in that area. Thus, the decision to apply nutrients should

depend on the background concentrations available at the contaminated site, as well as the impact

on ecological and health receptors. 

The study showed that better hydrocarbon biodegradation takes place in the upper intertidal zone

than in the lower intertidal zone due to the greater persistence of nutrients and highly aerobic

conditions. Hopane was confirmed as a useful biomarker for tracking biodegradation success in

the field. 

For the first time, first-order biodegradation rate constants were developed from field data for the

resolvable normal and branched alkanes and the important two- and three-ring PAH groups (and

at least one four-ring PAH group) present in light crude oil. The relative biodegradation rates of

homologous PAHs measured in the field were found to agree closely with those measured in the

laboratory, thus corroborating the rates as being due to biodegradation and not physical washout

or solubility differences. 

Lessons Learned

After a major spill has been beached, the first task is to measure the natural nutrient concentrations

in that environment to determine if they are already high enough to sustain significant intrinsic

biodegradation. Concentrations approaching 1.5 to 2.0 mg N/L in the interstitial pore water should

support near-optimum hydrocarbon biodegradative activity. A determination should be made as to

whether such nutrient levels are normal for the affected area for that time of the year. Oiled sandy
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

5-4

shorelines should only be treated with nutrients if concentrations are clearly limiting (i.e., well below

1 to 2 mg/L).

If the beach is treated with water-soluble nutrients applied by a spray irrigation system, they should

be applied daily if the area gets completely submerged by tides and waves, even during neap tides.

If the area is submerged only during spring tides, the intertidal coverage by water determines the

frequency of nutrient addition. The Delaware study did not include evaluation of either oleophilic

or slow release granular fertilizer for nutrient enhancement. For large expanses of contaminated

shoreline or areas with difficult access and control (e.g., heavy wave action), oleophilic fertilizers may

be more appropriate.

Degradation effectiveness should be monitored using specific analytes quantified by GC/MS and

then only when analytes are normalized to a recalcitrant compound like hopane. Total petroleum

hydrocarbon (TPH) measurements should not be used to monitor treatment effectiveness; they are

too variable and too much affected by biogenic organic matter that has nothing to do with the

hydrocarbons present.

Bioaugmentation is often unnecessary for accelerating biodegradation of an oil spill on a sandy

beach. Quantifying the hydrocarbon degrader populations in the impact zone is useful, however.

A treatment product should not be considered for use on a shoreline based only on results of

bioremediation studies in a terrestrial environment. The abiotic loss mechanisms that act upon

petroleum, nutrients, and microorganisms are substantially different on a beach than on dry land.

Estimated Cost of Bioremediation

A rough estimate of the costs of an oil spill bioremediation project has been calculated, based on

the Delaware study. The following assumptions have been made for this analysis: 

# The spill has contaminated a 27-mile-wide intertidal zone of a long stretch of

coarse sandy beach in an area that is easily accessible (unlike Prince William

Sound), such as the Atlantic, Pacific, or Gulf coasts.

# Free product and heavy concentrations have already been removed by physical

cleanup procedures.

# Pore water nutrient levels are well below the 1.5 to 2.0 mg N/L needed for

optimum biodegradation effectiveness.

# Nutrients are added daily via a sprinkler or irrigation system to maximize

bioremediation effectiveness. 

Based on these assumptions, an estimated 2 person-years per kilometer (i.e., one supervisor and

three laborers working full-time for approximately 3 months) would be required for cleanup.

Assuming a supervisor salary (with benefits) of $100,000 per year and a laborer salary of $50,000

per year, the labor cost would be $62,500. Equipment needs are estimated to be about $75,000,

chemicals $45,000, storage $2,500, and analytical needs $50,000. Total direct costs would thus
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

5-5

be approximately $235,000. Applying overhead at the rate of 100 percent yields a total cost of

approximately $470,000 per kilometer of beach contaminated. 

The above cost estimates are highly dependent on manpower for daily application of water-soluble

fertilizer. If slow-release granular fertilizer is used (thus mitigating the need for daily application), and

assuming target levels of nitrogen can be achieved for periods approaching a week, then the

manpower and equipment needs will likely be significantly lower than those estimated above.

Detailed economic analysis awaits data from further field evaluations.

Protocol Development

As a result of the Oil Pollution Act of 1990 (OPA), EPA instituted a research program to develop an

objective protocol assessing the bioremediation effectiveness and toxicity of commercial oil spill

bioremediation agents. A tiered approach was developed in which a product is subjected first to a

laboratory batch screening test and tested against a control for its ability to biodegrade crude oil

(10, 11). An acute toxicity test is also performed to assess the product's ability to induce mortality

in mysid shrimp species. The next tier involves further testing of the product compared with a control

in a flow-through microcosm. The final tier consists of an actual field trial of the product. The

laboratory screening test consists of shake flasks containing natural seawater, 5 g/L weathered

Alaska North Slope crude oil, and the product. Two controls are set up: a no-nutrient, no-product

control (i.e., natural seawater and weathered oil) and a nutrient control (natural seawater, weathered

oil, and nitrate and phosphate salts as nutrients). Triplicate flasks are sacrificed at days 0, 7, and 28

to determine the extent of biodegradation of the crude oil components. Measurements are made

by GC/MS. Alkane and aromatic hydrocarbon degraders are also measured by a most probable

number technique (12). For a product to be deemed effective, it must demonstrate statistically

significant removal of both alkane and aromatic hydrocarbons compared with the controls at the

conclusion of the exposure period. EPA is currently attempting to refine the protocol by changing

the natural seawater to a sterile artificial formulation and standardizing the microbial inoculum. Such

refinements would make the test more reproducible. The inoculum would be used as a positive

control for living products, whereas it would serve as the actual biodegrading population in the case

of a non-living product. Products that successfully demonstrate the ability to biodegrade both the

alkane and aromatic components of weathered crude oil are then placed on the National

Contingency Plan product schedule, which makes them eligible for use in an oil spill.

References

1. Venosa, A.D., M.T. Suidan, B.A. Wrenn, K.L. Strohmeier, J.R. Haines, B.L. Eberhart, D.

King, and E.L. Holder. 1996. Bioremediation of an experimental oil spill on the shoreline

of Delaware Bay. Environ. Sci. Technol. 30(5):1,164-1,175.

2. Bragg, J.R., R.C. Prince, E.J. Harner, and R.M. Atlas. 1994. Effectiveness of

bioremediation for the Exxon Valdez oil spill. Nature 368:413-418.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

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3. Halmo, G. 1985. Enhanced biodegradation of oil. In: Proceedings of the 1985

International Oil Spill Conference. American Petroleum Institute, Washington, DC. 

4. Rosenburg, E., R. Legmann, A. Kushmaro, R. Taube, R. Adler, and E.Z. Ron. 1992.

Petroleum bioremediation—A multiphase problem. Biodegradation 3:337-350.

5. Sendstad, E. 1980. Accelerated biodegradation of crude oil on Arctic shorelines. In:

Proceedings of the Third Arctic and Marine Oil Spill Program. Environment Canada.

6. Sveum, P. 1987. Accidentally spilled gas-oil in a shoreline sediment on Spitzbergen:

Natural fate and enhancement of biodegradation. In: Proceedings of the Tenth Arctic and

Marine Oilspill Program. Environment Canada.

7. Sveum, P., and A. Ladousse. 1989. Biodegradation of oil in the Arctic: Enhancement by

oil-soluble fertilizer application. In: Proceedings of the 1989 International Oil Spill

Conference. American Petroleum Institute, Washington, DC. 

8. Pritchard, P.H., and C.F. Costa. 1991. EPA's Alaska oil spill bioremediation project.

Environ. Sci. Technol. 25:372-379.

9. Pritchard, P.H., J.G. Mueller, J.C. Rogers, F.V. Kremer, and J.A. Glaser. 1992. Oilspill

bioremediation: Experiences, lessons, and results from the Exxon Valdez oil spill in Alaska.

Biodegradation 3:315-335.

10. Venosa, A.D., J.R. Haines, and B.L. Eberhart. 1996. In: Sheehan, D., ed. Protocols in

bioremediation. Totowa, NJ: Humana Press.

11. Venosa, A.D., J.R. Haines, W. Nisamaneepong, R. Govind, S. Pradhan, and B. Siddique.

1992. J. Ind. Microbiol. 10:13-23.

12. Wrenn, B.A., and A.D. Venosa. 1996. Canadian J. Microbiol. 42:252-258.
5-7

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Oil-Contaminated

Shorelines

Presented by

Gregory Sayles or Dolloff F. Bishop

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, Ohio
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation
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Land Treatment

Daniel Pope

Dynamac Corporation, Ada, OK

Definition of Land Treatment

Land treatment involves use of natural biological, chemical and physical processes in the soil to

transform organic contaminants of concern. Biological activity apparently accounts for most of the

transformation of organic contaminants in soil, although physical and chemical mechanisms may

provide significant loss pathways for some compounds under some conditions. Degradation by

ultraviolet light may serve as a loss pathway for certain hydrophobic compounds at the soil surface.

Volatilization of some low molecular weight compounds also takes place at the soil surface and

provides a significant loss pathway for such compounds. Certain chemical reactions such as

hydrolysis can play an important role in transformation of some compounds. Humification, the

addition of compounds to the humic materials in soil, can be an important route of transformation

for some polynuclear aromatic compounds. The relative importance of these processes varies

widely for different compounds under different circumstances. The land treatment concept serves

as the basis for design and operation of soil bioremediation technologies at a large number of waste

sites requiring cleanup.

In Situ and Ex Situ Land Treatment

Land treatment techniques for bioremediation purposes most often are used for treatment of

contaminated soil, but certain petroleum waste sludges have long been applied to soil for treatment.

Ideally, the contaminated soil can be treated in place (in situ). Often, however, the soil must be

excavated and moved to a location better suited to control of the land treatment process (ex situ).

In situ land treatment is limited by the depth of soil that can be effectively treated. In many soils,

effective oxygen diffusion sufficient for desirable rates of bioremediation extends to a range of only

a few inches to about 12 inches into the soil, although depths of 2 feet and greater have been

effectively treated in some cases. 

Ex situ treatment generally involves applications of lifts of contaminated soil to a prepared bed

reactor. This reactor is usually lined with clay and/or plastic liners, provided with irrigation,

drainage, and soil water monitoring systems, and surrounded with a berm. The lifts of contaminated

soil are usually placed on a bed of relatively porous, noncontaminated soil.

The land treatment process may be severely limited in clayey soils, especially in areas of high rainfall.

This limitation is primarily related to oxygen transfer limitations and substrate availability to the

microorganisms. Clayey soils should be applied in shallower lifts than sandy soils. Tilth ("workability"

of the soil) can often be improved by adding bulking agents.
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After application to the land treatment unit, each lift should be tilled at intervals to enhance oxygen

infiltration and contaminant mixing with the microorganisms. The soil should be near the lower end

of the recommended soil moisture percentage range before tilling. Tilling very wet or saturated soil

tends to destroy the soil structure, reduce oxygen and water intake, and cause reduced microbial

activity. Tilling more than is necessary for enhanced oxygen infiltration and contaminant mixing may

be counterproductive because tilling tends to destroy the soil structure and compact soil below the

tilling zone.

Timing of application of succeeding lifts should be based on reduction to defined levels of particular

compounds or categories of compounds in the preceding lift. For instance, the goal might be to

reduce total petroleum hydrocarbons (TPH) to less than a regulatory or risk- calculated limit in the

current lift before application of a new lift. Once desired target levels of compounds of interest are

established, data obtained from land treatment unit (LTU) monitoring activities can be statistically

analyzed to determine whether and when desired levels are reached and the LTU is ready for

application of another lift. 

Nutrients, Carbon Sources, and Other Additives

Fertilizers can be used to supply nutrients, and wood chips, sawdust, or straw can supply carbon.

Various animal manures are often used to supply both carbon sources and nutrients. High organic

levels in manures, wood chips, and the other organic amendments increase sorptive properties of

soil, thereby decreasing mobility of organic contaminants and possibly decreasing availability to the

microorganisms. Organic amendments will also increase the water-holding capacity of soil, which

can be desirable in sandy soils but can cause difficulty when land treatment is conducted in areas

of high rainfall and poor drainage. 

Agricultural fertilizer is usually supplied in prilled or pelleted form (the fertilizer compounds formed

into pellets with a clay binder) suitable for easy application over large areas. Completely watersoluble fertilizers can be applied through irrigation systems, allowing application rates to be closely

controlled, applications to be made as often as irrigation water is applied, and immediate availability

to the microorganisms. 

Bioaugmentation

Microorganism cultures are often sold for addition to bioremediation units. Two factors limit use

of these added microbial cultures in LTUs: 1) nonindigenous microorganisms rarely compete well

enough with indigenous populations to develop and sustain useful population levels, and 2) most

soils with long-term exposure to biodegradable wastes have indigenous microorganisms that are

effective degraders if the LTU is managed properly. 

Certain soil factors may interfere with microbiological activity in the LTU soil. High salt levels,

indicated by high electrical conductivity (EC) readings, may reduce or stop useful microbiological

activity. If levels are too high, it may be necessary to leach the soil with water to remove excess salts

before biodegradation can occur. High levels of sodium may be detrimental to soil structure. 
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6-3

Soil Moisture Control

Historically, it has been recommended that soil moisture be maintained at 40 to 70 percent of field

capacity; however, recent experience indicates that 70 to 80 percent of field capacity may be

optimum. A soil is at field capacity when soil micropores are filled with water and soil macropores

are filled with air. This condition allows soil microorganisms to get air and water, both of which are

necessary for aerobic biodegradation to occur. Maintaining soil at somewhat less than 100 percent

of field capacity allows more rapid movement of air into the soil, thus facilitating aerobic metabolism

without seriously reducing the supply of water to microorganisms. If soils are allowed to dry

excessively, microbial activity can be inhibited or stopped; if the wilting point is reached, cells may

lyse or rupture. Continuous maintenance of soil moisture at adequate levels is of utmost

importance. Either too little or too much soil moisture is deleterious to microbial activity. Surface

drainage of the LTU can be critical in high rainfall areas. If soil is saturated more than an hour or

two, aerobic microbial action is reduced. 

Underdrainage is generally provided by a sand layer or a geotextile/drainage net layer under the

LTU. The system should be designed so that excess water quickly drains away and thus microbial

activity is not inhibited. The interface between the lift and the drainage layer underneath should be

composed of well-graded materials so that the transition from the (usually) relatively fine soil texture

of the lift to the relatively coarse texture of the drainage layer is gradual rather than sudden. Grading

of the materials reduces the tendency for the soil lift to become saturated before drainage occurs,

which inhibits aerobic biological activity. 

Types and Concentrations of Contaminants Remediable by Land Treatment

The types of contaminants most commonly treated in LTUs are petroleum compounds and organic

wood preservatives. Historically, petroleum refineries have used land treatment to dispose of waste

sludges. Although waste petroleum sludges currently are not often applied to soil for treatment, the

technology has been applied to remediation of soil contaminated with many types of petroleum

products, including fuel, lubricating oil, and used petroleum products. Land treatment has

historically been used to remediate contaminated process waters from wood preserving operations.

This technology currently is not used for this purpose but is currently used to remediate soil

contaminated with wood preserving wastes. 

Other applications for land treatment technology include remediation of soil contaminated with coal

tar wastes, pesticides, and explosives. Since coal tar wastes are similar to creosote wastes (wood

preserving creosote is made from coal tar), such wastes are considered amenable to land treatment.

Land treatment appears to be potentially useful for certain pesticides, but the evidence for

applicability of this technology to explosives-contaminated soil is inconclusive. 

Levels of Contamination Susceptible to Land Treatment

The levels of petroleum product contamination amenable to land treatment vary by waste type and

site conditions. In many cases, soils with higher levels of contaminants than are recommended for
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

6-4

land treatment can be mixed with less contaminated soils to bring contamination levels down to

recommended starting levels for treatment. Levels of petroleum product contamination as high as

25 percent by weight of soil have been reported as treatable, although experience indicates that

levels 5 to 8 percent by weight or less are more readily treated. 

Soils contaminated with 15,000 to 20,000 mg/kg dry weight creosote wastes have been treated in

soil systems, although more usual starting levels are in the 5,000 to 10,000 mg/kg range.

Pentachlorophenol wastes are rarely treated at more than 1,000 mg/kg starting levels since

pentachlorophenol is quite toxic to microorganisms at the higher levels. 

The final levels attainable also vary by waste and site conditions. Generally, once total contaminant

levels are below 50 to 200 mg/kg polynuclear aromatic hydrocarbons, remediation by land

treatment is slow, and further treatment by conventional land treatment techniques may be

ineffective. For instance, land treatment of creosote wastes is generally considered successful if total

carcinogenic polynuclear aromatic hydrocarbons are reduced to below 50 to 100 mg/kg, and

specific components are reduced to their "land ban" levels (for instance, pyrene to 7 mg/kg).

Laboratory treatability studies may be used to assess the "best case" potential for final contaminant

levels, with the assumption that actual final levels in the field would rarely if ever be lower than those

found in laboratory study.

Costs for land treatment are estimated at between $20 to $200 per cubic yard.
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Land Treatment

Daniel Pope

Dynamac Corporation

Ada, OK

Land Treatment

Biological, chemical, physical

processes transform contaminants

Degradation by Biological

Activity

● Most transformation of organic

contaminants

● Physical, chemical mechanisms also

involved

Degradation by Ultraviolet

Light

● Soil surface

● Higher PAHs

Volatilization – Low Molecular

Weight Compounds

● BTEX

● Naphthalene

● Methyl naphthalenes

Hydrolysis – Pesticides

● Amides

● Triazines

● Carbamates

● Thiocarbamates

● Nitriles

● Esters

● Phenylureas
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Humification

● Polymerization of contaminants

● PAHs known to humify

Know Thy Waste

Relative importance of processes

varies widely for different

compounds under different

circumstances

Compounds Amenable to

Land Treatment – PAHs

● 2-ring PAHs – readily degraded,

volatile, leachable

● 3-ring PAHS – degradable, leachable

● 4-ring PAHS – fairly degradable,

leachable

● 5–6-ring PAHs – difficult to degrade

Compounds Amenable to

Land Treatment – Phenols

● Penta & Tetrachlorophenol

■ Difficult over 1,000 ppm

● Other phenolics

Compounds Amenable to Land

Treatment – Hydrocarbons

● Aliphatics 1–8 C chains

■ Degradable

■ Volatile

Compounds Amenable to Land

Treatment – Hydrocarbons

● Most 12–15+ C chains

■ Slower degradation

■ Relatively immobile

■ Relatively nontoxic
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Compounds Amenable to

Land Treatment – BTEX

● Degradable

● Volatile

Compounds Amenable to

Land Treatment

● Energetics – more often composted

● Phthalates

● Pesticides

Bioremediation—

What Is It?

● Two fundamental aspects of

bioremediation . . .

● Developing large populations of

microorganisms that can transform

pollutants

● Bringing microorganisms into

intimate contact with pollutants

Land Treatment

Technology

● Contaminated soil

● Sludge application to soil

In Situ – Ex Situ Land

Treatment

● The issue is control

● Control of runoff, leachate, volatiles

In Situ – Practical Soil Depth

● Based on effective oxygen diffusion

● Bioventing for greater depths
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

In Situ

● Treat surface soil, remove

● Treat surface soil, deep till

Semi In Situ

● Remove soil to depth

● Add lifts back to excavation

for treatment

Tillage Depth

● Most tractor-mounted tilling devices

till down to one foot

● Large tractors, specialized equipment

till to three feet or more

● Large augers move soil from 50-100

feet to surface, but practicality not

fully shown

Ex Situ

● Application of lifts of contaminated

soil to prepared-bed reactor

● Clay and/or plastic liners

● Bed of porous soil

● Irrigation, drainage, and soil water

monitoring systems

● Berm

Land Treatment – Lift

Depth

● Generally limited to 6–24 inches of

soil

● Usually 12 inches or less lift depth

● Refinery LTU 36 inches or more

Soil Type

● Limited in heavy clay soils, especially

in high rainfall areas

● Oxygen transfer limitations

● Substrate availability
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Soil Type – Working With

Heavy Soils

● Shallow lifts for easier tilling, better

diffusion

● Improve tilth with bulking agents

Improving Tilth – Bulking

Agents

● Organic matter (sawdust, compost,

manures, etc.)

● Add gypsum if soil has high sodium

content

Preparing Soil for

Application

● Screen to remove debris greater than

1 in. diameter

● Remove large debris that may adsorb

waste compounds

Tilling

● Enhances oxygen infiltration

● Mixes contaminants with

microorganisms

● Disperses contaminants

Tilling

● Lower end of soil moisture

percentage range before tilling

● Tilling very wet or saturated soil

tends to destroy soil structure,

reduce microbial activity

● Wait 24 hours after irrigation or a

significant rainfall event

Tilling Schedule

● Compromise of several antagonistic

factors

● Loosens soil for oxygen access

● Destroys soil structure

● Dries soil

● Mixes contaminants and bugs

● Equipment compacts soil
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Tilling – Mixing

● Mostly along line of travel

● Till in varying directions

Tilling Equipment

● Rotary tiller for tilling, mixing

purposes

● Disk harrow often used, may not mix

soil well

● Subsoil plow, chisel plow to break up

zone of compaction

Tilling

● Subsequent lifts tilled into top 2 in. or

3 in. of previous lift

● To mix populations of well

acclimated microorganisms

● Avoids sudden transition in

permeabilities if different soil types

being remediated

Lift Application Timing

● Based on reduction to defined levels

of particular compounds or

categories of compounds

● Usually more detailed sampling to

determine finish

Nutrients, Carbon Sources,

and Other Additives

Carbonaceous (Organic)

Amendments

● Animal manures

● Wood chips, sawdust

● Straw, hay
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Carbonaceous

Amendments

● Supply carbon and some nutrients

● Act as bulking agent, adsorbent

Carbonaceous Adsorbents

● Slow migration

● May sequester contaminants

● Increase permeability—Increased

oxygen, water flux

● Increase oxygen demand due to

microbes breaking down

● Increase water holding capacity

Carbonaceous Amendments—

Application Rates

● Must be balanced with nutrients

● 3–4% by weight of soil

Carbonaceous

Amendments

● Manures often mixed with bedding—

straw, sawdust, rice hulls

● Bedding acts as bulking agent, but

also has a nutrient demand

Carbonaceous

Amendments

● Should have moderately small

particle size

● Thoroughly mixed with soil

Fertilizers

● Can cause pH to drop

● Acid forming equivalent indicated

on bag
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Fertilizers – Soluble Forms

● Can be applied through irrigation systems

● Application rates may be closely controlled

● Applications can easily be made as often as

irrigation water is applied

● Immediately available to microorganisms

● Equipment meters concentrated nutrient

solutions into irrigation system on demand

Soil Nutrient Levels

● Nutrient requirements not thoroughly

studied

● Detailed information not available to

indicate optimal levels

● Difficult to show response in field

Soil Nutrient Levels

Desired levels based on

concentration in soil, or

concentration ratio of several

nutrients

Micronutrients

● Carbonaceous amendments may

contain some micronutrients

● Trace amounts in many packaged

inorganic fertilizers

● Commercially available as

micronutrient blends

● Apply specific micronutrients only if

treatability studies show response

Proprietary Micronutrients

● Usually expensive compared with

horticultural fertilizer sources

● Generally easily supplied with readily

available horticultural fertilizers

Complex Nutrients

● Vitamins, growth factors

● Need easily shown in lab culture,

with defined media

● Difficult to show effectiveness in

field
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Bioaugmentation

● Indigenous microorganisms isolated,

cultured

● Nonindigenous microorganisms

● Genetically engineered

microorganisms

Bioaugmentation

● Nonindigenous microbes rarely compete well

enough to develop, sustain useful population

● Most soils with long-term exposure to

biodegradable wastes have indigenous

microorganisms that are effective degraders

given proper management of the LTU

● Little data from well-designed experiments to

show efficacy

● Perhaps more useful as understanding increases

Soil Moisture Control

● 40–80% of field capacity

● Usually at high end of range

Field Capacity

● Soil micropores filled with water

● Soil macropores filled with air

● Microorganisms get air and water

Soil Moisture

Maintaining 40–80% of FC allows more

rapid movement of air into soil,

facilitating aerobic metabolism without

seriously reducing supply of water to

microorganisms

Soil Moisture

● Some evidence that continuous

maintenance at high levels better

● Some evidence that low end of range

good for some compounds

● Requires careful management to

maintain any given level
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Soil Moisture

● If soils dry excessively, microbial

activity seriously inhibited, stopped

● Maintenance at proper level is not

trivial

Measuring Soil Moisture

● Gravimetric—simple, accurate, slow

● Tensiometer—simple, fairly accurate for

many soils

● Gypsum blocks—good for undisturbed

soil

● Capacitance effect—accuracy questionable

● Neutron probe—accurate, but uses

radioactive material, expensive eqipment

Surface Drainage

● Critical in high rainfall areas

● Saturation greater than one hour

greatly reduces microbial action

● Surface should be sloped 0.5–1.0%

● Greater slopes—erosion hazard

● Design to allow collection, return of

eroded soil

Internal Drainage

● Sand/gravel layer

● Geotextile/drainage net layer

Internal Drainage

Initial lifts usually placed on bed of

sand, other porous soil, which

causes a perched water table to

develop

Perched Water Table

● Lift takes up water until field

capacity achieved

● Then begins to drain excess water

● Lower part of lift layer may remain

overly wet
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Internal Drainage

● The interface between lift & drainage

layer should have well-graded

materials

● The psoil particle size transition from

lift to drainage layer should be

gradual

● Water movement through interface

enhanced with gradual transition

Internal Drainage

● Good internal drainage reduces

tendency for soil lift to become

saturated

● Interface may be graded by tilling lift

into top of drainage layer

LTU Leachate and Runoff

● Recycled onto LTU

■ With or without treatment

● Treated (biological or adsorption) and

discharged

Disposal of Treated Soil

● Replace in excavation

● Disposal cell

LT as Part of a Treatment

Train

High organics (bulking agents,

contaminants) in soil may inhibit

subsequent solidification/stabilization

for metals treatment

LT Disadvantages

● Slow—takes a long time for treatment

● High contaminant concentrations may be hard

to treat

● Low contaminant concentrations may not show

significant reduction

● Final levels may not be achievable depending on

the requirements

● Space requirements are high

● Volatiles/dust/leachate control may be difficult
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

LT Costs

● Earthmoving—$1–2+ per yard

● Containment—berm

● Monitoring—usually major part of

expense

● Operations

● Volatiles control can be very

expensive
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

7-1

Land Treatment Unit Case Study: Champion International Superfund Site

Daniel Pope

Dynamac Corporation, Ada, OK

Introduction

The Champion International Superfund Site at Libby, Montana (referred to as the "Libby Site"), is

an operating lumber mill where wood preserving operations using creosote and

pentachlorophenol (PCP) were conducted from 1946 to 1969. Soil, sediments, and ground water

at the site were contaminated with creosote and PCP wood treating solutions and wastes.

Champion International uses three biological processes for environmental remediation at the Libby

site: 1) a prepared-bed, lined land treatment unit (LTU) for treatment of excavated soil; 2) an

abovegrade, fixed-film bioreactor for treatment of extracted ground water, and 3) an oxygen and

nutrient enhanced bioremediation system for in situ treatment of the upper aquifer. As part of the

U.S. Environmental Protection Agency's (EPA's) Bioremediation Field Initiative, a team consisting

of Utah State University, EPA's National Risk Management Research Laboratory (Ada, Oklahoma),

and Dynamac Corporation conducted a performance evaluation of bioremediation systems used

by Champion International at the Libby site. 

Objectives of the LTU performance evaluation were to: 

# Describe and summarize previous and current remediation activities. 

# Develop an evaluation plan, including statistical requirements for the number,

timing, and location of samples.

# Perform a laboratory evaluation of the potential for soil microorganisms to

bioremediate soil contaminants under site conditions of temperature and soil

moisture.

# Conduct a comprehensive field evaluation to assess treatment effectiveness,

treatment rate, and detoxification of contaminated soil in the LTU. 

SUMMARY OF REMEDIATION AND MONITORING ACTIVITIES CONDUCTED BY CHAMPION

INTERNATIONAL

When full-scale soil remediation began, approximately 75,000 cubic yards of contaminated soil and

sediment at the site was excavated down to the water table from the three primary source areas at
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

7-2

the site: a former tank farm, an unlined butt-dip area, and an unlined waste pit. Rocks larger than

1 inch in diameter were removed from the excavated material and used to construct subgrade

infiltration galleries upgradient from the waste pit area where substantial residual contamination

remained in the subsurface. Effluent from the abovegrade fixed-film bioreactor was applied to the

infiltration galleries to stimulate biodegradation of any contamination adhering to the rocks, and to

allow infiltration of treated water from the bioreactor back into the subsurface to stimulate

subsurface bioremediation. The excavated soil remaining after rocks were removed (about 45,000

cubic yards) was placed into the waste pit excavation, where it is pretreated by land treatment (tilling,

irrigation, nutrient addition) prior to placement in the LTU. 

The geometric means of initial soil concentrations from all three contaminated sites are as follows:

Total carcinogenic polynuclear aromatic hydrocarbons 189.0 mg/kg

(TCPAHs)

PCP 29.0 mg/kg

Note: Maximum concentrations greater than geometric mean by factors of 6 to 90.

Target remediation levels as specified in the record of decision for soil treated in the two LTUs are

as follows:

Naphthalene 8.0 mg/kg

Phenanthrene 8.0 mg/kg

Pyrene 7.3 mg/kg

TCPAHs 88.0 mg/kg

PCP 37.0 mg/kg

LTU Cell Design

The lined, prepared-bed LTU is composed of two cells with a total area inside the outer berm

perimeter of both cells of 2 acres. The berms allow containment, treatment, and ultimate disposal

of additional contaminated soils, if required.

The bottom of the LTU cells are sloped to a central gravel drain (2 percent slope), which is sloped

to a collection sump (1 percent slope) so drainage water can be removed as needed. Leachate is

removed from the collection sump by means of an automated pump and piping system. Beneath

the drainage system is a geotextile filter underlain by a high-density polyethylene liner, which in turn

is supported by a base layer of compacted soil. 
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

7-3

Monitoring

Monitoring, conducted by Champion International, involves periodic collection and analysis of

leachate, soil, ground-water, and air samples both inside and outside treatment cells during

operation and closure periods. 

Leachate monitoring involves sampling from LTU sumps on a quarterly basis and during rainfall

events. Monitoring of LTU soil involves operational, confirmation, and compliance sampling.

Operational sampling consists of onsite laboratory analysis of contaminants during lift treatment as

well as assessing nutrient and soil moisture requirements. After operational samples indicate

contaminant target levels have been met in a lift, confirmation samples are analyzed by an offsite

laboratory to confirm attainment of contaminant target levels. Compliance samples may include

previously collected confirmation samples or additional samples, if required, to fully demonstrate

that target levels have been reached. 

Ground-water monitoring includes six wells (four downgradient and two upgradient). Monitoring of

the ground-water wells around the LTU is performed semiannually. 

Ambient air is monitored for polynuclear aromatic hydrocarbons (PAHs) and PCP by an upwind and

downwind station to characterize concentrations due to unit operations and to protect workers'

health. Moisture is applied to LTU for dust control during operation. 

Land Treatment Operations

Contaminated soils are placed in the LTU cells in 6- to 12-inch lifts for treatment during the summer.

Water is applied to the LTU to maintain adequate moisture levels (approximately 40 to 70 percent

of field capacity) in the treatment zone and for dust control. 

Nutrients (nitrogen and phosphorus) are added to the LTU dissolved in irrigation water or as solid

fertilizers applied directly to the LTU. The nutrient requirement selected was a carbon:nitrogen ratio

in the soils of approximately 12-30:1 and a nitrogen:phosphorus ratio of approximately 10:1.

Nutrients are added as frequently as every other day, depending on soil moisture and nutrient needs.

The LTU is tilled at least weekly, using a tractor-mounted rototiller. Tilling is suspended if the LTU

contains ponded water. 

LAND TREATMENT PERFORMANCE EVALUATION

Introduction . Utah State University conducted a field and laboratory performance evaluation of the

LTUs. During the performance evaluation, soil in the two LTU cells was sampled at several depths

over a 2-year period. Concentrations of the 16 priority pollutant PAH compounds and PCP were

determined. The performance evaluation was based on: 1) the changes in concentration of soil

contaminants over time to evaluate the effectiveness of remediation, 2) changes in the concentration

of soil contaminants in a lift after application of additional lifts to evaluate downward migration of
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

7-4

contaminants, 3) changes in soil toxicity as determined by bioassays to evaluate toxicity reduction,

and 4) a laboratory study of chemical, physical, and biological processes affecting soil contaminant

concentrations to determine the mechanisms responsible for remediation. 

Results . Soil sampling indicated that land treatment was able to meet the treatment goals for

reduction of contaminant concentrations in the contaminated soil, and there was no evidence of

downward migration of target PAH compounds and PCP through the LTUs. In addition, pyrene, PCP,

and TCPAH concentrations continued to decrease with time after placement of lifts in both LTUs. 

Laboratory Assessment

Two laboratory evaluations of soil microbial metabolic potential were conducted to add information

concerning biodegradation versus physical/chemical mechanisms for disappearance of

phenanthrene and PCP, e.g., volatilization and mineralization. The first laboratory evaluation was

designed to determine rates of biological mineralization and volatilization as affected by

contaminant concentration, temperature, and soil moisture. The second evaluation was designed

to provide information addressing a mass balance of radiolabeled carbon that was used to evaluate

humification of the two chemicals.

Resu lts. The laboratory studies demonstrated that both PCP and phenanthrene were partially

metabolized to carbon dioxide in the contaminated soil matrix at the site. Both were also mineralized

with the indigenous soil microorganisms at temperatures and moisture levels representative of site

conditions. It appears that significant volatilization of PCP or phenanthrene at the full-scale site is

unlikely. The laboratory evaluation corroborates the interpretation that decreases in target chemical

concentrations are due to biological processes rather than physical/ chemical processes.

Laboratory evaluations demonstrated that not all of the parent compounds were mineralized within

soil in the laboratory microcosms. Rather, carbon in the parent compounds also became distributed

among air, solvent extract, and soil-bound phases. A major pathway for C for phenanthrene and 14

PCP was humification (binding to soil), such that the compound is not solvent-extractable from soil.

A significant fraction of C was solvent-extractable from the soil, either in the form of the parent

14

compound or intermediates. Mineralization represented the third most important fraction for C in 14

this laboratory study. Volatilization of phenanthrene and PCP over the 45-day evaluation was less

than 1 percent and therefore not considered to be an important route of compound removal from

soil.

Soil Toxicity Testing . The Microtox assay was used to measure general physiological toxicity, and

the Ames assay was used to measure mutagenicity of soil solvent extracts. Toxicity assays indicated

that soil within the LTUs was detoxified to background soil levels. Average Microtox toxicity

decreased from an EC value of 6.6 initially to nontoxic (greater than 100) for all soil samples 50

tested. The initial mutagenic potential of soil applied to LTU 1 was considered to be approximately

330 revertants per gram of soil (weighted activity). Results of mutagenicity testing for Lift 1 sampled

3 months after application and biological treatment indicated detoxification to soil background

levels (less than 150 revertants per gram of soil).
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

7-5

Conclusions

The field performance evaluation of two full-scale LTUs at the Libby, Montana, Superfund site

indicated that enhanced land treatment of soil contaminated with wood preservative chemicals was

effective and resulted in the treated soil meeting target remediation levels for target contaminants

as specified in the record of decision. Downward migration of target chemicals as a result of the

application of additional lifts was not observed. The contaminated soil was detoxified to background

levels as a result of the treatment, based on the results of toxicity and mutagenicity assays. 

In summary, results of the field performance of the LTUs at the Champion International Superfund

site in Libby, Montana, indicated that bioremediation using indigenous microorganisms was the

process that accomplished soil treatment. Soil treatment included degradation of target PAH

compounds and PCP in contaminated soil to target remediation levels and detoxification of soil. 
7-6

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Land Treatment Case

Study: Libby

Superfund Site

Daniel Pope

Dynamac Corporation

Ada, OK

Land Treatment Case Study:

Champion International

Superfund Site

● Currently an operating lumber mill

● Creosote/pentachlorophenol wood

preserving from 1946 to 1969

● Soil, sediments, & ground water

contaminated with creosote and

PCP wood treating solutions,

wastes

Biological Processes For

Remediation

● Prepared-bed, lined land treatment

unit (LTU) for soil

● Above grade, fixed-film bioreactor

for extracted GW

● Oxygen/nutrient enhanced

bioremediation for in situ

treatment of the upper aquifer

U.S. EPA Bioremediation Field

Initiative Performance

Evaluation

● Utah State University

● Dynamac Corporation

● NRMRL Ada Division

(RSKERL)

● Champion International

LTU Performance Evaluation

Objectives

● Document remediation

activities

● Laboratory evaluation of

bioremediation

● Field evaluation: treatment

effectiveness and rate,

detoxification of soil

Remediation/Monitoring

Activities Summary

As conducted by Champion

International
7-7

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Full-Scale Soil Remediation

● 75,000 yards contaminated

soil/sediment excavated

● Rocks >1 inch diameter removed

● Remaining soil (~45,000 yards)

replaced in excavation

● Pretreated by “in situ” LT prior to

placement in LTU

LTU Cell Design

● Lined, prepared-bed

land treatment unit

● Two cells ~1.0 acre

each

Monitoring

(Champion International)

● LTU soil

● LTU leachate

● Ground water (6 wells)

● LTU air emissions

Land Treatment Operations

● 6- to 12-inch lifts

● Water ~40 to 70% FC

● Weekly tilling

● Discontinued during winter

Nutrients

● Applied in irrigation water or

as solids

● C:N ratio 12-30:1

● N:P ratio 10:1

● Based on TOC, TKN, total

phosphorus

LTU Performance Evaluation

Utah State

● LTU cells sampled over two–

year period

● Concentrations of 16

priority pollutant PAHs and

PCP
7-8

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Performance Evaluation:

Contaminant Concentrations

● Contaminant concentration

changes over time

● Concentration changes in a

lift after application of

additional lifts

Performance Evaluation:

Toxicity Reduction

● Microtox assay – general

physiological toxicity

● Ames assay –

mutagenicity

Performance Evaluation:

Contaminant Fate

Lab studies of chemical,

physical, and biological

processes to determine

mechanisms responsible for

remediation

● Contaminant reduction goals

met

● No evidence of downward

migration of PAHs, PCP

● Pyrene, PCP, TCPAH, decreased

after lifts covered in both LTUs

Field Evaluation Results

Laboratory Study

Objectives

Determine fate of

14C-phenanthrene and 14CPCP in LTU soil, as affected

by soil moisture, temperature

Laboratory Study

Results

PCP, phenanthrene partially

metabolized with indigenous

soil microorganisms at

temperatures and moisture

levels representative of site

conditions
7-9

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Significant volatilization of

PCP or phenanthrene in lab

study did not occur

Laboratory Study

Results

● Not all of parent compounds

were mineralized within soil in

laboratory microcosms

● Carbon in parent compounds

became distributed among air,

solvent extract, and soil-bound

phases

Laboratory Study

Results

● Major pathway for phenanthrene,

PCP was humification

● Next significant pathway was

solvent-extractable from soil

parent compound or intermediates

● Mineralization was third most

important pathway

● Volatilization was less than 1%

Laboratory Study

Results Soil Toxicity Testing

● Microtox – general

physiological toxicity

● Ames assay –

mutagenicity of soil

solvent extracts

Average Microtox

Toxicity

● Initial EC50 value of 6.6

● After treatment, EC50

value >100 (nontoxic) for

all soil samples tested

Ames Test

● Initial mutagenic potential of

applied soil ~330 revertants

per gram of soil

● Lift 1 sampled after 3 months

treatment indicated

detoxification to soil

background levels (less than

150 revertants per gram of soil)
7-10

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Conclusions: Field

Performance Evaluation

Land treatment of soil

contaminated with wood

preservatives was effective and

resulted in the treated soil

meeting target remediation levels

for target contaminants as

specified in the Record of

Decision (ROD)

Conclusions: Field

Performance Evaluation

Downward migration of target

chemicals as a result of the

application of additional lifts

was not observed

Conclusions: Field

Performance Evaluation

Contaminated soil was

detoxified to background

levels as a result of the

treatment, based on results of

toxicity and mutagenicity

assays
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

8-1

Phytoremediation

Steve Rock

Office of Research and Development, National Risk Management Research Laboratory,

U.S. Environmental Protection Agency, Cincinnati, OH

Daniel Pope

Dynamac Corporation, Ada, OK

Phytoremediation is the use of higher plants to bioremediate contamination in soil, water, or

sediments. Variations of phytoremediation that have been used in the past include wetlands to treat

municipal sewage or neutralize acidic mine drainage. Currently, phytoremediation is proposed for

remediation of both organic and inorganic contaminants in soil, sediments and water. 

Phytoremediation, as with bioremediation using microorganisms, involves the use of natural

processes to change the form or location of contaminants. Roots of higher plants take up water,

nutrients, and other compounds from soil. Water moves throughout the plant, eventually going to

the leaves and out into the atmosphere in the process of transpiration. Ongoing processes of plant

metabolism use water, nutrients, carbon dioxide, and sunlight to synthesize organic compounds,

which are moved throughout the plant for use in growth and for storage of reserves. A large

community of microorganisms thrives in contact with the plant (particularly on the root system) and

is supported to a greater or lesser degree by products of the plant. Plants may transport oxygen

down to the root system and release some of the oxygen to the soil. As the roots grow through the

soil, they form channels that can increase soil aeration, particularly as the roots die and decay,

leaving voids. As with bioremediation using natural microbial processes, it is possible to use these

natural plant processes to remediate contaminants.

Much of the biodegradation associated with certain kinds of phytoremediation occurs in a zone

around the root system called the rhizosphere (Figure 1). The rhizosphere is a zone of enhanced

microbial activity at the interface between the root and the soil. The rhizosphere supports larger

microbial populations than surrounding soil and has different types of microorganisms than

surrounding soil. The enhanced microbial activity in the rhizosphere is thought to be responsible for

degradation of certain contaminants, particularly of some organic contaminants.

The rhizosphere is a narrow zone, with a depth from a few millimeters to perhaps a centimeter. The

actual depth of the rhizosphere is hard to measure, but the "rhizosphere effect" of enhanced

microbial activity appears to diminish rapidly with distance. Since the rhizosphere is closely involved

with phytoremediation, the degree of contact that the root system has with the soil is important.

Plant root systems vary considerably, but in general most root systems can be divided into two

classes: tap root systems, with large main roots emerging from the plant base and branching to

smaller and smaller roots; and fibrous root systems, with many small roots emerging from the plant

base and also branching to smaller and smaller roots. Fibrous root systems generally have more

surface area per length of root than taproot systems. Some plants, notably grasses, have very fine,

fibrous root systems that are highly ramified throughout the soil volume they occupy. This should
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

8-2

mean that the plant roots actually contact more of the soil, and therefore their affect on remediation

should be more uniform throughout the soil volume occupied.

Plants may transport oxygen into the subsurface; lower the water table by transpiration, thereby

pulling oxygen into the soil from the atmosphere; and increase hydraulic conductivity of the soil as

roots produce channels in soil. Flood-tolerant and wetland plants are especially efficient at

transporting oxygen into the subsurface. These processes are thought to enhance aerobic

biodegradation by increasing oxygen in the subsurface.

As plants transpire, the movement of water through the plant also carries along dissolved

components (Figure 2). Dissolved contaminants such as chlorinated solvents can be removed from

the soil in the transpiration stream and emitted to the atmosphere through the plant leaves. This type

of "remediation" could be undesirable, obviously.

Many plants transpire significant quantities of water under the right conditions, but certain plants,

called phreatophytes, which ordinarily grow their roots down to the water table, can transport

relatively large quantities of water from the soil to the atmosphere. Willow and poplar species are

well known examples. Many plants, particularly the phreatophytes, can significantly influence

ground-water levels, especially in soils of low permeability. Such plants could not only remediate

the ground water by the various mechanism already discussed but also could help protect ground

water by lowering the water table below contaminated zones.

Most plants grow roots down to about 2 meters deep or less, but some plants can reach far deeper

under good conditions. Obviously it might be desirable for phytoremediation to have plants that

grow dense, highly ramified, fibrous root systems down very deep. Research is needed to determine

the depth of influence of plant root systems, and ways to encourage deeper rooting and greater soil

volume coverage.

The community of microorganisms in the rhizosphere has been shown to be involved in degradation

of numerous contaminants, including pesticides, polynuclear aromatic hydrocarbons, petroleum

compounds, volatile organic chemicals, and inorganics. Also, plants can degrade contaminants

during plant metabolic activities; for instance, 2,4,6-trinitrotoluene has been shown to be degraded

by plant enzymes. Plants can use contaminants as nutrients; nitrate contamination of ground water

can serve as a nitrogen source for plants. 

Plants can adsorb or take up and accumulate contaminants either in their roots and other

belowground parts or in aboveground parts including stems, leaves, and fruits. Plants are not able

to take up all types of contaminants; small, low molecular weight polar compounds are favored for

uptake into the plant, but large, high molecular weight lipophilic compounds tend to be excluded.

Plants may extract metals from soil and accumulate them in tissues. Accumulators of lead,

cadmium, chromium, nickel, cobalt, copper, zinc, and selenium have been found (Table 1).

Location of the accumulation site in the plant is important. Accumulation of contaminants in the

root may pose problems with removal of the contaminant from the site, since it may be impractical

to harvest the root systems and separate them from the soil. Ideally, the plant would efficiently

extract the contaminant from the soil down to very low levels and accumulate the contaminant to

high concentrations in an aboveground plant part that could be easily harvested without harming

the plant.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

8-3

Applications and Examples 

In general, phytoremediation appears to be best suited for cleanups over a wide area, with fairly

shallow contaminants in low to medium concentrations. Using plants to remediate a site can be

much less expensive than conventional cleanup options because installation and maintenance costs

are typically very low. Public acceptance of phytoremediation can be very high, in part because

of the added benefits of parklike aesthetics, including providing bird and wildlife habitat. A planted

wetland or interceptor barrier of poplar trees can remediate a chronic problem for years with little

or no attention. The cleanup time can be longer than with some physical or chemical processes,

and like most bioremediation is typically measured in months and years.

Phytoremediation has been shown to reduce concentrations of hydrocarbons from spills and leaking

underground storage tanks; polychlorinated biphenyls from transformers; pentachlorophenol and

creosote from wood preserving sites; nitrates, pesticides, and herbicides from agricultural runoff; and

chlorinated solvents like trichloroethylene from industrial processes. Some plants can extract heavy

metals such as lead, chromium, and uranium. Study in this field is relatively new, with much of the

work done on the laboratory and pilot scale, though some field work is now under way.

Wetlands constructed with reeds and cattails are used to prevent acid mine drainage from polluting

streams. The biological processes in a wetland neutralize the acidity of the water and decrease the

mobility of the metals. Poplar and willow trees are planted as interceptor barriers to remediate

ground-water contamination or to protect surface water from agricultural runoff. The roots of these

trees can "pump and treat" hundreds of gallons of water each day. Contaminants may be degraded

by the microbial community that is supported by the trees or by the tree itself. Plants such as

mustard may be used for extraction of heavy metals by taking up the contaminants into the roots,

then translocating them to the shoots and leaves. Some plants may sequester metals in the root

structure but not move them further into the plant. Alfalfa, ryegrass, and other plants are used for

in situ soil remediation. These plants encourage biodegradation of organic contaminants by

microbes by providing oxygen, nutrients, enzymes, and other key elements in the root zone of

influence or rhizosphere.

Plants are limited as to the depths that they can effectively treat. Mustard plants grow down 12 to

18 inches. Ryegrass and fescue can extend roots a few feet. Alfalfa has been found with roots down

to 20 feet. Poplar tree roots can tap a water source 10 to 20 feet down, and some claim much

deeper root depth. 

Bibliography 

1. Anderson, T.A., E.A. Guthrie, and B.T. Walton. 1993. Bioremediation. Environ. Sci.

Technol. 27(13).

2. Aprill, W., and R.C. Sims. 1990. Evaluation of the use of prairie grasses for stimulating

polycyclic aromatic hydrocarbon treatment in soil. Chemosphere 20(1-2):253(13).
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

8-4

3. Baker, A.J.M., S.P. McGrath, C.M.D. Sidoli, and R.D. Reeves. 1994. The possibility of in

situ heavy metal decontamination of polluted soils using crops of metal-accumulating

plants. Resour. Conserv. Recycl. 11(1-4):41.

4. Baker, A.J.M., and R.R. Brooks. 1989. Terrestrial higher plants which hyperaccumulate

metallic elements: A review of their distribution, ecology, and phytochemistry. Biorecovery

1:81-126.

5. Banks, M.K., G.R. Fleming, A.P. Schwab, and B.A. Hetrick. 1994. Effects of the

rhizosphere microflora on heavy metal movement in soil. Chemosphere.

6. Banuelos, G.S., G. Cardon, B. Mackey, J. Ben-Asher, L. Wu, P. Beuselinck, S. Akohoue,

1993. Boron and selenium removal in boron-laden soils by four sprinkler-irrigated plant

species. J. Environ. Qual. 22(4):786.

7. Bollag, J.-M. 1992. Decontaminating soil with enzymes. Environ. Sci. Technol. 26(10).

8. Brooks, R.R. 1972. Geobotany and biogeochemistry in mineral exploration. New York,

NY: Harper and Row.

9. Brown, S.L., R.L. Chaney, J.S. Angle, and A.J.M. Baker. 1994. Phytoremediation potential

of Thlaspi caerulescens and bladder campion for zinc- and cadmium- contaminated soil.

J. Environ. Qual. 23(6):1,151. 

10. Chaney, R.L. 1983. Plant uptake of inorganic waste constituents. In: Parr, J.F., et al., ed.

Land treatment of hazardous wastes. Noyes Data Corp., Park Ridge, NJ. pp. 5,076.

11. Cunningham, S.-0., and W.R. Berti. 1993. Remediation of contaminated soils with green

plants: An overview. In Vitro Cell. Devel. Biol. Plant 29(4):227-232.

12. Dushenkov, V., P.B.A.N. Kumar, H. Motto, and I. Raskin. 1995. Rhizofiltration: The use

of plants to remove heavy metals from aqueous streams. Environ. Sci. Technol.

29(5):1,239.

13. Entry, J.A., N.C. Vance, M.A. Hamilton, and D. Zabowski. 1994. In situ remediation of

soil contaminated with low concentrations of radionuclides. In: In situ remediation:

Scientific basis for current and future technologies. Proceedings of the 33rd Hanford

Symposium on Health and the Environment, Pasco, WA, November 7-11. Battelle Press.

p. 1,055.

14. Erickson, L.E., M.K. Banks, L.C. Davis, A.P. Schwab, N. Muralidharan, K. Reilley, and J.C.

Tracy. 1994. Using vegetation to enhance in situ bioremediation. Environ. Prog.

13:226-231.

15. Hinchman, R., and C. Negri. No date. The grass can be cleaner on the other side of the

fence. Logos 12(2):8.
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16. Lee, E., and M.K. Banks. 1993. Bioremediation of petroleum-contaminated soil using

vegetation: A microbial study. J. Environ. Sci. Health Environ. Sci. Eng. 28(10):2,187.

17. Licht, L.A., and J.L. Schnoor. 1990. Poplar tree buffer strips grown in riparian zones for

biomass production and nonpoint source pollution control. In: Proceedings of the

American Society of Agricultural Engineers, Paper 902057. pp. 1-21.

18. Pierzynski, G.M., J.L. Schnoor, M.K. Banks, J.C. Tracy, L. Licht, and L.E. Erickson. 1994.

Vegetative remediation at superfund sites. In: Hester, R.E., and R.M. Harrison, eds. Mining

and its environmental impact—issues in environmental science and technology, Vol. 1.

Royal Society of Chemistry. pp. 46-69.

19. Raskin, I., P.B.A.N. Kumar, S. Dushenkov, and D.E. Salt. 1994. Bioconcentration of heavy

metals by plants. Current Opinion in Biotechnol. 5:285.

20. Schnoor, J.L., L. Licht, S.C. McCutcheon, N.L. Wolfe, and L.H. Carreira. 1995.

Phytoremediation of organic and nutrient contaminants. Environ. Sci. Technol. 29(7):318A.

21. Stomp, A.M., K.H. Han, S. Wilbert, and M.P. Gordon. 1993. Genetic improvement of tree

species for remediation of hazardous wastes. In Vitro Cell. Devel. Biol. Plant

23F(4):227-232.

22. Walton, B.T., and T.A. Anderson. 1990. Microbial degradation of trichloroethylene in the

rhizosphere: Potential application to biological remediation of waste sites. Appl. and

Environ. Microbiol. 4:1,012-1,016.
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Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Phytoremediation

Daniel Pope

Dynamac Corporation

Ada, OK

Growing plants to clean

contamination from soil,

water, or sediments

Early Indications of

Phytoremediation Potential

● Plants have been used for

prospecting for

minerals—Geobotany

● Wetlands have been found to

neutralize acidic mine drainage

Certain plants can help

degrade contaminants,

others can take up

contaminants

Figure 1. Hypothetical

Mechanism

Figure 2.

Diagram of

Phytoremediation
8-7

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Table 1. Examples of Metal

Hyperaccumulators

Metal in Dry

Weight of Original

Metal Plant Species Leaves (%) Location

ZN Thlaspi calaminare <3 Germany

Viola species 1 Europe

Cu Aeolanthus biformifolius 1 Zaire

Ni Phyllanthus serpentinus 3.8 New Caledonia

Alyssum bertoloni >3 Southern

and 50 other alyssum Europe and

species Turkey

Sebertia acuminata 25 (in latex) New Caledonia

Stackhousia tryonii 4.1 Australia

Pb Brassuca juncea <3.5 India

Co Haumaniastrum robertii 1 Zaire

Mature cottonwood or

poplar will pump and

treat 25 to 300 gallons of

water per day

Phytoremediation Project for

the Chevron Ogden Terminal

by Phytokinetics Treating TPH in soil with

grass and alfalfa; Treating

TPH in ground water with

poplar and juniper

Site Map

Phytoremediation uses

slightly modified standard

agronomic practices
8-8

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Treatability study in

greenhouse to determine

best species for site

Schematic

of Soil

Column

Phytoremediation in Soil Is

Best Applied to:

● Soil: widespread, fairly shallow,

low to medium concentration

contamination

● Ground water: shallow (to 20'

easily, some claim deeper)

Treatment

Depth Grasses

18–24 in.

Mustard

18 in.

Alfalfa

48 in.

Poplar Trees

to 20 ft

Advantages of

Phytoremediation

● Less expensive with low

installation and maintenance

cost

● High public acceptance

● Can clean chronic pollution

sources (i.e., acid mine seeps)

Disadvantages of

Phytoremediation

● At least 2–3 years for cleanup

● Most contaminants not tested

extensively except for

hydrocarbons, pesticides, and

agricultural nutrients
8-9

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Field Experience

Field-scale demonstrations on

hazardous waste are underway in:

Oregon Utah California

Texas Ohio Virginia Maryland
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

9-1

Development and Application of Composting Techniques for Treatment of

Soils Contaminated With Hazardous Waste

Carl L. Potter

Office of Research and Development, National Risk Management Research Laboratory,

U.S. Environmental Protection Agency, Cincinnati, OH

Introduction

Historically, composting has been used to degrade solid waste materials such as leaf litter, sewage

sludge, and food wastes. More recently, composting has been investigated as a remediation

technology for hazardous wastes (1). Laboratory and field-scale work has been conducted to

determine the fate of polycyclic aromatic hydrocarbons (2) and explosives (3) in the composting

environment. Composting is not generally employed to treat heavy metals or other inorganics,

although it may be applicable to inorganic cyanides. Other studies have indicated that composting

is potentially effective in degrading or transforming petroleum hydrocarbons (4, 5) and pesticides

(6) to environmentally acceptable or less mobile compounds.

Process Description

Optimum conditions for composting may vary depending on a number of factors, but generally 40

to 60 percent moisture content, a carbon-to-nitrogen ratio of 20:1 to 30:1, and aerobic conditions

are considered best. Bulking agents may consist of sawdust, corn cobs, straw, hay, alfalfa, peanut

hulls, or other organic materials.

The aerobic compost process passes through four major microbiological phases, identified by

temperature: mesophilic (35E to 55EC), thermophilic (55E to 75EC), cooling, and maturation. The

greatest microbial diversity has been observed in the mesophilic phase. Microbes found in the

thermophilic phase have been spore-forming bacteria (Bacillus spp.) (7) and thermophilic fungi (8,

9). Microbial recolonization during the cooling phase brings the appearance of mesophilic fungi

whose spores withstood the high temperatures of the thermophilic phase. Composting can be

anaerobic, but most methods use aerobic conditions. 

Composting can be performed in windrows, where material is put into rows and periodically turned;

aerated static piles, where perforated pipes within the pile supply air; and vessels, where material

is periodically mixed inside an aerated containment vessel.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

9-2

Future Research

Despite promising studies, the ability of composting to completely degrade synthetic organic

compounds has not been fully demonstrated. Although composting systems have been used to

biodegrade some hazardous compound, few studies (mostly bench-scale) have provided mass

balance closures or fully investigated all of the intermediate products, final products, and byproducts

of the composting process. The lack of mass balance closure and conclusive evidence of the fate

of contaminants in field-scale applications is not unique to composting. Many other technologies

(both ex situ and in situ) lack conclusive evidence of contaminant fate in field-scale applications.

Future investigations will include technical developments necessary to improve composting

applications for degradation of hazardous waste. This will involve increased application of pilot-scale

composting systems in addition to ongoing research in bench-top composters. Emphasis will be

placed on developing techniques for trapping volatile organic compounds from pilot-scale systems,

determining mass balance of contaminant degradation in the compost, and identifying microbial

species responsible for biodegradation of contaminants.

Future studies will also attempt to validate extrapolation of results from bench-top to pilot-scale and

field demonstration levels. Maintaining a bench-top system at optimum conditions is relatively easy

compared with a large-scale composter where optimum conditions will not prevail at all times. The

degree of variance from optimal conditions requires investigation and approximation in small-scale

systems.

References

1. Ziegenfuss, P.S., and T.R. Williams. 1991. Hazardous materials composting. J. Haz. Mat.

28:91-99.

2. U.S. EPA. 1995. On-scene coordinator's report: Removal action at the Indiana

Woodtreating Corporation Site, Bloomington. Site ID# R.D. Draft.

3. U.S. Army Corps of Engineers, Toxic and Hazardous Materials Agency. 1991.

Optimization of composting for explosives contaminated soil. Final Report No. CETHA-TSCR-91053. November. 

4. U.S. EPA. 1995. Bioremediation in the field. EPA/540/N-95/500. August.

5. Moore, R.E. 1992. Enhanced bioactivity treats hydrocarbon-contaminated soils. Natl.

Environ. J. January/February:34-37.

6. Michel, F.C., C.A. Reddy, and L.J. Forney. 1995. Microbial degradation and humification

of the lawn care pesticide 2,4-dichlorphenoxyacetic acid during the composting of yard

trimmings. Appl. Environ. Microbiol. July:2,566-2,571.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

9-3

7. Nakasaki, K., M. Sasaki, M. Shoda, and H. Kubota. 1985. Change in microbial numbers

during thermophilic composting of sewage sludge with reference to CO evolution rate. 2

Appl. Environ. Microbiol. 49(1):37-41.

8. Strom, P.F. 1985. Identification of thermophilic bacteria in solid-waste composting. Appl.

Environ. Microbiol. 50(4):906-913.

9. Fogarty, A.M., and O.H. Tuovinen. 1991. Microbiological degradation of pesticides in

yard waste composting. Microbiol. Rev. June:225-233.
9-4

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Composting

Presented by

Gregory Sayles or Dolloff F. Bishop

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, Ohio
9-5

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Windrow Compost System

Windrow

Mobile Composter

Schematic Diagram of

Extended Aerated Pile

Unscreened

or Screened

Compost

Bulking Materials and Sludge

Perforated

Pipe

Trap for

Water

Fan

Filter Pile

Screened

Compost

Composting Extended Piles with Forced Aeration
9-6

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Static Pile Composter

Nutrients

Aeration

Microorganisms

Soil

Side View

Top View Asphalt Plastic Piping

(Compatible with Contaminants)

Visqueen

Cover

In-Vessel

Composter

CO2

Mixer Infeed

Composting

Mix

Air Outfeed
9-7

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation
9-8

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Conclusions

● Composting reduced soil concentrations

of PAHs over a 10-week treatment period

● 30% bulking as effective as 50% for

remediation of PAH during first 10 weeks

● PAH degraders withstood temperature as

high as 56°C

Field Example

Indiana Woodtreating Corp. Site

● 22,000 tons of PAHcontaminated soil

● Soil screened to

remove rocks >3

inches

Indiana Woodtreating

Corp. Site

● Each 100 tons mixed with:

■ 5 rolls straw

■ 5 bails horse manure

■ 200 lbs. urea fertilizer

■ 100 lbs. ammonium nitrate

fertilizer (34-0-0)

● Soil treated in 9 piles

Indiana Woodtreating

Corp. Site

● Initial total soil

PAH (TPAH): 20,410 mg/kg

● Action levels:

TPAH 500 mg/kg

Each carcinogenic PAH 100

Indiana Woodtreating

Corp. Site

Results of Test

● After 1 year of

composting: TPAH <500 mg/kg

● Additional 1 year

of treatment

using land

farming: TPAH <100 mg/kg
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

10-1

Biopile Treatment of Soils Contaminated With Hazardous Waste

Carl L. Potter

Office of Research and Development, National Risk Management Research Laboratory, 

U.S. Environmental Protection Agency, Cincinnati, OH

Introduction

Biopile systems offer the potential for cost-effective treatment of contaminated soils. Like

composting, biopiles provide favorable environments for indigenous microorganisms to degrade

contaminants present in the soil matrix. Although similar to compost piles, these systems differ in

that lesser quantities of bulking agents are used in biopile units. Air is supplied to the biopile system

through a system of piping and pumps that either forces air into the pile under positive pressure or

draws air through the pile under negative pressure (1). Depending on the contaminants in the soil,

conditions are established in the biopile to favor either anaerobic or aerobic microorganisms. In

some cases, exogenous microbes, such as fungi, may be added to the biopile to enhance

contaminant degradation.

Field studies have indicated biopile successes in remediation of soils contaminated with

pentachlorophenol (2) and petroleum hydrocarbons (3). Costs of soil bioremediation using biopiles

range from $30 to $100 per ton of soil, depending on soil conditions and the biodegradability of

contaminants.

Process Description

Biopile structure resembles a static pile compost system. Conceptually, one may think of a biopile

as an ex situ bioventing system in that aeration usually involves forcing air through the soil by

injection or extraction through perforated pipes. Volatile organic compound emissions can be

controlled by aerating the pile with negative pressure and venting off gases into a small compost pile

or biofilter (1).

Optimum conditions for biopiles vary depending on the type of soil, climate conditions, and the

chemical and biological attributes of the soil. Because biopile treatment is an ex situ technology,

most conditions can be controlled to achieve an acceptable range of conditions. Generally,

moisture content between 40 and 85 percent of soil field capacity, a carbon-to-nitrogen ratio of

10:1 to 100:1, and pH between 6 and 8 are acceptable depending on soil conditions. Organic

amendments can be used to increase the water-holding capacity of poor soils. 

Wood chips may be added as bulking agents to increase soil porosity and promote aeration and

irrigation. Sawdust or straw can be added to supply carbon. Animal manure (1 to 4 percent w/w)

can supply both carbon and nutrients.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

10-2

Future Studies

Future studies are needed to evaluate the applicability of biopile technology and to optimize systems

for treating an increased variety of contaminants. Alternating between anaerobic and aerobic

conditions may provide a mechanism for degrading heavily chlorinated organic compounds via

reductive dehalogenation combined with oxidative mineralization (4). 

Also, soil microbiology and fungal treatment will receive increased focus in the future. Fungal

technology appears promising for biodegradation of recalcitrant contaminants (5). Fungi do not

generally metabolize contaminants; degradation occurs extracellularly by enzymes excreted by the

fungi. Much research remains to be done to identify the fungal strains most capable of degrading

specific contaminants.

References

1. Lei, J., J.-L. Sansregret, and C. Benoit. 1994. Biopiles and biofilters combined for soil

cleanup. Poll. Eng. June:56-58.

2. McGinnis, B., R.R. DuPont, and K.E. Everhart. 1992. Determination of respiration rates

in soil piles to evaluate aeration efficiency and biological activity. Presented at the 85th

Annual Meeting of the Air and Waste Management Association, Kansas City, MO, June 21-

26. 

3. Moore, R.E. 1992. Enhanced bioactivity treats hydrocarbon-contaminated soils. Natl.

Environ. J. January/February:34-37.

4. Sims, J.L., J.M. Suflita, and H.H. Russell. 1991. Reductive dehalogenation of organic

contaminants in soils and ground waters. In: EPA Ground Water. EPA/540/4-90/054.

5. Glaser, J.A., and R.T. Lamar. 1995. Lignin-degrading fungi as degraders of

pentachlorophenol and creosote in soil. In: Bioremediation: Science and Applications.

SSSA Special Pub. No. 43:117-133.
10-3

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Biopiles

Aerated Static Soil Piles for

Treatment of Shallow

Contaminated Soil

Presented by

Gregory Sayles or Dolloff F. Bishop

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, Ohio

Schematic Diagram of

Extended Aerated Pile

Unscreened

or Screened

Soil

Nutrient Materials

Perforated

Pipe

Trap for

Water

Fan

Filter Pile

Screened

Compost

Extended Soil Piles With Forced Aeration

Biopile Systems

● Potential to provide cost-effective

treatment

● Provide a favorable environment

for indigenous aerobic or

anaerobic microorganisms

● Similar to compost piles

● Air delivery system

● Nutrient enhanced

Biopile Design

Pile Size

● Height = 3 to 10 feet

● Width is unrestrited unless pile is turned

■ 6 to 8 feet if turned

Land Requirements

● Amount of soil treated/Pile height

● Additional land required for:

■ Berms

■ Access

■ Sloping terrain

Aeration Equipment

● Blowers or fans

● Aeration piping in pile lifts

● Turning equipment if pile is turned

Biopile Construction

● Site preparation

■ Clearing and grading

● Berms, liners, and covers (if needed)

● Piping

■ Moisture addition

■ Nutrient addition

■ Aeration (if forced air)

Biopile Design (continued)

Leachate Management

● Collection

● Treatment

Soil Pretreatment

● Shredding

● Blending

■ Amendments

■ Bulking agents (increase porosity)

■ pH adjustment

Biopile Design (continued)
10-4

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Biopile Soil Conditions

Moisture 40% < Field capacity < 85%

pH 6 < pH < 8

Temperature 10°C < Temperature < 45°C

C:N:P 10:1:0.5 < C:N:P < 100:10:1

Heavy metals <2,500 ppm

Economic Considerations

Electricity input

● 2 hp blower running at 2 psi

● $50–$75 per month per pile

Analytical Monitoring

● Chemical

● Biological

Bioremediation Cost

● Type of contaminants (biodegradability)

● Contaminant concentrations (time required)

● Typically $50 to $100/ton of soil

Advantages of Biopiles

● Simple to design and implement

● Low cost ($50–$100/ton)

● Require less land area than land

farming

● VOC emissions can be controlled

Field Example

● Former wood treating site in

southeastern U.S.

● PCP-contaminated soil

● Biopiles compared to land

treatment in an effort to save

space on site
10-5

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Conclusion

Vented soil piles are as effective if

not more effective than landfarms
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

11-1

Effective Treatment of Hazardous Waste Constituents in Soil by

Lignin-Degrading Fungi

John A. Glaser

Office of Research and Development, National Risk Management Research Laboratory,

U.S. Environmental Protection Agency, Cincinnati, OH

Introduction

The diversity of fungi and their remarkable ability to degrade complex and persistent natural

materials (Table 1) such as lignin exemplify the host of useful features (1) found with these

organisms. In contrast to bacteria, fungi are able to extend the location of their biomass through

hyphal growth in search of growth substrates. Lignin-degrading fungi have been investigated for their

enzymatic activity to degrade aromatic organic chemicals, which are structurally related to the

composition of lignin. Enzymes involved in lignin breakdown are extracellular and have low substrate

specificity. Fungi can thoroughly colonize soil and show exceptional tolerance to high concentrations

of toxic pollutants. Chemical structural similarities and expected reactivities between lignin and

organic pollutants have fostered the consideration of these fungi as potential pollutant degraders.

White rot fungi are unique in their ability to transform all components of native lignin to carbon

dioxide and water. Lignin is constructed of an amorphous polymeric network that resists attack by

many microbes. Three major classes of oxidative enzymes designated, lignin peroxidases (LIPs),

manganese-dependent peroxidases (MnPs), and laccases, play an important role in the fungal

degradation of lignin. All three enzymes can oxidize phenolic compounds, thereby creating phenoxy

radicals. Nonphenolic aromatic compounds, however, are oxidized via cation radicals. Laccase can

oxidize nonphenolic compounds with relatively low ionization potential, while nonphenolics with high

oxidation potential are readily oxidized by LIPs and MnPs.

Pollutant Degradation

Extensive lists of xenobiotic organic chemicals currently considered degradable by lignin-degrading

fungi have been compiled. Contaminant categories to which lignin-degrading fungi have been

applied are wood-treating/town gas chemicals, munitions, and pesticides and other chlorinated

organic chemicals. Fungal bioremediation is an emerging technology that has been applied in the

field only to wood treating wastes (pentachlorophenol and creosote). Application to other

contaminants requires field evaluation.

Field-Scale Evaluation

Application of fungal treatment in beds of contaminated soil (2) was studied at an Oshkosh,

Wisconsin site (Figure 1). The contamination was a wood preservative formulation composed of 5
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

11-2

percent pentachlorophenol (PCP) in mineral spirits. Soil concentrations of 1 to 4,435 mg/kg to

depths of 30 cm were determined through extensive sampling. Blended soil, with the larger stones

and rocks removed, was added to each soil bed. Two fungi (Phanerochaete chrysosporium and P.

sordida) were selected as candidate treatment species (Table 2) for the evaluation. The fungi were

added to the contaminated area using spore inoculated/infested wood chips with the appropriate

fungal strain. The pentachlorophenol concentration (Table 3) was depleted by 82 percent for P.

chrysosporium and 85 percent for P. sordida, after 56 days of treatment, despite temperatures that

dipped below the temperature range considered optimal for these fungi. P. sordida is a known soil

inhabitant and can tolerate lower soil temperatures than P. chrysosporium. P. sordida is known to

have a lower optimum temperature (30EC) than P. chrysosporium (40EC).

Some of the decrease in PCP is by methylation-producing pentachloroanisole (PCA), the methyl

ether of PCP (Table 4). PCA accumulation in the treatment plots was monitored and did not increase

with time, suggesting that degradation of PCA occurs in the inoculated soil. Transformation of PCP

to PCA is evident in both liquid and soil cultures and seems to compete with other PCP

transformation reactions (i.e., oxidation). In laboratory soil cultures (3) inoculated with P.

chrysosporium, the amount of soil-bound versus an organic extractable PCP-transformation product,

later identified as PCA, was greatly influenced by soil type. PCP oxidation may be enhanced further

by identifying the soil conditions that favor oxidation over transformation to PCA.

Another treatment effectiveness study (Figure 2) for fungal treatment of PCP-contaminated soil (Table

5) was conducted at an abandoned wood treating site at Brookhaven, Mississippi. The field study

(Figure 3) was a two-phase field assessment. The first phase (4) was designed (Table 6) to evaluate

the ability of three different fungal species to deplete PCP in soil. P. sordida was superior in its ability

to deplete PCP in soil. The results for depletion of PCP by P. sordida paralleled the results of the

Wisconsin study, where the inoculation with either P. chrysosporium or P. sordida was applied to soil

contaminated with 250 to 400 Fg/g PCP. In the Brookhaven study, P. sordida treatment (Figure 4)

resulted in an overall decrease of 88 to 91 percent at PCP concentrations of 672 mg/kg in 6.5

weeks. P. chrysosporium treatment reduced PCP by 67 to 72 percent in multiple soil beds at PCP

concentrations greater than 1,000 mg/kg.

The Brookhaven site was also contaminated with 4,017 Fg/g of total polynuclear aromatic

hydrocarbons (PAHs), other components of creosote. The effects of solid-phase bioremediation with

P. sordida (with two control treatments) on soil concentrations of 14 priority pollutant PAHs (5) were

determined over a 56-day period.

Depletion of both three- and four-ring PAH analyses (Table 7) in P. sordida-treated soil was greater

than in the controls. Concentrations of the three-ring analyses decreased by an average of 31

percent after 7 days and by an average of 911 after 56 days. Four-ring analyses were more

persistent; losses first became apparent between 14 and 28 days of treatment, and an average of

45 percent was depleted after 56 days. Five- and six-ring analyses were the most recalcitrant

species, persisting at original levels throughout the course of the study. The persistence of these

compounds in soil is due to their low bioavailability when bound to soil particles. Depletion of fivering analyses of PAHs, however, have been reported by some researchers under conditions providing

a higher fungus:contaminant ratio than that used in this evaluation.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

11-3

A larger scale demonstration (Figure 5) of the P. sordida treatment (6) was conducted as the second

phase of the study. Inoculation of the soil with a 10-percent dry weight inoculum consisting of fungal

hyphae and growth substrate reduced PCP soil concentrations of greater than 1,000 mg/kg by 64

percent after 20 months of treatment (Figure 6). The two control soil beds showed reductions of 18

and 26 percent of the PCP soil concentration.

Low initial amounts (Table 8) of fungal biomass, measured by ergosterol analysis, may have

contributed to the reduced performance. Heavy rains and weather-related modification to the tilling

schedule may also have limited the performance of the P. sordida treatment.

P. chrysosporium ATCC 24725-based treatment was applied to 6,000 cubic meters of soil

contaminated with a mixture of chlorophenols, known as KY-5, at a site in Finland (7, 8). Initial

concentrations of total chlorinated phenols decreased with depth of excavated soil layers ranging

from 203 to 38 mg/kg. Contaminant composition of the constructed fungal treatment piles varied

with the order of excavation. Soil contaminant reduction depended on the initial contaminant

concentration. Concentrations of total chlorinated phenols between 173 and 203 mg/kg were

reduced by 85 and 90 percent after 20 months of treatment (Table 9). After only 12 weeks,

chlorophenol concentrations of 38 to 84 mg/kg were reduced by 80 to 90 percent to target

endpoints of less than 10 mg/kg. One of the piles produced poor contaminant depletion kinetics,

which was attributed to soil processing and pile construction.

Conclusions

Removal of PCP has now been demonstrated (Table 6) in a strongly acidic (pH 3.8) Mississippi clay

soil and in alkaline (pH 9.6) Wisconsin sandy gravel soil. This strongly supports the potential of fungi

for treating organic pollutants in a wide range of soils having varied physical and chemical

characteristics.

In the Mississippi test, P. sordida was capable of reducing an initial soil PCP concentration of 672

mg/kg by 89 percent using a 101 inoculum loading level by dry weight. The depletion of three-ring

and four-ring analyses of PAHs (total measured PAHs, 4,017 ppm) by P. sordida was also

promising, with reductions of 85 to 95 percent and 24 to 72 percent, respectively. These

percentage depletions for PCP and the PAH analyses were, in the Mississippi test, obtained after

only 56 days of experimentation.

References

1. Glaser, J.A., and R.T. Lamar. 1995. Lignin-degrading fungi as degraders of

pentachlorophenol and creosote in soil. In: Bioremediation: Science and applications. SSSA

Special Publication 43. Soil Science Society of America. pp. 117-133.

2. Lamar, R.T., and D.D. Dietrich. 1990. In situ depletion of pentachlorophenol from

contaminated soil by Phanerochaete ssp. Appl. Environ. Microbiol. 56:3,093-3,100.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

11-4

3. Lamar, R.T., J.A. Glaser, and T.K. Kirk. 1990. Fate of pentachlorophenol (PCP) in sterile

soils inoculated with the white-rot basidiomycete Phanerochaete chrysosporium:

Mineralization, volatilization and depletion of PCP. Soil Biol. Biochem. 22:443-440.

4. Lamar, R.T., J.W. Evans, and J.A. Glaser. 1993. Solid-phase treatment of

pentachlorophenol-contaminated soil using lignin-degrading fungi. Environ. Sci. Technol.

27:2,566-2,571.

5. Davis, M.W., et al. 1993. Field evaluation of the lignin-degrading fungus Phanerochaete

sordida to treat creosote-contaminated soil. Environ. Sci. Technol. 27:2,572-2,576.

6. Lamar, R.T., et al. 1994. Treatment of a pentachlorophenol- and creosote-contaminated

soil using the lignin-degrading fungus Phanerochaete sordida: A field demonstration. Soil

Biol. Biochem. 26:1,603-1,611.

7. Holroyd, M.L., and P. Caunt. 1994. Fungal processing: A second generation biological

treatment for the degradation of recalcitrant organics in soil. Land Contamin. Reclam.

2:183-188.

8. Holroyd, M.L., and P. Caunt. 1995. Large-scale soil bioremediation using white rot fungi.

In: Hinchee, R.E., J. Fredrickson, and B.C. Alleman, eds. Bioaugmentation for site

remediation. Columbus, OH: Battelle Press. pp. 181-187.
11-5

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Effective Treatment of

Hazardous Waste

Constituents in Soil by

Lignin-Degrading Fungi

Presented by

Gregory Sayles or Dolloff F. Bishop

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, Ohio

Table 1. Rationale for

Fungal Biotreatment

● Enzyme systems capable of degrading

complex natural aromatic polymers

● Chemical structure of natural

polymers resemble many organic

pollutants

● Fungi have the ability to reach remote

areas of the soil by extension of

hyphae

Selection Criteria

● Powerful oxidizing enzymes

■ Extracellular

■ Broad range substrate specificity

■ Multiplicity of isoenzymes

● Ability to move throughout the soil

● Genetic Stability

Classes of Oxidative

Enzymes

● Lignin peroxidases

(LIPS)

● Manganeses–dependent

peroxidases (Mn Ps)

● Laccases

Contaminant Categories Where

Lignin-Degrading Fungi

Applied

● Wood treating wastes*

● Town gas chemicals

● Munitions

● Pesticides and other

chlorinated organics

* Only waste having significant field testing

Figure 1. Wisconsin Site Layout
11-6

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Wisconsin Soil

Characteristics

Characteristic Value

Texture Gravel/sand

pH 9.6

Pollutant conc. 250–400 mg/kg

CEC 17.22

Total carbon (%) 8.95

Sulfur (%) 0.14

Table 2. Wisconsin

Treatment Systems

 Inocula Sterile Organic

Conditions P. chrysosp. P. sordida chips matter

Treatment A1 + – + +

A2 – + + +

Controls B – – + +

C – – + –

D – – – +

E – – – –

Table 3. Wisconsin PCP

Decrease

 Percent PCP Decrease

Conditions Day 8 Day 15 Day 29 Day 46

A1 9.1 33.3 70.6 82.3

A2 9.7 42.2 75.9 85.8

B 4.9 13.7 20.9 27.5

C 0.5 –10.0 7.1 16.2

D 15.3 26.1 10.7 3.0

E 10.9 13.8 23.8 19.1

 Percent PCP Converted to PCA

Conditions Day 1 Day 15 Day 29 Day 46

A1 1.3 13.1 13.0 14.1

A2 0.8 6.6 9.4 9.1

B 0.8 1.4 1.1 0.7

C 1.3 2.3 1.4 1.5

D 0.5 0.9 0.6 0.6

E 0.6 0.9 0.8 0.7

Table 4. Wisconsin PCA

Conversion

Figure 2. Brookhaven Site Location

Central

Brookhaven

Table 5. Mississippi Soil

Characteristics

Characteristic Value

Texture Sandy Clay

pH 3.8

Pollutant conc. PCP 429-5,200 mg/kg

(ave.) 2,355 mg/kg

Total carbon (%) 2.2

Total nitrogen (%) 0.04
11-7

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Figure 3. Unit Processes in Site

Preparation Table 6. Mississippi

Experimental Design

Amendment Quantity (dry wt)

P. chrysosporium 5.0% and 10.0%

P. sordida 10.0%

P. chrysosp./T. hirsuta 5.0% each

T. hirsuta 10.0%

P. chrysosporium 13.0%

P. chrysosporium 10.0%; 3.0% (day 14)

No treatment, wood chip,

 and inoculum controls —, —, 10.0%

Figure 4. Treatment

Performance Table 7. Transformation of

PAHs

 % Decrease

Init. Conc. No Treatment Carrier P. sordida

Compound (mg/kg) Control Control Treatment

Acenapthene 429 49 68 95

Phenanthrene 941 69 49 90

Anthracene 684 57 48 285

Fluoranthene 972 23 42 72

Chrysene 90 6 14 233

Figure 5. Demo Treatment Plot

Perspective

Figure 6. Pentachlorophenol

Depletion

Demonstration Study
11-8

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Table 8. Ergosterol

Evaluation

 Conc. (mg/kg)

Found Expected

Inoculum 241 —

Raw soil 0.2 —

Inoculated soil 4 24

Table 9. Transformation of

Chlorinated Phenols

Finland Field Application (20 Month Treatment)

Treatment Init. TOLX Conc.* Init. TCP Conc.* P. chrysosporium

Bed (mg/kg) (mg/kg) Pile pH Treatment Removal

A 2,727 203 7.1 85%

B — 173 — 94%

C — 84 — —

D 816 38 7.7 —

*TOLX = Toluene extract; TCP = Total Chlorophenols

Fungal Treatment

Summary

● Treatment of pentachlorophenol occurred for

concentrations greater than 1,000 mg/kg

● Consistent transformations values for PCP of 80

to 90% occurred for the Wisconsin and

Mississippi sites

● Soil pH does not apparently affect the fungal

treatment because pH values for the sites ranged

from 3.5 to 9.2

● Fungal treatment in 56 days efficiently

transformed three-ring PAHs by 85-95%; four-ring

PAHs by 24-72%
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

12-1

Slurry Bioreactors for Treatment of Contaminated Soils, Sludges, and

Sediments

Paul McCauley and John Glaser

Office of Research and Development, National Risk Management Research Laboratory,

U.S. Environmental Protection Agency, Cincinnati, OH

Introduction

A slurry bioreactor may be defined as a containment vessel and apparatus used to create a threephase (solid, liquid, and gas) mixing condition to hasten the biodegradation of soil-bound and

water-soluble contamination as a water slurry of the contaminated soil, sediment, or sludge and

biomass (usually indigenous bacteria) capable of degrading targeted contaminants. 

Advantages and Limitations

Bioremediation of contaminated soils, sludges, and sediments using slurry bioreactors offers several

advantages over other remediation technologies:

# Intimate contact between microbiota and contaminants combined with process

controls such as (but not limited to) pH, temperature, and nutrients provide

conditions favorable for rapid remediation of targeted contaminants.

# Since most reactor vessels fully contain the contaminated solid and liquid fractions,

they offer almost unlimited treatment flexibility. Nutrient amendments, which in

some cases may not be permitted in situ (such as ammonium and nitrate), may be

used in a slurry bioreactor. Other amendments that can be used in slurry

bioreactors include designer bacteria, surfactants, and enzyme inducers. Slurry

bioreactors may be fitted to provide sequential anaerobic/aerobic treatment

conditions. Slurry bioreactors may fit into various treatment trains, which must

include particle size separation (most slurry bioreactors do not accept particles

larger than 1⁄4 inch in diameter) and commonly include soil washing. Slurry

bioreactors can be operated in batch mode (at least 10 percent of the slurry should

be reserved for seeding subsequent batches), or several bioreactors can be

sequentially linked for continuous or semicontinuous operation.

# Most bioreactor vessels fully contain the contaminated solid and liquid fractions and

can be designed to contain volatile contaminants; they offer a high degree of safety

as related to contaminant containment.

# Slurry bioreactors require a relatively small space compared to technologies such

as land treatment, biopiles, and composting. Many slurry bioreactors may be

mounted on trailers and transported for use at several sites.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

12-2

Slurry bioreactors also have limitations:

# Bioslurry is an ex situ process, which by definition requires excavation and transport

(even if only a few feet) of the contaminated waste.

# Reactor mixers consume energy. 

# Slurry bioreactors generally will not accept particles larger than 1⁄4 inch in diameter,

requiring soil sieving or some other type of particle size separation. Sand particles

are highly abrasive in slurry bioreactors, shorten their operating life, and generally

contain a small fraction of the contamination. Operators often choose

hydrocycloning for sand fraction rejection.

# Bioslurrys require dewatering after remediation is terminated.

# There is a limited history of full-scale bioslurry operations. Although there are many

pilot studies, slurry bioreactors are not easily scaled upward in size. Some

investigation or experimentation may be required to achieve optimal operating

conditions in a full-scale operation. These limitations will increase the cost of

remediation by slurry bioreactors.

Waste Streams

Contaminants that have been successfully remediated using slurry bioreactors include wood treating

waste, oil separator sludge, munitions, pesticides (not including highly chlorinated pesticides), and

halogenated aromatic hydrocarbons. Slurry bioreactors have been used most frequently to remediate

creosote.

Case Study

OHM, Inc., conducted large-scale slurry bioreactor remediation of creosote-contaminated lagoon

solids stabilized with fly ash (total polycyclic aromatic hydrocarbons [PAHs] of 11 g/kg). Extensive

classification of contaminated solids was accomplished and included screening and hydrocycloning.

Slurry bioreactors with a 750,000-liter operating capacity were used to treat a 20-percent slurry.

The results were mixed with 82 to 99 percent remediation of the three- to four-ring PAHs and 34

to 78 percent remediation of the five- to six-ring PAHs.

Bibliography

1. Berg, J.D., T. Bennett, B.S. Nesgard, and A.S. Eikum. 1993. Slurry phase biotreatment of

creosote-contaminated soil. In: Speaker abstracts: In Situ and On-Site Bioreclamation, the

Second International Symposium, San Diego, CA.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

12-3

2. Cioffi, J., W.R. Mahaffey, and T.M. Whitlock. 1991. Successful solid-phase bioremediation

of petroleum-contaminated soil. Remediation 373-389.

3. Glaser, J.A., and P.T. McCauley. 1993. Soil slurry bioreactors: A perspective. In: Speaker

abstracts: In Situ and On-Site bioreclamation, the Second International Symposium, San

Diego, CA.

4. Griffin, E.A., G. Brox, and M. Brown. 1990. Bioreactor development with respect to process

constraints imposed by bio-oxidation and waste remediation. Appl. Biochem. Biotechnol.

24/25:627-635.

5. Irvine, R.L., J.P. Earley, and P.S. Yocum. 1992. Slurry reactors for assessing the treatability

of contaminated soil. In: Deutsche Gesellschaft fur Chemisches Appartwesen. Frankfurt,

Germany: Chemische Technik und Biotechnologie e.V. pp. 187-194.

6. Jerger, D., D.J. Cady, S.A. Bentjen, and J.H. Exner. 1993. Full-scale bioslurry reactor

treatment of creosote-contaminated material at southeastern wood preserving Superfund

site. In: Speaker abstracts: In Situ and On-Site Bioreclamation, the Second International

Symposium, San Diego, CA.

7. Luyben, K.Ch.A.M., and R.J. Kleijntjens. 1992. Bioreactor design for soil decontamination.

In: Deutsche Gesellschaft fur Chemisches Appartwesen. Frankfurt, Germany: Chemische

Technik und Biotechnologie e.V. pp. 195-204.

8. Mahaffey, W.R., and R.A. Sanford. 1991. Bioremediation of PCP-contaminated soil: Bench

to full-scale implementation. Remediation 305-323.

9. Ross, D. 1990. Slurry-phase bioremediation: Case studies and cost comparisons.

Remediation 617N.

10. Smith, J.R. 1991. Summary of environmental fate mechanisms influencing bioremediation

of PAH-contaminated soils, technical report. Remediation Technologies, Inc., Pittsburgh, PA.

11. Smith, J.R. 1989. Adsorption/Desorption of polynuclear aromatic hydrocarbons in

soil-water systems. Technology Transfer Seminar on Manufactured Gas Plant Sites,

Pittsburgh, PA.

12. Stroo, H.F. 1989. Biological treatment of petroleum sludges in liquid/solid contact reactors.

EWM World 3:9-12.

13. Stroo, H.F., J.R. Smith, M.F. Torpy, M.P. Coover, and R.A. Kabrick. No date.

Bioremediation of hydrocarbon-contaminated solids using liquid/solids contact reactors.

Technical report. Remediation Technologies, Inc., Kent, WA.

14. U.S. EPA. 1992. Contaminants and remedial options at wood preserving sites.

EPA/600/R-92/182. Cincinnati, OH.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

12-4

15. U.S. EPA. 1990. Engineering bulletin: Slurry biodegradation. EPA/540/2-90/076.

Cincinnati, OH.

16. U.S. EPA. 1989. Innovative technology: Slurry-phase biodegradation. OSWER Directive

9200.5-252FS.
12-5

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Slurry

Bioreactors

Presented by

Gregory Sayles or Dolloff F. Bishop

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, Ohio

Slurry Bioreactors

For the treatment of

contaminated soils,

sludges, and sediments

A Slurry Bioreactor

Water

and

Soil

1. Enhanced process control

2. Faster rates of biodegradation of contaminants are

possible

3. Better physical contact between pollutants and

microorganisms

4. Distribution of nutrients, gases (air, oxygen), and

other materials for support of biological process

is greatly improved

5. Optimal soil, sediment, or sludge particle size

distribution can be selected

6. Liquid phase organic solubilities may be enhanced

by surfactant application

Advantages of Slurry Bioremediation

Bioreactor Feed Characteristics

● Solids particle size: <200 mesh

● Solids content in slurry:

10-30% (w/w)

● Total organics: <10% (w/w), i.e.,

no free product

● pH 4.5–9.0

1. Particle size distribution

2. Texture/composition (silt, clay, sand)

3. Soil nutrients (nitrogen, phosphorous)

4. pH

5. Cation exchange capacity (CEC)

6. Metals (speciated)

7. Total organic carbon

Contaminated Soil Characterization

Requirements
12-6

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Process Components

Small

Clean Big Soil

Big Big

Soils

or

Sludges

Pre-Slurry

Treatment

if Needed

HydroCyclone

Size

Classification

Process Components (continued)

Water

Small

Particles

Big

Particles

Clean

Soil

Clean

Water

Slurry

Bioreactor

Treatment

if needed

S/W

Separator

Reactor Configurations

● Batch (most common)

● Sequenced batch

■ Anaerobic—aerobic

■ Long-short residence time

Types of In-Vessel Mixing

● Impeller

● Airlift (rising air bubbles

induce slurry circulation)

● Combination of above

Slurry Bioreactor Mixing

Air Air

Candidate Waste Streams

● Soils, sediments, and sludges

associated with:

■ Wood treating waste (PAHs, PCP)

■ Oil/water separators

■ Munitions

■ Pesticides

■ Halogenated aromatic hydrocarbons
12-7

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Creosote

Contaminated

Soils and Sludges

Cape Fear

Wood Preserving

Fayetteville, NC

Full Scale

Predesign

Site Contamination Status

Fennema

Excavating

Byron Center, MI

Pri Mart #7

Buchanan, MT

Soil Contaminated

With Fuel

Hydrocarbons (PAHs)

Soil Contaminated

With Fuel

Hydrocarbons (PAHs)

Full Scale

Underway

Full Scale

 Underway

Examples of Slurry Bioreactor Use in

the U.S.

Examples of Slurry Bioreactor Use in

the U.S. (continued)

Soil Contaminated

With Fuel

Hydrocarbons (PAHs)

Wseco Oil #37

Muskegon, MI

Full Scale

 Underway

Site Contamination Status

Moss-American

Milwaukee, WI

Lone Star Army

Ammunition Plant

Texarkana, TX

Creosote

Contaminated Soils

and Sludges

TNT, TPHs

Full Scale

Predesign

Full Scale

Predesign

Sheridan

Disposal Services

Hempstead, TX

PCBs and Other

Assorted Organic

Pollutants

Full Scale

Predesign

Field Example: Southern Wood

Preserving, Canton, MS

● Creosote contaminated lagoon

solids, stabilized with fly ash

● pH 6–8

● Used extensive size classification

● Bioreactor uses impeller and

airlift mixing

Canton Site Layout

Water

Treatment

Slurry Reactors

Soil

Prep

Soil Drying Bed

Source 

Soil

Contaminated Material

Size Fractions

 Quantity

Fraction Size (yd3) Tons

Large Debris + 6 inch 150 165

Power Screen Rejects –6 + 1/2 inch 300 330

Shaker Screen Rejects –1/2 + 12 mesh 1,500 1,825

Hydrocyclone Rejects –12 + 200 mesh 1,500 1,825

Material for Treatment –200 mesh 7,050 9,995

TOTAL 10,500 14,140

OHM Canton Site Reactor

Air 

Supply

Impeller

Floating Mixer

Soil 

Slurry

Diffuser

Assembly
12-8

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Diffuser 

System

Floating 

Mixer

Topside Detail

OHM Canton Site Reactor Reactor Operating

Conditions

Volume (L) 750,000

Impeller Speed (RPM) 900

Air Flow Rate (Scfm) 350+/–100

Solids Loading % 20

Reactor Operating

Conditions (continued)

Temperature (C) 30+/–10

pH (S.U.) 7.2+/–1.0

DO (mg/L) >2.0

Ammonia Nitrogen (mg/L) 60+/–20

Phosphorous (mg/L) 20+/–10

Retention Time ?

Canton Site Treatment Results

PAH Treatment

Treatment

Initial Final Effectiveness

3 RING

Acenaphthene 909 ± 230 6 ± 3 99

Acenalthylene 93 ± 81d 15 ± 5 82

Anthracene 1,950 ± 530 121 ± 59 94

Fluorene 630 ± 283 14 ± 6 97

Phenanthrene 1,031 ± 661 34 ± 23 96

Canton Site Treatment

Results (continued)

Treatment

Initial Final Effectiveness

4 RING

Benzo(a)anthracene 280 ± 51 12 ± 5 95

Chrysene 296 ± 59 36 ± 11 90

Fluoranthene 1,708 ± 395 32 ± 7 98

Pyrene 1,148 ± 252 33 ± 12 97

Canton Site Treatment Results

PAH Treatment

Treatment

Initial Final Effectiveness

5 & 6 RING

Benzo(b)fluoranthene 321 ± 34 208 ± 54 52

Benzo(k)fluoranthene Combined with Benzo(b)fluoranthene

Benzo(g,h,i)perylene 92 ± 82 18 ± 12 43

Benzo(a)pyrene 130 ± 52 79 ± 15 34

Dibenzo(a,h)anthracene 92 ± 82 9 ± 6 78

Indeno(2,3-cd)pyrene 94 ± 79 31 ± 5 46
12-9

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Canton Site: Cost of Operation Only

Cost Soil Slurry

Category Preparation Treatment

Labor/Equipment $30–35 $10–15

Supplies/Utilities $20–25 $25–30

Analytical Support <$5 $5–10

TOTAL $50–60 $40–55

Cost for Full-Scale Slurry-Phase Bioremediation of RCRA 

K001 Waste Per Ton of Contaminated Soil

Canton Site: Cost of Project

Components

Unit Task Cost*

Treatability Testing $200,000

Predesign Engineering $100,000

Slurry Treatment $800,000

Slurry Dewatering $700,000

Site Preparation and Closure $400,000

Administration and Support $500,000

TOTAL (Price per ton) $190–200

Project Costs for Full-Scale Application of Slurry Treatment

to K001 Contaminated Soil 

*Cost rounded to nearest $100,000.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

13-1

Fixed Film Bioreactors

Dolloff F. Bishop and Richard C. Brenner

Office of Research and Development, National Risk Management Research Laboratory,

U.S. Environmental Protection Agency, Cincinnati, OH

Introduction

Fixed film bioreactors have become conventional technology for treating biodegradable

contaminants in air and water. Principal fixed film bioreactor applications include treatment of

industrial wastewaters, leachates or ground water, and air emissions of volatile organic compounds

(VOCs). In the reactors, biological activity usually converts contaminants to innocuous end products

such as carbon dioxide, methane, and water. Conventional fixed film reactor approaches involve

aerobic, aerobic co-metabolic (with aliphatic and aromatic organic inducers), and anaerobic

metabolism. Emerging reactor approaches also include sequential anaerobic/aerobic metabolism.

Fixed film bioreactors use either fixed, expanded, or fluidized beds of inert or adsorptive media to

support the biofilm's biodegradation of contaminants. Practical inert media include plastic, stone,

sand, wood, and ceramics. Contaminant removal from the air or water is achieved through biofilm

sorption. Adsorptive media, typically peat or granular activated carbon (GAC), remove contaminants

from the air or water through both biosorption and physical adsorption. While highly efficient

adsorptive media such as GAC are expensive, the high adsorptive capacity provides improved

protection to the biofilms by limiting microbial inhibition from toxic contaminants while increasing

contaminant removal efficiencies, especially during treatment startup. GAC media also improve

biosystem response to widely varying contaminant concentrations. 

Representative Reactor Systems

Many contaminants can be biodegraded using aerobic metabolic or co-metabolic pathways. A few,

however, require anaerobic conditions for efficient biodegradation. Selection and design of reactor

systems depend on several factors: contaminant biodegradation kinetics, contaminant sorptive

properties, metabolic or co-metabolic pathways of the individual contaminants, contaminant

concentration(s), and reactor system temperature and pH. Representative reactor systems include

aerobic fluidized-bed GAC filters (1, 2), anaerobic expanded- or fluidized-bed GAC filters (3-5) for

aqueous streams, and biofilters (6-8) for contaminated air.

Aerobic fluidized-bed GAC filters (Figure 1) are best suited for low to moderate concentrations of

contaminants such as typically found in ground water and leachates. These filters can treat slowly

aerobically degradable, poorly biosorbable, or inhibitory contaminants. Some contaminants will

require the addition of appropriate co-metabolites for efficient biodegradation. Where only

aerobically degradable (metabolic and co-metabolic) and noninhibitory contaminants are found in

the aqueous stream, however, fixed film bioreactors with inert media may be used.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

13-2

Envirex Ltd. and Envirogen Ltd. employ, before the inlet to the bioreactor, efficient pure oxygen

contacting approaches, with oxygen recycle that limits stripping of VOCs into the gas phase and

prevents difficult-to-control three-phase flow in the bioreactor. With aqueous stream recycle,

transferred dissolved oxygen is sufficient to meet the biological oxygen demand (BOD) of groundwater contaminants.

Anaerobic expanded- or fluidized-bed GAC filters (Figure 2) are best applied to moderate to highstrength aqueous waste streams such as leachates and industrial wastewaters. In these waste

streams, most contaminants are at least slowly anaerobically biodegradable. Highly halogenated

contaminants and aromatic contaminants with multiple nitro groups (munitions), however, are

recalcitrant or require a co-metabolite for aerobic degradation. The presence of these compounds

requires or favors anaerobic biotreatment. A significant advantage of anaerobic fixed film

bioreactors is that oxygen does not have to be transferred to the aqueous stream, producing

substantial operating cost savings, especially for high BOD streams. A major disadvantage is that

slow anaerobic degradation rates for many compounds mean bigger reactors are required.

Air biofilters use two alternative reactor approaches: biofilters (Figure 3) with natural media (e.g.,

peat, compost, wood bark) and trickling biofilters (Figure 4) with inert or adsorptive media and

continuous recycling of nutrients and buffer solutions. Commercial peat and compost biofilters

require efficient air humification to maintain biofilm activity and to prevent irreversible channeling

of the bed, which causes bypassing of VOCs into the filter's effluent air stream. High contaminant

concentrations (greater than 100 parts per million volume) at ambient temperatures produce

plugging of commercial biofilters by excess biomass. Periodic (1- to 5-year) media replacement in

commercial biofilters is also required because of consumption of available nutrients and

deterioration of media structure.

Trickling biofilters, an emerging technology, use recycling of nutrient and buffer solutions to support

metabolic activity and maintain desired reactor pH. These biofilters can treat higher loadings (800

to 1,000 parts per million volume) but require media cleaning at the high loadings to prevent filter

plugging and excessive pressure loss. Cleaning of ceramic pellet media through regular hydraulic

backwashing has been successfully demonstrated at pilot scale. Cleaning of complex media

structures is under study.

Novel media designs (Figure 5) to permit treatment of all VOCs have also been evaluated, typically

at bench scale. Carbon coating of inert media or carbon pellets produces improved filter

performance for slightly soluble VOCs. VOC permeable silica gel pellets with retarded oxygen

transport and with encapsulated biomass produce sequential anaerobic/aerobic treatment. Partial

dehalogenation of perchloroethylene (PCE) and trichlorethylene (TCE) occurs in the pellet core.

Then, aerobic degradation of the daughter products (e.g., vinyl chloride) occurs in the outer zone

of the pellet. Sodium formate is added to the nutrient and buffer solution to provide an energy

source for the dehalogenation.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

13-3

Performance and Costs

Aerobic fluidized-bed GAC bioreactors treating typical contaminant concentrations in ground water

efficiently remove most contaminants. As an example, in a reactor (Table 1) with a 5-minute

hydraulic residence time (HRT), concentrations of benzene, toluene, ethylbenzene, and xylenes (BTEX)

were reduced (1) from 5,420 to 64 parts per billion (98.9 percent removal). Benzene removal

exceeded 99.9 percent (less than 1 part per billion residual benzene). Anaerobic fluidized-bed GAC

bioreactors (5) treating moderate- to high-strength leachate (Table 2) produced highly efficient

removals (98 to 99 percent of chlorinated aliphatic VOCs, 85 to 97 percent of aromatic and ketone

VOCs, and 97 to 99 percent removal of semivolatile organic compounds) at HRTs of 3 to 12 hours.

Commercial biofilters (Table 3) with natural media (6) very efficiently remove soluble aerobically

degradable VOCs, such as alcohols, ketones, and phenols; efficiently remove moderately soluble

aerobically degradable VOCs, such as BTEX; and minimally remove slightly soluble or aerobically

recalcitrant VOCs, such as pentane, cyclohexane, PCE, and TCE. Trickling biofilters with adequate

retention time and appropriate media very efficiently treat all types of VOCs (Table 4). Examples of

performance with hydraulic backwashing to control pressure losses are shown in Figures 6 through

8.

The costs of these fixed film systems (Figures 9 through 12) vary depending on the application

characteristics. Capital costs are generally competitive with alternative technologies such as activated

carbon adsorption, but operating costs, especially long term, are substantially lower than those of

alternative technologies. 

References

1. Hickey, R.F., et al. 1990. Combined biological fluid bed-carbon adsorption system for BTEX

contaminated ground-water remediation. Paper presented at the Fourth National Outdoor

Action Conference on Aquifer Restoration, Groundwater Monitoring and Geophysical

Methods, Las Vegas, NV.

2. Hickey, R.F., et al. 1993. Applications of the GAC-FBR to gas industry wastewater streams.

Paper presented at the Sixth International IGT Symposium on Gas, Oil and Environmental

Biotechnology, Colorado Springs, CO.

3. Suidan, M.T., et al. Anaerobic treatment of a high strength industrial waste bearing

inhibitory concentrations of 1,1,1-trichloroethane. Water Sci. Tech. 23:1,385-1,393.

4. Suidan, M.T., et al. 1987. Anaerobic treatment of coal gasification wastewater. Water Sci.

Tech. 19:229-236.

5. Suidan, M.T., and R.C. Brenner. 1996. Expanded-bed GAC anaerobic bioreactors— an

innovative technology for treatment of hazardous and inhibitory wastes. In: Sikdar, S., and

R. Levine, eds. Bioremediation: Principles and practices. Lancaster, PA: Technomic

Publishing Company. In press.
Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

13-4

6. Leson, G. 1996. Biofilters in practice. In: Sikdar, S., and R. Levine, eds. Bioremediation:

Principles and practices. Lancaster, PA: Technomic Publishing Company. In press. 

7. Govind, R., and D.F. Bishop. 1996. Biofiltration for treatment of volatile organic

compounds (VOCs) in air. In: Sikdar, S., and R. Levine, eds. Bioremediation: Principles and

practices. Lancaster, PA: Technomic Publishing Company. In press.

8. Leson, G., and A.M. Winer. 1991. Biofiltration: An innovative air pollution control

technology for VOC emissions. J. Air Waste Mgmt. Assoc. 41:1,045. 
13-5

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Fixed Film

Bioreactors

Dolloff F. Bishop or Gregory Sayles

Office of Research and Development

National Risk Management Research Laboratory

U.S. Environmental Protection Agency

Cincinnati, OH

Fixed Film Bioreactors

for Air and Water

● Fixed, expanded, and fluidized

beds

● Aerobic metabolism

● Aerobic co-metabolic metabolism

● Anaerobic metabolism

● Sequential anaerobic/aerobic

metabolism

Fixed Film Support Media

● Inert media – plastic, stone, sand,

wood, ceramics, and glass

● Adsorptive media – granular activated

carbon, peat compost, resins

● Contaminant removal – inert media by

biosorption and biodegradation,

adsorptive media by biosorption,

physical adsorption and

biodegradation

Bioreactor Selection and

Design Criteria

● Contaminant biodegradation

kinetics

● Contaminant sorptive properties

● Contaminant metabolic pathways

● Contaminant concentrations

● Reactor system temperature and pH

Figure 1. Aerobic Fluidized-Bed

GAC Filter GAC-Fluid Bed Advantages

● Low ppb residuals in effluents

● Small size

● No off gas

● Good stability

● No carbon regeneration
13-6

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Figure 2. Anaerobic Expanded

or Fluidized-Bed GAC Filter

Figure 3. Commercial

Biofilters

Figure 4. Trickling Biofilters Commercial Biofilter

Characteristics

● VOC destruction unlike some

control technologies

● Some VOC poorly removed

● Low energy usage

● Efficient moisture control essential

● Plugging at high VOC loading

● Periodic media replacement

Trickling Biofilter

Characteristics

● Destruction of all VOCs

● Recycling of nutrient and buffer

solution

● Low energy usage

● Media cleaning at high VOC

loadings

● No media replacement

Figure 5. Novel Media Designs

Porous Ceramic and

Carbon Coated Media Silica Gel Pellets

Aerobic Zone

Wire Mesh Anaerobic Zone
13-7

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Table 1. BTEX Removal in a

Fluidized-Bed GAC Reactor

Influent Effluent

Compound (ppb) (ppb) % Removal

Benzene 1,100 >1 >99.9

Ethylbenzene 137 >1 >99.9

Toluene 1,079 1.3 99.9

P,M Xylenes 751 5.1 99.3

O-Xylenes 234 0.7 99.7

Table 2. Anaerobic GAC

Bioreactor Performance

Influent

Compound Conc (mg/L) % Removal

Perchloroethylene 20 >99

Chlorobenzene 1.1–20 >85

Penta chlorophenol 1.3–20 >99

Methyl Isobutyl-Ketone 10 >94

Naphthelene 30 >99

Table 3. Commercial Biofilter

Performance

Compound Removal*

Aliphatic hydrocarbons Low-moderate

Aromatic hydrocarbons Moderate-high

Alcohols, aldehyeds, and High

 ketones

Sulfur compounds Moderate-high

Chlorinated hydrocarbons Low-moderate

 (low concentrations)

*High = >95%, Moderate = 85-95%, and Low = >85%

Table 4. Trickling Biofilter

Performance

Influent Conc.

Compound (ppmv) % Removal

Toluene 430 >99

Methylene Chloride 150 >99

Trichloroethylene 25 ~35 (>99)*

Ethylbenzene 20 >99

Chlorobenzene 40 >95

*Addition of co-metabolite phenol to nutrient and buffer

solution.

Figure 6. Biofilter Performance on

BTEX Removal

Figure 7. Biofilter Performance on

Individual BTEX Components
13-8

Seminar Series on Bioremediation of Hazardous Waste Sites: Practical Approaches to Implementation

Figure 8. Typical Toluene Removal Recovery

Following Biofilter Backwashing Cycle

Figure 9. Life Cycle Cost

Comparison

Figure 10. Cost Comparison

Figure 11. Comparison of Total Capital

Investment (TCI) for Biofilters (Three

Residence Times) and RTO

Figure 12. Comparison of Energy

Cost for Biofilters and RTO