GROUNDBREAKING TECHNOLOGY: IN-SITU ANAEROBIC BIOREMEDIATION

FOR TREATMENT OF CONTAMINATED SOIL AND GROUNDWATER

Kelly A. Fernandes, Undergraduate Student, Northeastern University, Boston, MA, 02115,
USA
Abstract
Anaerobic in-situ bioremediation is a technique often used to cleanse contaminated soil and
groundwater. "Anaerobic in-situ bioremediation" is a phrase with distinct terms all having
relevance in the application of this technique. "Anaerobic mplies the absence of dissolved
oxygen, while "in-situ" simply means that the environmental cleansing occurs with out
removing, displacing, or significantly disturbing the specimen or surrounding area.
"Bioremediation" is a term used to describe the biological use of microbes or plants to
detoxify the environment. In order to properly implement this complex process, one must
have an understanding of microbiology, biochemistry, genetics, metabolic processes, and
structure and function of natural microbial communities.
Keywords: bioremediation, groundwater treatment, in-situ
BACKGROUND
d anaerobic in-situ bioremediation has been an accepted practice in groundwater clean up only
since the 1980's, but has been proven to be very effective for treating soil and groundwater
contamination all over the world. Anaerobic in-situ bioremediation makes use of organisms to
remove pollutants from contaminated sites. The application of biological treatment will lead to
clean up of the hazardous chemicals, which are usually organics. Typically indigenous
microorganisms consume and transform the compound until the pollutant is completely
degraded, and the wastes are harmless. Bacterial or fungal strains can also be implemented
rather than indigenous microorganisms. In conclusion, in-situ bioremediation is responsible
for satisfying the following objective:
Transforming toxic materials into non-toxic chemicals and compounds without
producing additional toxins in the process.
Bioremediation treats several classes of chemicals: Fuel Hyrdrocarbons, PAH's
(polychlorinated aromatic hyrdrocarbons), crude oil compounds, coal compounds, PCBs
(polychlorinated biphenyls), chlorinated solvents (i.e. TCE), pesticides, and many more.
Anaerobic bioremediation is particularly useful in the biodegradation of persistent wastes and
chemical compounds, like TCE, PCB's, and Fuel Hydrocarbons.
There are several reasons why bioremediation is the preferred method for soil and
groundwater contamination treatment. First, bioremediation is usually a more cost- effective
solution compared to other alternatives. Bioremediation costs about that of incineration,
and saves approximately 60% to 90% of landfill costs. In addition to these cost benefits,
contaminated soil and groundwater can be treated simultaneously, further reducing possible
operating costs. Employing an in-situ approach to the bioremediation is also an advantage in
itself. In-situ treatment reduces contaminant exposure to personnel. Also, contaminant
transportation and disposal accidents are completely eliminated when implementing in-situ
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treatment. Over all, in-situ bioremediation has few environmental impacts and worries, since
complete degradation of contaminants occurs in the subsurface. Therefore, making in-situ
bioremediation an ideal method of contamination clean up for leaking underground storage
tanks, where fuels and oils are the contaminants, like many environmental issues that we are
encountering in the twenty-first century.
Techniques
There are two techniques being currently used around the world for the implementation of in-
situ bioremediation. These two different techniques are dependent on several factors. One of
paramount importance are microorganisms. Existing anaerobes in the subsurface, versus the
lack there of, can determine the technique used for this process. Biostimulation is the process
where existing anaerobes are stimulated with enzymes and catalysts to speed up metabolic
processes. When useful anaerobes are not present in the subsurface, they must be added. This
is called bioaugmentation. Both of these techniques are successful and efficient. The two
techniques must only be differentiated between because of existing conditions in the
contaminated zone. This type of data would be collected in the initial stage of bioremediation:
Site Investigation and Feasibility Studies (see below).
THEORY
Most bioremediation projects rely on the introduction of an "electron acceptor" into the
contaminated zone to stimulate the growth of microorganisms which will digest contaminants
and render non-toxic wastes. There are several chemicals and compounds that can serve as
electron acceptors (energy sources) and allow the organisms to metabolically degrade the
contaminants. Oxidation and reduction (redox) processes occur in the subsurface and consume
organic matter. Often times in anaerobic conditions, denitrification is the most efficient
bioremediation process used. See Table 1 below.
G is a measure of the energy yield for these processes. It is clear that aerobic oxidation has a
high-energy yield, but very low solubility in water, resulting in a slower reaction. To
remediate a large volume would require limitless amounts of atmospheric oxygen to get
acceptable results, which would be very expensive. Denitrification has a high-energy yield as
well, and NO
-
3
solubility in water is very high, requiring much less of a nitrate solution to
receive acceptable results. In addition, aerobic remediation isn't always a possibility depending
on the contaminant. Anaerobic treatment is much more effective when degrading persistent
wastes. In most situations, the preferable method for anaerobic bioremediation is by
denitrification. The following equation is more appropriate for modeling the denitrification
process (the above equations depict ideal laboratory conditions and distilled water):
C
6
H
12
O
6
+ 4NO
3

-
6CO
2
+ 6H
2
O + 2N
2

Denitrification is the bacterial mediated reduction of nitrate and nitrite to nitrogen gas. Most
denitrifying bacteria are anaerobes and will only proceed in the absence of dissolved oxygen
(this condition is typical in the subsurface).
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Table 1: Redox reactions with accompanying data relative to bioremediation
Source: Proceedings of the 1998 Petroleum Hydrocarbons and Organic Chemicals in
Groundwater: Prevention, Detection, and Remediation, 1998.

Redox Reaction Simplified
Equation
Electron Acceptor G (kJ) Solubility (mg/L)

Aerobic
Oxidation
CH
2
O+O
2
=CO
2
+
H
2
O
O
2
502 11
Denitrification CH
2
O+NO
3
-
+H
+
=N
2
+HCO
3
+ H
+
+ H
2
0
N0
3
-
477 930000
Manganese
Reduction
CH
2
O+2MnO
2
+
3H
+
= 2Mn
2
+HCO
3
+ 2H
2
O
MnO
2
340 Insoluble
Iron Reduction CH
2
O+ 7H
+
+

4Fe(OH)
3
=4Fe
2+

+HCO
3
+ 10H
2
O
Fe(OH)
3
116 Insoluble
Sulfate
Reduction
CH
2
O+SO
2-
4
= HS
-

+ HC0
3
+ H
+
S0
4
-
105 2000
Methane
Fermentation
CH
2
O + CO
2
= CH
4

+ CO
2
CO
2
93 Miscible
DESIGN PROCESS
Prior to design and implementation of a remediation project there are two very important
stages: Compliance Analysis, and Site Investigation and Feasibility Studies. Both of these
steps of the general design procedure also outline the relevant information and data needed for
design.
Compliance Analysis involves examining the regulations governing environmental quality,
clean up techniques, and remediation goals or MCLs for the region.
Site Investigation and Feasibility Studies both involve the gathering of initial data. Site
investigations tend to be more general and less "scientific." Feasibility Studies use site-specific
data to determine whether this particular type of treatment is even a viable option, while Site
Investigation just involves gathering the initial findings about the site: Where? What? How?
The following is typical data acquired both in Site Investigation and Feasibility Studies:
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Type and volume of material to be treated
Transport and fate characteristics of waste
Source of pollution
Concentrations and distribution of contaminant
Microbiology background:
Naturally occurring bacteria in subsurface
If not, how to select anaerobes
What type of catalyst for microorganisms
Biodegradability of contaminants
Soil type and properties
Environment suitable for growth of bacteria
Chemical reactivity of contaminants with respect to:
Electron acceptor
Enzymes
Site topography
Groundwater flow
Geologic properties
Role of Bacteria
It is only appropriate to now discuss the importance and characteristics of bacteria. How is it
that a simple, single-celled organism can eliminate such hazardous wastes? First, only special
types of bacteria that can carry on bacterial digestion in some way can be considered for this
type of treatment. Some of these bacteria are found only in a specific environment and require
specialized types of food. In order to properly carry out waste digestion, bacteria must:
Consume organic wastes.
Digest the waste completely and quickly with out producing odors and toxic gas.
Not be pathogenic.
Grow and reproduce in the given environmental conditions of the contaminant.
Under ideal conditions, bacteria can produce new generations every 20-30 minutes,
reproducing logarithmically. Eventually, these special bacteria species completely deplete
their food source (the contaminant), sometimes with the added help of enzymes as catalysts.
For these reasons, it is necessary to have a strong background in microbiology. Identifying the
appropriate bacteria, either to add to the subsurface, or naturally occurring, is an important
step in the remediation process.
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How to Obtain Information Needed for Design
With regard to obtaining information for the compliance analysis, much of data can either be
obtained by referencing town or city by-laws, as well as state and local Departments of
Environmental Protection and by using microbiology, chemistry, geology, and geotechnical
texts and readings for research.
For the remaining data required for design, See Table 2 below.

Table 2: Parameters and techniques for acquiring data, necessary prior to design and
implementation.

Perameter How/Technique Relevant Info & Data

Source of Pollution monitoring wells, inspection
industries in area, varying
concentrations
Type & Volume of
Material to be Treated
monitoring wells, soil/water
analysis & total plate
counts(from lab)
depth, lateral distances of
contamination
Transport & Fate
Characteristics
soil properties (boring
tests/logs ), NAPL (light or
dense)
presence of confining layers vs.
porous media/permeability
Concentrations &
Distributions
monitoring wells, soil/water
analysis (from lab)
monitoring wells, soil/water
analysis (from lab)
Biodegradability
chemical properties,
biodegration potential
presence & accessibility of readily
transformed chemical bonds
Soil Type & Properties
boring tests/logs, geotechnical
references

Chemical Reactivity organic chemistry references
other similar Bioremediation
projects may lend useful insight
Site
GeologylHydrogeology
survey data, contour maps,
boring logs/tests, water table
records
survey data, contour maps, boring
logs/tests, water table records
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EXPERIMENTAL PROCEDURES
Theoretical and practical values of how much of the electron acceptor will be needed to
degrade the contaminant can vary significantly. For example, theoretically it is
known that one mole of a nitrate solution can degrade one mole of hydrocarbons. Practically,
however this is not true. There are competing biochemical reactions occurring in the
subsurface, so one mole of nitrate will not react with one mole of hydrocarbons. There is no
one way to model the subsurface and these reactions occurring in the laboratory. However,
due to prior implementation of bioremediation projects, and smaller pilot scale experiments in
the field, it has been determined that a more appropriate and practical figure is ten to fifteen
moles of nitrate will react with one mole of hydrocarbons, to result in complete degradation. If
the ratio of electron acceptor to contaminant for complete degradation can not be
approximately predetermined, the monitoring system -will be useful in noticing little change in
contaminant concentrations. If this is the case, more may be needed to completely degrade the
contaminant.
Also, total heterotrophic plate counts, experimentally determined at the site, are very useful in
examining the current microbial activity in the subsurface. This will lead to the decision of
either stimulating an existing bacterial colony, or in the lack there-of, implementing them to
the subsurface.
Research is also currently being conducted on the efficiency and effectiveness of
bioremediation for other contaminants, as well as varying microorganisms. Algae and
genetically engineered microorganisms may serve as prime candidates in the future for
bioremediation projects. With further experimentation and pilot scale projects, we may also
see degradation of heavy metals as a possibility.
CONCLUSION
In-situ bioremediation is a preferred approach to detoxifying contaminated soil and
groundwater. It incorporates microorganisms, nutrients, and enzymes in a natural process in
which the organisms consume and digest organic compounds and render the waste completely
non-toxic. Usual by-products of bioremediation include water, carbon dioxide, methane, and
hydrogen gas, all of which in small amounts, lead to minimal environmental impacts and
hazards. Bioaugmentation (adding microbes and nutrients to the subsurface) and
biostimulation (adding nutrients and enzymes to stimulate biological activity to existing
microorganisms) are the two typical techniques used to implement the bioremediation process.
The advantages of in-situ bioremediation outweigh the disadvantages: it is cost effective and
presents less danger to personnel on-site. It also eliminates transportation and disposal
accidents, resulting in less environmental impacts. Bioremediation is especially useful for
degrading petroleum and hydrocarbon contamination. Degradation of metals, in addition to the
use of algae and genetically engineered organisms is also currently being researched. In-situ
bioremediation is clearly successful and cost-effective. It has been used around the world since
only the 1980's, but in that time has demonstrated its overwhelming role in the clean up of
hazardous contamination..