Bioremediation

Chapter 8

What is bioremediation?

The use of bacteria and fungi and plants to break down or degrade toxic

chemical compounds that have accumulated in the environment

What are environmental contaminants?

• Pollutants– naturally-occurring

compounds in the environment that are present in unnaturally high concentrations.

– Examples:• crude oil

• refined oil

• phosphates

• heavy metals

• Xenobiotics– chemically synthesized

compounds that have never occurred in nature.

– Examples:• pesticides

• herbicides

• plastics

Early examples of bioremediation

• Outhouse→Centralized engineered wastewater treatment systems– Microorganisms oxidize organic waste

molecules to carbon dioxide and water– Why do we want to use engineered man-

made for this?

More recent examples

• By 1970s it became apparent that we were polluting the environment faster than the natural microbial processes could degrade the pollutants– Congress established the Environmental Protection

Agency• Identified “Superfund Sites” that had priority over other

polluted systems for special funding and cleanup in 1980– 1 in 5 Americans lives within 3-4 miles of a polluted site treated

by the EPA

– Not much progress has been made even though $billions has been spent

Sources of Contamination

• Industrial spills and leaks

• Surface impoundments• Storage tanks and

pipes• Landfills• Burial areas and dumps• Injection wells

ConfiningUnit

Water table

Saline Water

Lateralintrusion of saline water

Ocean

Municipalwater well

Abandonedoil well

DeepAquifer

pond

Infiltration ofpesticides andfertilizers from

farmlands

Brine leakage from ruptured well casing

septic tank leakage

Fresh water

Accidental fuel spill

Municipal landfill

Leakage from hazardous waste site

Contaminated shallow

well

Leakingpetroleum

tank

ConfiningUnit

Groundwater contamination

• Groundwater constitutes 96% of available freshwater in U.S.

• 95% of potable water in rural areas of U.S. comes from groundwater

• In 1988, EPA confirmed that 26 states had various amounts of 44 different pesticides in their groundwater

• Cost of cleanup is in the $ trillions

• Issues that are still hotly debated– How clean is clean?

Most recent

• National Institute of Environmental Health Sciences established the Environmental Genome Project– Study impact of environmental chemicals on

human disease• Identify genes and their products that are sensitive

to toxic chemicals in the environment• Identify genes that encode for products that

detoxify the chemicals

What types of treatment technologies are in use to remove contaminants from the

environment?

• Soil vapor extraction• air sparging• bioremediation• thermal desorption• soil washing• chemical dehalogenation• soil extraction• in situ soil flushing

What Makes Bioremediation a Promising Approach?

• permanence– contaminant is degraded

• potentially low cost– 60-90% less than other technologies

Why use Bioremediation?

• No additional disposal costs

• Low maintenance• Does not create an

eyesore• Capable of impacting

source zones and thus, decreasing site clean-up time

Economics of in-situ vs. ex-situ remediation of contaminated soils

• Cost of treating contaminated soil in place $80-$100 per ton

• Cost of excavating and trucking contaminated soil off for incineration is $400 per ton.

• Over 90% of the chemical substances classified as hazardous today can be biodegraded.

Fundamentals of Biodegradation

• All organics are biodegradable, BUT biodegradation requires specific conditions

• There is no Superbug

• Contaminants must be bioavailable

• Biodegradation rate and extent is controlled by a “limiting factor”

Contaminants Potentially Amenable to Bioremediation________________________________________________________________________________________

Readilydegradable_____________

Somewhatdegradable_____________

Difficult todegrade_____________

Generallyrecalcitrant_____________

fuel oils, gasoline creosote, coaltars

chlorinatedsolvents (TCE)

dioxins

ketones andalcohols

pentachloro-phenol (PCP)

some pesticidesand herbicides

polychlorinatedbiphenyls (PCB)

monocyclicaromatics

bicyclic aromatics(naphthalene)

What challenges exist for bioremediation of pollutants and xenobiotics?

• Pollutants– may exist at high, toxic

concentrations– degradation may

depend on another nutrient that is in limiting supply

• Xenobiotics– microbes may not yet

have evolved biochemical pathways to degrade compounds

– may require a consortium of microbial populations

Fundamentals of cleanup reactions

• Aerobic metabolism– Microbes use O2 in their metabolism to

degrade contaminants

• Anaerobic metabolism– Microbes substitute another chemical for O2 to

degrade contaminants• Nitrate, iron, sulfate, carbon dioxide, uranium,

technicium, perchlorate

ATP

ACETATE

CO2

Fe(III)

Fe(II)

Metabolism of a Pollutant-degrading Bacterium

*Benzoate*Benzoate*Toluene*Toluene*Phenol*Phenol*p*p-Cresol-Cresol*Benzene*Benzene

*U(VI)*U(VI)*Co(III)*Co(III)*Cr(VI)*Cr(VI)*Se(VI)*Se(VI)*Pb(II)*Pb(II)*Tc(VII)*Tc(VII)

*CCl*CCl44

*Cl-ethenes*Cl-ethenes*Cl-aromatics*Cl-aromatics*Nitro-aromatics*Nitro-aromatics

Uranium reduction leads to uranium precipitation and immobilization

U6+sol U4+

insol

U6+sol U6+

sol U4+insol

Volatile organic compounds (VOC)

• These are major contributors to air pollution– Paint industry– Pharmaceutical industry– bakeries– printers– dry cleaners– auto body shops

Cometabolism

• Bacterium uses some other carbon and energy source to partially degrade contaminant (organic aromatic ring compound) bacterium

cornstarch CO2 + H2O

contaminant

degradationproducts

Hard to degrade contaminants

• Chlorinated hydrocarbons– solvents– lubricants– plasticizers– insulators– herbicides and pesticides.

Chemicals that exist as ring structures, which havechlorine atoms attached are referred to as halogenatedaromatics and are often toxic.

Examples:chlorobenzoates2, -4-dichlorophenoxyacetatechlorobenzenes

Removal of the Cl atom is the rate-limiting step in thedegradation of the halogenated aromatic. Adioxygenase enzyme is used to replace the Cl atomwith an OH group. The process requires the presenceof oxygen.

Degradation of chlorinated hydrocarbons

• Degradation of organic toxins requires the participation of entire biochemical pathways involving many enzymes coded for by many genes.

• Some of the genes exist on the chromosome while other genes reside on plasmids.

• Phenol-degrading dmp operon is regulated by DmpR, a NtrC-like positive regulator.

(m o n o o xy g e n a s e )

O H

ph e n o l

O H

O H

c a te c h o l

C O O H

C O O H

c is , c is -m u c o n a te

p h e A

d m p X

p h e n o lh y d ro la s e

p h e B

d m p B

d io xy g e n a s e

s u c c in a te a n da c e ty l C o A

CO2 + H2O

The layout of the genes involved in chlorocatechol-degradation on the plasmid is similar to the layout of the catechol-degrading genes on the chromosome

COOH

Cl

OHOH

OH

COOH

Cl

COOH

Cl

o

C=O

COOH

COOHCOOH

O

COOHCOOH

O

succinate

3-chlorobenzoate 3-chlorocatechol 2-chloro-cis,cis-muconate

dieneiactone

maleylacetate

3-ketoadipate

A B

D

A, pyrocatechase II

B, muconate lactonizing enzyme II

D, hydrolase II

Genetic engineering of bacteria to remove toxic metals from the environment

Hg2+-metallothein

Hg2+→HgoHg2+

New gene/enzyme

New gene/transport proteinsE. coli bacterium

Hgo (less toxic form of metal)

Phytoremediation

• ≈350 plant species naturally take up toxic materials– Sunflowers used to remove radioactive

cesium and strontium from Chrenobyl site– Water hyacinths used to remove arsenic from

water supplies in Bangladesh, India

Phytoremediation

• Drawbacks– Only surface soil (root zone) can be treated– Cleanup takes several years

Transgenic plants

Royal DemolitioneXplosive

Stimulates plant growth!

Gene from bacterium moved to plant genome

Careers in Bioremediation

• Outdoor inspection • Lab testing• Administration

Company employeeGovernmentEmployee

Regulatory oversight

Summary

• Many factors control biodegradability of a contaminant in the environment

• Before attempting to employ bioremediation technology, one needs to conduct a thorough characterization of the environment where the contaminant exists, including the microbiology, geochemistry, mineralogy, geophysics, and hydrology of the system