BIOREMEDIATION OF

HYDROCARBONS
AND XENOBIOTICS

by
DR. S.KARPAGAM
Associate Professor of Botany
QUEEN MARY’S COLLEGE
3 E’s
Water molecule
Dihydrogen Monoxide
DHMO
Dihydrogen Oxide,
Hydrogen Hydroxide,
Hydronium Hydroxide,
Hydric acid.
water
Main source
of fresh
water
RAIN
 On 14 November 2008, the Moon Impact Probe
separated from the Chandrayaan struck the south pole to
place its flag on the Moon. The probe impacted near
Shackleton Crater at 20:31 ejecting underground soil
that could be analysed for the presence of lunar water
ice.

The estimated cost for the project was 386

crore (US$80 million).
 The Cassini spacecraft also carries a set
of sensitive magnetometers, a Visual
and Infrared Mapping Spectrometer
(VIMS) that will enable scientists to
detect subtle changes in the brightness
of surfaces in specific colours which are
signatures of particular materials such as
methane or water ice

ESTIMATED COST 3.5 Million dollars

ANCIENT CIVILIZATIONS
GREAT BARRIER REEF
New islands emerge from
coral reefs
Marine ecosystem is unique not only by
its vast area but also by its influence on
the climatic conditions of the earth as a
whole, and weather conditions of
regional areas.

Sewage from cities, towns, toxic heavy

metals, pesticides, hazardous chemicals,
petroleum hydrocarbons are opened
into the sea.
Marine ecosystem
Species diversity
Unexplored regions
Thermal vents
Carbon sink
MARINE
POLLUTION

Sewage from cities

Industrial effluents
Oil slick
Pollution from
warships
Military activities
Oil spill
A xenobiotic is a chemical which is
found in an organism but which is
not normally produced or expected
to be present in it. It can also cover
substances which are present in
much higher concentrations than are
usual.
However, the term xenobiotics is very
often used in the context of pollutants
such as dioxins and
polychlorinated biphenyls and their
effect on the biota, because xenobiotics
are understood as substances foreign
to an entire biological system, i.e.
artificial substances, which did not exist
in nature before their synthesis by
humans.
Inorganic water pollutants include:
*Acidity caused by industrial discharges (especially
sulfur dioxide from power plants)
*Ammonia from food processing waste
*Chemical waste as industrial by-products

*Fertilizers containing nitrates and *phosphates--which

are found in stormwater runoff from agriculture

*Heavy metals from motor vehicles (via urban

stormwater runoff) and acid mine drainage

*Silt (sediment) in runoff from construction sites,

logging, slash and burn practices or land clearing sites
Organic water pollutants include:
*Detergents
*Disinfection by-products found in
chemically disinfected drinking water, such
as chloroform
*Food processing waste, which can include
oxygen-demanding substances, grease
*Insecticides and herbicides, a huge range
of organohalides and other chemical
compounds
Contd…
*Petroleum hydrocarbons, including fuels (gasoline,
diesel fuel, jet fuels, and fuel oil) and lubricants (motor
oil), and fuel combustion byproducts,.
*Volatile organic compounds (VOCs), such as industrial
solvents, from improper storage.
*Chlorinated solvents, which are dense non-aqueous
phase liquids (DNAPLs), may fall to the bottom of
reservoirs, since they don't mix well with water and are
denser.
*Various chemical compounds found in personal hygiene
and cosmetic products
CHEMICAL TESTING
Water samples may be examined using the
principles of analytical chemistry.
pH,
Biochemical oxygen demand
Chemical oxygen demand
nutrient estimation (nitrate phosphorus)
Metals (copper, zinc, cadmium, lead mercury),
hydrocarbons (oil and grease, petroleum ),
pesticides.
Bacteriological water analysis is a
method of analysing water to estimate
the numbers of bacteria present
Multiple tube method
ATP Testing
Plate count
Membrane filtration
Pathogen analysis
Chemicals released into the
environment forced indigenous
microflora to
-degrade compounds by producing
enzymes
-detoxify by deriving metabolic energy
Petroleum hydrocarbons
-alkanes, aromatics, resins,
asphaltenes
-single carbon to 40 carbons
-ranges from gas like methane
to liquids to solids
n-alkanes are rapidly degraded
by
β- oxidation
to form
2C shorter fatty acid
and liberate CO2 and acetyl CoA
B-Oxidation of hydrocarbons
AROMATIC HYDROCARBONS
Benzene ring hydroxylated
ring cleavage two types
- ortho cleavage
- meta cleavage
Petroleum hydrocarbons are
degraded by Bacteria
Pseudomonas spp.
Micrococcus
Achromobacter
Nocardia
Arthrobacter
Acinetobacter
Brevibacterium
Corynebacterium
Rhodotorula
Hydrocarbons are degraded by
fungi
Aspergillus
Penicillium
Fusarium
Gliocladium
Trichoderma
Candida
Saccharomyces
Rhodotorula
Sporobolomyces
 BIOREMEDIATION IN-SITU
OR EX-SITU
Bioventing,
Landfarming,
Bioreactor,
Composting,
Bioaugmentation,
Rhizofiltration,
Biostimulation.
*Mycoremediation - use of fungal
mycelia in bioremediation.

Mycofilteration - use fungal mycelia to

filter toxic waste

*95% of PAH (Polycyclic aromatic

hydrocarbons) was reduced to non-
toxic components in mycelia inoculated
setups
A.M. CHAKRABARTY

Superbug
OCT PLASMID
alk BAC,
alkR
Degrades C6 to C10
n-alkanes
The sewage, river and floodwater discharged into the
ocean carried plenty of terrestrial bacterial species along
with it.

It was thought that they rapidly disappeared when

released in to sea (Carlucci and Pramer, 1959).

Colwell and coworkers (1985) established that when

enteric bacteria are introduced into sea water, they enter
the viable non culturable state (VNC).

The VNC cells are viable with metabolic activity and

were even found to have retained their virulence
(Grimes and Colwell, 1986).
Van Overbeek and coworkers reported that
P. fluorescens R2f cells could be detected
with the immunofluorescence technique
even in samples where no colonies were
recovered through the conventional culture
techniques.

The terrestrial bacteria when discharged in

to sea water do not die immediately and act
as donors of plasmids to indigenous marine
bacteria.
The donor bacteria was isolated from soil
samples collected from petroleum polluted sites
around Madras Refineries.
Potential hydrocarbon degrading culture of
Pseudomonas fluorescens SKP3 was isolated by
enrichment culture technique.
The bacteria was resistant to wide range of
antibiotics.

The bacteria showed resistance to heavy metals

also.
Table showing the antibiotic resistance/ sensitivity pattern
of donor
Antibiotic MIC (μg/ml) LC (μg/ml)
Chloramphenica 575 <900
lGentamycin 70 95
Rifampicin 15 20
Ampicillin 375 575
Kanamycin 65 85
Mitomycin C 5 10
Tetracycline 40 55
Streptomycin 325 570
Benzylpenicillin 275 675
Nalidixic acid - 5
Amoxycillin 550 >900
Erythromycin 375 685
Penicillin G 575 >900
Cephalexin 525 755
Vancomycin 725 >900
Cloxycillin 775 >900
Plasmid DNA was isolated by the
Casse et al. (1979) method.

Conjugation was carried out by

Broth mating method
Plate mating method
Filter mating method
The survival rate of non-halotolerant
bacteria in marine water is not clearly
known.
In the present study, the survival rate of
the donor is 3 to 4 weeks.
Therefore, it would act as a potential
donor of the plasmid to the marine
bacteria.
The frequency of transfer is influenced
by several environmental factors, like
temperature and availability of
nutrients.

When the pool of resistance genes

increases in the environment it would
bring havoc to human health care.
3 R’s

Reduce
Reuse
Recycle
Kulithalai
Municipality

*Solid Waste
management
*Segregation at
source
*Composting of
degradable waste
*Recycling of non-
degradable waste