1. Nano bioremediation
Submitted to-
Submitted by-
Shreya M. Modi.
Student of M.phil/p.hD in nanoscience

2. ABSTRACT
Nanoscience, Nanotechnology and their applications have altered the face of Science and
technology. Environmental hazards, Population and pollution is increasing day by day,
and it is the great challenge to science and society to solve these problems. Till now,
microorganisms are used widely in the process of remediation, but now a days
application of nanotechnology and nanoparticals have become boon to solve all these
problems. Nanoparticals have more advancement than microorganisms. In this term
paper I have just tried to explain the advancement of Nano bioremediation. Using the
nanoparticals and nanotechnological instruments it is possible to carry out genetic and
protein engineering of microbial cells which can be used for bioremediation.
Immobilization of microbial cells and enzymes with nanoparticals also enhance the
process of remediation. Apart from this, some microorganisms have ability to synthesize
the nanoparticals which are helpful to the process and also can be recovered for their
application in other field. As well as Nanophytoremediation also enhance the
remediation process.
Key Words- Nanotechnology, Nanoparticals, Bioremediation, Immobilization
Nanophytoremediation

3. NANOSCIENCE AND NANOTECHNOLOGY
Science require measurement. Measurement is the language of science. Nanoscale
implies a scale of measurement that exists at the level of the nanometer.
Nanoscience is the study of atoms, molecules, and objects whose size is on the nanometer
scale ( 1 - 100 nanometers ).
Nanotechnology originates from the Greek word ‘dwarf’.[3]
By definition, nanotechnology is the science of microengineering. Microengineering is
the science of engineering that deals with particle manipulation if those particles are
smaller than 100 nanometers. [1]
Nanotechnology is a broad and interdisciplinary field dealing with structures and
particles at the Nano scale. Nanotechnology can be defined as “Research and technology
development at the atomic, molecular, or macromolecular levels using a length scale of
approximately one to one hundred nanometers in any dimension; the creation and use of
structures, devices and systems that have novel properties and functions because of their
small size; and the ability to control or manipulate matter on an atomicscale” (US EPA
2007, p 5). [2]
The goal of nanotechnogy is to direct atoms and molecules to form desired structures or
patterns with novel functionality at the nanoscale, the physical, chemical, and biological
properties of materials differ in fundamental and valuable ways from the properties of
individual's atoms and molecules or bulk matter.[3]
Nanoparticals have significant properties like-
-Higher Surface Area
-Highly active surface bonds
-Smaller size of nanoparticals etc… make them more reactive and more sensitive to the
environment and other fields.
Nanotechnology has ability to image, measure, model, and manipulate matter on the
Nano scale to exploit those properties and functions and also has ability to integrate those
properties and functions into systems spanning from nano- to macro-scopic scales.
Nanotechnology has been contributing to commercial products for many years. For
example, nanometer sized carbon improves the mechanical properties of fibers;
nanometer silver particals initiates photographic film development.[3]

4. [4]
Nanotechnology can be applied in so many fields.
Now a days the field of nanotechnology is going to become omnipresent as
microorganisms, because it is applied in almost every field.
INTRODUCTION
The advancement of science and technology have altered our life completely. Both
population and pollution is growing very fast. Sax(1974) stated that “The communal
activities of man as asocial being have created a new order of by products which
increased in volume at rate faster than population and has resulted in increasing
contamination of the environment where natural purifying activities can no longer keep
up with it.”[5] The remediation of contaminants by use of existing technologies is not
effective and efficient to clean up the environment, but now a days the nanotechnology
can be applied in the process of remediation. Nanotechnology itself and nanoparticals
have potential property to solve the environmental problems. They also enhance
bioremediation by modifying the activity of microorganisms. In this term paper I have
only just tried to give something about Nano bioremediation.
REMEDIATION
The act or process of correcting fault or deficiency is known as remediation.
ENVIRONMENTAL REMEDIATION
It deals with the removal of pollution or contaminants from environmental media such as
soil, groundwater, sediment or surface water for the protection of environment and living
beings.
BIOREMEDIATION

5. [6]
Bioremediation comes from two words bios means life and remediate means to decipher
an issue. The degradation of noxious waste from the environment using microorganisms
is called as bioremediation. Microorganisms like bacteria, fungi, algae etc take part in
bioremediation. There are many forms of bioremediation they are given as bioleaching,
bio-venting, phyto- remediation, land-farming, composting, rhizo-filtration, bio-
absorption, bio-augmentation, myco-remediation and bio-reacting. There may be natural
or intrinsic bioremediation.[7,8]
NANOBIOREMEDIATION
Nano + Bio + Remediation
The use of nanoscience, nanoparticles and nanotechnology to enhance the microbial
activity to remove pollutants, they also enhance Nanobioremediation.
Nanobioremediation has the potential not only to reduce the overall coasts of cleaning up
large-scale contaminated sites, but it can also reduce clean up time.
GENETIC MODIFICATION OF MICROBES
Microorganisms have so many advantages for this purpose because they possess many
important properties like-
Reproduce very rapidly, can be grown in small or vast quantities, easily broken down
capacity, etc......[9]
Bionanotechnology can be observed as "Nanotechnology through Biotechnology" [10]
that is, the bio-fabrication of nano-objects, or bi-functional macromolecules usable as
tools to construct or manipulate nano-objects. Because of their wide physiological
diversity, small size, genetic manipulability and controlled culturability, microbial cells
ar e ideal producers of a diversity of nanostructures, materials and instruments for Nano
sciences, ranging from fully natural products such as viruses, polymers and
magnetosomes, to engineered proteins or protein constructs such as virus-like particles
(VLPs), and peptide-displaying phages or cells and tailored metal particles
Nanotechnology play very important role in the genetic engineering of microbial gene to
enhance its capacity for multipurpose use.
Deinococcus radiodurans.
This bacterium is currently the most radioactive-resistant organism known on Earth.
Its tremendous ability to withstand high doses of radiation well beyond any naturally-
occurring levels on the planet have caused it to become the focus of a radioactive waste
clean-up initiative funded by the US Department of Energy (DOE)[11,12,13,15]
D. radiodurans shows remarkable genome plasticity. It is able to maintain, replicate and
express extremely large segments of foreign DNA inserted into its genome by tandem
duplication [13,14]. This capability has been exploited recently to show that it can
accommodate and functionally express highly amplified DNA duplication insertions
encoding bioremediation functions

6. While incapable of degrading actual radioactive elements, genetic engineering of this
organism to include genes from other organisms for the degradation or immobilization of
major heavy metal and organic solvent contaminants found in radioactive dumpsites
could aid the clean-up effort of these sites at a significantly reduced cost[15]
For example, the highly characterized merAlocus from Escherichia coli has been cloned
into D. radiodurans [ 11]. merA encodes mercuric ion reductase (MerA), which reduces
highly toxic, thiol-reactive mercuric ion, Hg(II), to much less toxic and nearly inert
elemental and volatile Hg(0). Four different D. radiodurans expression systems were
developed and used to regulate merA expression by varying its cellular gene dosage. [16]
Engineered D. radiodurans strains expressing mer functions could resist and
reduce toxic Hg(II) to volatile elemental Hg(0) in the presence of high-level
chronic radiation. Hg(II)-reducing and toluene-metabolizing D. radiodurans
strain is also reported.[15]
Other metal reducing/resistance functions that have been cloned into D. radiodurans and
are being studied include genes from the following organisms that are specific for the
indicated metal ions:
Desulfovibrio vulgaris (cytc3), U(VI);
Ralstoniaeutrophus CH34 (czc), Cd(II), Zn(II), and Co(II); and
Bacillus thuringiensis, Cr(VI).
for introducing into a single D. radiodurans host the many different bioremediating gene
systems that will be necessary for cleanup of heterogenous radioactive waste
environments. These type of genetic engineering of microorganisms are very beneficial
because if we use different organisms for different waste clean up, we must have to add
some nutrients, growth factors etc... to fulfill their growth requirements. But here, by
applying nanobioremediation single type of organism can carry out clean up of many
waste products.
NANOSCALE BIOPOLYMERS WITH CUSTOMIZABLE
PROPERTIES FOR HEAVY METAL REMEDIATION
Metal chelatingpolymers require toxic solvents for synthesis and require ultrafiltration for
their separation from the solution.
• One way of solving this problem to develop metal binding materials that can be
recovered by changing the environment
like- pH, Temperature etc.. Around them.
One such material is nanoscale modified biopolymers which can be manufactured by
genetic and protein engineering of microorganisms which can control the size and
arrangement at the molecular level.[17]
This table contain some examples of modified microorganisms using nanotechnology
instrument.[18]
Microorganisms Modification Contaminants References
Pseudomonas sp. Pathway mono/dichlorobenzoates REINEKE and
B13 KNACKMUSS,
1979, 1980

7. P. putida Pathway 4-ethylbenzoate RAMOS et al.,
1987
P. putida KT2442 Pathway RAMOS et al., 1987 PANKE et al.,
1998
PANKE et al., 1998 Pathway chloro-, ROJO et al., 1987
methylbenzoates
C. testosteroni VP44 Substrate o-, p- HRYWNA et al.,
Specificity monochlorobiphenyls 1999
Pseudomonas sp. Substrate PCB ERICKSON and
LB400 Specificity MONDELLO,
1993
E. coli S ubstrate PCB, benzene, toluene KUMAMMRU et
JM109(pSHF1003) Specificity al., 1998
E. coli Regulation TCE, toluene WINTER et al.,
FM5/pKY287 1989
DECOLORIZATION OF THE DYE CONGORED BY Aspergillus
nigerSILVER NANOPARTICALS
Removal of dyes from industrial waste waters is of global concern because dyes cause
many problems in aqueous environments. Dyes may significantly affect photosynthetic
activity in aquatic life because of reduced light penetration and may also be toxic to some
aquatic life due to the presence of aromatics, metals, chlorides, etc. [19]
The A.niger is allowed to grow in the presence of AgNO3 and incubated in dark, it will
form silver nanoparticals within 48 hours which enhance the degradation process of
Congo red dye.[20]
A significant decolorization rate was observed for the dye Congo red. The Aspergillus
niger silver nanoparticle effectively decolorized85.8%of dye within 24 hour incubation
and the dye was fully decolorized within 48 hour of incubation. Whereas the plain
culture (Aspergillus niger) was able to degrade only 76%of dye at the same incubation
conditions and complete decolorization was observed after 48 hour incubation.(Graph.1)

8. Graph.1. % of decolorization of the dye Congo red by Aspergillus niger silver
nanoparticle and Aspergillus niger (plain culture)
REMOVAL OF PHENOLIC POLLUTANTS FROM MUNICIPAL
WASTE WATER IMMOBILIZED LACCASE ENZYMES USING
NANOPARTICALS
4
Laccase is generally found in higher plants and fungi but recently it was found in some
bacteria such as S.lavendulae, S.cyaneus, and Marinomonas mediterranea[21,22,23]
Endocrine disrupting compounds (EDCs) can cause adverse health effects like
developmental disorders, birth defects or cancer.
One major pathway for EDCs to be released into the environment is through wastewater
treatment plant effluents. Consequently, removal of EDCs from wastewater is of concern.
Many EDCs in wastewater are phenolics e.g. bisphenol A (BPA). It has been proposed
that laccase–an enzyme using molecular oxygen as substrate to oxidize phenolic
moieties–could be utilized for the removal of phenolic contaminants from wastewater. In
the present work laccase of a Thielavia genus has been immobilized on fumed silica
nanoparticles. The stability and activity of the resulting biocatalysts regarding the
removal of bisphenol A from biologically treated wastewater was assessed and compared
to the activity and stability of free laccase enzymes. Stability of the immobilized laccase
was considerably higher than that of the free enzyme. Approximately 75% of the initial
BPA was transformed within 2 hours. The ability to significantly eliminate BPA at
environmentally relevant concentrations as well as the increased stability of the
immobilized over the free enzymes shows the large potential for laccase-nanoparticle
conjugates in municipal wastewater treatment for the elimination of phenolic
contaminants.[24]
Apart from this,
Mesoporous carbon materials, with their properties such as a large specific surface
area, a high pore volume, a porosity made up of uniformed mesopores with tunable sizes
and higher hydrothermal resistance compared with mesoporous silica materials and other
materials, have been considered as highly suitable candidates for laccase enzyme
isolated from Trametes versicolor and molecule immobilization [25]
Magnetic bio-separation technology is a promising technology in the support systems
for enzyme immobilization, since on the basis of magnetic properties, compared with
conventional filtering separation, rapid separation and easy recovery could be reached in
external magnetic field, and the capital and operation costs could also be reduced [26]
The use of laccase enzyme instead of whole organism is very much beneficial process
because enzyme can be harvested and reused after the process and no need to remove
microbial cells.

9. NANOTECH COATING CAN ENHANCE ELECTRICITY OUTPUT
FROM WASTE WATER
Engineers at Oregon State University have discovered that the proper nanotech coating
could increase the electricity output of wastewater-to-energy production by more than
20 times.
.In producing power from wastewater, bacteria are placed in an anode chamber – where
they form a biofilm, consume nutrients and grow – to release electrons
The researchers then experimented with the use of new coatings on the anodes of
microbial electrochemical cells to generate more electricity from sewage. They found that
coating graphite anodes with a nanoparticle layer of gold can increase electricity
production by 20 times, while coatings with palladium produced an increase as well,
but not nearly as much.[27]
Use of silver nanoparticles to control biofilm formation in aqueous environment and UF
membrane apparatus.
Due to increasing tolerance of the biofilm community to antibiotics, biocides and
mechanical stress, it has become just as difficult to completely eradicate mature biofilms
as it is to completely avoid the presence of planktonic cells, the origin of the biofilm in
the water. Common treatments to prevent or remove bio fouling include using
disinfection, minimizing nutrients in the feed or altering surface materials to prevent
bacterial attachment, or clean-in-place (CIP) to remove mature biofilm by chemical or
Mechanical shear.
Nanoparticles are collection in aggregate of atoms in the range of 1-100 nm with unique
structure and properties, which are widely used in an increase amount of applications.
Silver nanoparticles (Ag-NPs) in particular, provide effective growth inhibition of
various microorganisms in suspension and on solid medium.
In addition, a few types of filtration membranes and devices like catheter incorporating
silver nanoparticles have demonstrated anti-biofouling properties.[28]
NANOBIOREMEDIATION TO CLEAN UP OIL SPILL
Immobilization cells of Ps. mendocina H3, Ps. pseudoalcaligenes H7, Ps. stutzeri H10,
Ps.alcaligenes H15, Ps. pseudoalcaligenes H16, Ps. mallei 36K and Micrococcus luteus
37 was demonstrated high sorbtional activity carriers.
The degrees of attached microbial cells were reached 80-90%.
In depend from strain of microorganisms attached to carbonized nanoparticles new
nanobiopreparates possesses important properties and may be sorbents of different
metals, oxidizer oil, aromatic carbohydrates, toluene, herbicide, pesticide and other.
For bioremediation of oil contaminated soil is important that carbonizated sorbents itself
may sorbs oil drops for further oxidation carbohydrates of oil by microbial cells, to be
source of mineral compounds and improve condition of soils.
Us were investigated oil-oxidative activity nanobiopreparates receiving by
immobilization specific microorganism’s cells on particles carbonizated rice hunk with

10. nanosize. According results were received different physical and chemical methods ex-
situ bioremediation of oil-contamination soil by new nanobioprepates was discovered that
their destructive activity marked above than free microbial cells.[29]
DEGRADATION OF HYDROPHOBIC COMPOUND ENHANCED
BY NANOPARTICALS.
Nanoparticals are also being used to increase the bioavailability of hydrophobic ogranic
compounds for their enhanced bioremediation.
Polymeric nanoparticals prepared from a poly(ethylene) glycol Modified Urethane
Acrylite(PMUA) precursor was applied to enhance the bioavailability of Polynuclear
Aromatic Hydrocarbons(PAHs) in soil and aqueous solutions.
Due to the hydrophobicity of interior regions of PMUA there is increased affinity
between PAHs and released into the aqueous phase and enhances the rate of
Mineralization.Subsequently the released PAHs can be treated by natural attenuation or
pump and treat process in which polymeric nanoparticals can be recovered and recycled
after microbial degradation of PAHs.[30]
IMMOBILIZATION OF MICROBIAL CELLS USING
NANOPARTICLES
Immobilized microbial cells are frequently used in bioconversions, biotransformation,
and biosynthesis processes due to their better operational stability, easier separation from
products for possible reuse, and satisfactory efficiency in catalysis compared to free
cells.[31]
• Further nanoparticles can also be used to immobilize bacterial cells which are
capable of degrading specific toxic compounds or to biorecover certain
compounds.
• In one of the study,
• Magnetic nanoparticles (Fe3O4) were functionalized with ammonium oleate and
coated on the surface of Pseudomonas delafieldii.
• On application of external magnetic field to the microbial cells, the nanopartical
coated cells concentrate on particular site of the reactor wall separating them from
the whole solution and enabling recycling of the cells for the treatment of the
same compound.
• These coated cells were applied for the desulfurization of organic sulfur from
the fossil fuel.[i.e.-dibenzothiophene] in a bioreactor and were observed to be as
efficient as the non-nanoparticle coated microbial cells
• Apart from this,

11. • Biodesulfurization (BDS) of dibenzothiophene (DBT) was carried out by
Rhodococcus erythropolis IGST8 decorated with magnetic Fe3O4 nanoparticles,
synthesized in-house by a chemical method, with an average size of 45–50 nm, in
order to facilitate the post-reaction separation of the bacteria from the reaction
mixture.[32]
• Scanning electron microscopy (SEM) showed that the magnetic nanoparticles
substantially coated the surfaces of the bacteria. It was found that the decorated
cells had a 56% higher DBT desulfurization activity in basic salt medium (BSM)
compared to the nondecorated cells.
• We propose that this is due to permeabilization of the bacterial membrane,
facilitating the entry and exit of reactant and product respectively. Model
experiments with black lipid membranes (BLM) demonstrated that the
nanoparticles indeed enhance membrane permeability.
PHYTOREMEDIATION
Phytoremediation, so called phytotechnology, is a relatively new technology involved the
plants which play a role in remediation of contaminated environment. It is a green
technology and environmental friendly.
There are several types of plants to remedy and take up contaminants from soil, surface
water, ground water, and sediment. Phytoremediation has been used to take up heavy
metals, organic compounds and toxic chemicals such as 2,4,6-trinitrotoluene (TNT),
trichloroethylene, benzene, toluene, ethyl benzene, xylene, lead, mercury, arsenic and
radionuclides from contaminated environment.
Nano-phytoremediation for degradation and removal of TNT-contaminated soil has
obviously more effective than either nanoremediation or phytoremediation.[33,34]
Regarding the time points of the complete TNT remediation and half life of TNT, the
highest removal efficiency of nano-phytoremediation was found in soil with the
TNT/nZVI ratio of 1/10 (100 mg/kg initial TNT concentration) in treated potting soil by
Panicum maximum.[35]
MICROBIAL PRODUCTION OF SELENIUM NANOPARTICLES
USED FOR WASTE WATER TREATMENT AND OHER
APPLICATION
Specialized microorganisms, so called dissimilatory metal reducers, can indeed be used
to convert water soluble, toxic selenium compounds (selenite, selenate) to water
insoluble, non-toxic elemental selenium. However, the separation of this solid produced
from the aqueous phase is challenging due to the fact that the solid products formed are–
although pure elemental selenium- of nanoparticle size. So far, this circumvented
recovery by simple (and thus cheap) gravitational settling.
Here we show that the Nano particulate size and the poor settle ability of the solid
products formed is due to an organic polymer fraction associated. We used capillary
liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-
MS/MS) to identify proteins associated. We could demonstrate that these proteins
strongly associated to selenium surfaces, not only microbially produced but also by

12. chemical synthesis. Furthermore, we studied the influence of the organic polymers
associated on the colloidal stability of the Nano particulate suspensions by means of
electrophoretic measurements (i.e. zeta –potential). The results gained can be directly
used to enable selenium nanoparticle recovery by cheap gravitational settling. This
represents and vital way point towards the recovery of nanoparticle elemental selenium
from industrial "WASTE" water.[36]
These selenium nanoparticles are used to cure Selenium deficiency in several
CONCLUSION
According to above all application of Nanobioremediation it can be definitely concluded
that, Nanoparticals, Nanotechnological instrument play efficient role in the process of
Nanobioremediation. By applying the nanobioremediation to environment hazards, it can
clean them Faster and Safer than other methods and technology. We can say that,
Nanobioremediation Maintain all three criteria.

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