Green nanotechnology – A new hope for medical biology

Abstract

The development of eco-friendly technologies in material synthesis is of considerable importance to expand their biological applications. Nowadays, a variety of green nanoparticles with well-defined chemical composition, size, and morphology have been synthesized by different methods and their applications in many cutting-edge technological areas have been explored. This review highlights the classification of nanoparticles giving special emphasis on biosynthesis of metal nanoparticle by viable organisms. It also focuses on the applications of these biosynthesized nanoparticles in a wide spectrum of potential areas of medical biology including catalysis, targeted drug delivery, cancer treatment, antibacterial agents and as biosensors.

Full text

environmental toxicology and pharmacology 36 (2013) 997–1014
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Review
Green nanotechnology–Anewhope for
medical biology
Debjani Nath∗, Pratyusha Banerjee
Cytogenetics and Molecular Biology Laboratory, Department of Zoology, University of Kalyani, Nadia, West Bengal,
India
article info
Article history:
Received 26 June 2013
Received in revised form
29 August 2013
Accepted 4 September 2013
Available online 13 September 2013
Keywords:
Nanoparticles
Green synthesis
Chemical synthesis
Biological application
abstract
The development of eco-friendly technologies in material synthesis is of considerable impor-
tance to expand their biological applications. Nowadays, a variety of green nanoparticles
with well-defined chemical composition, size, and morphology have been synthesized by
different methods and their applications in many cutting-edge technological areas have
been explored. This review highlights the classification of nanoparticles giving special
emphasis on biosynthesis of metal nanoparticle by viable organisms. It also focuses on the
applications of these biosynthesized nanoparticles in a wide spectrum of potential areas of
medical biology including catalysis, targeted drug delivery, cancer treatment, antibacterial
agents and as biosensors.
© 2013 Elsevier B.V. All rights reserved.
Contents
1. Introduction .................................................................................................................. 998
2. Classification of nanoparticles .............................................................................................. 999
2.1. Liposomes ............................................................................................................. 999
2.2. Superparamagnetic nanoparticles ................................................................................... 999
2.3. Fullerenes: buckyballs and carbon nanotubes....................................................................... 999
2.4. Dendrimer............................................................................................................ 1001
2.5. Quantum dots........................................................................................................ 1001
2.6. Liquid crystals ....................................................................................................... 1001
3. Metal nanoparticles and characterization methods ...................................................................... 1001
4. Synthesis of metal nanoparticles by traditional physical and chemical methods ....................................... 1001
4.1. Laser ablation ........................................................................................................ 1001
4.2. Inert gas condensation .............................................................................................. 1002
4.3. Sol–gel method....................................................................................................... 1002
∗Corresponding author. Tel.: +91 339433384571.
E-mail addresses: nath debjani@yahoo.co.in,bdytdebnath@yahoo.com (D. Nath).
1382-6689/$ – see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.etap.2013.09.002

998 environmental toxicology and pharmacology 36 (2013) 997–1014
4.4. Hydrothermal and solvothermal synthesis......................................................................... 1002
4.5. Colloidal methods ................................................................................................... 1003
5. Bio-inspired green nanomaterial synthesis ............................................................................... 1003
5.1. Use of bacteria to synthesize nanoparticles ........................................................................ 1003
5.2. Use of actinomycetes to synthesize nanoparticles................................................................. 1004
5.3. Use of fungi to synthesize nanoparticles ........................................................................... 1005
5.4. Use of plants to synthesize nanoparticles .......................................................................... 1006
6. Bio-inspired green nanoparticles over chemically synthesized nanoparticles .......................................... 1008
7. Applications of metal nanoparticles in medical biology .................................................................. 1008
7.1. Catalysis.............................................................................................................. 1009
7.2. Biological application of metal nanoparticles ...................................................................... 1009
7.2.1. Labelling..................................................................................................... 1009
7.2.2. Sensors ...................................................................................................... 1009
7.2.3. Drug delivery ................................................................................................ 1010
7.2.4. Cancer therapy.............................................................................................. 1010
7.3. Environmental cleanup as defense against environmental challenge to medical biology ....................... 1010
8. Conclusions ................................................................................................................. 1011
Conflict of interest .......................................................................................................... 1011
Acknowledgements......................................................................................................... 1011
References................................................................................................................... 1011
1. Introduction
Nanomaterials, with its characteristic dimension at the range
of 1–100 nm, are at the leading edge of nanoscience and nano-
technology. In recent years nanomaterials, specifically metal
nanoparticles, have received particular interest in diverse field
of applied science ranging from material science to biotech-
nology (Guo et al., 2005; Daniel and Astruc, 2004; Huang
et al., 2007). Although widespread interest in nanomateri-
als is recent, the concept was actually introduced over 40
years back. With the advancement of science and technol-
ogy, the morphology of this material, which contains metallic
nanoparticles, has been understood. Because of extremely
small size and high surface volume ratio of nanoparticles,
the physicochemical properties of nanoparticles-containing
materials are quite different to those of the bulk materials
(El-Sayed, 2001). Thus, nanomaterials have potential appli-
cations in electronics and photonics, catalysis, information
storage, chemical sensing and imaging, environmental reme-
diation, drug delivery and biological labelling (Guo et al.,
2005; Daniel and Astruc, 2004; Huang et al., 2007). It is well
known that the optical, electronic, and catalytic properties
of metal nanoparticles are greatly influenced by their size,
shape, and crystal structure. For example, silver (Ag) and gold
(Au) nanocrystals of different shapes possess unique optical
scattering responses (Daniel and Astruc, 2004; Roduner, 2006).
Whereas highly symmetric spherical particles exhibit a single
scattering peak, anisotropic shapes such as rods, triangular
prisms, and cubes exhibit multiple scattering peaks in the
visible wavelengths due to highly localized charge polariza-
tions at corners and edges (Mie, 1908). Thus, synthesis of metal
nanoparticles with defined morphology gained much interest.
A variety of strategies have been developed for the synthesis
of metal nanoparticles (MNPs) and nanomaterials.
Optimizing the nanomaterial synthesis has now become a
prolific area of investigation. Metal nanoparticles are partic-
ularly unique in nanoscale system because of the ease in its
synthesis and chemical modifications. Hazardous substances
such as sodium borohydride, tetrakishhydroxymethylphos-
phonium chloride (THPC), poly-N-vinyl pyrrolidone (PVP), and
hydroxylamine have been used for the synthesis of nanopar-
ticles in the traditional wet methods. Other dry methods such
as UV irradiation, aerosol and lithography are also not consid-
ered environment-friendly. The use of such toxic chemicals is
still the subject of paramount concern because toxic chemicals
on the surface of nanoparticles and non-polar solvents limit
their applications in clinical fields. Therefore, the biosynthesis
of clean, biocompatible, non-toxic and environment-friendly
nanoparticles produced both extracellularly and intracel-
lularly deserves merit (Karazhanov and Raveendran, 2003;
Sharma et al., 2009; Narayanan and Sakthivel, 2010).
Despite significant private and public investment, progress
moving nanomaterials from the laboratory to industrial pro-
duction has been slow and difficult. Two challenges that have
slowed the development, are the poor understanding of the
new hazards introduced by nanotechnology (Senjen, 2007) and
lack of appropriate policies to manage any new risks. Scien-
tists, engineers and entrepreneurs, however, continue to move
forward, grappling with challenges that range from the tech-
nical to the regulatory and everywhere in between. Just as the
concepts of nanoscale invention have required new insights
from scientists, they are also demanding new approaches
to managing, producing, funding and deploying novel tech-
nologies into the larger chemical sector. In this case, there
is an unusual opportunity to use science, engineering and
policy knowledge to design novel products that are benign
as possible to human and environmental health. Recognition
of this opportunity has led to the development of the “green
nanoscience” concept (McKenzie and Hutchison, 2004; Dahl
et al., 2007).
Green nanotechnology has drawn on the field of green
chemistry, and the framework of the Principles of Green
Chemistry (Anastas and Warner, 1998) features significantly
in work to design new nanotechnologies for joint economic,

environmental toxicology and pharmacology 36 (2013) 997–1014 999
social, and health/environmental benefit (Hutchison, 2008).
These efforts have been aided by awareness throughout the
nanotech community that they need to address the poten-
tial negative impacts of nano from the outset (Albrecht et al.,
2006). Nanotechnology presents an opportunity to develop a
new technology, and a new industry in a sustainable way from
the outset. We are at a unique point where we have more
understanding of how to go about this than at any time in the
past. This new emerging science and associated technologies
do not have to follow the path that has been typical of many
past innovations in the chemical industry that, despite pro-
viding significant benefits, also turned out to have significant,
unanticipated costs to human health and the environment.
The development and commercialization of viable green nan-
otechnologies is difficult, and the barriers will require effort
from the scientists, researchers, government and communi-
ties.
In the first part of this review, special emphasis is given on
the classification of nanoparticles and different methodolo-
gies for the characterization of it, in the second part different
strategies for the synthesis of MNPs have been discussed,
with particular emphasis on biosynthesis by viable organisms.
Lastly it focuses on the application of nanoparticles in differ-
ent fields along with an idea of current state of green nano
research.
2. Classification of nanoparticles
A broad library of nanoparticles consisting of different physi-
cal and chemical properties has been constructed. Due to new
discoveries made in nanotechnology, however, classifications
of nanoparticles are constantly changing. Currently nanopar-
ticles can be separated into several different classes.
•There are the nanoclusters that are defined as semi crys-
talline nanostructures with dimensions within 1–10 nm and
a narrow size distribution.
•There are the nanopowders that result from the aggregation
of non-crystalline nanomaterials with dimensions between
10 and 100 nm.
•The nanocrystals that are single crystalline nanomaterials
with dimensions between 100 and 1000 nm.
These examples are only the most basic classifica-
tion of nanoparticles; others include nanorods, nanocups,
nanospheres, nanodiamonds, nanostars and the quantum
dots (Chakrabarti et al., 2004). Some examples of general
and multi-functional classes of nanoparticles (Fig. 1) used in
biotechnology and particularly in the area of nanomedicine
are listed below.
2.1. Liposomes
An initial study on liposomes or lipid vesicles originated in
the 1960s when there was a need to understand new types
of polymer nanocontainers. Initially, liposomes were used as
model systems to study biological membranes but by 1970 had
been developed into a medium for the transportation of drugs
(Graff et al., 2004). Liposomes, in general, had been reported
to be highly useful in biophysics as a good model system in
understanding the properties of cell membrane and chan-
nels. In chemistry, they served as an excellent illustration of
catalysis, energy conversion and photosynthesis while in bio-
chemistry they improved the understanding of the biological
function of proteins specifically in secretion, trafficking and
signalling, gene delivery and other functions in cell biology
(Graff et al., 2004). Liposomes have various extensive appli-
cations in the pharmaceutical industry as directed-delivery
agent for drugs such as anticancer, anti-fungal and vaccines.
They are also useful in cosmetics in the manufacturing of
shampoos and other skin care products. They are very impor-
tant tools in diagnostics as they are able to degrade in the cells
after delivery (Graff et al., 2004; Salata, 2004). Liposomes were
said to be the first synthesized nanoparticles (Fig. 1A) used
for drug delivery but a major limitation was their tendency to
fuse together in aqueous environments and release contents
before getting to the target site. This has lead to the search for
either a replacement or a method of stabilization using newer
substitute nanoparticles.
2.2. Superparamagnetic nanoparticles
Superparamagnetic nanoparticles (Fig. 1B) are a class of
inorganic based particles having an iron oxide core [super-
paramagnetic iron oxide nanoparticles (SPION)] coated by
either inorganic materials (silica, gold) or organic materials
like phospholipids, fatty acids, polysaccharides, peptides, sur-
factants, polymers (Gupta and Curtis, 2004; Liu et al., 2007).
An important property of SPIONs that makes them unique
compared to other nanoparticles is their induced magnetiza-
tion, i.e. they are able to attract to a magnetic field without
retaining residual magnetism after the removal of the field.
This property makes them attractive for many applications,
ranging from various selective bio-separations and contrast
enhancing agents for MRI in drug delivery systems, magnetic
hyperthermia (local heat source in the case of tumour ther-
apy) and magnetically assisted transfection of cells (Neuberger
et al., 2005; Aptekar et al., 2009).
2.3. Fullerenes: buckyballs and carbon nanotubes
Fullerenes are molecules made exclusively of carbon and they
exist in different forms such as hollow spheres, ellipsoids or
tubes. The spherical forms of fullerenes are referred to as
buckyballs, and tubular forms as carbon nanotubes (CNTs) or
buckytubes. Fullerenes possess a structure similar to that of
graphite that is composed of stacked sheets of graphene and
often linked by hexagonal, pentagonal or heptagonal (not very
common) rings (Theodore and Kunz, 2005; Kroto and Walton,
2011; Prasad and Jemmis, 2008). Discovery of fullerenes has
increased the number of known allotropes of carbon, which
was previously limited to graphite, diamond, and amorphous
carbon.
Due to their novel characteristics, a great deal of research
on buckyballs and CNTs has been carried out especially
in the various technological fields of material sciences,
nanotechnology and electronics. Bulky ball, also known as,
Buckminsterfullerene or C60 is the smallest form of fullerenes
and the most abundant in nature, as it is mostly found in

1000 environmental toxicology and pharmacology 36 (2013) 997–1014
Fig. 1 – Different classes of nanoparticles. (A) Nanoparticle in liposomes; (B) superparamagnetic nanoparticle; (C) nanotube;
(D) dendrimer; (E) quantum dots with changing optical properties. (For interpretation of the references to colour in text, the
reader is referred to the web version of the article.)
soot, though a second type of buckyball composed of boron
atoms (i.e. boron bulky ball or B80) instead of carbon has
been described (Gopakumar et al., 2008) in which each boron
atoms that makes up the B80 structure forms about 5–6
bonds and is believed to make the structure more stable than
the C60. CNTs are also allotropes of carbon that, as men-
tioned earlier, are cylindrical in structure. Nanotubes with
length-to-diameter ratio of 132 000 000:1 have been fabricated
and this makes them much larger than any other material.
CNT have unique properties that make them important to
nanoscience/nanotechnology and other fields of science like
material science, optics and even architecture. A possible use
of them in the construction of body armour has been reported
(Mintmire et al., 1992). They display exceptional strength
and peculiar electrical properties. In addition to this, CNTs
have also been found to be excellent conductors of heat and
are referred to as thermal conductors. The major drawback,
however, in their widespread application is their potential to
possess some toxic properties. CNTs (Fig. 1C) are classified
into various forms including single walled carbon nanotubes
(SWCNT), double walled carbon nanotubes (DWCNT) and
multi walled carbon nanotubes (MWCNT) (Konishi et al., 1995;
Jian et al., 2006; Clourier et al., 2006; Langmuir et al., 2009).
The uniqueness in the properties of each type of CNTs is what
determines their applications in different scientific areas. For
example, the SWNT which is the most studied of all the CNTs
have particularly strong electric properties which is lacking in
the other types (Martel et al., 2001; Theodore and Kunz, 2005).

environmental toxicology and pharmacology 36 (2013) 997–1014 1001
2.4. Dendrimer
Dendrimers are highly branched structures with uniform size,
radial symmetry and assume a circular shape in solution (Graff
et al., 2004). Dendrimers are built layer-by-layer from core to
periphery by repetitive covalent bond-forming reactions. The
density of the dendrimers increased for every layer formed in
each step as a result of the geometric growth at each branch-
ing point (Tomalia, 2005). By choosing the final reagent, it was
possible to design dendritic molecules with different active
surface groups (Fig. 1D).
The first dendritic molecule was synthesized in about 1980,
but interest in them only developed during the 2000s due
to increased discovery of various applications especially in
the biotechnological areas. In nanomedicine, dendrimers had
been found to be an invaluable tool in attaching fluorescent
dyes, enzymes cell identification tags and other molecules
because of the many molecular “hooks” present on their sur-
face. Production of these molecules, however, can be quite
challenging and expensive and a drawback for their large scale
application. Nevertheless, their high stability and the possibil-
ity of functionalizing them with biomolecules like antibodies
and receptors, makes them a very important medium in target
drug delivery (Smith and Diederich, 2000).
2.5. Quantum dots
Quantum dots (QDs) or nanocrystals are semiconductor
nanoparticles that can emit light in all colours of the spectrum
depending on their size (Fig. 1E). The size of QDs decreases as
they get closer to the blue-end of the spectrum, and increase
as they proceed to the red end.
They have unique properties such that they can even be
tuned beyond visible light, into the infra-red or into the ultra-
violet spectrum and were able to confine conduction band
electrons, valence band holes, or excitations in all three spa-
tial directions (Murray et al., 2000). QDs are valuable tools
in biotechnology most especially in cellular imaging and
labelling as they are believed to be an excellent alternative
to conventional fluorescent dyes used in imaging.
2.6. Liquid crystals
These are pharmaceuticals made from liquid organic crys-
talline materials and are designed to mimic naturally
occurring biomolecules like proteins or lipids. They are
regarded as a very safe and specific vehicle of drug delivery
as they can target a particular area of the body where tissues
are inflamed and are capable of detecting tumours.
3. Metal nanoparticles and characterization
methods
Nanoparticles of metallic origin have been shown to exhibit
unusual properties that they normally will not display in
their bulk form (Elechiguerra et al., 2005; Blackman, 2009).
Due to their huge potential and benefits to nanotechnology,
they have come under intense scrutiny as far as applications
across various disciplines are concerned. In biochemistry, for
example, they are considered to be better catalysts (Astruc,
2008) and good biological and chemical sensors (Nam et al.,
2003; Nie et al., 2007); in information systems, their size and
magnetic properties are being explored in the production of
data storage devices where the issue of miniaturization is
posing an overwhelming challenge (Mayes and Mann, 2004);
in medicine their potential as drug delivery agents has being
reported (Nie et al., 2007).
Nanotechnology, being an interdisciplinary field of science,
will have many characterization and analytical techniques
available (Table 1) in the elucidation of nanomaterials (Gabor
et al., 2008). These characterization methods are based on
two physical processes: the first being primary (1◦) analytical
probes such as photons, neutrons, ions and electrons, which
may be combined with input stresses like magnetic and elec-
tric fields and mechanical stress.
Second, the measurable secondary (2◦) effect obtained e.g.
the release or absorption of electrons, electromagnetic radi-
ation, volume change, mechanical distortion and third, the
choice of the investigating medium, energy, temperature,
time, intensity, phase or angle. During characterization, the
1◦probe, which may be either a beam of electron or a photon
of light, interacts with the analyte or matter causing a change
in its equilibrium and responds in order to gain its previous
state of equilibrium thus modifying the 1◦probe. Examples of
alteration produced as a result of interaction of 1◦probe with
matter are excitation of electrons, phonons, excitons or plas-
mons. Modification of the 1◦probe as a result of this produces
a2
◦effect which is the measured signal (Kelsall et al., 2005).
4. Synthesis of metal nanoparticles by
traditional physical and chemical methods
Synthesis of MNPs is carried out by several physical and
chemical methods that include laser ablation (Mafuné et al.,
2001), ion sputtering (Raffi et al., 2007), solvothermal synthe-
sis (Rosemary and Pradeep, 2003), chemical reduction (Chaki
et al., 2002), and sol–gel (Shukla and Seal, 1999) method.
Basically, there are two approaches for nanoparticle synthe-
sis, the top-down and bottom-up. Top-down approaches seek
to create nanoscale objects by using larger, externally con-
trolled microscopic devices to direct their assembly, while
bottom-up approaches adopt molecular components that
are built up into more complex assemblies. The top-down
approach often uses microfabrication techniques where exter-
nally controlled tools are used to cut, mill, and shape materials
into the desired shape and size. Micropatterning techniques,
such as photolithography and inkjet printing are well known
examples of top-down approach. On the other hand, bottom-
up approaches use the self-assembled properties of single
molecules into some useful conformation.
4.1. Laser ablation
Laser ablation (Mafuné et al., 2001) enables to obtain colloidal
nanoparticles solutions in a variety of solvents. Nanoparti-
cles are formed during the condensation of a plasma plume
produced by the laser ablation of a bulk metal plate dipped
in a liquid solution. This technique is considered as a ‘green

1002 environmental toxicology and pharmacology 36 (2013) 997–1014
Table 1 – Characterization of metal nanoparticles.
Characterization of size, shape, and surface
properties of nanoparticle
Characterization of chemical properties of nanoparticle
Single-particle
techniques
Ensemble analytical
techniques
Single-particle techniques Ensemble analytical
techniques
1. Scanning Electron
Microscopy (SEM)
1. Dynamic Light
Scattering (DLS)
1. Surface composition 1. Atomic/chemical structure
1.1. Electron spectroscopy for
Chemical Analysis (ESCA) or
X-ray photoelectron
spectroscopy (XPS)
1.1. Fourier Transform
Infrared Spectroscopy
1.2. Raman Scattering (RS)
1.3. X-ray Absorption
Spectroscopy (XAS)
1.4. Circular dichroism
2. Transmission Electron
Microscopy (TEM)
2. Laser Diffraction/Static Light
Scattering
2. Surface charge
2.1. Zeta potential
3. Atomic Force
Microscopy (AFM)
3. Field Flow
Fractionation (FFF)
3. Surface reactivity
3.1. Comparative
microcalorimetry
4. Centrifugal sedimentation
5. Specific Surface Area (BET)
6. Time of Flight Mass
Spectroscopy
technique’ alternative to the chemical reduction method for
obtaining noble MNPs. However, the main drawbacks of this
methodology are the high energy required per unit of MNPs
produced and the little control over the growth rate of the
MNPs.
4.2. Inert gas condensation
Inert gas condensation (IGC) is the most widely used meth-
ods for MNPs synthesis at laboratory-scale. Gleiter (1989)
introduced the IGC technique in nanotechnology bysynthesiz-
ing iron nanoparticles. In IGC, metals are evaporated in ultra
high vacuum chamber filled with helium or argon gas at typ-
ical pressure of few hundreds Pascal’s. The evaporated metal
atoms lose their kinetic energy by collisions with the gas,
and condense into small particles. These particles then grow
by Brownian coagulation and coalescence and finally form
nano-crystals. Recent application of this technique includes
size-controlled synthesis of Au/Pd NPs (Pérez-Tijerina et al.,
2008) and hetero-sized Au nanoclusters for non-volatile mem-
ory cell applications (Kang et al., 2011).
4.3. Sol–gel method
The sol–gel process is a wet-chemical technique developed
recently in nanomaterial synthesis. The inorganic nanostruc-
tures are formed by the sol–gel process through formation of
colloidal suspension (sol) and gelation of the sol to integrated
network in continuous liquid phase (gel). Size and stability
control quantum-confined semiconductor, metal, and metal
oxide nanoparticles has been achieved by inverted micelles
(Gacoin et al., 1997), polymer blends (Yuan et al., 1992), block
copolymers (Sankaran et al., 1993), porous glasses (Justus et al.,
1992), and ex situ particle-capping techniques (Olshavsky and
Allcock, 1997). However, the fundamental problem of aqueous
sol–gel chemistry is the complexity of process and the fact
that the as-synthesized precipitates are generally amorphous.
In non-aqueous sol–gel chemistry the transformation of the
precursor takes place in an organic solvent. The nonaqueous
(or non-hydrolytic) processes are able to overcome some of
the major limitations of aqueous systems, and thus repre-
sent a powerful and versatile alternative. The advantages are
a direct consequence of the manifold role of the organic com-
ponents in the reaction system (e.g., solvent, organic ligand of
the precursor molecule, surfactants, or in situ formed organic
condensation products). Nowadays, the family of metal oxide
nanoparticles is synthesized by non-aqueous processes and
ranges from simple binary metal oxides to more complex
ternary, multi-metal and doped systems.
4.4. Hydrothermal and solvothermal synthesis
The hydrothermal and solvothermal synthesis of inorganic
materials is an important methodology in nanomaterial
synthesis. In hydrothermal method, the synthetic process
occurs in aqueous solution above the boiling point of water,
whereas in solvothermal method the reaction is carried
out in organic solvents at temperatures (200–300◦C) higher
than their boiling points. Though development of hydrother-
mal and solvothermal synthesis has a history of 100 years,
recently this technique has been applied in material synthesis
process. Normally, hydrothermal and solvothermal reactions
are conducted in a specially sealed container or high pressure
autoclave under subcritical or supercritical solvent conditions.
Under such conditions, the solubility of reactants increases
significantly, enabling reaction to take place at lower temper-
ature. Among numerous examples, TiO2photocatalysts were
synthesized through hydrothermal process (Ren et al., 2007).
Because low cost and energy consumption, hydrothermal
process can be scale-up for industrial production. Solvother-
mal process enables to choose among numerous solvents
or mixture thereof, thus increasing the versatility of the

environmental toxicology and pharmacology 36 (2013) 997–1014 1003
synthesis. For example, well-faceted nanocrystals of TiO2with
high reactivity were synthesized in a mixture of the solvents
hydrogen fluoride (HF) and 2-propanol (Yang et al., 2008).
4.5. Colloidal methods
The crystallographic control over the nucleation and growth
of noble-metal nanoparticles has most widely been achieved
using colloidal methods (Taoet al., 2008; Turkevitchet al., 1951;
Frens, 1972; Brust and Kiely, 2002). In general, metal nanopar-
ticles are synthesized by reducing metal salt with chemical
reducing agents like borodride, hydrazine, citrate, etc., fol-
lowed by surface modification with suitable capping ligands to
prevent aggregation and confer additional surface properties.
Occasional use of organic solvents in this synthetic process
often raises environmental questions. At the same time, these
approaches produce multi-shaped nanoparticles requiring
purification by differential centrifugation and consequently
have low yield. Thus, the development of reliable experimen-
tal protocols for the synthesis of nanomaterials over a range of
chemical compositions, sizes, and high monodispersity is one
of the challenging issues in current nanotechnology. In this
context, current drive is focused on the development of green
and biosynthetic technologies for production of nanocrystals
with desired size and shape.
5. Bio-inspired green nanomaterial
synthesis
Nature has devised various processes for the synthesis of
nano- and micro-length scaled inorganic materials which
have contributed to the development of relatively new (Fig. 2)
and largely unexplored area of research based on the biosyn-
thesis of nanomaterials (Mohanpuria et al., 2008). Biosynthesis
of nanoparticles is a kind of bottom up approach where the
main reaction is reduction/oxidation (Durán et al., 2011). The
microbial enzymes or the plant phytochemicals with anti
oxidant or reducing properties are usually responsible for
reduction of metal compounds into their respective nanopar-
ticles.
5.1. Use of bacteria to synthesize nanoparticles
Bacteria play a crucial role in metal biogeochemical cycling
and mineral formation in surface and subsurface environ-
ments (Lowenstam, 1981; Southam and Saunders, 2005). The
use of microbial cells for the synthesis of nanosized materi-
als (Fig. 3) has emerged as a novel approach for the synthesis
of metal nanoparticles. Although the efforts directed towards
the biosynthesis of nanomaterials are recent, the interac-
tions between microorganisms and metals have been well
documented and the ability of microorganisms to extract
and/or accumulate metals is employed in commercial biotech-
nological processes such as bioleaching and bioremediation
(Gericke and Pinches, 2006). Bacteria are known to produce
inorganic materials either intra cellularly or extra cellularly.
Microorganisms are considered as a potential biofactory for
the synthesis of nanoparticles like gold, silver and cadmium
sulphide.
Among the microorganisms, prokaryotic bacteria have
received the most attention in the area of metal nanoparticle
biosynthesis. The formation of extracellular and intracellular
metal nanoparticles by bacteria like Escherichia coli,Pseu-
domonas stutzeri,Pseudomonas aeruginosa,Plectonema boryanum,
Salmonells typlus,Staphylococcus currens,Vibrio cholerae, etc.,
have been reported (Klaus et al., 1999;Beveridge and Murray,
1976; Southam and Beveridge, 1994).
Some well known examples of bacteria synthesizing inor-
ganic materials include magnetotactic bacteria (synthesizing
magnetic nanoparticles) and S layer bacteria which produce
gypsum and calcium carbonate layers (Shankar et al., 2004).
Some microorganisms can survive and grow even at high
metal ion concentration due to their resistance to the metal.
The mechanisms involve: efflux systems, alteration of sol-
ubility and toxicity via reduction or oxidation, biosorption,
bioaccumulation, extra cellular complexation or precipita-
tion of metals and lack of specific metal transport systems
(Husseiny et al., 2007). For e.g. P. stutzeri AG 259 isolated from
silver mines has been shown to produce silver nanoparticles
(Mohanpuria et al., 2008).
Many microorganisms are known to produce nanostruc-
tured mineral crystals and metallic nanoparticles with
properties similar to chemically synthesized materials, while
exercising strict control over size, shape and composition of
the particles. Examples include the formation of magnetic
nanoparticles by magnetotactic bacteria, the production of sil-
ver nanoparticles within the periplasmic space of P. stutzeri
and the formation of palladium nanoparticles using sulphate
reducing bacteria in the presence of an exogenous electron
donor (Gericke and Pinches, 2006).
Though it is widely believed that the enzymes of the orga-
nisms play a major role in the bioreduction process, some
studies have indicated it otherwise. Studies indicate that some
microorganisms could reduce silver ions where the processes
of bioreduction were probably non enzymatic. For e.g. dried
cells of Bacillus megaterium D01, Lactobacillus sp. A09 were
shown to reduce silver ions by the interaction of the silver
ions with the groups on the microbial cell wall (Fu et al., 2000,
2006).
Silver nanoparticles in the size range of 10–15 nm were
produced by treating dried cells of Corynebacterium sp. SH09
with diamine silver complex. The ionized carboxyl group of
amino acid residues and the amide of peptide chains were the
main groups trapping [Ag(NH3)2+] onto the cell wall and some
reducing groups such as aldehyde and ketone were involved
in subsequent bioreduction. But it was found that the reaction
progressed slowly and could be accelerated in the presence of
OH−(Fu et al., 2006).
The supernatant of gram positive, thermophilic bacterium
Bacillus licheniformis synthesized AgNPs in the range of 50nm
(Kalishwaralal et al., 2008). Formation of nanoscale elemental
silver particles through enzymatic reduction was reported in
Geobacter sulfurreducens (Law et al., 2008).
Kalimuthu et al. (2008) studied AgNPs synthesis using
bacteria B. licheniformis, isolated from sewage collected from
municipal wastes, and ultrasonically lysed bacterial cell. The
synthesized AgNPs had average particle size of around 50nm.
Recently, a rapid method for synthesizing small (1–7 nm)
monodiperse AgNPs has been described by electrochemically

1004 environmental toxicology and pharmacology 36 (2013) 997–1014
Fig. 2 – A schematic representation of bioinspired green nanomaterial synthesis.
active biofilm (EAB) using sodium acetate as an electron donor
(Kalathil et al., 2011).
The formation of AuNP was indicated by the change in
reaction mixture, which turned to light yellow after 1 h. The
AuNPs were synthesized adopting similar procedure using two
P. aeruginosa isolates (Husseiny et al., 2007). The synthesis of
stable gold nanocubes by the reduction of aqueous AgCl4−
solution by B. licheniformis was reported by Kalishwaralal et al.
(2009). Biofilm formation of gram-negative ␤-proteobacterium
Cupriavidus metallidurans is very common on Au grains. The
isolated C. metallidurans from soils and sediments from tem-
perate and tropical Australian sites interacted with Au3+ ions
and form AuNPs distributed homogenously throughout cell
wall (Reith et al., 2009). The AuNPs were also synthesized on
the surface of Rhodopseudomans capsulate by interaction bacte-
rial cells with HAuCl4 solution (He et al., 2007). The aqueous
chloroaurate ions were reduced after 48h of incubation and
transformed to AuNPs. The pH value of the solution controlled
the shape of AuNPs.
In the case of bacteria, most metal ions are toxic and there-
fore the reduction of ions or the formation of water insoluble
complexes is a defense mechanism developed by the bacteria
to overcome such toxicity (Sastry et al., 2003).
5.2. Use of actinomycetes to synthesize nanoparticles
Actinomycetes are microorganisms that share important
characteristics of fungi and prokaryotes such as bacteria. Even
Fig. 3 – Hypothetical diagram of possible mechanism of synthesis of metal nanoparticles by bacteria.

environmental toxicology and pharmacology 36 (2013) 997–1014 1005
though they are classified as prokaryotes, they were originally
designated as ray fungi. Focus on actinomycetes has primar-
ily centred on their exceptional ability to produce secondary
metabolites such as antibiotics. It has been observed that a
novel alkalothermophilic actinomycete, Thermomonospora sp.
synthesized gold nanoparticles extracellularly when exposed
to gold ions under alkaline conditions (Sastry et al., 2003). In
an effort to elucidate the mechanism or the processes favou-
ring the formation of nanoparticles with desired features,
Ahmad et al. (2003), studied the formation of monodisperse
gold nanoparticles by Thermomonospora sp. and concluded that
extreme biological conditions such as alkaline and slightly
elevated temperature conditions were favourable for the for-
mation of monodisperse particles. Based on this hypothesis,
alkalotolerant actinomycete Rhodococcus sp. has been used for
the intracellular synthesis of monodisperse gold nanoparti-
cles by Ahmad et al. (2003). In this study it was observed that
the concentration of nanoparticles were more on the cytoplas-
mic membrane. This could have been due to the reduction
of metal ions by the enzymes present in the cell wall and
on the cytoplasmic membrane but not in the cytosol. The
metal ions were also found to be non toxic to the cells which
continued to multiply even after the formation of the nanopar-
ticles.
It has been observed that a novel alkalothermophilic acti-
nomycete, Thermomonospora sp. synthesized golden nanopar-
ticles extracellularly when exposed to gold ions under alkaline
conditions (Sastry et al., 2003). The use of algae for the biosyn-
thesis of nanoparticles is a largely unexplored area. There is
very little literature supporting its use in nanoparticle forma-
tion. Recently stable gold nanoparticles have been synthesized
using the marine alga Sargassum wightii. Nanoparticles with a
size range between 8 nm and 12 nm were obtained using the
seaweed. An important potential benefit of the method of syn-
thesis was that the nanoparticles were quite stable in solution
(Singaravelu et al., 2007).
5.3. Use of fungi to synthesize nanoparticles
The fungal mediated MNP synthesis is a relatively recent
research area. Fungi have been widely used for the biosynthe-
sis of nanoparticles and the mechanistic aspects governing
the nanoparticle formation have also been documented for
a few of them. In addition to monodispersity, nanoparticles
with well defined dimensions can be obtained using fungi.
Compared to bacteria, fungi could be used as a source for the
production of large amount of nanoparticles. This is due to
the fact that fungi secrete more amounts of proteins which
directly translate to higher productivity of nanoparticle for-
mation (Mohanpuria et al., 2008).
Yeast, belonging to the class ascomycetes of fungi has
shown to have good potential for the synthesis of nanoparti-
cles. Gold nanoparticles have been synthesized intracellularly
using the fungi Verticillium luteoalbum. The rate of particle
formation and the size of the nanoparticles could to an
extent be manipulated by controlling physical parameters
such as pH, temperature, concentration of metal (gold) and
exposure time. A biological process with this ability to strictly
control the shape of the particles would be a considerable
advantage (Gericke and Pinches, 2006). Yeast, belonging to the
class ascomycetes has shown to have good potential for the
synthesis of nanoparticles. Cells of Schizosaccharomyces pombe
were found to synthesize semiconductor CdS nanocrystals
and the productivity was maximum during the mid log
phase of growth. Addition of Cd in the initial exponential
phase of yeast growth affected the metabolism of the organ-
ism (Kaushik and Dhiman, 2002). Baker’s yeast (Saccharomyces
cerevisiae) has been reported to be a potential candidate for the
transformation of Sb2O3nanoparticles and the tolerance of
the organism towards Sb2O3has also been assessed. Particles
with a size range of 2–10 nm were obtained in this condition.
Extracellular secretion of the microorganisms offers the
advantage of obtaining large quantities of materials in
a relatively pure state, free from other cellular proteins
associated with the organism with relatively simpler down-
stream processing. Mycelia free spent medium of the fungus,
Cladosporium cladosporioides was used to synthesize silver
nanoparticles extracellularly. It was hypothesized that pro-
teins, polysaccharides and organic acids released by the
fungus were able to differentiate different crystal shapes and
were able to direct their growth into extended spherical crys-
tals (Balaji et al., 2009). The extracellular synthesis of AgNPs
by a marine fungus Penicillium fellutanum, isolated from costal
mangrove sediment, has been described by Kathiresan et al.
(2009). The extracellular synthesis of stable AgNPs using the
fungus Penicillium brevicompactum WA 2315 was demonstrated
by Shaligram et al. (2009). A single pot green chemical synthe-
sis of AuNP by fungal strain Rhizopus oryzae has been reported
by Das et al. (2009).
Fusarium oxysporum has been reported to synthesize silver
nanoparticles extracellularly. Studies indicate that a nitrate
reductase was responsible for the reduction of silver ions and
the corresponding formation of silver nanoparticles. However
Fusarium moniliformae did not produce nanoparticles either
intracellularly or extracellularly even though they had intra-
cellular and extracellular reductases in the same fashion as
F. oxysporum. This indicates that probably the reductases in
F. moniliformae were necessary for the reduction of Fe(III) to
Fe(II) and not for Ag(I) to Ag(0) (Duran et al., 2005). Aspergillus
flavus has been found to accumulate silver nanoparticles on
the surface of its cell wall when challenged with silver nitrate
solution. Monodisperse silver nanoparticles with a size range
of 8.92 ±1.61 nm were obtained and it was also found that
a protein from the fungi acted as a capping agent on the
nanoparticles (Vigneshwaran et al., 2007).
Aspergillus fumigatus has been studied as a potential candi-
date for the extracellular biosynthesis of silver nanoparticles.
The advantage of using this organism was that the synthesis
process was quite rapid with the nanoparticles being formed
within minutes of the silver ion coming in contact with the
cell filtrate. Particles with a size range of 5–25nm could be
obtained using this organism (Bhainsa and D’Souza, 2006).
In addition to the synthesis of silver nanoparticles, F. oxys-
porum has also been used to synthesize zirconia nanoparticles.
It has been reported that cationic proteins with a molecu-
lar weight of 24–28 kDa (similar in nature to silicatein) were
responsible for the synthesis of the nanoparticles (Bansal
et al., 2004).
Instead of fungi culture, isolated proteins from them have
also been used successfully in nanoparticles production.

1006 environmental toxicology and pharmacology 36 (2013) 997–1014
Nanocrystalline zirconia was produced at room temperature
by cationic proteins while were similar to silicatein secreted
by F. oxysporum (Mohanpuria et al., 2008).
The use of specific enzymes secreted by fungi in the synthe-
sis of nanoparticles appears to be promising. Understanding
the nature of the biogenic nanoparticle would be equally
important. This would lead to the possibility of genetically
engineering microorganisms to over express specific reducing
molecules and capping agents and thereby control the size
and shape of the biogenic nanoparticles (Balaji et al., 2009).
The exposure of Verticillium sp. to silver ions resulted in
a similar intracellular growth of silver nanoparticles (AgNPs)
(Mukherjee et al., 2001). The intracellular formation mecha-
nism of AuNPs and AgNPs has not been understood. However,
it has been postulated that the gold and silver ions initially
bind on the fungal cell surface through electrostatic interac-
tion. The adsorbed metal ions are then reduced by enzymes
present in the cell wall, leading to the formation of the metal
nuclei, which subsequently grow through further reduction
of metal ions. Absar et al. (2005) reported the extra- and
intracellular biosynthesis of AuNPs by fungus Trichothecium
sp. They observed that Trichothecium sp. reacted with gold
ions during stationary phase and forms extracellular AuNPs
of various morphologies, such as spherical, rod-like and tri-
angular. However, under shaking conditions, the same fungal
biomass forms intracellular AuNPs under shaking conditions.
It was postulated that under shaking condition fungi secretes
enzymes and proteins into the medium, however in shaking
conditions these enzymes and proteins are not being released,
thus resulting in the formation of extracellular or intracellu-
lar AuNP, respectively. Fungal templates have been used for
noble-MNP synthesis (Bigall et al., 2008). Fungal cells were
grown in presence of AuNP. Growth of a variety of fungi,
such as Penicillium citreonigrum,Trametes versicolor,Fusarium
sp., Phanaerochaete crysosporium,Trichoderma viride,Neurospora
crassa,Nematolona frowardii, and Bjerkandera adusta was tested
in citrate-stabilized colloidal medium containing different
noble-metal nanoparticles.
Thermomonos sp. reduced the gold ions extracellularly,
yielding ANPs (Sastry et al., 2003). Even the edible mushroom
Volvariella volvacea can produce Au and Ag NPs through metal
reducing compounds. The mushroom was boiled initially in
water and then filtered. The filtrate was cooled to room tem-
perature and used as reducing agent for AuNPs synthesis.
Following reduction purple coloured AuNPs was formed. The
mushroom biomass also prevents NPs aggregation after their
formation (Philip, 2009).
Microbiological methods generate nanoparticles at a much
slower rate than that observed when plant extracts are used.
This is one of the major drawbacks of biological synthesis of
nanoparticles using microorganisms and must be corrected if
it must compete with other methods. Different microorgan-
isms that are capable of synthesizing different nanoparticles
are listed in Table 2.
5.4. Use of plants to synthesize nanoparticles
The advantage of using plants for the synthesis of nanoparti-
cles is that they are easily available, safe to handle and possess
a broad variability of metabolites that may aid in reduction.
A number of plants (Fig. 4) are being currently investi-
gated for their role in the synthesis of nanoparticles. Gold
nanoparticles with a size range of 2–20 nm have been syn-
thesized using the live alfalfa plants (Torresday et al., 2002).
Nanoparticles of silver, nickel, cobalt, zinc and copper have
also been synthesized inside the live plants of Brassica juncea
(Indian mustard), Medicago sativa (Alfa lfa) and Heliantus
annus (Sunflower). Certain plants are known to accumulate
higher concentrations of metals compared to others and
such plants are termed as hyperaccumulators. Of the plants
investigated, B. juncea had better metal accumulating abil-
ity and later assimilating it as nanoparticles (Bali et al.,
2006).
Recently much work has been done with regard to plant
assisted reduction of metal nanoparticles and the respective
role of phytochemicals. The main phytochemicals respon-
sible have been identified as terpenoids, flavones, ketones,
aldehydes, amides and carboxylic acids in the light of IR spec-
troscopic studies. The main water soluble phytochemicals
are flavones, organic acids and quinones which are respon-
sible for immediate reduction. The phytochemicals present
in Bryophyllum sp. (Xerophytes), Cyprus sp. (Mesophytes) and
Hydrilla sp. (Hydrophytes) were studied for their role in the
synthesis of silver nanoparticles. The Xerophytes were found
to contain emodin, an anthraquinone which could undergo
redial tautomerization leading to the formation of silver
nanoparticles.
The Mesophytes contain three types of benzoquinones,
namely, cyperoquinone, dietchequinone and remirin. It was
suggested that gentle warming followed by subsequent incu-
bation resulted in the activation of quinones leading to
particle size reduction. Catechol and protocatechaldehyde
were reported in the hydrophytes studied along with other
phytochemicals. It was reported that catechol under alka-
line conditions gets transformed into protocatechaldehyde
and finally into protocatecheuic acid. Both these processes
liberated hydrogen and it was suggested that it played a
role in the synthesis of the nanoparticles. The size of the
nanoparticles synthesized using xerophytes, mesophytes and
hydrophytes were in the range of 2–5 nm (Jha and Prasad,
2009).
Recently gold nanoparticles have been synthesized using
the extracts of Magnolia kobus and Diopyros kaki leaf extracts.
The effect of temperature on nanoparticle formation was
investigated and it was reported that polydispersed particles
with a size range of 5–300 nm was obtained at lower temper-
ature while a higher temperature supported the formation
of smaller and spherical particles (Song et al., 2009). Name
of some plants and synthesized nanoparticles are listed in
Table 3.
While fungi and bacteria require a comparatively longer
incubation time for the reduction of metal ions, water sol-
uble phytochemicals do it in a much lesser time. Therefore
compared to bacteria and fungi, plants are better candidates
for the synthesis of nanoparticles. Recently, Fahmy et al.
introduced fully green nanotechnology as a gateway to ben-
eficiation of natural cellulose fibres (Fahmy and Mobarak,
2008, 2011). Taking use of plant tissue culture techniques and
downstream processing procedures, it is possible to synthe-
size metallic as well as oxide nanoparticles on an industrial

environmental toxicology and pharmacology 36 (2013) 997–1014 1007
Table 2 – List of microorganisms and synthesized metal nanoparticles.
Microorganisms Products Size (nm) Shape
Sargassum wightii Au 8–12 Planar
Rhodococcus sp. Au 5–15 Spherical
Shewanella oneidensis Au 12 ±5 Spherical
Plectonemaboryanum Au <10–25 Cubic
Plectonema boryanum UTEX 485 Au 10 nm to 6 ␮m Octahedral
Candida utilis Au Not available Not available
V. luteoalbum Au Not available Not available
Escherichia coli Au 20–30 Triangles, hexagons
Yarrowia lipolytica Au 15 Triangles
Pseudomonas aeruginosa Au 15–30 Not available
Rhodopseudomonas capsulate Au 10–20 Spherical
Shewanella algae Au 10–20 Not available
Brevibacterium casei Au, Ag 10–50 Spherical
Trichoderma viride Ag 5–40 Spherical
Phaenerochaete chrysosporium Ag 50–200 Pyramidal
Bacillus licheniformis Ag 50 Not available
Escherichia coli Ag 50 Not available
Corynebacterium glutamicum Ag 5–50 Irregular
Trichoderma viride Ag 2–4 Not available
Ureibacillus thermosphaericus Au 50–70 Not available
Bacillus cereus Ag 4–5 Spherical
Aspergillus flavus Ag 8.92 ±1.61 Spherical
Aspergillus fumigatus Ag 5–25 Spherical
Verticillium sp. Ag 25 ±8 Spherical
Fusarium oxysporum Ag 5–50 Spherical
Neurospora crassa Au, Au/Ag 32, 20–50 Spherical
Shewanella algae Pt 5 Not available
Enterobacter sp. Hg 2–5 Spherical
Shewanella sp. Se 181 ±40 Spherical
Escherichia coli Cd/Te 2.0–3.2 Spherical
Yeast Au/Ag 9–25 Irregular polygonal
Fusarium oxysporum Au-Ag alloy 8–14 Spherical
Pyrobaculum Islandicum U(VI), Tc(VII), Cr(VI), Co(III), Mn(IV) N/A Spherical
Desulfovibrio desulfuricans Pd 50 Spherical
Fig. 4 – A schematic representation of plant as a source of green nanosynthesis, its characterization and biomedical
application.

1008 environmental toxicology and pharmacology 36 (2013) 997–1014
Table 3 – Name of some plants and synthesized nanoparticles.
Plants Nanoparticles Shape Size
Alfalfa plant (Medicago sativa) Au and Ag Spherical and triangular 20–40 nm
Avena sativa Au Spherical 25–85 nm
Azadirachta indica Ag, Au and Ag/Au bimetallic Spherical 50–100 nm
Aloe vera Ag Spherical 15–15.6 nm
Emblica officinalis Ag & Au Mainly spherical 10–20 nm and 15–25 nm respectively
Cinnamomum camphora Au & Ag Mainly spherical 55–80 nm
Tamarind leaf extract Au nanotriangles Mainly spherical 20–40 nm
Capsicum annum Ag Nanoparticles Mainly spherical 16–40 nm
Medicago sativa Ti/Ni bimetallic Mainly spherical 2–6 nm
Medicago sativa Zn Spherical 2–5.6 nm
scale once issues like the metabolic status of the plant etc. are
properly addressed.
Recently, scientists in India have reported the green syn-
thesis of silver nanoparticles using the leaves of the obnoxious
weed, Parthenium hysterophorus. Particles in the size range of
30–80 nm were obtained after 10min of reaction. The use of
this noxious weed has an added advantage in that it can
be used by nanotechnology processing industries (Parasar
et al., 2009). Mentha piperita leaf extract has also been used
recently for the synthesis of silver nanoparticles. Nanopar-
ticles in the size range of 10–25 nm were obtained within
15 min of the reaction (Parasar et al., 2009). Azadirachta
indica leaf extract has also been used for the synthesis of
silver, gold and bimetallic (silver and gold) nanoparticles
(Tripathi et al., 2009). Studies indicated that the reducing
phytochemicals in the neem leaf consisted mainly of ter-
penoids. It was found that these reducing components also
served as capping and stabilizing agents in addition to reduc-
tion as revealed from FT IR studies. The major advantage
of using the neem leaves is that it is a commonly avail-
able medicinal plant and the antibacterial activity of the
biosynthesized silver nanoparticle might have been enhanced
as it was capped with the neem leaf extract. The major
chemical constituents in the extract were identified as nim-
bin and quercetin (Shankar et al., 2004; Tripathi et al.,
2009).
6. Bio-inspired green nanoparticles over
chemically synthesized nanoparticles
In keeping with global efforts to reduce generation of haz-
ardous waste and to develop energy-effective production
routes, ‘green’ chemistry and biochemical processes are
progressively integrating with modern developments in sci-
ence and technology. Hence, any synthetic route or chemical
process should address the fundamental principles of ‘green
chemistry’ by using environmentally benign solvents and
nontoxic chemicals (Anastas and Warner, 1998). The green
synthesis of MNPs should involve three main steps based
on green chemistry perspectives, namely (1) the selection
of a biocompatible and nontoxic solvent medium, (2) the
selection of environmentally benign reducing agents, and (3)
the selection of nontoxic substances for stabilization of the
nanoparticles. Employing these principles in nanoscience will
facilitate the production and processing of inherently safer
nanomaterials and nanostructured devices. Green nanotech-
nology (Dahl et al., 2007) thus aims to the application of green
chemistry principles in designing nanoscale products, and
the development of nanomaterial production methods with
reduced hazardous waste generation and safer applications.
Further, biochemical processes can occur at low tempera-
tures, because of the high specificity of the biocatalysts. Hence,
a synthetic route that include one or more biological steps
will result in consistent energy saving and lower environmen-
tal impact with respect to conventional methods. To optimize
safer nanoparticle production, it would be desirable to employ
bio-based methods, which could minimize the hazardous
conditions of material fabrication. Taking inspiration from
nature, where living organisms produce inorganic materials
through biologically guided process known as biominer-
alization, should be adopted as a superior approach to
nanomaterials assembly (Mann, 1993). The biomineralization
processes exploit biomolecular templates that interact with
the inorganic material at nanoscale, resulting in extremely
efficient and highly controlled syntheses. The structures of
these biocomposite materials are highly controlled both at
nano- and macroscale level, resulting in complex architec-
tures that provide multifunctional properties.
Simpler organisms, such as bacteria, algae, and fungi, have
also developed highly specialized strategies for biominerals
synthesis through hundreds of millions of years of evolution.
The role of the templating molecule in biomineralization is to
provide a synthetic microenvironment in which the inorganic
phase morphology is tightly controlled by a range of low-range
interactions.
7. Applications of metal nanoparticles in
medical biology
The reason why these nanoparticles are attractive for medi-
cal purposes is based on their important and unique features
such as surface to mass ratio that is much larger than that
of other particles, their quantum properties and their abil-
ities to adsorb and carry other compounds such as drugs,
probes and protein. In the ever expanding field of nanomate-
rial research, metal nanoparticle received particular attention
due to their wide application (Fig. 5) in catalysis, electronics,
sensing, photonics, environmental cleanup,imaging, and drug
delivery (Guo et al., 2005; Daniel and Astruc, 2004; Huang et al.,
2007).

environmental toxicology and pharmacology 36 (2013) 997–1014 1009
Fig. 5 – A schematic representation on biological
application of green nanomaterials.
7.1. Catalysis
The application of nanoparticles as catalysts is a rapidly
growing field in nanoscience and technology. The proper-
ties of noble metal nanoparticles make them ideal materials
for nanocatalysis, where reaction yield and selectivity are
dependent on the nature of the catalyst surface. Compared
to bulk materials, nanoparticles have high surface-area-to
volume ratio and thus found to exhibit higher turnover fre-
quencies. The catalytic activity of Au, Ag and Pt in the
decomposition of H2O2to oxygen is well known. Additionally
they also catalyze luminal-H2O2systems. It was observed that
the chemiluminescence emission from the luminal-H2O2sys-
tem was greatly enhanced by addition of Ag colloid (Guo et al.,
2008). The catalytic application of Ag in oxidation of ethylene
to ethylene oxide and methanol to formaldehyde is also the
most popular.
More interestingly, the nanoparticles shows shaped con-
trolled catalytic activity. The shape-controlled catalytic
properties have recently been observed in benzene hydro-
genation by Pt catalyst (Brown and Hutchison, 1997). Among
different Pt nanocrystals (cubes, tetrahedral, and spheres)
tetrahedral nanocrystals, completely bound by crystal facets,
exhibited highest catalytic activity whereas cubic nanocrys-
tals exhibit the lowest activity. However, it is not clearly
understood whether this observation is truly a shape-
dependent effect, as in solution the surface reconstruction
and shape changes of NPs is evident. To study the dynamics
of adsorbents on solution phase nanocrystalline structures is
required for direct surface measurements analogous to those
commonly used for single-crystalline studies.
Among other metal catalysts, Au has potentially more
advantages due to its lower cost and greater stability. Au
is substantially cheaper and more plentiful then Pt. AuNP,
less than 5 nm supported on base metal oxide or carbon
demonstrated very high activity (Hvolbek et al., 2007). High
activity of AuNP for the oxidation of many compounds, par-
ticularly CO and trimethylamine are also observed. AuNP
based gas sensors have recently been developed for detec-
ting a number of gases, including CO and NOx (Thompson,
2007). Very recently Zeng et al. (2010) also demonstrated shape
controlled catalytic activity of AuNPs for well known p-nitro
phenol reduction in presence of sodium borohydride. Gold
nanoboxes among other nanostructure (nanocages, and solid
nanoparticles) have highest catalytic activity. The good intrin-
sic electrical connection across the entire surface of an Au
nanocage makes it a much better catalyst than small Au solid
nanoparticles for the redox reaction.
7.2. Biological application of metal nanoparticles
The application of metal nanoparticles in biological science
showed very rapid progress in the area of labelling, delivery,
heating, and sensing in the past decades. The SPER optical
properties of colloidal AuNPs directed towards recent biomed-
ical applications with an emphasis on cancer diagnostics and
therapeutics.
7.2.1. Labelling
For labelling, electron absorbing properties of the metal
nanoparticles are exploited to generate contrast. The AuNPs
strongly absorb electrons, thus make them suitable as a con-
trasting agent in TEM. Besides, nanoparticles have the same
size domain as proteins that make nanomaterials suitable for
bio tagging or labelling (Sperling et al., 2008). Due to their
small size and functionalising properties, i.e. with antibod-
ies (immunostaining), AuNPs provide extremely high spatial
resolution and applied in many labelling applications (Salata,
2004). Additionally optical detection techniques are wide
spread in biological research because of change of their opti-
cal or fluorescence properties. Similarly, the particles’ optical
properties – strong absorption, scattering and especially plas-
mon resonance – make them of value for a large variety of
light-based techniques including combined schemes such as
photothermal or photo-acoustic imaging. In addition, AuNP
can be radioactively labelled by neutron activation, which
allows for very sensitive detection, and used as an X-ray con-
trast agent.
7.2.2. Sensors
Metal nanoparticles can also be used as sensors. The opti-
cal and electronic sensing of biomaterials on surfaces is a
common practice in analytical biochemistry. Thus, the immo-
bilization of biomolecule–NP conjugates on surfaces provides
a general route for the development of optical or electronic
biosensors. Metal NPs such as Au or Ag NPs exhibit plasmon
absorbance bands in the visible spectral region that are
controlled by the size of the respective particles. Their optical
properties can change upon binding to certain molecules,
allowing the detection and quantification of analytes (Huo,
2007). The absorption spectra of AuNP change drastically
when several particles come close to each other. Numerous
studies on the labelling of bioassays and the staining of

1010 environmental toxicology and pharmacology 36 (2013) 997–1014
biological tissues by metal particles as a means to image and
visualize biological processes have been reported. The spec-
tral shifts which originate from adjacent or aggregated metal
nanoparticles, such as Au NPs, are of increasing interest in the
development of optical biosensors based on biomolecule–NP
hybrid systems. As an example, NPs that were functionalized
with two kinds of nucleic acid, which were complementary
to two segments of an analyzed DNA, were hybridized with
the analyzed DNA. This led to the aggregation of the NPs
and to the detection of a red shifted interparticle plasmon
absorbance of the nanoparticle aggregate.
7.2.3. Drug delivery
Because of nontoxicity and nonimmunogenicity AuNPs is
ideal for preparation of drug delivery scaffold. Functionaliza-
tion property of AuNP also makes it an excellent potential
vehicle for the drug delivery. Functionalized AuNP represent
highly attractive and promising candidates in the applica-
tions of drug delivery. Aubin-Tam and Hamad-Schifferli (2008)
recently developed drug delivery system with AuNPs and
infrared light. This delivery system released multiple drugs in
a controlled fashion. They demonstrated that nanoparticles
of different shapes respond to different infrared wavelengths.
For example, nanobones and nanocapsules melt at light
wavelengths of 1100 and 800nm, respectively. Thus exci-
tation at one wavelength could selectively melt one type
of Au nanorods and selectively release one type of DNA
strand. Brown et al. (2010) also reported AuNPs for the
improved anticancer drug delivery of the active component
of oxaliplatin. Naked AuNPs were functionalized with a thi-
olated poly(ethylene glycol) (PEG) monolayer capped with a
carboxylate group. [Pt(1R,2Rdiaminocyclohexane)(H2O)2]2NO3
was added to the PEG surface and yielding a supramolecular
complex with drug molecules. The cytotoxicity, drug uptake,
and localization in the A549 lung epithelial cancer cell line
and the colon cancer cell lines HCT116, HCT15, HT29, and
RKO were examined for platinum-tethered nanoparticles. The
platinum-tethered nanoparticles showed significant improve-
ment in cytotoxicity than oxaliplatin alone in all of the cell
lines and an unusual ability to penetrate the nucleus in the
lung cancer cells.
7.2.4. Cancer therapy
Nanotechnology is one of the most popular research areas,
especially with regard to biomedical applications. Nanopar-
ticles have very good opportunity in the form of targeted
drug therapies (Ghosh et al., 2008). Nanoparticles also carry
the potential for targeted and time-release drugs. A potent
dose of drugs could be delivered to a specific area but engi-
neered to release over a planned period to ensure maximum
effectiveness and the patient’s safety. The strong light absorb-
ing properties of AuNPs makes it suitable as heat mediating
objects; the absorbed light energy is dissipated into the
surroundings of the particles’, generating an elevated temper-
ature in their vicinity. This effect can be used to open polymer
microcapsules, for example, for drug delivery purposes and
even destroys the cancerous cells. The nanoparticles are func-
tionalized with antibody specific to the cancerous cells. The
functionalized nanoparticles specifically bind with the tar-
geting cells, which was then killed by hyper thermal therapy
through heating the particle-loaded tissue. However, for such
in vivo applications, the potential cytotoxicity of the nanopar-
ticles might become an issue and should be investigated with
care. Due to biocompatibility, hyper thermal activity AuNPs
find wide application now-a-days in killing of malignant
cancerous cells (Dickerson et al., 2008). Recently, Melancon
et al. (2008) demonstrated destruction of cancerous cell by
photothermal effect of AuNPs. The hollow gold nanoshells
(HAuNS; average diameter, ∼30 nm) were covalently attached
to monoclonal antibody directed to the epidermal growth
factor receptor (EGFR). The resulting anti-EGFR-HAuNS exhib-
ited excellent colloidal stability and efficient photothermal
effect in the near-infrared region. Anti-EGFR-HAuNS then
bound in EGFR-positive A431 tumour cells. Irradiation of A431
cells treated with anti-EGFR-HAuNS with near-infrared laser
resulted in selective destruction of these cells.
AuNPs has also been applied to amplify the biorecogni-
tion of the anticancer drug (Shen et al., 2008). Dacarbazine
[5-(3,3-dimethy-1-triazenyl) imidazole-4-carboxamide; DTIC]
is a commonly used anticancer drug. AuNPs were stabilized
by PPh3 with negative charge. The oxidized DTIC is positive
charged. Thus, DTIC could be easily assembled onto the sur-
face of AuNPs. The specific interactions between anticancer
drug DTIC and DNA or DNA bases were facilitated by AuNPs.
7.3. Environmental cleanup as defense against
environmental challenge to medical biology
Although MNPs are increasingly being employed in different
emergent areas, their use in environmental biotechnology is
still limited. One of the key environmental challenges is the
contamination of water bodies by different chemicals due
to diverse anthropogenic and industrial activities. The most
interesting application of MNPs is purification of drinking
water contaminated with heavy metals and pesticides. Cur-
rent limitations in removal of heavy metals have been tried to
overcome through adsorption process on MNPs due to alloy
formation. Au and mercury exist in several phases such as
Au3Hg, AuHg, and AuHg3.
The interaction of AgNPs with Hg2+ ions was investigated
because enhanced ability of Ag to form alloy in different
phases. It was found that the surface plasmon of AgNPs blue
shifted along with a decrease in the intensity, immediately
after the addition of Hg2+ ions (Bootharaju and Pradeep, 2010).
Partial oxidation of AgNPs to silver ions is responsible for the
decrease in intensity. The shift is attributed to the incorpo-
ration of mercury into the AgNPs. The mercury nanoparticle
solutions exhibited plasmon absorption band below 300nm.
The Hg–Ag alloy nanoparticles, prepared by simultaneous
reduction with sodium borohydride, exhibited a plasmon in
the region of 300–400 nm. The potential of AgNPs to reduce a
number of heavy metals can also be looked at as a method to
prepare alloy nanoparticles; e.g., Ag–Hg bimetallic nanoparti-
cles.
Recently colorimetric detections of heavy metals like
arsenic, mercury, lead, etc., have also been tried by using
MNPs. One of the important properties exhibited by function-
alized MNPs surfaces is the detection of heavy metals. In one
such method, heavy metal specific biomolecule functional-
ized AuNP can be utilized. An example of this approach is

environmental toxicology and pharmacology 36 (2013) 997–1014 1011
the interaction of metal ions with nucleotides: Hg2+ promoted
formation of thymine–thymine base pairs (Ono and Togashi,
2004). In a similar approach, ligands functionalized MNPs have
been used for specific detection of metal ions. This ligand-
metal ion complexation leads to observable optical changes
at concentrations in the ppm level. Examples of such ligands
are gallic acid (Pb2+), cysteine (Hg2+,Cu
2+), and mercaptounde-
canoic acid (Pb2+,Cd
2+,Hg
2+). Carboxylate group modified
surface of AuNP can be induced to aggregate in the presence of
Hg2+ and pyridinedicarboxylic acid, which is manifested in the
form of colorimetric response, fluorescence quenching and
enhancement of hyper-Rayleigh scattering intensity (Huang
et al., 2007; Darbha et al., 2008).
The removal of pesticides by MNPs is a new addition to
this field. Among other contaminants, presence of pesticide
residue in potable water above permissible limit is of great
concern to public health. This happens due to indiscriminate
use of pesticide, specially belonging to organophosphorus
groups, in agricultural practices. It is essential to reduce
the concentration of pesticide in potable water but difficult
to achieve by conventional chemical methods due to wide
variation of their chemical structures. To meet these environ-
mental challenges, very recently researchers are focusing on
the development of methods based on nanotechnology. Very
recently, Das et al. (2009) demonstrated adsorption of differ-
ent organophosphorous pesticides on AuNPs surface. AuNPs
was synthesized on the surface of the R. oryzae mycelia in a
single set. The AuNPs adsorbed on mycelia were then used
for adsorption of different organophosphorous pesticides.
Following adsorption of these pesticides the surface mor-
phology conspicuously changed compared the unadsorbed
nanomaterial as depicted from atomic force microscopic
images.
8. Conclusions
In this review, we provided an account of the biological meth-
ods for MNPs (green nano) synthesis, as well as their most
promising applications in biomedical devices and in environ-
mental processes. From the considerations as outlined in this
review, it emerges that biosynthesis represents a promising
route for MNPs production. In fact, biosynthesis results in
low energy use and environmental impact, with respect to
conventional chemical synthesis methods. Further, the high
specificity of biomolecules involved in the biosynthesis pro-
cess may enable an efficient control of MNPs size and shape,
whose tight control is critical to optimize MNP-based devices
and applications. Current nanotoxicological research aims to
identify the physico-chemical characteristics of NPs respon-
sible for the observed health effects. These results could
be incorporated in the design of new engineered NPs. The
challenge is to produce new nanomaterials that are without
adverse characteristics and still fulfil the industrial require-
ments. This approach would have the advantage of initiating
a sustainable and safe nanotechnology.
Biosynthetic MNPs have been observed in numerous
fungal and bacterial species, and the molecular machinery
needed for MNPs biosynthesis overlap significantly with
the developed and optimized system for bioreduction and
detoxification of soluble metals. However, most published
studies deal with MNP biosynthesis in viable microorgan-
ism, and the complexity of the system makes it difficult to
identify the exact nature of the multiple biological agents
responsible for the biosynthetic process. Further research
with cell-free extract and biological fractions may lead to this
identification and thorough understanding of the complex
regulatory processes underlying the expression of metal
reducing agents. Many green nanomaterials require new
commercial production techniques, which increases the need
for basic research, engineering research, and coordination of
the two between the industrial and research communities.
Toxicology and analysis protocols need to be developed and
constantly updated to reflect advances in the science.
Little development in the field is being specifically targeted
towards sustainability. This is a problem because the opportu-
nity to develop green processes fromthe beginning is not being
taken. The transformation to sustainable development is an
enormous economic opportunity. Moreover, past achievement
aimed at being able to watch the next generation prosper has
come via terribly destructive methods. What is needed now is
redevelopment – a paradigm shift in the approaches in provid-
ing aids to medical sciences. Clearly if nanotechnology is to be
the key to the future, it should be developed with sustainability
in mind from the outset.
Conflict of interest
There is no conflict of interest regarding the publication of this
review in any journal between the authors, funding agencies
or the host institute.
Acknowledgements
Authors are grateful to the Vice-Chancellor, University of
Kalyani, Kalyani, Nadia, for his interest and support in this
work. Authors are thankful to DST Purse Project, DST India,
for their cooperation and support.
references
Absar, A., Satyajyoti, S., Khan, M.I., Rajiv, K., Sastry, M., 2005.
Extra-/intracellular biosynthesis of gold nanoparticles by an
alkalotolerant fungus, Trichothecium sp. J. Biomed.
Nanotechnnol. 1, 47–53.
Ahmad, A., Mukherjee, P., Senapati, S., Mandal, D., Khan, M.I.,
Kumar, R., Sastry, M., 2003. Extracellular biosynthesis of silver
nanoparticles using the fungus Fusarium oxysporum. Colloid
Surf. B: Biointerfaces 28, 313–318.
Albrecht, M.A., Evans, C.W., Raston, C.L., 2006. Green chemistry
and the health implications of nanoparticles. Green Chem.,
http://dx.doi.org/10.1039/b517131h (critical review).
Anastas, P.T., Warner, J.C., 1998. Green Chemistry: Theory and
Practice. Oxford University Press, Oxford, England/New York.
Aptekar, J.W., Cassidy, M.C., Johnson, A.C., Barton, R.A., Lee, M.Y.,
Ogier, A.C., Vo, C., Anahtar, M.N., Ren, Y., Bhatia, S.N.,
Ramanathan, C., Cory, D.G., Hill, A.L., Mair, R.W., Rosen, M.S.,
Walsworth, R.L., Marcus, C.M., 2009. Silicon nanoparticles as
hyperpolarized magnetic resonance imaging agents. ACS
Nano 3 (12), 4003–4008.

1012 environmental toxicology and pharmacology 36 (2013) 997–1014
Astruc, D.D., 2008. Transition metal nanoparticles in catalysis:
from historical background to the state of the art. In: Astruc,
D.D. (Ed.), Nanoparticles and Catalysis. Verlag GmbH & Co.
KGaA, Weinheim, pp. 1–48.
Aubin-Tam, M.-E., Hamad-Schifferli, K., 2008. Structure and
function of nanoparticle–protein conjugates. Biomed. Mater.
3, 034001 (invited contribution).
Balaji, D.S., Basavaraja, S., Deshpande, R., Mahesh, D.B.,
Prabhakar, B.K., Venkataraman, A., 2009. Extracellular
biosynthesis of functionalized silver nanoparticles by strains
of Cladosporium cladosporioides fungus. Colloid Surf. B:
Biointerfaces 68, 88–92.
Bali, R., Razak, N., Lumb, A., Harris, A.T., 2006. The synthesis of
metal nanoparticles inside live plants. IEEE Xplore,
http://dx.doi.org/10.1109/ICONN.2006.340592.
Bansal, V., Rautaray, D., Ahmad, A., Sastry, M., 2004. Biosynthesis
of zirconia nanoparticles using the fungus Fusarium
oxysporum. J. Mater. Chem. 14, 3303–3305.
Beveridge, T.J., Murray, R.G.J., 1976. Uptake and retention of
metals by cell walls of Bacillus subtilis. Bacteriol. Sep. 127 (3),
1502–1518.
Bhainsa, K.C., D’Souza, S.F., 2006. Extracellular biosynthesis of
silver nanoparticles using the fungus Aspergillus fumigatus.
Colloid Surf. B: Biointerfaces 47 (2), 160–164.
Bigall, N.C., Reitzig, M., Naumann, W., Simon, P., van Pée, K.H.,
Eychmüller, A., 2008. Fungal templates for noble metal
nanoparticles and their application in catalysis. Angew.
Chem. Int. Ed. 47, 7876–7879.
Blackman, J.A., 2009. Metallic nanoparticles. In: Misra, P., Black,
J.A. (Eds.), Handbook of Metal Physics. Elsevier BV,
Amsterdam, pp. 4–6.
Bootharaju, M.S., Pradeep, T., 2010. Uptake of toxic metal ions
from water by naked and monolayer protected silver
nanoparticles: an X-ray photoelectron spectroscopic
investigation. J. Phys. Chem. C 114,
8328–8336.
Brown, L.O., Hutchison, J.E., 1997. Convenient preparation of
stable, narrow-dispersity, gold nanocrystals by ligand
exchange reactions. J. Am. Chem. Soc. 119, 12384–12385.
Brown, S.D., Nativo, P., Smith, J.A., Stirling, D., Edwards, P.R.,
Venugopal, B., Flint, D.J., Plumb, J.A., Graham, D., Wheate, N.J.,
2010. Gold nanoparticles for the improved anticancer drug
delivery of the active component of oxaliplatin. J. Am. Chem.
Soc. 132, 4678–4684.
Brust, M., Kiely, C.J., 2002. Some recent advances in nanostructure
preparation from gold and silver: a short topical review.
Colloid Surf. A: Physicochem. Eng. Asp. 202, 175–186.
Chaki, N.K., Sundrik, S.G., Sonawane, H.R., Vijayamohanan, K.,
2002. J. Chem. Soc. Chem. Commun., 76.
Chakrabarti, S., Fathpour, S., Moazzami, K., Phillips, J., Lei, Y.,
Browning, N., Bhattacharya, P., 2004. Pulsed laser annealing of
self-organized InAs/GaAs quantum dots. J. Electron. Mater. 33,
L5.
Clourier, S.G., Eakin, J.N., Guico, R.S., Sousa, M.E., Crawford, G.P.,
Xu, J., 2006. Molecular self organization in cylindrical
nanocavities. Phys. Rev. E 75, 051703.
Dahl, J.A., Maddux, B.L.S., Hutchison, J.E., 2007. Toward greener
nanosynthesis. Chem. Rev. 107, 2228–2269.
Daniel, M.C., Astruc, D., 2004. Gold nanoparticles: assembly,
supramolecular chemistry, quantum-size-related properties,
and applications towards biology, catalysis, and
nanotechnology. Chem. Rev. 104, 293–346.
Darbha, G.K., Rai, U.S., Singh, A.K., Roy, P.C., 2008. Gold nanorod
based sensing of sequence specific HIV-1 virus DNA using
hyper Rayleigh scattering spectroscopy. Eur. J. Chem. 14 (13),
3896–3903.
Das, S.K., Das, A.R., Guha, A.K., 2009. Gold nanoparticles:
microbial synthesis and application in water hygiene
management. Langmuir 25, 8192–8199.
Dickerson, E.B., Dreaden, E.C., Huang, X., El-Sayed, I.H., Chu, H.,
Pushpanketh, S., McDonald, J.F., El-Sayed, M.A., 2008. Gold
nanorod assisted near-infrared plasmonic photothermal
therapy (PPPT) of aquamous cell carcinoma in mice. Cancer
Lett. 269, 57–66.
Duran, N., Marcato, D.P., Alves, L.O., De Souza, G., Esposito, E.,
2005. Mechanical aspect of biosynthesis of silver
nanoparticles by several Fusarium oxysporum strains. J.
Nanobiotechnol. 117.
Durán, N., Marcato, P.D., Durán, M., Yadav, A., Gade, A., Rai, M.,
2011. Mechanistic aspects in the biogenic synthesis of
extracellular metal nanoparticles by peptides, bacteria,
fungi, and plants. Appl. Microbiol. Biotechnol. 90,
1609–1624.
Elechiguerra, J.L., Burt, J.L., Morones, J.R., Camacho-Bragado, A.,
Gao, X., Lara, H.H., acaman, M.J., 2005. Interaction of silver
nanoparticles with HIV-1. J. Nanobiotechnol. (3), 6.
El-Sayed, M.A., 2001. Some interesting properties of metals
confined in time and nanometer space of different shapes.
Acc. Chem. Res. 34, 257–264.
Fahmy, T.Y.A., Mobarak, F., 2008. Vaccination of biological
cellulose fibers with glucose: a gateway to novel
nanocomposites. Int. J. Biol. Macromol. 42 (1), 52.
Fahmy, T.Y.A., Mobarak, F., 2011. Green nanotechnology: a short
cut to beneficiation of natural fibers. Int. J. Biol. Macromol. 48
(1), 134.
Frens, G., 1972. Particle size and sol stability in metal colloids.
Colloid Polym. Sci. 250, 736–741.
Fu, J.K., Liu, Y., Gu, P., Tang, D.L., Lin, Z.Y., Yao, B.X., Weng, S.Z.,
2000. Spectroscopic characterization on the biosorption and
bioreduction of Ag(I) by Lactobacillus sp. A09. Acta
Physico-Chim. Sin. 16 (9), 770–782 (in Chinese).
Fu, M., Li, Q., Sun, D., Lu, Y., He, N., Deng, X., Wang, H., Huang, J.,
2006. Rapid preparation process of silver nanoparticles by
bioreduction and their characterizations. Chin. J. Chem. Eng.
14 (1), 114–117.
Gabor, L.H., Joydeep, D., Harry, F.T., Anil, K.R., 2008.
Characterization methods. In: Gabor, L.H., Joydeep, D., Harry,
F.T., Anil, K.R. (Eds.), Introduction to Nanoscience. CRC Press,
Taylor & Francis Group, Boca Raton, pp. 108–175.
Gacoin, T., Malier, L., Boilot, J.P., 1997. Sol–gel transition in CdS
colloids. Chem. Mater. 9, 1502.
Gericke, M., Pinches, A., 2006. Biological synthesis of metal
nanoparticles. Hydrometallurgy 83, 132–140.
Ghosh, P., Han, G., De, M., Kim, K.C., Rotello, M.V., 2008. Gold
nanoparticles in delivery applications. Adv. Drug Delivery Rev.
60, 1307–1315.
Gleiter, H., 1989. Nan crystalline materials. Prog. Mater. Sci. 33,
223–315.
Gopakumar, G., Nguyen, M.T., Ceulemans, A., 2008. The boron
buckyball has an unexpected Thsymmetry. Chem. Phys. Lett.
450, 475–477.
Graff, A., Benito, S.M., Verbert, C., Meier, W., 2004. Polymer
nanocontainers nanobiotechnology. In: Niemeyer, C.M.,
Mirkin, C.A., Concepts (Eds.), Applications and Perspectives.
WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim, pp.
168–170.
Guo, R., Song, Y., Wang, G., Murray, R.W., 2005. Does core size
matter in the kinetics of ligand exchanges of
monolayer-protected Au clusters? J. Am. Chem. Soc. 127,
2752–2757.
Guo, J.Z., Cui, H., Zhou, W., Wang, W., 2008. Ag
nanoparticle-catalyzed chemiluminescent reaction between
luminal and hydrogen peroxide. J. Photochem. Photobiol. A:
Chem. 193, 89–96.
Gupta, A.K., Curtis, A.S.G., 2004. Lactoferrin and ceruloplasmin
derivatized superparamagnetic iron oxide nanoparticles for
targeting cell surface receptors. Biomaterials 25 (15),
3039–3040.

environmental toxicology and pharmacology 36 (2013) 997–1014 1013
He, S., Guo, Z., Zhang, Y., Zhang, S., Wang, J., Gu, N., 2007.
Biosynthesis of gold nanoparticles using the bacteria
Rhodopseudomonas capsulate. Mater. Lett. 61, 3984–3987.
Huang, C.C., Yang, Z., Lee, K.H., Chang, H.T., 2007. Synthesis of
highly fluorescent gold nanoparticles for sensing mercury(II).
Angew. Chem. Int. Ed. 46, 6824–6828.
Huo, Q., 2007. A perspective on bioconjugated nanoparticles and
quantum dots. Colloid Surf. B: Biointerfaces 59, 1–10.
Husseiny, I.M., El-Aziz, A.M., Badr, Y., Mahmoud, T., 2007.
Biosynthesis of gold nanoparticles using Pseudomonas
aeruginosa. Spectrochim. Acta A 67, 1003–1006.
Hutchison, J.E., 2008. Greener nanoscience: a proactive approach
to advancing applications and reducing implications of
nanotechnology. ACS Nano 2, 395–402.
Hvolbek, W., Janssens, V.T., Clausen, S.B., Falsig, H., Christensen,
H.C., Norskov, K.J., 2007. Catalytic activity of Au nanoparticles.
Nano Today 2, 14–18.
Jha, A.K., Prasad, K., 2009. A green low-cost biosynthesis of Sb2O3
nanoparticles. J. Biochem. Eng. 43, 303–306.
Jian, K., Hurt, R.H., Shelton, B.W., Crawford, G.P., 2006.
Visualization of the liquid crystal director field within carbon
nanotube cavities. Appl. Phys. Lett. 88, 163110.
Justus, B.L., Tonucci, R.J., Berry, A.D., 1992. Nonlinear optical
properties of quantum confined GaAs nanocrystals in Vycor
glass. Appl. Phys. Lett. 61, 3151–3153.
Kalathil, S., Lee, J., Cho, M.H., 2011. Electrochemically active
biofilm-mediated synthesis of silver nanoparticles in water.
Green Chem., http://dx.doi.org/10.1039/c1gc15309a.
Kalimuthu, K., Babu, S.R., Venkataraman, D., Bilal, M.,
Gurunathan, S., 2008. Biosynthes is of silver nanoparticles by
Bacillus licheniformis. Colloid Surf. B: Biointerfaces 65,
150–153.
Kalishwaralal, K., Deepak, V., Ramkumarpndian, S., Nellaiah, H.,
Sangiliyandi, G., 2008. Extracellular biosynthesis of silver
nanoparticles by the culture supernatant of Bacillus
licheniformis. Mater. Lett. 62, 4411–4413.
Kalishwaralal, K., Deepak, V., Ram Kumar, P.S., Gurunathan, S.,
2009. Biosynthesis of gold nanocubes from Bacillus
lichemiformis. Bioresour. Technol. 100, 5356–5358.
Kang, S., Kang, M.H., Lee, E., Seo, S., Ahn, C.W.H., 2011. Facile,
hetero-sized nanocluster array fabrication for investigating
the nanostructure-dependence of nonvolatile memory
characteristics. Nanotechnology 22, 254018,
http://dx.doi.org/10.1088/0957-4484/22/25/254018.
Karazhanov, S.Zh., Raveendran, P., 2003. Ab initio study of double
oxides ZnX2O4(X = Al, Ga, In) having spinel structure. J. Am.
Chem. Soc. 125, 13940.
Kathiresan, K., Manivannan, S., Nabeel, A.M., Dhivya, B., 2009.
Studies on silver nanoparticles synthesized by a marine
fungus Penicillum fellutanum isolated from coastal
mangrove sediment. Colloid Surf. B: Biointerfaces 71,
133–137.
Kaushik, P., Dhiman, A.K., 2002. Medicinal Plants and Raw Drugs
of India, p. XII+623. Retrieved from: http://www.
ve dicbooks.net/medicinal-plants-and-raw-drugs-of-india-
p-886.html
Kelsall, R.W., Hamley, I.W., Geoghegan, M., 2005. Nanoscale
Science and Technology. John Wiley & Sons Ltd., London, pp.
40–58.
Klaus, T., Joerger, R., Olsson, E., Granqvist, C.G., 1999. Silver-based
crystalline nanoparticles, microbially fabricated. Proc. Natl.
Acad. Sci. U. S. A. 23 (9624), 13611–13614.
Konishi, Y., Yoshida, S., Asai, S., 1995. Bioleaching of pyrite by
acidophilic thermophile Acidianus brierleyi. Biotechnol. Bioeng.
48 (6), 592–600.
Kroto, H.W., Walton, D.R.M., 2011. Fullerene. Encyclopædia
Britannica Online.
http://www.britannica.com/EBchecked/topic/221916/fullerene
(accessed 04.02.11).
Langmuir, X., Li, Q., Xie, J., Jin, Z., Wang, J., Li, Y., Jiang, K., Fan, S.,
2009. Fabrication of ultralong and electrically uniform
single-walled carbon nanotubes on clean substrates. Nano
Lett. 9 (9), 3137–3141.
Law, N., Ansari, S., Livens, F.R., Renshaw, J.C., Lloyd, J.R., 2008.
Formation of nanoscale elemental silver particles via
enzymatic reduction by Geobacter sulfurreducens. Appl.
Environ. Microbiol. 74, 7090–7093.
Liu, H., Salabas, E.L., Schuth, F., 2007. Magnetic nanoparticles:
synthesis, protection, functionalization, and application.
Angew. Chem. Int. Ed. 46, 1222–1244.
Lowenstam, H.A., 1981. Minerals formed by organisms. Science
211, 1126–1131.
Mafuné, F., Kohno, J., Takeda, Y., Kondow, T., 2001. Formation of
gold nanoparticles by laser ablation in aqueous solution of
surfactant. J. Phys. Chem. B 105 (22), 5114–5120.
Mann, S., 1993. Molecular tectonics in biomineralization and
biomimetic materials chemistry. Nature 365, 499–505.
Martel, R., Derycke, V., Lavoie, C., Appenzeller, J., Chan, K.K.,
Tersoff, J., Avouris, P., 2001. Ambipolar electrical transport in
semiconducting single-wall carbon nanotubes. Phys. Rev. Lett.
87, 256805–256809.
Mayes, E.L., Mann, S., 2004. Mineralization in nanstructured
biocompartments: ferritin for high-density data storage. In:
Niemeyer, C.M., Mirkin, C.A. (Eds.), Nanobiotechnology:
Concepts, Applications and Perspectives. WILEYVCH Verlag
GmbH & Co. KGaA, Weinheim, pp. 278–287.
McKenzie, L.C., Hutchison, J.E., 2004. Green nanoscience: an
integrated approach to greener products, processes, and
applications. Chim. Oggi 25, 8.
Melancon, M.P., Lu, W., Yang, Z., Zhang, R., Cheng, Z., Elliot, A.M.,
Stafford, J., Olson, T., Zhang, J.Z., Li, C., 2008. In vitro and
in vivo targeting of hollow gold nanoshells directed at
epidermal growth factor receptor for photothermal ablation
therapy. Mol. Cancer Ther. 7, 1730–1739.
Mie, G., 1908. Beiträge zur Optik trüber Medien, speziell
kolloidaler Metallösungen. Ann. Phys. 25, 377–445.
Mintmire, J.W., Dunlap, B.I., White, C.T., 1992. Are fullerene
tubules metallic? Phys. Rev. Lett. 68 (5), 631–634D.
Mohanpuria, P., Rana, K.N., Yadav, S.K., 2008. Biosynthesis of
nanoparticles: technological concepts and future
applications. J. Nano Res. 10, 507–517.
Mukherjee, P., Ahmad, A., Mandal, D., Senapati, S., Sainkar, S.R.,
Khan, M.I., Parischa, R., Ajayakumar, P.V., Alam, M., Kumar, R.,
Sastry, M., 2001. Fungus mediated synthesis of silver
nanoparticles and their immobilization in the mycelia matrix:
a novel biological approach to nanoparticle synthesis. Nano
Lett. 1, 515–519.
Murray, C.B., Kagan, C.R., Bawendi, M.G., 2000. Synthesis and
characterization of monodisperse nanocrystals and
close-packed nanocrystal assemblies. Annu. Rev. Mater. Sci.
30 (1), 545–610.
Nam, J.M., Thaxton, C.S., Mirkin, C.A., 2003. Nanoparticle-based
bio-bar codes for the ultrasensitive detection of proteins.
Science 301, 1884–1886.
Narayanan, K.B., Sakthivel, N., 2010. Biological synthesis of metal
nanoparticles by microbes. J. Colloid Interface Sci. 156 (1),
6–8.
Neuberger, T., Schöpf, B., Hofmann, H., Hofmann, M., Von
Rechenberg, B., 2005. Superparamagnetic nanoparticles for
biomedical applications: possibilities and limitations of a new
drug delivery system. J. Magn. Magn. Mater. 293 (1), 483–496.
Nie, S., Xing, Y., Kim, G.J., Simons, J.W., 2007. Nanotechnology
applications in Cancer. Annu. Rev. Biomed. Eng. 9, 257–288.
Olshavsky, M.A., Allcock, H.R., 1997. Small scale system for
in-vivo drug delivery. Chem. Mater. 9, 1367.
Ono, A., Togashi, H., 2004. Highly selective oligonucleotide-based
sensor for mercury (II) in aqueous solutions. Angew. Chem.
Int. Ed. 43, 4300–4302.

1014 environmental toxicology and pharmacology 36 (2013) 997–1014
Parasar, U.K., Saxena, P.S., Srivastava, A., 2009. Bioinspired
synthesis of silver nanoparticles digest. J. Nanomater.
Biostruct. 4 (March (1)), 159–166.
Pérez-Tijerina, E., Gracia-Pinilla, M.A., Mejía-Rosales, S.,
Ortiz-Méndez, U., Torres, A., José-Yacamán, M., 2008. Highly
size-controlled synthesis of Au/Pd nanoparticles by inert-gas
condensation. Faraday Discuss. 138, 353–362.
Philip, D., 2009. Biosynthesis of Au, Ag and Au-Ag nanoparticles
using edible mushroom extract. Spectrochim. Acta A 73,
374–381.
Prasad, Jemmis, E., 2008. Stuffing improves the stability of
fullerene-like boron clusters. Phys. Rev. Lett. 100 (16),
165504–165508.
Raffi, M., Rumaiz, A.K., Hasan, M.M., Shah, S.I., 2007. Studies of
the growth parameters for silver nanoparticle synthesis by
inert gas condensation. J. Mater. Res. 22, 3378–3384.
Reith, F., Etschmann, B., Gross, C., Moors, H., Benotmane
Monsieurs, M.A.P., Grass, G., Doonan, C., Vogt, S., Lai, B.,
Martinez-Criado, G., George, G.N., Nies, D.H., Mergeay, M.,
Pring, A., Southam, G., Brugger, J., 2009. Mechanisms of gold
biomineralization in the bacterium Cupriavidus metallidurans.
Proc. Natl. Acad. Sci. U. S. A. 160, 17757–17762.
Ren, W., Ai, Z., Jia, F., Zhang, L., Fan, X., Zou, Z., 2007. Low
temperature preparation and visible light photocatalytic
activity of mesoporous carbon-doped crystalline TiO2. Appl.
Catal. B 69, 138–144.
Roduner, E., 2006. Physics and chemistry of nanostructures: why
nano is different. In: Nanoscience and Nanotechnologies.
EOLSS.
Rosemary, M.J., Pradeep, T., 2003. Solvothermal synthesis of silver
nanoparticles from thiolates. J. Colloid Interface Sci. 268,
81–84.
Salata, O.V., 2004. Applications of nanoparticles in biology and
medicine. J. Nanobiotechnol. 2, 3–9.
Sankaran, V., Yue, J., Cahen, R.E., Schrock, R.R., Silbey, R.J., 1993.
Advanced drug delivery advices. Chem. Mater. 5, 1133.
Sastry, M., Ahmad, A., Khan, I.M., Kumar, R., 2003. Biosynthesis of
metal nanoparticles using fungi and actinomycete. Curr. Sci.
85 (2), 162–170.
Senjen, R., 2007. Nano silver-a threat to soil, water and human
health? Friends of the earth Australia, Available at:
http://nano.foe.org.au/node/189
Shaligram, S.N., Bule, M., Bhambure, R., Singhal, S.R., Singh, K.S.,
Szakacs, G., Pandey, A., 2009. Biosynthesis of silver
nanoparticles using aqueous extract from the compactin
producing fungal. Process Biochem. 44, 939–943.
Shankar, S.S., Rai, A., Ahmad, A., Sastry, M., 2004. Rapid synthesis
of Au, Ag and bimetallic Au core–Ag shell nanoparticles using
neem (Azadirachta indica) leaf broth. J. Colloid Interface Sci.
275, 496–502.
Sharma, V.K., Yngard, R.A., Lin, Y., 2009. Silver nanoparticles:
green synthesis and their antimicrobial activities. Adv. Colloid
Interface Sci. 145, 83–96.
Shen, Q., Wang, X., Fu, D., 2008. The amplification effect of
functionalized gold nanoparticles on the binding of cancer
drug decarbazine to DNA and DNA bases. Appl. Surf. Sci. 255,
577–580.
Shukla, S., Seal, S., 1999. Cluster size effect observed for gold
nanoparticles synthesized by sol–gel technique as studied by
X-ray photoelectron spectroscopy. Nano Struct. Mater. 11,
1181–1193.
Singaravelu, G., Arockiyamari, J., Ganesh Kumar, V., ovindaraju,
K., 2007. A novel extracellular biosynthesis of monodisperse
gold nanoparticles using marine algae. Colloid Surf. B:
Biointerfaces 57, 97–101.
Smith, D.K., Diederich, F., 2000. Supramolecular dendrimer
chemistry: a journey through the branched architecture. Top.
Curr. Chem. 210, 183–227.
Song, J.Y., Jang, H.K., Kim, B.S., 2009. Biological synthesis of gold
nanoparticles using Magnolia kobus and Diopyros kaki leaf
extracts. Process Biochem. 44, 1133–1138.
Southam, G., Beveridge, T.J., 1994. The in vitro formation of
placer gold by bacteria. Geochim. Cosmochim. Acta 58,
4527–4530.
Southam, G., Saunders, J.A., 2005. The geomicrobiology of ore
deposits. Econ. Geol. 100, 1067–1084.
Sperling, A.R., Gil, R.P., Zhang, F., Zanella, M., Parak, J.W., 2008.
Biological application of gold nanoparticles. Chem. Soc. Rev.
37, 1896–1908.
Tao, A.R., Habas, S., Yang, P., 2008. Shape control of colloidal
metal nanocrystals. Small 4, 310–325.
Theodore, L., Kunz, R.G., 2005. Nanotechnology: prime materials
and manufacturing methods. In: Theodore, L., Kunz, R.G.
(Eds.), Nanotechnology: Environmental Implications and
Solutions. John Wiley& Sons, Inc., Hoboken, NJ, pp. 70–79.
Thompson, T.D., 2007. Using gold nanoparticles for catalysis.
Nano Today 2, 40–43.
Tomalia, D.A., 2005. Birth of a new macromolecular architecture:
dendrimers as quantized building blocks for nanoscale
synthetic polymer chemistry. Prog. Polym. Sci. 30, 294–324.
Torresday, J.L.G., Parsons, J.G., Gomez, E., Peralta-Videa, J., Troian,
H.E., Santiago, I.P., Yacaman, M.J., 2002. Formation and growth
of Au nanoparticles inside live alfa alfa plants. Nanoletters 2
(4), 397–401.
Tripathi, A., Chandrasekaran, N., Raichur, A.M., Mukherjee, A.,
2009. Antibacterial applications of silver nanoparticles
synthesized by aqueous extract of Azadirachta indica (Neem)
leaves. J. Biomed. Nanotechnol. 5, 93–98.
Turkevitch, J., Stevenson, P.C., Hillier, J., 1951. Nucleation and
growth process in the synthesis of colloidal gold. Discuss.
Faraday Soc. 11, 55–75.
Vigneshwaran, N., Kathe, A.A., Varadarajan, P.V., Nachane, R.P.,
Balasubramanya, R.H., 2007. Silver-protein (core–shell)
nanoparticle production using spent mushroom substrate.
Langmuir 23 (13), 7113–7117.
Yang, H.G., Sun, C.H., Qiao, S.H., Zou, J., Liu, G., Smith, S.C.,
Cheng, H.M., Lu, G.Q., 2008. Anatase TiO2single crystals with
a large percentage of reactive facets. Nature 453, 638–641.
Yuan, Y., Fendler, J., Cabasso, I., 1992. Preparation and
characterization of stable aqueous higher-order fullerenes.
Chem. Mater. 4, 312.
Zeng, J., Zhang, Q., Chen, J., Xia, Y., 2010. A comparative study of
the catalytic properties of Au-based nanocages, nanoboxes,
and nanoparticles. Nano Lett. 10, 30–35.