by Charusheela Rameteke
The synthesis of nanomaterials has been the area of utmost importance in the
nanotechnology community due to their extraordinary properties and the
Due to an increasing demand in nanomaterials, researchers are taking a
massive interest in the synthesis of nanomaterials as well as assessing new
Although a number of approaches can be employed for the synthesis of
nanomaterials (i.e. chemical, physical or biological), there is an avoidable
need for the use of stabilizing agent during the synthesis process. Excellent
stability and homogeneity of nanomaterials can be achieved through the use of
Nucleic Acid is a macromolecule
composed of chains of monomeric nucleotides. In biochemistry these molecules
carry genetic information or form structures within cells. The most common
nucleic acids are deoxyribonucleic acid (DNA) and ribonucleic acid
Deoxyribonucleic acid (DNA) is a nucleic acid that
contains the genetic instructions used in the development and functioning of all
known living organisms and some viruses.
Ribonucleic acid (RNA) is a biologically important type
of molecule that consists of a long chain of nucleotide units. RNA is very
similar to DNA, but differs in a few important structural details.
There are several physicochemical procedures available for the stabilization
of nanomaterials that operate via one of the four principles: electrostatic,
steric, electrosteric stabilizations and stabilization by a ligand or a
solvent1. Although, the physicochemical procedures
for stabilization of nanomaterials are well established, they possess a major
disadvantage of being capital intensive and hazardous for the environment as
they involve the use of hazardous chemicals and practices.
The use of biological systems for the synthesis of nanomaterials has been
well reported since the ancient times2. The
nanomaterial synthesizing biocomponents have an added advantage of providing
excellent stability to the nanomaterial being synthesized. Despite the fact that
the machineries and the underlying mechanisms for the synthesis of nanomaterials
has been extensively studied by researchers, the stabilizing entities in the
nanomaterial synthesis protocols remain unexplored.
This article provides an introductory overview on the stabilization of
nanomaterial by biological entities. Depending on the site of stabilization,
they have been categorized into two groups: in vivo stabilizing entities
and in vitro stabilizing entities (Figure 1).
Figure 1: Biological
entities in stabilization of
In vitro Stabilization
When nanomaterials are synthesized in close proximity to any biosystems, i.e.
bacteria, fungi, plants or their extracts, the stabilization is classified as
in vitro stabilization. The stabilization is achieved through a single
unit of biomolecule that "caps" the synthesized nanomaterial. Biomolecules such
as proteins, peptides and a special class of metal binding molecules referred to
as phytochelatins are used for the in vitro stabilization of synthesized
In case of bacterial and fungal nanoparticles synthesis, stabilization has
been attributed to specific proteins either found intracellularly or released
into the medium extracellulary3,4.
After the synthesis of nanomaterials, these stabilizing proteins are supposed to
neutralize the charges over the metallic nanomaterial by providing appropriate
seal form a stable structure the nanomaterials.
When nanomaterials are synthesized in close proximity to bacteria and fungi,
stabilization is achieved through the use of a specific proteins which is found
either intracellularly or released into the medium extracellulary3,4. These stabilizing proteins neutralize
the charges over the metallic nanomaterial after nanomaterials has been
The interaction of protein with different metals has been well documented in
the literature and thorough study of critical stability constants of the
reactants supports the finding that proteins are responsible for stabilization
of nanomaterials. Metal-protein bonds provide the molecule more stability as
compared to its free form5. On comparing the
critical stability constants with reference to proteinecious structure with free
amino group, sulfahydryl and disulfide group, the entities with free amino
groups provide more stability to the metal species on nanoscale.
In case of yeast and algae, stabilization is brought about by a special class
of peptides called phytochelatins6,7. Phytochelatins are functional analogues of metallothiones
consisting of three amino acids namely glycine, cysteine and glutamine.
Stabilization is achieved through the binding of metal ions by thiolate
coordination to form metal complexes8.
Recently, it has been reported in photosynthetic algae, stabilization of
nanoparticle has been achieved by the use of siderophore, a special class of
molecules which are secreted by the algal cells in response to the presence of
iron in the extracellular medium9. In plant, the
stabilization mechanisms of nanomaterials are not inclusive as per literature
reports10. However, Shankar et al., (2004) have
suggested flavanone and terpenoid constituents of the leaf broth to be the
surface active molecules stabilizing the metallic nanoparticles11. On another note, as phytochelatins being mainly found
in plants can also be suggested to be contributing in nanomatrerials'
All these entities discussed under the category of in vitro
stabilization can be extracted from the biological systems and also be used
in vivo for nanomaterials stabilization12.
In vivo Stabilization
This category includes those biomolecules which can be exploited for
nanomaterials stabilization but only outside the living system. These
biomolecule entities provide a support template for nanomaterials stabilization.
Therefore, it may also be regarded as template assisted stabilization.
Viruses as Template
Viruses; to be more specific, viral scaffolds are used as a template in the
nucleation and assembly of inorganic materials.Viral scaffolds have a very
organized nano-structure and the materials encaged within the structure are also
on the nanoscale. For example, cowpea chorotic mottle virus (CCMV) has a protein
shell and RNA in its cavity. This RNA in the core can be removed and the cavity
created can be used to grow nanocrystals. The charge formed inside this cavity
after removal of RNA is neutralized leading to stabilization of the
Adopting this approach, it has been reported that CCMV has been used for the
formation of polytungstate nanoparticles13.
Likewise, cowpea mosaic virus has been used as nucleation cages for the
mineralization of inorganic materials and the tobacco mosaic virus has also been
used for the synthesis and stabilization of PbS and CdS14. In this case, charge replacement has been followed by
consequent charge neutralization over encavitated metal ion.
Another example of template assisted biomolecules that has drawn much
attention is cell surface protein layer or S-layers, found in many bacteria and
archaebacteria. These are organic templates that allow the synthesis of
inorganic nanocrystalline superlattices with a broad range of particle sizes
(3-15 nm in diameter), interparticle spacings (up to 30 nm) and lattice
symmetries (oblique, square or hexagonal)15.
The S-layer protein has an extraordinary property of reassemble into a
hierarchical structure forming a layer when put onto any substrate and thus acts
as a suitable template for nanoparticles formation. This property has been well
demonstrated in acidophilic bacterium Sulfolobus acidocaldarius and
The S-layer proteins from the respective bacteria were deposited on the solid
support and were then exposed to a metal-salt solution. This results in the
formation of regularly arranged nanoparticles with unusual properties, which
differed drastically from that of the bulk material. Further, it may be expected
that, in near future, modification of S-layer proteins by recombinant DNA
technologies will significantly influence the development of the applied S-layer
research15,17. In context of the
above discussion, S-layers seem to provide a best template for nanomaterials
As mentioned earlier, a protein from mammalian origin, ferritin has also been
used as a template for nanomaterial formation. It has been previously used for
the synthesis as well as stabilization of iron phosphate, iron arsenate, iron
vanadate, iron molybdate, cadmium sulfide, and zinc selenite
nanoparticles18-20. Stabilization of
nanopartiles inside ferritin cage is also presumed to take place due to charge
Theoretical computations have shown that the potential of the outer surface
of ferritin is net positive while the inner surface has a negative net charge.
The inner and outer surfaces are connected by channels, which provide a path for
cations to come into the cavity. Cations in the cavity can then bind to the
inner surface. The binding sites become nuclei for crystallization, but
crystallization ends when the cavity is filled with crystals21.
Apart from all the above listed stabilizing entities, there is another class
of macromolecules that can act as templates for the stabilization of
naoparticles. Single stranded DNA and DNA duplex as well as RNAs with chemically
modified bases can be used for binding of functionalized nanoparticles.
The advantage of this method is that nanoparticles can potentially be placed
wherever a modified base is inserted during the synthesis of DNA. In double
stranded DNA, the bases are separated by about 0.34 nm, allowing the placement
of particles with sub nanometer precision. Thus the arrangement of particles is
dependent only on the design of the DNA template and the size of the
Engineered DNA strands can act as an excellent template since the
intermolecular distance of the nanoparticles can be varied at will. Besides, the
two DNA strand synthesized can be joined to each other to get a longer DNA
strand. Various researchers have succeeded to stabilize the nanoparticles on DNA
subsurfaces in this manner23-26.
From the discussion above on in vitro and in vivo
by bio-entities stabilization of nanomaterials, it can be concluded that the
rationality of stabilization lies in overall charge of the stabilizing molecules
and its potential in effective charge neutralization. Therefore it is necessary
to understand the structure of the constituents of microorganism involved in
capping of the metals and its spatial arrangement around the metal.
Investigations into the nature of linkages established between the metal and
stabilizing entity will also serve a lot in understanding the mechanism of
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Copyright AZoNano.com, Charusheela Rameteke (National
Environmental Engineering Research Institute)