There is 1 related live offer.

Save 25% on magneTherm

Biological Entities in Stabilization of Nanomaterials

by Charusheela Rameteke

Charusheela Rameteke, Tapan Chakrabarti and Ram Avatar Pandey, Environmental Biotechnology Division, National Environmental Engineering Research Institute, India
Corresponding author:,

The synthesis of nanomaterials has been the area of utmost importance in the nanotechnology community due to their extraordinary properties and the underlying applications.

Due to an increasing demand in nanomaterials, researchers are taking a massive interest in the synthesis of nanomaterials as well as assessing new synthesis techniques.

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 stabilizing agents.

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 (RNA)

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 nanomaterials

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 nanomaterials.

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 synthesized.

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' stabilization.

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 material.

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 Sposarcina ureae16.

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 stabilization.


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 distribution only.

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.

Nucleic Acids

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 particle22.

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 stabilization.


1. Roucoux A, Schulz J and Patin H (2002) Reduced transition metal colloids: A novel family of reusable catalysts? Chem. Rev. 102:3757-3778.
2. Lowenstam HA (1981) Minerals formed by organisms. Science 211:1126-1131.
3. Ahmad A, Mukherjee P, Mandal D, Senapati S, Khan MI, Kumar R and Sastry M (2002) Enzyme mediated extracellular synthesis of CdS nanoparticles by the fungus, Fusarium oxysporum. J Am Chem Soc 124:12108-12109.
4. Mukherjee P, Ahmad A, Mandal D, Senapati S, Sainkar SR, Khan MI, Ramani R, Parischa R, Ajayakumar PV, Alam M, Sastry M and Kumar R (2001b) Bioreduction of AuCl4- ions by the fungus, Verticillium sp. and surface trapping of the gold nanoparticles formed. Angew Chem Int Ed 40:3585-3588.
5. Martell AE and Smith RM (1974) Critical stability constants Vol I, Plenum Press, NEW York & London, PP-469.
6. Gekeler W, Grill E, Winnacker E and Zenk MH (1988) Algae sequester heavy metals via synthesis of phytochelatin complexes. Archivves of microbiology 159:197-202.
7. Dameron CT, Reese RN, Mehra RK, Kortan AR, Carroll PJ, Steigerwald ML, Brus LE and Winge DR (1989) Biosynthesis of cadmium sulfide quantum semiconductor crystallites. Nature 338:596-597.
8. Cobbett CS (2000) Phytochelatins and their oles in heavy metal detoxification. Plant Physiol, 123:825-832.
9. Brayner R, Yepremian C, Djediat C, Coradin T, Herbst F, Livage J, Fievet F and Coute A (2009) Photosynthetic Microorganism-Mediated Synthesis of Akaganeite (ß-FeOOH) Nanorods.Langmuir, 25(17):10062-10067.
10. Kumar V and Yadav SK (2009) Plant-mediated synthesis of silver and gold nanoparticles and their applications. J Chem Technol Biotechnol 84:151-157.
11. Shankar SS, Rai A, Ahmad A and Sastry M (2004) Rapid synthesis of Au, Ag, and biometallic Au core-Ag shell nanoparticles using Neem (Azadirachta indica) leaf broth. J. colloid and interface. Science 275:496-502.
12. Kumar SA, Abyaneh MK, Gosavi SW, Kulkarni SK, Pasricha R, Ahmad A and Khan MI (2007) Nitrate reductase-mediated synthesis of silver nanoparticles from AgNO3. Biotechnol Lett, 29:439-445.
13. Douglas T and Young M (1998) Host-guest encapsulation of materials by assembled virus protein cages. Nature 393:152-155.
14. Xu W (2005) Biotemplates for Nanoparticle Synthesis Literature Seminar (available online at
15. Sara M and Sleytr UB (2000) Minireview S-layer proteins. J Bacteriol.182:859-868.
16. Schuster B, Gufler PC, Pum D and Sleytr UB (2004) S-layer proteins as supporting scaffolding for functional lipid membranes. IEEE Trans Nanobioscience 3(1):16-21
17. Pum D, Sleytr UB (1999) The application of bacterial S-layers in molecular nanotechnology. Trends Biotechnol 17:8-12.
18. Iwahori K, Yoshizawa K, Muraoka M and Yamashita I (2005) Fabrication of ZnSe nanoparticles in the apoferritin cavity by designing a slow chemical reaction system. Inorg. Chem 44:6393-6400.
19. Iwahori K and Yamashita I (2008) Size-controlled one pot synthesis of fluorescent cadmium sulfide semiconductor nanoparticles in an apoferitin cavity. Nanotechnolgy 19:1-7.
20. Polanams J, Ray A D and Watt R K (2005) Nanophase iron phosphate, iron arsenate, iron vanadate and iron molybdate minerals synthesized within the protein cage of ferritin. Inorg.Chem 44:3203-3209.
21. Douglas T and Ripoll DR (1998) Calculated electrostatic gradients in recombinant human H-chain ferritin. Protein Science 7:1083-1091.
22. Wells JC, Stevenson KA, Muralidharan G, Thundat TG, Maya L and Barhen1 J. (2001) Programmed Assembly of Quantum-Dot Arrays on DNA Templates: Hardware for Quantum Computing? (available online at
23. Wong KKW and Mann S (1996) Biomimetic synthesis of cadmium sulfide-ferritin nanocomposites. Adv. Mater. 8:928-932.
24. Yang J, Lee JY, Too H, Chow G and Gan LM (2006) Single stranded DNA stabilization and assembly of Au nanoparticles of different sizes. Chemical Physics 323:304-312.
25. Mitchell GP, Mirkin CA and Letsinger RL (1999) Programmed assembly of DNA functionalized quantum dots. J. Am. Chem. Soc 121:8122-8123.
26. Han G, Martin CT and Rotello VM (2006) Stability of gold nanoparticle-bound DNA toward biological, physical, and chemical agents. Chem Biol Drug Des 67:78-82.

Copyright, Charusheela Rameteke (National Environmental Engineering Research Institute)

Date Added: Mar 23, 2010 | Updated: Jun 11, 2013
Tell Us What You Think

Do you have a review, update or anything you would like to add to this article?

Leave your feedback