FORM-2
THE PATENTS ACT, 1970
(39 of 1970)
&
THE PATENTS RULES, 2003
COMPLETE
Specification
(See section 10 and rule 13)
A PROCESS OF MAKING SILVER NANO SCALE PARTICLES
AGHARKAR RESEARCH INSTITUTE OF MAHARASHTRA ASSOCIATION FOR THE CULTIVATION OF SCIENCE
a Society registered under the Indian Societies Act of G. G. Agarkar Road, Pune 411 004. Maharashtra, India,
THE FOLLOWING SPECIFICATION PARTICULARLY DESCRIBES
THE INVENTION AND THE MANNER IN WHICH IT IS TO BE PERFORMED:-

This invention relates to a process of making Silver nano scale particles.
Field of the invention.
This invention relates to nano particles and particularly, it relates to a method of making submicronic Silver particles.
Background of the invention
Nano particles are part of an emerging science called 'nano technology*. The word nano technology comes from the Greek prefix 'nano' meaning "one billionth". In modern scientific parlance, a nanometer is one billionth of a meter, about the length of ten hydrogen atoms placed side by side in a line. The smallest things that an unaided human eye can see are 10,000 nanometers across. Nano particles are typically and generally spherical in shape.
Nanoscience , simply, is the study of the fundamental principles of structures with at least one dimension roughly between 1 and 100 nanometers and Nanotechnology is the application of these nanostructures into useful nano scale devices.
Nano scale particles of substances exhibit properties unlike the properties of their macro counterparts often with stunning new results. Nano scale is unique because it is the size scale where the familiar day-to-day properties of materials like conductivity, hardness or melting point meet the more exotic properties of the atomic and molecular world such as wave-particle duality and quantum effects. At the nano scale, the most fundamental properties of the materials and machines depend on their size in a way they don't at any other scale. For e.g. a nano scale wire or circuit component does not necessarily obey Ohm's law. Nano-scale particles have unique physical properties (e.g. optical, dielectric, magnetic, mechanical), transport properties (e.g., thermal, atomic diffusion) and processing characteristics (e.g., faster sintering kinetics, super-plastic forming).
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Physicist Richard Feynman first described the possibility of molecular engineering. In 1959 Feynman gave a lecture at the California Institute of Technology called "There's Plenty of Room at the Bottom" where he observed that the principles of physics do not deny the possibility of manipulating things atom by atom. He suggested using small machines to make even tinier machines, and so on down to the atomic level itself. Nano technology as it is understood now though, is the brainchild of Feynman's one-time student K. Eric Drexler. Drexler presented his key ideas in a paper on molecular engineering published in 1981, and expanded these in his books Engines of Creation and Nano systems: Molecular Machinery, Manufacturing and Computation, which describes the principles and mechanisms of molecular nano technology.
In 1981 the invention of the Scanning Tunneling Microscope or STM, by Gerd Binnig and Heinrich Rohrer at IBM's Zurich Research Labs, and the Atomic Force Microscope (AFM) five years later, made it possible to not only take photos of individual atoms, but to actual move a single atom around. Soon after, John Foster of IBM Almaden labs was able to spell "IBM" out of 35 xenon atoms on a nickel surface, using a scanning tunneling microscope to push the atoms into place.
A nanometer is a magical point on the dimensional scale. Nano structures are at the confluence of the smallest of Human-made devices and the largest molecules of the living things. Nano technology exploits the new physical, chemical and biological properties of systems that are intermediate in size, between isolated atoms/molecules and bulk materials, where the transitional properties between the two limits can be controlled.
The synthesis and characterization of nano particles has received attention in recent years because of the possibility of their widespread use in industry and chemistry. Nanotechnology is gaining importance in areas such as biomedical sciences, optics, electronics, magnetics, mechanics, ceramics, catalysis and energy science. However, the preparation of such nano structured materials poses several unique challenges. A range of nano particles has been produced by physical, chemical and biological methods.
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Two approaches have been adopted for nano fabrication - The Top down processes,
which include the methods of synthesis that carve out or add aggregates of molecules to a
surface. The second is the bottom up approach, which assembles atoms or molecules into nano structures.
PHYSICAL methods include Electron beam lithography, Scanning probe method, Soft lithography, Microcontact printing, Micromoulding.
In Electron Beam Lithography, an electron beam scans the surface of a semiconductor containing a buried layer of quantum well material. The resist gets removed where the beam has drawn a pattern.
Soft lithography
is an extension of the previous technique and overcomes the impracticability of applying electron beam lithography to large scale manufacturing by making a mould or a stamp, which can be used repeatedly to produce nanosructures.
In Micro contact printing, the PDMS stamp is inked with a solution consisting of organic molecules called thiols and then pressed against a thin film of Silver on a silicon plate. The thiols form a self-assembled monolayer on the Silver surface that reproduces the stamp pattern; features in the pattern can be as small as 50 nm.
In Micromoulding, the PDMS stamp is placed on a hard surface, and a liquid polymer flows into the recesses between the surface and the stamp. The polymer solidifies into the desired pattern, which may contain features smaller than 10 nm.
Scanning probe microscope can image the surface of conducting materials with atomic scale detail. Hence single atoms can be placed at selected positions and structures can be built to a particular pattern atom by atom. It can also be used to make scratches on a surface and if the current flowing from the tip of the STM is increased the microscope becomes a very small source for an electron beam which can be used to write nanometer
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scale patterns. The STM tip can also push individual atoms around on a surface to build rings and wires that are only one atom wide.
In Sonochemical method, an acoustic cavitation process is used to generate a transient localized hot zone with extremely high temperature gradient and pressure (Suslick et al. 1996). Such sudden changes in temperature and pressure bring about the destruction of the sonochemical precursor (e.g., organometallic solution) and the formation of nanoparticles.
Hydrodynamic cavitation consists of synthesis of Nanoparticles by creation and release of gas bubbles inside sol-gel solutions (Sunstrom et al. 1996).
High energy ball milling is already a commercial technology, but has been considered dirty because of contamination problems from ball-milling processes. However, the availability of tungsten carbide components and the use of inert atmosphere and/or high vacuum processes have reduced impurities to acceptable levels for many industrial applications. Common drawbacks include the low surface area, the highly polydisperse size distributions, and the partially amorphous state of the as prepared powders.
CHEMICAL methods include Wet chemical preparation, Surface passivation, Core shell synthesis, Organometallic precursor, Sol get method,. Langmuir- lodgett method, Precipitation in structured media, Zeolites, Micelles and inverse micelles formation.
A number of chemical strategies are now available for the construction of higher order structures. Organic molecules can be linked together by molecular recognition. For example, synergistic noncovalent donor acceptor interactions can give rise to intertwined rings (catenanes) . Liquid crystal polymers having self-organized structures can be formed from organic molecules containing head groups capable of complementary hydrogen bonding interactions. Organic molecules can be assembled around metal ions such as Cu (1) that provide stereo chemical constricts in the construction of double helices. The synthesis of inorganic clusters, by contrast, is usually dependent on
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passivating the surface of a growing aggregate by capping the surface sites with stabilizing ligands.
Wet Chemical Preparation method involves the reaction between a metal ion and the desired anion under controlled conditions to generate nanocrystals of desired size.
BIOLOGICAL methods include Biomineralization using Bacteria, Yeast, Fungi, Plants and Biotemplating using Ferritin, Lumazine synthase, Virus Surface layers DNA etc. A few attempts have been made to synthesize sulfides, typically cadmium sulfide (CdS) using microorganisms. It was shown that CdS nanoparticles can be synthesized in the yeasts Candida glabrata and Schizosaccharomyces pombe . These nanoparticles are coated with short peptides known as phytochelatins . which have the general structure (y-Glu-Cys)n-Gly where n varies from 2-6. The nanoparticles are size reproducible, more monodisperse, and have greater stability than synthetically produced nanoparticles . Further work on microbial synthesis of CdS nanoparticles is scant and is limited to studies on characterization and efficient production in batch cultivation .
U.S. Pat. No. 5,876,480 & 6,054,495 describe a process for creating unagglomerated metal na«t»-particles, comprising the steps of
(a) forming a dispersion in an aqueous or polar solvent, the dispersion including unpolymerized lipid vesicles, the unpolymerized lipid vesicles each comprising at least one lipid bilayer. the lipid bilayer including a negatively charged lipid that has an anionic binding group, and the lipid vesicles having catalytic first metal ions bound thereto by ionic bonding,
(b) combining the dispersion of step (a) with a metallization bath containing free second metal ions to form a mixture, and
(c) incubating the mixture of step (b) at a temperature sufficient to reduce said free second metal ions and to form unagglomerated metal na«o-particles having an average diameter between about 1-100 nm.
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U.S. Pat. No. 6,068.800 describes a process and apparatus for producing nano-scale particles using the interaction between a laser beam and a liquid precursor solution using
either a solid substrate or a plasma during the laser-liquid interaction.
US Patent no 5,618,475 and 5.665,277 relate to production of particulates having nanoparticle dimensions, such as about 100 nanometers diameter or less, and, more particularly, to apparatus and method for producing nanoparticles of metals, alloys, intermetallics, ceramics, and other materials by quench condensation of a high temperature vapor generated by an evaporator having features effective to isolate evaporation conditions from downstream conditions and to concurrently evaporate materials of dissimilar vapor pressures.
US Patent no 5,736,073 describes a process of production of nanometer particles by directed vapor deposition of electron beam evaporant onto a substrate.
US Patent no 6,706,902 The continuous process according to the invention includes impregnating support materials and, after thermal activation, drying the support materials by spraying or by fluidized bed technology leads to form precious metal-containing support compositions that are active in the catalysis of oxidation reactions.
US Patent 6,562,403 is broadly concerned with chemical methods of forming ligated nanoparticle colloidal dispersions and recovered ligated nanoparticles which may be in superlattice form.
US Patent no 5,698.483 A process for producing nano size powders comprising the steps of mixing an aqueous continuous phase comprising at least one metal cation salt with a hydrophilic organic polymeric disperse phase, forming a metal cation salt/polymer gel, and heat treating the gel at a temperature sufficient to drive off water and organics within the gel, leaving as a residue a nanometer particle-size powder.
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The main disadvantages of these methods are that they are expensive and technically difficult and too slow for mass production. Most of the techniques are also capital intensive as well as inefficient in materials and energy use. The known methods are difficult to control in order to acquire a desired size and shape of the nano-scale particles to be produced. Many of these synthesis techniques also require the use of a vacuum unit and involve environmental concerns about chemical waste disposal.
Physical and chemical methods in the manufacture of nano particles involve controlling crystallite size by restraining the reaction environment. However, problems occur with general instability of the product and in achieving monodisperse size. The dispersion of nano particles usually display very intense color due to plasmon resonance absorption, which can be attributed to the collective oscillation of conduction electrons, induced by the presence of an electromagnetic field.
Other problem areas in the above mentioned methods are uniform distribution of particles, morphology and crystallinity, particle agglomeration during and after synthesis and separation of these particles from the reactant.
Nano particles are extremely reactive as the coordination of surface atoms in nano particle is incomplete, and can lead to particle agglomeration to minimize total surface or interfacial energy of the system. This problem is overcome by passivating the bare surface atoms with protecting groups. Capping or passivating the particle not only prevents agglomeration, it also protects the particles from its surrounding environment, and provides electronic stabilization to the surface. The capping agent usually takes the form of a Lewis base compound covalently bound to surface metal atoms.
Chemical techniques have therefore been developed to passivate or stabilize these nano particles. It is desirable that nano particles are protected from the environment but are ;till allowed to maintain their intrinsic properties. It has been shown that the size, norphology, stability and properties (chemical and physical) of these nanoparticles have
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strong dependence on the specificity of the preparation method and experimental conditions.
The stabilizing of nano particles in a sub micronic regime requires an agent that can bind to the cluster surface and thereby uncontrolled growth or agglomeration of the cluster or discrete particles into larger particles is prevented. The simplest method involves the use of a solvent that acts as a stabilizer of the small clusters. Unagglomerated nano particles can also be made by the use of polymeric surfactants ands stabilizers added to a reaction designed to precipitate a bulk material. The polymer attaches to the surface of the growing clusters and either by steric or electrostatic repulsion prevents further growth of the nano clusters. Commonly used chemical stabilizers include sodium polyphosphate and anionic agents such as thiolates.
Most capping reactions involve additional steps and the capping agents are generally toxic substances.
SUMMARY OF THE INVENTION
Silver nano particles have tremendous industrial application as catalysts, coating materials in electronics, in optics and especially in medical therapeutics.
It is an object of this invention to manufacture Silver submicronic scale particles, particularly nano-scale particles using an inexpensive 'green' environmental friendly process in which the Silver particles are stable and do not agglomerate and are biocompatible.
According to this invention there is provided a process for making Silver sub micron scale particles comprising the steps of:
(a) dissolving a salt of the Silver in water having conductivity less than 3 micro
Siemens to obtain a solution in which the concentration of Silver ions is in the
range of 20, 000 to 50, 000 ppm ,
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(b) preparing a fresh filtered aqueous solution of biological tissue extract;
(c) diluting the aqueous solution with deionized water in the ratio ranging from 1:5 to 1: 50 to form a solution having an open circuit potential between +0.02 and +0.2 volt, total organic carbon content of at least 7500 ppm and a pH between 5.5 to 7.5;
(d) maintaining the said aqueous solution under continuous agitation at a temperature between 20 and 30 degrees Celsius;
(e) inoculating a minute quantity of the Silver salt solution in the said aqueous extract solution under continuous agitation such that the final concentration of the metal ion in the reaction mixture is in the range of 50 to 300 ppm;
(f) continuing the agitation for a period of 30 minutes to 3 hours under well illuminated conditions to obtain a colloidal suspension of Silver nano particles;
(g) separating the nano particles from colloidal suspension by a known process such as centrifugation.
Typically, the biological tissue extract is an extract of macerated plant cells selected from a group of tissues comprising living tissue of leaves, fruits, stems, roots and flowers and parts thereof.
Alternatively, the biological tissue extract is an extract of macerated animal cells selected from a group of tissues comprising living tissue of worms, insects, fishes, mollusks, crustaceans, and higher animals
Still alternatively, the biological tissue extract is an extract of macerated microbial cells selected from a group of tissues comprising living tissue of bacteria, fungi, yeasts, viruses, protozoa and algae.
The theoretical considerations underlying the invention are as follows:
Metal nano clusters are optically transparent and acts as dipoles. Conduction and valence bands of metal nano clusters lie closely and electron movement occurs quite freely. The
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potential applications of these systems are mainly associated with unusual dependence of the optical and electronic properties on the particle size. Silver particles having 5-80 nm
sizes show a sharp absorption band at 410-430 nm.
In nature, the chemical composition of cells across flora and fauna are remarkably similar. Thus living plants cells are very similar chemically to animal cells and microbial cells.
All cells contain biomolecules such as polysaccharides, proteins, lipids and nucleic acids which are made up of building blocks such as monosaccharides, amino acids, fatty acids, nucleotides. In addition many dynamically acting biomolecules such as glutathione, cytochromes, ubiquinone, NADH, FADH, pyruvic acid, citric acid, maleic acids, glycerol are also present. These biomolecules have various reactive groups such as sulfhydryl, amino , imino, carboxyl, hydroxy!, and the like.
When living cells are macerated in water the biomolecules are released in the water. These biomolecules collectively contain all the reactive groups where the elements are in specific proportion to each other. It has been found that these biomolecules have the surprising collective ability to not only reduce metal ions such as Silver but also to act sterically on metal nano particles formed upon reduction so as to stabilize them. The binding interaction between biomolecules is relatively weak as compared with the interaction between these particles and typical chemical capping agents such thiols.
The synthesis of a particle, requires a two-step process, i.e., nucleation followed by successive growth of particles. In accordance with the present invention where the solution of biological tissue extract is added during the synthesis in the first step, part of metal ions in solution gets adsorbed on free nucleophilic groups (-SH, OH, NH2) present on the surface of biomolecules in the aqueous solution and get reduced. The reduced metal atoms thus created act as nucleation centers and facilitate further reduction of metal ions present in the bulk solution. The atomic coalescence leads to the formation of metal
clusters and can be controlled by the natural ligands and surfactants forming part of
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biomolecular mass. Thus biomolecules act as both reducing as well as stabilizing agent. It has been found that a threshold concentration value of biomolecular mass is required to
seed the process of nano particle formation and stabilization. This is expressed in terms of the total organic carbon content of the biomolecular mass.
It has also been found by experimentation that the reduction potential plays an important and crucial part in the formation and stabilization of nano particles, particularly, Silver nano particles. The electromotive force exhibited by 1 M concentration of a reducing agent and its oxidized form at 25° C and pH 7.0 is called its standard reduction potential. It is a measure of the relative tendency of the reducing agent to lose electrons.
The reduction potential is measured in positive or negative volts on a scale in which the positive sign denotes a lower reduction potential than the negative sign. Therefore a substance with a standard reduction potential of + 0.1 volt has a higher reduction potential than a substance with a standard reduction potential of+0.2 volts and therefore substance with a standard reduction potential of+0.1 volts will reduce the substance having a reduction potential of+0.2 volts. The standard reduction potentials of some of the biomolecules, such as NADH, ubiquinone, cytochrome bk are -0.32Volt, -0.05 Volt and + 0.03 Volt. On the other hand, the standard reduction potentials for conversion of ions such as Silver to their solid state is +1.5 Volt. Thus the findings in accordance with this invention is that many biomolecules which have an effective standard reduction potential higher than that required for the conversion Silver ions to solid state particles , can effectively reduce these ions in solution. It is not possible to measure the standard reduction potential of the solutions formed in this invention because the molar concentrations of individual components of the biomolecular mass is unknown. However, dynamically the open circuit potential of the solution can be determined easily. At specific molar concentrations the open circuit potential has a direct relationship with the standard reduction potential. The open circuit potential indicates the initial empirical redox state of the solution.
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For the process of our invention, it is critical that the aqueous biological tissue extract solution has an open circuit potential +0.02 to + 0. 2 Volt, a pH between 5.5 and 7.5 and
temperatures between 20 to 30 degrees Celsius are also important parameters. Also the total organic carbon content of the solution must be greater than 7,500 ppm. Purity of water having conductivity less than 3 micro Siemens is also of significant importance in optimizing the process of particle formation. Concentration of metal ions in the reacting solution should lie between 50 to 300 ppm; metal ions in the mother solution being 20,000 to 50, 000 ppm.
Silver is known to undergo photocatalytic reduction in the presence of organic compounds. Hence, the presence of strong light assists in the formation of submicronic scale particles in accordance with this invention .
It has been found in the course of experimentation that bio molecules normally found in plant cells, animal and microbial cells in the intact plant, animal or microbe have a reducing potential and are their reducing ability very slowly decreases when exposed to air under ambient conditions and rapidly to denaturing treatment. The reducing ability is therefore inversely related to the freshness of the tissue. It has also been found that the reducing ability varies from tissue to tissue and is inversely proportional to the duration of exposure to air eventually tending to zero.
The invention will now be described with reference to the accompanying examples which in no manner limit the ambit and scope of this invention.
Example 1
Aloe-Vera extract:
To start with salt of Silver nitrate was used . A stock solution of 100 mM was prepared in deionized water of conductivity 2.7 microSiemens.
Fresh leafs of aloe vera (20-25cm) were peeled and extract was collected by crushing. Further it was subjected to centrifugation at 4000 rpm for 10 min to remove big pieces.
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The clear gel was collected. The aloe vera extract was diluted as 1:5, . The open circuit potential was measured as follows:
An aliquot of 7 ml were removed and checked for open circuit potential at 25 C on
Electrochemical analyzer (CH Instruments 600B, USA) using a three-electrode system. Ag/AgCI (aq) was used as reference electrode. Glassy carbon as working electrode (diameter 3 mm) and Pt wire (length 4 cm) as counter electrode. The value found was + 0.15 Volt. Similarly pH of free flowing solution was checked using Digital pH meter (control Dynamics, India) and it was found to be 5.6.
The concentration of total organic carbon was measured using Beckman TOC analyzer and was found to be 22,180 ppm.
The extract solution was kept continuous stirred and was challenged with one drop of Silver salt solution to get final concentration of 1 mM.
The color of the solution started to change from colour less to brownish black after three incubation under bright light conditions. Formation of nano particles was monitored by measurement of UV-visible spectra in Diode array spectrophotometer (Ocean optics ODI base 32). When the UV-Visible spectrum was taken characteristic plasmon peak of Silver nanoparticles at 420 nm was observed (Fig 1 of the accompanying drawings).
Silver submicronic particles in the size range of 5 to 30 nm were formed . Transmission electron microscopy (TEM) study of particles was carried out at 200 kV using Philips electron microscope equipped with field emission gun, i.e., CM200 FEG. TEM specimen was prepared by pipetting 2 uL of colloid solution onto a carbon coated copper grid. Atomic force microscopy (AFM) imaging was performed on a Nanonics MultiView 1000 AFM head with E scanner (Nanonics Imaging Ltd., Jerusalem, Israel). Scanning was performed in non-contact mode. Images were obtained with 20 nm radius with resonance frequency of 80 kHz. AFM tips obtained from Nanonics Imaging. AFM images were captured, processed and analyzed with QUARTZ software, Version 1.00 (Cavendish
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Instruments Ltd., UK). For specimen. 5 ul of sample was placed on a 1-cm2 glass slide (thickness 0.5 mm) and dried in laminar airflow before imaging.
TEM image of Silver nano particles also confirms the formation particles in the range of 5-30 nm (Fig 2 of the accompanying drawings). AFM images of nano particles confirm their formation. (Fig 3 of the accompanying drawings).
Example 2
Azadirachla indica plant extract preparation:
To start with salt of Silver nitrate was used . A stock solution of 100 mM was prepared in
deionized water of conductivity 2.7 microSiemens.
Neem fruits (ripe ones) were collected . To get extract, fruits (145 gm) were washed with deionized water and fruit pulp was extracted by separating outer covering from seeds. Seeds with mucilaginous substance (92gm) were kept in 150 ml of deionized water and mashed to get water-soluble extracts. The extract was filtered through muslin cloth and diluted 50 fold with deionized water.
An aliquot of 7 ml were removed and checked for open circuit potential as in example 1. The value found was + 0.09 Volt. Similarly pH of free flowing solution was checked using Digital pH meter (control Dynamics, India) and it was found to be 6.7.
The concentration of total organic carbon was measured using Beckman TOC analyzer and was found to be 8,180 ppm.
The process of example 1 was repeated. Stable Silver nano particles were formed that showed stability over a period of sixty days.
Example 3
To start with salt of Silver nitrate was used . A stock solution of 100 mM was prepared in
deionized water of conductivity 2.7 microSiemens.
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Argyeria speciosa leaf extract preparation: The leaf extract was prepared by grinding 49gm of leaves (7 in no), in distilled water; the final volume was made up to 150 ml. This extract was filtered through muslin cloth and further centrifuged at 5000 rpm for 5 min to remove suspended materials before use. The solution was diluted 35 fold with deionized water.
The open circuit potential and pH were measured as in example 1 and were found to be 0.11 volts and pH 6.3 respectively. The concentration of total organic carbon was measured using Beckman TOC analyzer and was found to be 12.180 ppm. The extract was used for the reduction of Silver salts as described in example 1. Silver nano particles formed showed stability over a period of sixty days.
Example 4
Bitter gourd extract:
Bitter gourd (20gm) was soaked overnight in water, ground in a mixer, and diluted with 150 ml of deionized water; The suspension extract was filtered through muslin cloth and further centrifuged at 5000 rpm for 5 min to remove suspended materials before use. The solution was diluted 25 fold with deionized water.
The open circuit potential and pH were measured as in example 1 and were found to be 0.1 volt and pH 6.1 respectively. The concentration of total organic carbon was measured using Beckman TOC analyzer and was found to be 14,180 ppm.
The extract was used for the reduction of Silver salts as described in example 1. Silver nano particles formed showed stability over a period of sixty days.
Example 5
Using Labconco, USA water pro system with pre-filter, carbon filter and reverse osmosis membrane water was collected. The said water had the conductivity of 2.7 micro Siemens
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as measured by the online digital meter fitted in the instrument. Silver nitrate solution was prepared as per example 1.
Simultaneously whole leaves and buds of ocimum sanctum (55.37 gm wet wt) were washed, peeled and macerated with 150 ml of deionized water in a blender (500 rpm) for 10 minutes to get a homogenous viscous suspension. This viscous suspension was filtered through Whatman No 1 filter paper under vacuum to obtain a clear 165 ml of viscous solution. From this stock an aliquot of 10 ml was diluted upto 250 ml by deionized water and mixed thoroughly by shaking to get a free flowing solution.
The open circuit potential and pH were measured as in example 1 and were found to be 0.09 volt and pH 6.0 respectively. The concentration of total organic carbon was measured using Beckman TOC analyzer and was found to be 13,180 ppm.
25 ml of free flowing solution was taken in 150 ml Erlenmeyer flask. 25 ml of free flowing solution was taken in 150 ml Erlenmeyer flask, which was kept on rotary shaker set at 25 C and 120 rpm. 50 ul of silver nitrate solution (34,875) was introduced into flask under agitation using a micropipette. Care was taken to ensure that drops came into direct contact with solutions and not onto sides of the flask. Immediately the color of solution changed to purple within 2 minutes of agitation. The metal concentration was checked by Atomic Absorption spectrometry (Perkin Elmer, USA Model 2380) using Silver hollow cathode lamp (242.8 nm) and it was found to be 47 ppm. Agitation of Erlenmeyer flask was continued for 5 minutes, at the end of 5 minutes colloidal suspension was scanned from 200-800 nm using Diode Array spectrophotometer (Ocean Optics, USA). A peak at 410 nm was detected . This peak was characteristic plasmon peak for Silver nano particles, typically having average diameter of 15-80 nm.
Example 6
The water having conductivity of 2.7 microSiemens was used. The stock solution of
silver nitrate was prepared as mentioned in example 1 and the concentration was found to
be 34,875 ppm.
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Simultaneously whole flowers of Calotropis procera (Common name Arkapatri; 47.00 gm wet wt) were macerated with 150 ml of deionized water as described in example 1 and filtered through Whatman No 1 filter paper under vacuum to obtain a clear 155 ml of solution. From this stock an aliquot of 10 ml was diluted upto 100 ml by deionized water and mixed thoroughly by shaking. The open circuit potential and pH was measured as described in example 1 and found to be + 0.09 Volts and 6.0 respectively. Total organic carbon was 18,230 ppm.
25 ml of solution was challenged with 50 ul of silver nitrate stock solution (34,875) under similar conditions as described in example 1.
After 3 h of incubation in well illuminated conditions different aliquot of samples were removed and checked for various parameters as described in example 1. Characteristic plasmon peak of nano Silver at 410 nm was observed indicating an average diameter in the range of 15 -80 nm. The stability of nano particles was checked in detail as described in example 1 and it was observed that the Silver nano particles are stable under normal and moderately harsh environmental conditions.
Example 7
The water having conductivity of 2.7 microSiemens was used. The stock solution of silver nitrate was prepared as mentioned in example 1 and the concentration was found to be 34,875 ppm
In yet another sets of examples fresh earth worm extracts, e coli extracts, fish extracts,
goat meat extracts, yeast, algae extracts were separately
macerated/homogenized/sonicated with 150 ml of deionized water in each case as described in example 1. The respective suspensions were filtered through Whatman No 1 filter paper under vacuum or centrifuged at 6000 G to obtain clear solutions. The open circuit potentials, pHs, and total organic contents were measured . and the solutions were diluted in the range of 1:5 to 1: 50.
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25 ml of each of the solutions were challenged with 50 to 150 ul of silver nitrate stock solution (34,875 ppm) under similar conditions as described in example 1 the temperature was maintained at 25 to 27 degrees C. After 3 h of incubation and stirring different aliquot of samples were removed and checked for various parameters as described in example 1. Characteristic plasmon peak of Silver nano particles at 550 nm were observed indicating an average diameter in the range of 30-250 nm. The stability of nano particles was checked in detail and it was observed that the Silver nano particles are stable under normal and moderately harsh environmental conditions.
This shows that aqueous solution of biological tissue extracts are excellent reducing agents for making sub micron scale Silver particles. The particles thus formed are stable in moderately harsh conditions.
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We Claim:
[1] A process for making Silver sub micron scale particles comprising the steps of:
(a) dissolving a salt of the Silver in water having conductivity less than 3 micro
Siemens to obtain a solution in which the concentration of Silver ions is in the
range of 20, 000 to 50 000 ppm ,
(b) preparing a fresh filtered aqueous solution of biological tissue extract;
(c) diluting the aqueous solution with deionized water in the ratio ranging from 1:5 to 1:50 to form a solution having an open circuit potential between + 0.2 and +0.2 volt and a pH between 5.5 to 7.5 and total organic carbon content at least 7,500 ppm;
(d) maintaining the said aqueous solution under continuous agitation at a temperature between 20 and 30 degrees Celsius;
(e) inoculating a minute quantity of the Silver salt solution in the said aqueous extract solution under continuous agitation such that the final concentration of the metal ion in the reaction mixture is in the range of 50 to 300 ppm;
(f) continuing the agitation for a period of 30 minutes to 3 hours in well illuminated conditions ; to obtain a colloidal suspension of Silver nano particles;
(g) separating the nano particles from colloidal suspension by a known process such as centrifugation.
[2] A process for making Silver sub micron scale particles as claimed in claim 1, in which the biological tissue extract is an extract of macerated plant cells.
[3] A process for making Silver sub micron scale particles as claimed in claim 1, in which the biological tissue extract is an extract of macerated cells of at least one plant tissue selected from a group of tissues comprising living tissue of grasses, leaves, fruits, stems, roots and flowers and parts thereof.
[4] A process for making Silver sub micron scale particles as claimed in claim 1, in
which the biological tissue extract is an extract of macerated animal cells.
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[5] A process for making Silver sub micron scale particles as claimed in claim 1, in which the biological tissue extract is an extract of macerated cells of at least one animal tissue selected from a group of tissues consisting of tissues of worms, insects, fishes, mollusks, crustaceans, and higher animals.
[6] A process for making Silver sub micron scale particles as claimed in claim 1, in which the biological tissue extract is an extract of macerated microbial cells.
[7] A process for making Silver sub micron scale particles as claimed in claim 1, in which the biological tissue extract is an extract of macerated cells of microbial tissue selected from a group of microbes which include bacteria, fungi, yeasts, viruses, protozoa and algae.

Dated this 11th day of May 2005

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ABSTRACT
A method of making Silver nano particles is disclosed. An aqueous solution of Silver is reduced with the help of an aqueous extract of biological tissue cells having specific characteristics .