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"Synthesis Of Concentrated Colloids Of Silver And/Or Gold Nanoparticles Using Alkyl Polyglucosides"

Abstract: A method of making concentrated and stable silver and/or gold nanoparticles using a water-soluble reducing sugar based nonionic surfactants, alkyl polyglucosides (APGs) of Formula I. Formula I The synthesized nanoparticles of silver and/or gold are used in preservation of personal care and home care products.

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Patent Information

Application #
Filing Date
18 February 2015
Publication Number
11/2015
Publication Type
INA
Invention Field
PHARMACEUTICALS
Status
Email
Parent Application
Patent Number
Legal Status
Grant Date
2019-07-20
Renewal Date

Applicants

GALAXY SURFACTANTS LTD.
C-49/2, TTC INDUSTRIAL AREA, PAWNE, NAVI MUMBAI-400 703, MAHARASHTRA, INDIA.

Inventors

1. KOSHTI, NIRMAL
68, DAHILA COURT PISCATAWAY NEW JERSEY NJ 08854 USA
2. PAREKH, VISHAL
608/S-2, UDYOG NAGAR CHS, KAMLA VIHAR COMPLEX, MAHAVIR NAGAR KANDIVALI (W)-400067 MAHARASHTRA, INDIA.
3. SAWANT, BHAGYESH JAGANNATH
202, NAV-GOKUL-DHAM CHS 'F' CABIN ROAD, KATEMANVALI, KALYAN (E)-421306 MAHARASHTRA, INDIA.

Specification

FORM 2 THE PATENT ACT, 1970 (39 of 1970) & The Patents Rules, 2003 COMPLETE SPECIFICATION (See section 10 and rulel3) l.TITLE OF THE INVENTION: SYNTHESIS OF CONCENTRATED COLLOIDS OF SILVER AND/OR GOLD NANOPARTICLES USING ALKYL POLYGLUCOSIDES 2. APPLICANT (a) NAME: GALAXY SURFACTANTS LTD. (b) NATIONALITY: An Indian Company incorporated under the Indian Companies ACT, 1956 (c) ADDRESS: C-49/2, TTC Industrial Area, Pawne, Navi Mumbai - 400703 Maharashtra, India 3. PREAMBLE TO THE DESCRIPTION The following specification particularly describes the invention and the manner in which it is to be performed. FIELD OF INVENTION The present invention relates to the method of synthesizing nanoparticles of silver and/or gold. It relates to the process of making stable nanoparticles of silver and/or gold using non-ionic sugar based water-soluble surfactants such as alkyl polyglucosides (APGs). The present invention relates to a simple, rapid and cost-effective method of synthesizing stable colloids of silver and/or gold nanoparticles. The present invention relates to synthesis of colloids of silver and/or gold nanoparticles with high concentrations without using any additional stabilizer. Further, the APG mediated as well as stabilized nanoparticles of silver and/or gold are used for the preservation of personal and homecare products. BACKGROUND OF INVENTION In past couple of decades, research on various nanomaterials has been the subject of huge interest since they have exhibited extremely fascinating and useful properties that have been exploited to improve or even revolutionize many sectors of information technology, energy, environmental science and medicine. Metallic nanomaterials such as silver (Ag) in particular, have found use in a broad range of applications such as catalysis, electronics, bio-sensing, medicine, water treatment etc. (Moreno-Manas M and Pleixats R, Ace. Chem. Res. 638, 36 (2003); Markin C A, Letsinger R L, Mucic R C and Storhoff J J, Nature, 607, 382 (1996); Han M, Gao X, Su J Z and Nie S, Nature Biotechnol. 631, 19 (2001), Kamat P V J. Phys. Chem., 7729, B106 (2002)). Silver nanoparticles have unique optical and electrical properties and thus colloidal silver is one of the widely used substrates for Surface Enhanced Raman Spectroscopy (SERS) for single molecule detection (Nie S and Emory S R, Science, 1102, 275 (1997); Dick L A, McFarland A D, Haynes C L and P van Duyne R, J. Phys. Chem. 853, B106 (2002)). The applications of silver nanoparticles in the medical field include both diagnostic as well as therapeutic. Antibacterial effects of silver species have been known from centuries. Recently, the antimicrobial properties of silver nanoparticles have been documented (Sanpui P, Murugadoss A Durga Prasad P V, Ghosh S S and Chattopadhyay A, IntJ. Food Microbiol. 142, 124(2008), Ruparelia J P, Chatterjee A K, Duttagupta S P and Mukherji S, Acta Biomaterialia 707, 43 (2008); Kumar A, Vemula P K, Ajayan P M and John G, Natl. Mater., 236, 3 (2008)). In most therapeutic applications, it is the antimicrobial activity as well as the anti-inflammatory activity that are being exploited. The exact mechanism for their antimicrobial action is not understood and it is possible that one or more than one mechanisms could be in operation. Adsorption to cell wall and pitting the same is proposed. It is also been suggested that the generation of free radicals from metal nanoparticles to cause the damage to cell membrane of bacteria to make it porous. It is also proposed that the Ag ions that are formed could be doing the damage by interacting with thiol groups of many vital enzymes. In recent years, there have been a few references in literature where silver in the form of Ag nanoparticles is documented for its potential applications in personal care. Silver nanoparticles have been exploited by their incorporation into foot powders, sprays, soaps, socks, shoe insoles and a wide range of fabrics (Edwards-Jones, V, The benefits of silver in hygiene, personal care and healthcare. Letters in Applied Microbiology, (2009), 49: 147-152). German patent application DE 10340277 discloses a personal care product with metallic silver agglomerates for treating skin disorders by providing antimicrobial effect. European patent application EP 1685824 (2005) discloses a topical cosmetic formulation for concealing wrinkles and eliminating damages resulted from a variety of skin disorders such as acne. The topical composition taught by this patent includes elemental silver, an electrolyte and hydrophobic particles. European patent EP 1303283 (2000) discloses the use of noble metals, particularly nano-crystalline silver in a topical formulation for the treatment of hyper-proliferative skin diseases such as psoriasis. It teaches the use of nanocrystalline silver for slow release from coatings on substrates such as cellulose, cotton or polyester. It also teaches the other ways of slow release of silver in the form of powders or ointments/creams. Antimicrobial activity of nanoparticles of both silver and gold against Gram negative E. coli and Gram positive BCG has been studied in detail by Zhou et al. (Journal of Nanobiotechnology, 10, 19 (2012)). The results indicate that the size, shape and capping agent influence the antimicrobial efficacy. Gold nanoparticles have been shown to be very effective when capped with polyamine hydrochloride. Preparation of stable gold nanoparticles by using stabilizing agent, a mercapto compound, triethylene glycol mono-11-mercaptoundecyl ether has been reported in a recent publication (P/iys. Chem. Chem. Phys., 10175-10179, 11(2009). Production of nanoparticles involves physical, chemical processes as well as bioprocess. Conventionally, silver nanoparticles have been prepared by various methods such as co-precipitation in aqueous solutions, electrochemical methods, aerosol, reverse micro-emulsion, chemical liquid deposition, and photochemical reduction, chemical reduction in solution and UV irradiation. There are several reports in literature wherein stable nanoparticles have been synthesized using microemulsion technique. (M. A. Lopez-Quintela et al.; Current Opinion in Colloid and Interface Science, 264, 9 (2004). These microemulsions employ organic solvent, water and a surface active agent that can be cationic, anionic or nonionic in nature. The review article covers all types of surfactants used for creating nanoparticles through microemulsion. Preparation of stable silver and silver-gold bimetallic nanoparticles in water-in-oil type of microemulsion using nonionic Triton X 100 has been reported (Angshuman Pal et al, African Physical Review Special issue (Microfluidics): 0001 (2007)). The major limitation of this microemulsion technique is the isolation of nanoparticles from the solvent (water-immiscible solvents). Cetyl trimethyl ammonium bromide (CTAB, a cationic surfactant), AOT (fatty alkyl sulphosuccinate, anionic surfactant) and Triton X 100 (Nonyl phenol ethoxylate , non ionic surfactant) are the examples of commonly used surfactants in W/O type of microemulsions. Amongst nonionic surfactants, similar to Triton X, other ethoxylated nonionics that are employed for creating microemulsions are Brij types and Tween types. Nonethoxylated type nonionic surfactants that are used in W/O type of microemulsion for creating nanoparticles of Cadmium sulfide and Palladium sulfide are sugar esters (P. S. Khiew et al. J. Crystal Growth, 35-43, 254 (2003)). The chemical reduction methods are based on reduction of silver or gold salt with reducing agents such as sodium citrate, sodium borohydride, hydroxylamine hydrochloride, hydrazine, ethylenediaminetetraacetic acid (EDTA) and ascorbic acid etc. Also, the synthesis of silver/gold nanoparticles is required to be carried out at nano-molar concentrations as synthesis carried out at higher concentrations leads to irreversible aggregations thereb y limiting the scope of synthesis of nanoparticles compositions of higher concentration. Further, a stabilizing agent is additionally required to be added to the colloid formed to prevent the aggregation of the formed nanoparticles. It is equally important that the reducing and the stabilizing agent (capping agent) be non-toxic, cost-effective and universally accepted to most of the applications of metal nanoparticles including both medical and non-medical. The stabilizers employed in the prior art are either electrostatic stabilizers such sodium citrate, Cetyltrimethylammonium bromide (CTAB) or are polymeric stabilizers (steric stabilization) such as poly(vinyl-pyrrolidone) (PVP), polyvinyl alcohol, bovine serum albumin (BSA), starch, cellulose, tannic acid etc. There is a need of the prior art of a multifunctional (reduction and stabilization) molecule to allow a better control over the particle size distribution, stability and commercial production (concentrated form and cost-effective) of nanoparticles for a variety of application. The inventors of the present invention have surprisingly developed a simple process of synthesizing stable, concentrated nanoparticles of silver and/or gold with size of less than 50 nm wherein the molecule used as reducing agent is also stabilizing the composition, thereby obviating the need of any additional/external stabilizer. OBJECT OF INVENTION i) It is an objective of the present invention to develop a facile synthesis of stable nanoparticles of silver and/or gold. ii) It is also an objective of the present invention to use water-soluble, reducing sugar based non-ionic surfactants for synthesizing the colloids of silver and/or gold nanoparticles from their respective ionic salts and stabilizing the same. iii) Another objective of the present invention is to synthesize stable colloidal solutions of silver and/or gold nanoparticles with concentration up to 5000 ppm. iv) Another objective of the present invention is to synthesize colloids of nanoparticles of silver and/or gold with narrow particle size distribution and broad spectrum of antimicrobial activity. v) Another objective is to synthesize stable silver and/or gold nanoparticles that are suitable for the preservation of home and personal care products. SUMMARY OF INVENTION The present invention describes a simple, rapid and cost-effective method of making stable nano sized particles of silver or gold by adding aqueous solution of their respective metal salt to a solution of nonionic, reducing sugar based water-soluble surfactants at temperature of 70 to 100 °C and alkaline pH. The present invention describes the use of nonionic, reducing sugar based water-soluble surfactants, alkyl polyglucosides (APGs) of Formula I, that serve the dual purpose of reducing ionic silver/gold compounds to metallic silver/gold nanoparticles and stabilizing the same. HO Formula I The synthesized stable silver nanoparticles exhibit broad range antimicrobial activity against both Gram negative and Gram positive bacteria as well as against yeasts and molds. The stable silver or gold nanoparticles compositions of the present invention find application in preservation of both personal and homecare formulations. DETAILED DESCRIPTION OF INVENTION In the present invention, the inventors report a novel and efficient method of producing highly concentrated colloids of silver or gold nanoparticles using one of the most widely used classes of commercially available water-soluble surfactants, alkyl polyglucosides (APGs) of Formula I. Formula I The present invention describes a simple, rapid and cost-effective method of making stable nano sized particles of silver or gold by adding aqueous solution of their respective metal salt to a solution of alkyl polyglucosides (APGs) at a temperature of 70 to 100 °C and alkaline pH. In a typical procedure, aqueous solution of APG with pH between 8.5 and 9.0 is preheated to 90 °C and the aqueous solution of silver/gold salt is added slowly to it. A small drop in pH during the reaction is adjusted by addition of alkali and pH is maintained in the range of 8.0 to 9.5. The nanoparticle formation is extremely facile and a gram molar scale synthesis does not take more than 30 mins to complete the reduction and subsequent stabilization. In an embodiment, the stoichiometry of alkylpolyglucoside (APG) is 1-2 equivalents with respect to silver salt (Ag salt: APG:: 1: 1.0 to 2.0). It is observed that if the degree of saccharide polymerization (n) in the reducing sugar based surfactants is higher, then lower equivalence of APG can be used to effect the reduction as well as stabilization of the nanoparticles formed. In an embodiment, metal salts of silver and gold are AgNO3 and HAuCl4 respectively. The process is described in detailed in examples 1, 7, 8 & 9. These examples demonstrate extremely facile synthesis of stable nanoparticles on 1 mmol to 15 mmol scale. In terms of concentration, scale of synthesis revealed by these examples has been from 50 ppm to 5500 ppm of final concentration of nanosized silver and gold. The colloidal silver particle compositions have been studied for their stability for over 90 days at room temperature (25 °C) and there is no indication of instability observed by spectroscopic and particle size measurement. The colloid of nanoparticle formation is evident from the color formation Fig 1(a) that has ɧ max of 413 nm in case of silver nanoparticles and ɧ max of 548 nm for gold nanoparticles indicating the Surface Plasmon Resonance (Fig 1(b) and Fig 2). '*%s% $**$ 1000 ppm Ag 200 ppm 100 ppm Ag Ag 50 ppm 25 ppm Ag Ag Fig 1(a) 0.8 0.6 0.4 0.2 300 400 500 600 700 800 Wavelength (nm) Fig 1 (b) 300 400 500 600 Wavelength (nm) 700 800 Fig.2: UV visible spectra of gold nanoparticles Particle size of silver nanoparticles is measured by dynamic light scattering (DLS) (Brookhaven Instrument model 90 Plus Particle Size Analyzer.) and average particle size was found to be in the zone of 10-50 nm with a low polydispersity of index (Fig 3 and Table 1). SU) S-B-BlCmtatd! SLN»B-fi(Cffl!4«ie 25 0 ! / y ; ; A! ! : / / : / / !// \ : : \ '• ; \ • '-:V : : 10 !02 10s 104 105 10* D.5 ' ' £00.d Diameter (nm) t«S0M qt)-85822 I Ret. lot - mi CumM- 5173 RsnW ?0 32 Fig. 3 Table 1: Particle size and size distribution of Ag nanoparticles recorded by DLS Run Effective diameter Polydispersity 1 32.2 0.219 2 31.1 0.227 3 31.6 0.214 Dm (nm) Dso (nm) D,0 (nm) SPAN=(D9o-D10/D5o) 11.7 20.1 34.7 1.144 Analysis of the particle size revealed that the synthesized nanoparticles had effective hydrodynamic diameter of 35 nm and had low polydispersity index indicating narrow particle size distribution, i.e the uniformity in size of the synthesized nanoparticles. (Table-1 & Figure 3). Zeta potential numbers of the samples of example 1 and 6 are approximately around -ve 20 Mv to -25 Mv indicating that the colloids are quite stable. Morphology of the synthesized nanoparticles is studied by electron microscopy (Fig. Fig.4: TEM of the synthesized APG- capped silver nanoparticles. Transmission electron microscopy is performed on TEM instrument (Philips make, Model- CM200, Operating voltages: 20-200kv Resolution : 2.4 A0). The TEM images show that particles are having near spherical shape and have particle size in range of 10-35nm.) Alkyl polyglycosides (Formula I) surfactants were developed in nineties from naturally occurring renewable raw materials, namely, fatty alcohols and sugars. Fatty alcohols are derived from vegetable oils and hence alkyl polyglycosides are referred to as green surfactants. When the sugar portion in alkyl polyglycoside is glucose then it is known as alkyl polyglucoside. These surface active molecules are completely biodegradable and eco-friendly and that makes them truly "Green Surfactant". In addition to being green in terms of ecotoxicity and synthesis, these are quite mild on human skin (unlike soap and other anionic surfactants like sodium dodecyl sulphate) and have excellent detergency. Commercially they are available from on metric tonne scale and Elotant Milcoside and Plantaren/AGNIQUE are some of the common trade names from two of the major producers in the world. Henkel, Germany is the pioneer of this class of surfactants and ample research is reported over last three decades by the pioneers and others for their suitability in personal care applications. Excellent performance as surfactants along with environmental advantages and low oral and dermal toxicity, low aquatic toxicity and biodegradability make them truly green surfactant for personal care application (green.surfacatants@cognis.com). Another huge advantage of alkyl polyglycoside is their non-ionic nature. Unlike other ionic surfactants, cationic or anionic, the non-ionic alkyl polyglycosides / polyglucosides are compatible with other ingredients in any composition and stable over wide range of pH. The hydrophobicity of the surfactants can be modified by varying the degree of polymerization of sugar units (Formula I) and hydrophilicity is adjusted by varying the long alkyl chain of saturated or unsaturated hydrocarbon chain available in nature, particularly a variety of vegetable oils. The glycosidic linkage can be the ones that occur in nature that 1, 4 or 1, 6 etc. Polysaccharides for alkyl polyglycoside can be derived from hydrolyzed starch or hydrolyzed cellulose where 1,4 linkage could be a or β. Lauryl polyglucoside of Formula II depicts one such linkage through anomeric carbon wherein two sugar units have been linked to a long alkyl chain of twelve carbons. Formula II The alkyl polyglycosides of the present invention used for the manufacture of stable silver nanoparticles are the ones wherein the glycoside portion of the surfactants are the reducing sugars. Examples of reducing sugars are glucose, glyceraldehyde, galactose, disaccharides like lactose, maltose have a reducing form as one of the units has open chain with aldehyde group. In glucose polymers such as starch or starch derivatives lik e glucose syrup, maltodexrin, and dextrin the macromoleucles that begin with reducing sugar. A reducing sugar is any sugar that has either aldehyde group or is capable of forming one in solution through isomerism. The cyclic hemiacetal form of aldoses can open to reveal an aldehyde and contain ketoses that can undergo tautomerization to become aldoses. However, acetals, including those found in oligosaccharide linkages can't easily become free aldoses to act like reducing sugars. In view of above, it is a surprising finding of the present invention that alkyl polyglucosides (Formula I) act as very efficient stoichiometric reducing agents to reduce Ag (+ 1) to Ag (0) or to reduce Au (+3) to Au(0). While creating silver or gold nanoparticles using alkyl polyglycoside, complete absence of free glucose in APGs has been completely established. Thus, there is no free glucose or reducing sugar in the alkyl polyglucoside and the reduction of Ag +1 to Ag (0) or to reduce Au (+3) to Au(0) is effected by saccharide portion of APG. The role of surfactant with reducing sugar was also established by using isolated biosurfactants that are commercially available. Sophorolipid (Sophogreen, from Soliance ) is glucolipid as shown in Formula III. Instead of using yeast fermented mixture as reported in the literature (Journal of Chemical Science, 515-520, vol 120, no 6, (2008)), isolated pure sophorolipid has been successfully used to create silver nanoparticles from silver nitrate as shown in Example 2. AcO AcO Formula III, sophorolipid, Formula III, lactonic form Sophorolipid acid form The nanoparticles thus produced have been found to be reasonably stable (Example 2). The major disadvantage of sophorolipids is the cost of manufacture and the scale of manufacture as compared with cost of manufacture of commercial alkyl polyglucosides. Secondly, the lactonic form (Formula III) hydrolyzes in alkaline condition and both the forms actually serve as anionic surfactants (end carboxylic acid groups can form the alkali metal salt). Alkyl polyglycosides (Formula I) in addition to cost advantage (economy of scale) have distinct advantage of being non-ionic under all circumstances and hence do not interact with other strongly ionic materials. It has also been demonstrated that by using reducing sugar, glucose, it is possible to reduce Ag (+1) salt in aqueous medium but nanoparticles generated are not stable and that can be easily seen from the experiment described in Example 4 & 6. Interestingly, another good water-soluble nonionic surfactant, lauryl alcohol ethoxylate 7 EO (Formula IV), prepared by ethoxylation of lauryl alcohol has been found to be incapable of stabilizing the colloid of silver nanoparticles as exemplified in Example 3. Formula IV Lauryl alcohol ethoxylate 7 EO (Formula IV) has seven ethereal oxygen linkages and terminal hydroxyl group, however, it does not provide effective capping/stability to glucose reduced silver nanoparticles. Similarly, Example 5 and 6 demonstrate that anionic surfactant, sodium lauryl ether sulphate (2 EO) that has terminal anionic sulphate group (Formula V) and ethereal linkages is also incapable of capping the nanoparticles produced from silver nitrate and glucose. Formula V The probable mechanism of synthesis and stabilization of silver nanoparticles can be explained as follows: Addition of potassium hydroxide or other suitable alkali to a solution of alkyl polyglucosides and silver nitrate leads to precipitation of silver ions (Ag+) of silver nitrate in form of silver oxide and silver hydroxide. Gradual addition of silver nitrate and the presence of APG helps in keeping the size of precipitated silver oxide and silver hydroxide to submicron scale. Under the maintained alkaline conditions, the hydrophihc portion of Alkyl polyglucosides i.e glycoside unit acts as reducing agent which carries out reduction of silver oxide and silver hydroxide to metallic silver at elevated temperatures of 80-95 °C. As soon the new surface/ nuclei of metallic silver is generated by reduction silver hydroxide, the surface active nature of APG molecules drives them to adsorb its hydrophobic portion towards the formed silver metal particles. Such micelle type aggregation of APG prevents the coalescence of nuclei and also limits the diffusion of metallic silver atoms into the surface of nuclei which not only limits the particle size even if the starting concentration of silver nitrate is high, but also keeps the formed silver nanoparticles stable over a period of time as the aggregation of particles is prevented. Another scenario that can be envisaged is the ligation of metal nanoparticles by hydroxyl groups of the head group of the nonionic surfactant micelles. Since the surfactant concentration is in stoichiometric ratio and above its CMC (critical micelle concentration), it is very likely that the head group of APG coordinates the newly formed metal nanoparticles and forms stable chelates. It is also likely to reduce precious metals like Ag and Au from their +1 and + 3 oxidation states using derivatives of APG with reducible sugar units in the molecule, for example 'poly suga quats' series made from APGs by Colonial chemicals CCI, USA. Metal salts of Silver and Gold: In the examples given in experimental section AgNO3 is used as source for silver ions. For gold ions HAuCl4 is used. Gold nanoparticles have been synthesized with equal ease as described in the Example No 9. Antimicrobial properties of Silver and gold nanoparticles: Silver nanoparticles synthesized using APG exhibited broad spectrum antimicrobial activity in concentration range of 25 - 45 ppm, against both gram positive bacteria such as Staphylococcus aureus, and gram negative bacteria such as Escherichia coli, Pseudomonas aeruginosa. APG-capped silver nanoparticles also demonstrated antimicrobial activity against acne causing bacteria such as Propionibacterium acnes and also against dandruff causing fungi such as Malassezia furfur and against other 18 commonly known yeasts and moulds. The gold nanoparticles were not as efficient as silver nanoparticles as indicated by higher MIC values of gold nanoparticles. The Minimum inhibitory concentration (MIC) and Minimum Bactericidal concentration (MBC) values of the synthesized APG -capped silver nanoparticles are presented in Table 2. Similarly, a pilot run of APG -capped gold nanoparticles at 400 ppm concentration indicated inhibitory activity on bacteria of Table 2 which seem to agree with the literature report (Chandran et al, Arabian Journal of Chemistry, In press, (2014).). Table 2: Antimicrobial activity of silver nanoparticles Microorganism MIC values in ppm MBC values in ppm Staphylococcus aureus ATCC 6538 25 45 (Escherichia coRATCC 8739 20 40

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1 ABSTRACT1.jpg 2018-08-11
2 507-MUM-2015-Other Patent Document-180215.pdf 2018-08-11
3 507-MUM-2015-Form 9-180215.pdf 2018-08-11
4 507-MUM-2015-Form 3-180215.pdf 2018-08-11
5 507-MUM-2015-Form 2(Title Page)-180215.pdf 2018-08-11
6 507-MUM-2015-Form 18-180215.pdf 2018-08-11
7 507-MUM-2015-Form 1-180215.pdf 2018-08-11
8 507-MUM-2015-FER.pdf 2018-08-11
9 507-MUM-2015-OTHERS [06-10-2018(online)].pdf 2018-10-06
10 507-MUM-2015-FER_SER_REPLY [06-10-2018(online)].pdf 2018-10-06
11 507-MUM-2015-CORRESPONDENCE [06-10-2018(online)].pdf 2018-10-06
12 507-MUM-2015-FER (By Post).pdf 2018-12-10
13 507-MUM-2015-RELEVANT DOCUMENTS [19-07-2019(online)].pdf 2019-07-19
14 507-MUM-2015-FORM-26 [19-07-2019(online)].pdf 2019-07-19
15 507-MUM-2015-FORM 13 [19-07-2019(online)].pdf 2019-07-19
16 507-MUM-2015-PatentCertificate20-07-2019.pdf 2019-07-20
17 507-MUM-2015-IntimationOfGrant20-07-2019.pdf 2019-07-20
18 507-MUM-2015-ORIGINAL UR 6(1A) FORM 26-290719.pdf 2019-12-21
19 507-MUM-2015-RELEVANT DOCUMENTS [18-03-2020(online)].pdf 2020-03-18
20 507-MUM-2015-RELEVANT DOCUMENTS [15-07-2021(online)].pdf 2021-07-15
21 507-MUM-2015-RELEVANT DOCUMENTS [05-08-2022(online)].pdf 2022-08-05
22 507-MUM-2015-RELEVANT DOCUMENTS [26-09-2023(online)].pdf 2023-09-26

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