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Process For The Preparation Of Stable Isomaltooligosaccharides

Abstract: The present invention relates to a process for preparing stable isomaitooligosaccharides comprising an enzymatic transfer reaction in the presence of glucansucrase between maltose and isomalto-oligosaccharides, wherein the said process comprises of about 0.025% to about 0.05% v/v solution of glucansucrase and about 20% to about 60% w/v solution of maltose.

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Patent Information

Application #
Filing Date
26 February 2009
Publication Number
47/2010
Publication Type
INA
Invention Field
CHEMICAL
Status
Email
Parent Application

Applicants

ALKEM LABORATORIES LTD.
DEVASHISH, ALKEM HOUSE, SENAPATI BAPAT MARG, LOWER PAREL,MUMBAI 400 013, MAHARASHTRA, INDIA.

Inventors

1. ANIL KUMAR PANDEY
Alkem Laboratories Ltd., C17/7, M.I.C.C.,Industrial Area, Tluka-Panvel, Dist-Riagad, Taioja-410208, Maharashtra.
2. ARUN KUSHAWAHA
Alkem laboratories Ltd., C 17/7,M.I.D.C, Industrial Area, Taluka Panvel, Dist-Raigad, Toloja-410208,Maharashtra.
3. SANJAY JAISWAL
Alkem laboratories Ltd., C17/7,M.I.D.C, Industrial Area, Taluka-Panvel,Dist-Raigad, Talija-410208, Maharashtra.
4. SAMPRADA SINGH
Devashish, Alkem House, Senapati bapat Marg,Lower Parel, Mumbai-400 013

Specification

FORM 2 THE PATENTS ACT. 1970 (39 OF 1970) & PATENTS RULES. 2006 PROVISIONAL SPECIFICATION (SECTION 10; RULE 13) "PROCESS FOR THE PREPARATION OF STABLE ISOMALTOOLIGOSACCHARIDES,, ALKEM LABORATORIES LIMITED, A COMPANY INCORPORATED UNDER THE COMPANIES ACT, 1956; HAVING ITS CORPORATE. OFFICE AT ALKEM HOUSE, DEVASHISH, ADJACENT TO MATULYA CENTRE, S.B.MARG, LOWER PAREL, MUMBAI - 400013, MAHARASHTRA, INDIA THE FOLLOWING SPECIFICATION DESCRIBES THE NATURE OF THE INVENTION AND THE MANNER IN WHICH IT IS TO BE PERFORMED FIELD OF THE INVENTION The present invention relates to a process for preparing stable isomaltooligosaccharides. BACKGROUND OF THE INVENTION Oligosaccharides are an important group of polymeric carbohydrates that are found either free or in combined forms in all living organisms. Structurally, oligosaccharides are composed of 2-10 monosaccharide residues linked by giycosidic bonds mat are readily hydrolyzed to their constituent monosaccharides either by acids or by specific enzymes. Oligosaccharides are of various types given in Table I, namely Starch related Oligosaccharides, Sucrose related Oligosaccharides, Lactose related Oligosaccharides and others. Starch oligosaccharides, which represent the fragments of original polysaccharides, are composed of a-D-glucopyranosyl units linked by a-1,4 and/or a* 1,6 bonds. Oligosaccharides containing only a-1,4 glucosidic linkages are called malto-oligosaccharides, while those containing both a-1,4 and a-1,6 glucosidic linkages are called branched-oligosaccharides or isomalto-oligosaccharides. Table 1 Raw material Product Starch related Oligosaccharides Malto-oligosaccharides: maltose ~ maltoheptaose, Isomalto-oligosaccharides: isomaltose, panose- isomaltotriose Cyclodextrins (CDs): a-CD, p-CD, y-CD, HP- p-CE>, branched CDs Others: maltitol, gentio-oligosaccharides, trehalose, nigerose Sucrose related Oligosaccharides Glycosyl sucrose, fructo-oligosaccharides, palatinose (isomaltulose), lactosucrose. xylosucrose, raffinose, stachyose, trehalulose Lactose related Oligosaccharides Galacto-oligosaccharides, lactosucrose, lactulose, lactitol Others Xylo-oligosaccharides, agaro-oligosaccharides, manno-oligosaccharides, chitin/chitosan oligosaccharides, etc. Isomalto-oligosaccharides (IMO) such as isomaltose, panose, isomaltotriose, isomaltotetraose, etc. are obtained from various sources and in different ways, including (a) extraction from plants, (b) enzymatic or acidic decomposition of natural polymers or (c) enzymatic synthesis. The latter method provides diversity in product and comprises reversion of hydrolysis, transhydrolytic reactions exploiting transferase activity of hydrolases, and transfer reactions catalysed by transferases using carbohydrate substrates such as sucrose, maltose, starch and dextran. Isomaltooiigosaccharides are known as growth-promoting saccharides for bifido bacterium and they will stimulate, in the large intestine, the growth of bacteria that are supposed to be beneficial for the health of the individual. The genus bifidobacteria, in particular, is known to have a positive influence on the general health condition. It has also fee function of tooth-decay prevention in the food containing sucrose and as it is hardly metabolized by yeast and lactobacillus. So it is one land of additives with low calories and helps to prevent weight gain. Isomaltooligosaccharide is a prebiotic and has prebiotic fibre. Isomaltooligosaccharide is used in a wide range of products like beverage industry, brewing industry, confectioneries, baking industry, health food industries and in various functional foods which may require high operational temperature and low pH. John fi Pazur et al., Journal of Biological chemistry, page no.265-272, 1951, describes conversion of maltose to 4-a-isomaltosyl-n-glucose (panose), and 6-a -isomaltosyl-D-glucose (dextrantriose), and the tetrasaccharide 4- a -dextrantriosyl-n-glucose (isomaltooligosaccharide) in presence of transglucosidase which is obtained from Aspergillus niger. Inventors have used 2% of maltose solution and 10% of enzyme solution for the conversion of maltose to isomaltooligosaccharide. Goulas AthanasioS K. et al., Biotechnology and Bioengineering, Volume 88, Issue 6, page no. 778 - 787,2004, describes a continuous synthesis process for the production of isomaltooHgosaccharides from sucrose, using the enzymes dextransucrase and dextranase in a recycle ultrafiltration using a variety of membranes and the parameters affecting reactor stability, productivity, ana product molecular weight distribution were investigated. Dextransucrase inactivation was shown to occur mainly on the substrate availability during continuous isomaltooligosaccharide synthesis. Dey-Chyi Sheu et al., Biotechnology Techniques, Volume 11, Number 5, page no. 287-291, May. 1997, describes partially purified alpha-glucosidase from Aspergillus carbonarious, immobilized on glutaraldelryde-activated chitosan beads in a packed bed reactor, to obtain 60% (wAv) of isomaltooiigosaccharides using 30% (Wv) maltose soiulion. 46% (Ww) of isomaltooiigosaccharides was produced when intact myceiia was attached with polyethyieneimine-glutaraldehyde. using 30% (w/v) maltose solution. Lucia Fernandez-Arrojo et al., Process Biochemistry Volume 42, Issue 11, page no. 1530-1536; November 2007, describes the general process of preparing prebiotic isomaltooligosaccharides by transglycosylation of a novel a-glucosidase from the basidiomycetous yeast Xanthophyiiomyces dendrorhous (formerly Phajjia rhodozymd) was using maltose as glucosyl donor. Aziz Tanriseven et al.. Biotechnology Techniques. Volume 13. Number 3. page no. 207-210. March, 1999, describes a process of conversion of sucrose to fructooligosaccharides and isomaltooligosaccharides. The sugar mixture contained ketose (33.5%), nystose (13.3%), fructofuranosyl nystose (5.7%), glucose (20.9%), and unreacted sucrose (26.6%). The unreacted sucrose was converted to isomaltooligosaccharides by reacting the sugar mixture with leuconostoc mesenteroides B-512FM dextransucrase. US4673643 claims a process for the manufacture of iso-m3Uooligassaccharides monovalent haptens, comprising adding more than 300 mmol glucose per 1000 U a (l-6)~D-glucosyl transferase to an aqueous solution of D-glucose, at temperatures from 265 to 310 K and at a pH value of 4.5 to 8. EP1835025A1 claims an industrially viable process of obtaining prebiotic oligosaccharides using a new Xanthophyiiomyces dendrorhous enzyme, characterized by showing u> glucosidase activity. A process of obtaining an enzymatic product with a-glucosidase activity as well as the substantially pure enzyme with a-glucosidase activity, are also provided. The enzymatic product and the enzyme have a high action spectrum and a high specific activity as positive aspects. W02 004068966 AI application claims a process for preparing isomalto-oligosaccharides with elongated chain and low glycemic index wherein, the said process comprises an enzymatic transfer reaction between sucrose and isomalto-oligosaccharides present in carbohydrate syrup. The enzymatic transfer reaction takes place in the presence of glucansucrase, wherein the glucansucrase is preferably selected from the group consisting of dextransucrase, alternansucrase, mutansucrase and mixtures thereof. The prior art processes disclosed do not mention stability details of isomaltooligosaccharides produced and the fate of end products at different conditions of pH and temperature. Isomaltooligosaccharides is an essential ingredient in health food industries as food or food supplement and may require withstanding various pH and terflperature conditions. There is therefore a need for a process to make stable isomaltooligosaccharides, which is able to withstand variations in pH and temperature. The inventors have surprisingly found that use of specific amounts of the substrate and enzyme gives stable isomaltooligosaccharides, which withstand a wide range of pH and temperature conditions as required in food preparations, without being much affected and a process for making the same. SUMMARY OF THE INVENTION The present invention provides a process to make stable isomaltooligosaccharides. The present invention provides a process to make stable isomaltooligosaccharides.. which is able to withstand variations of pH and temperature. The present invention provides a process to make stable isomaltooligosaccharides. using specific amounts of the substrate and enzyme. The present invention provides a process for preparing stable isomaltooligosaccharides characterized in that said process comprises an enzymatic transfer reaction between maltose and isomaltooligosaccharides. The invention further provides a process for preparing stable isomaltooligosaccharides characterized in that said process comprises an enzymatic transfer reaction in the presence of glucansucrase between maltose and isomaltooligosaccharides. The invention further provides a process for preparing stable isomaltooligosaccharides characterized in that said process comprises an enzymatic transfer reaction in the presence of transglucosidase-L between maltose and isomaltooligosaccharides The invention can be summarized as follows: A. A process for preparing stable isomaltooligosaccharides wherein the said process comprises an enzymatic transfer reaction of about 0.025% to about 0.05% v/v solution of trans glucosidase-L with about 20% to about 60% w/v solution of maltose. B. A process for preparing stable isomaltooligosaccharides wherein the said process comprises: (i) Mixing about 0.025% to about 0.05% v/v solution of transglucosidase-L with about 20% to about 60% w/v solution of maltose; (ii) Transferring the mixture of (i) to a container at a specific pH and temperature: (iii) Raising the temperature of the system of (ii) to inactivate the enzyme (iv) Filtering the solution from container of (ii) and concentrating the stable isomaltooligosaccharides; and (v) Heating the solution and filling in an appropriate container. C. A process for preparing stable isomaltooligosaccharides characterized in that said process comprises: (i) Mixing about 0.025% to about 0.05% v/v solution of transglucosidase-L with about 20% to about 60% w/v solution of maltose in phosphate buffer solution of pH of about 4.5 to about 6.5; (ii) Transferring the mixture of (i) to a container wherein temperature of the mixture is maintained at 40-60°C and pH of the mixture from about 4.5 to about 6.5; (iii) Raising the temperature of the system of (ii) up to 70°C; (iv) Filtering the solution and concentrating the stable isomaltooligosaccharides; and (v) Heating the solution at 70°C for 1 hr and filling the liquid in an appropriate container, D. The process according to step A wherein solution of maltose in phosphate buffer solution is about 30% w/v. E. The process according to step A wherein transglucosidase-L is used in amounts ranging from about 0.03% to about 0.04%v/v. F. The process according to step D wherein the pH of the solution is selected from about 5.0 to about 6.0 and temperature is from about 50°C to about 58°C. BRIEF DESCRIPTION OF DRAWING FIGURES Further objects of the present invention together with additional features contributing thereto and advantages accruing there from will be apparent from the following description of preferred embodiments of the invention which are shown in the accompanying drawing figures wherein: Table 1 describes details various kinds of oligosaccharides. Table 2 gives the three month Stability data chart of three batches of isomaltooligosaccharides. Table 3 gives the details of Studies of effect of pH and temperature on Isomaltooligosaccharides (MO) DETAILED DESCRIPTION OF THE INVENTION Before the present process and methods are described, it is to be understood that this invention is not limited to particular compounds, formulas or steps described, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present invention will be limited only by the appended claims. Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges is also encompassed within the invention, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either both of those included limits are also included in the invention. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention, the preferred methods and materials are now described. All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited. It must be noted that as used herein and in the appended claims, the singular forms "a", "and", and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a compound" includes a plurality of such compounds and reference to "the step" includes reference to one or more step and equivalents thereof known to those skilled in the art and so forth. The publications discussed herein are provided solely for their disclosure prior to the filing date of the present applicatioa Nothing herein is to be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the dates of publication provided may be different from the actual publication dates which may need to be independently confirmed. The term "stable" as used herein indicates no change in the percentages of total isomaltooligosaccharides on dry basis, when kept at 40°C±2°C/75%RH±5%RH for three months. The present inventors have found a process to make stable isomaltooligosaccharides which is able to withstand variations in pH and temperature, using specific amounts of the substrate and enzyme. The present invention provides a process for preparing stable isomaltooligosaccharides characterized in that said process comprises an enzymatic transfer reaction between maltose and isomalto-oligosaccharides. The invention further provides a process for preparing stable isomaltooligosaccharides characterized in that said process comprises an enzymatic transfer reaction in the presence of glucansucrase between maltose and isomalto-oligosaccharides. Accordingly one aspect of this invention provides a process for preparing stable isomalto¬oligosaccharides wherein the said process comprises an enzymatic transfer reaction of about 0.025% to about 0.05% v/v solution of transglucosidase-L with about 20% to about 60% w/v solution of maltose. Accordingly another aspect of this invention provides a process for preparing stable isomalto¬oligosaccharides characterized in that said process comprises:

Documents

Application Documents

# Name Date
1 433-mum-2009-abstract(25-2-2010).doc 2018-08-10
1 433-MUM-2009-FORM 5(25-2-2010).pdf 2018-08-10
2 433-mum-2009-form 2.pdf 2018-08-10
2 433-MUM-2009-ABSTRACT(25-2-2010).pdf 2018-08-10
3 433-MUM-2009-ASSIGNMENT(9-4-2012).pdf 2018-08-10
4 433-mum-2009-form 2(title page).pdf 2018-08-10
5 433-MUM-2009-FORM 2(TITLE PAGE)-(25-2-2010).pdf 2018-08-10
5 433-MUM-2009-CLAIMS(25-2-2010).pdf 2018-08-10
6 433-mum-2009-form 2(25-2-2010).pdf 2018-08-10
6 433-MUM-2009-CORRESPONDENCE(25-2-2010).pdf 2018-08-10
7 433-MUM-2009-CORRESPONDENCE(9-4-2012).pdf 2018-08-10
8 433-mum-2009-form 1.pdf 2018-08-10
8 433-mum-2009-correspondence.pdf 2018-08-10
9 433-mum-2009-description(provisional).pdf 2018-08-10
9 433-MUM-2009-DESCRIPTION(COMPLETE)-(25-2-2010).pdf 2018-08-10
11 433-mum-2009-description(provisional).pdf 2018-08-10
11 433-MUM-2009-DESCRIPTION(COMPLETE)-(25-2-2010).pdf 2018-08-10
12 433-mum-2009-form 1.pdf 2018-08-10
12 433-mum-2009-correspondence.pdf 2018-08-10
13 433-MUM-2009-CORRESPONDENCE(9-4-2012).pdf 2018-08-10
14 433-mum-2009-form 2(25-2-2010).pdf 2018-08-10
14 433-MUM-2009-CORRESPONDENCE(25-2-2010).pdf 2018-08-10
15 433-MUM-2009-FORM 2(TITLE PAGE)-(25-2-2010).pdf 2018-08-10
15 433-MUM-2009-CLAIMS(25-2-2010).pdf 2018-08-10
16 433-mum-2009-form 2(title page).pdf 2018-08-10
17 433-MUM-2009-ASSIGNMENT(9-4-2012).pdf 2018-08-10
18 433-mum-2009-form 2.pdf 2018-08-10
18 433-MUM-2009-ABSTRACT(25-2-2010).pdf 2018-08-10
19 433-MUM-2009-FORM 5(25-2-2010).pdf 2018-08-10