Drug – bio-affecting and body treating compositions – Plant material or plant extract of undetermined constitution... – Containing or obtained from a stem – stalk – branch – or twig
Reexamination Certificate
2001-02-15
2002-04-30
Lilling, Herbert J. (Department: 1651)
Drug, bio-affecting and body treating compositions
Plant material or plant extract of undetermined constitution...
Containing or obtained from a stem, stalk, branch, or twig,
C424S725000, C435S204000
Reexamination Certificate
active
06379721
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to a process for the preparation of &agr;-amylase, useful for starch saccharification from a novel plant source
Tinospora cordifolia
Miers belonging to Menispermaceae group of plants. The present invention particularly relates to a process for the preparation of a novel &agr;-amylase, which saccharify starch mainly into maltose and glucose.
BACKGROUND OF THE INVENTION AND PRIOR ART DISCUSSION
Starch degrading &agr;-amylase enzyme have large numbers of biotechnological applications e.g. in the productions of syrups containing oligosaccharides, maltose and glucose from corn and other starchy materials, fermentable carbohydrate for ethanol production by saccharification of corn or other starchy materials and as digestive enzyme in pharmaceutical preparations, as thinning agent in lowering viscosity of commercial starch preparations, improving enzyme in bread making and in many other applications. The traditional process of acid catalyzed saccharification of starch into syrup or fermentable sugars has largely been replaced for many advantageous reasons by the enzymatic processes using &agr;-amylase. References may be made to H. C. Barford, Cereal Foods World 21, 588, 1976 and L. A. Underkofler, L. J. Denault and E. F. Hon, Die Starke 17, 179, 1965.
The enzyme &agr;-amylase (&agr;-1,4 D-glucan glucanohydrolase Enzyme commission number 3.2.1.1) is widely produced by different microorganisms and is also present in some cereals. It hydrolyses &agr;-1,4-glucosidic linkages present in amylose, amylopectin and glycogen in an endo-fashion. But it does not hydrolyze &agr;-1,6-glucosidic branch point present in amylopectin. Saccharification of amylose and amylopectin by &agr;-amylase produces, maltose, maltotriose and glucose and a series of branched &agr;-limit dextrins respectively. The amount and nature of &agr;-limit dextrins varied with the nature of &agr;-amylase obtained from different sources.
The enzyme produced by microorganism may be saccharifying or liquefying in nature. Saccharifying enzyme produces more reducing sugar from starch than the liquefying enzyme, but later lower viscosity of starch more quickly than the former enzyme.
Bacillus subtilis
var
amylosaccharficus, Bacillus subtilis
Marburg,
Bacillus subtilis
Natto are potential producers of saccharifying &agr;-amylase. Reference may be made to H. Matsuzaki, K. Tamane, K. Yamaguchi, Y. Naguta and B. Maru Biochimica Biophysica Acta 365, 235, 1974.
Bacillus amyloliqurefaciens
produces large amount of liquefying &agr;-amylase. Reference may be made to N. G. Welkar and L. L. Campbell. Journal of Bacteriology 94, 1131, 1967, Saccharifying &agr;-amylase is immunologically different from liquefying enzyme and in having maltase activity. Reference may be made to H. Yoshida, K. Hiromi and S. Ono. Journal of Biochemistry, Tokyo, 62, 439, 1967. Saccharifying &agr;-amylase has tremendous uses in the production of syrup and fermentable carbohydrate from starchy raw materials and in medicine. Grain starch as carbon source, is used to the extent of 50% of total substrate for ethanol production. Liquefying amylase is used for quickly reducing viscosity of starch solution. Germinating cereals also contain saccharifying &agr;-amylase. Traditional starch hydrolysis is usually conducted using barley malt. Malt is obtained mainly from barley, sometimes also from wheat and oats. The cereals are allowed to germinate for a limited period of time and then dried under suitable condition to terminate growth of embryo. The ground powder of the dried cereals is used as the source of enzyme. In beer preparation, malt is used both as source of enzyme and source of carbohydrate for fermentation. In the preparation of whisky and other liquors, gelatinized starch is saccharified by malt enzyme and the hydrolysed product is fermented.
As an alternative to barley malt, saccharifying &agr;-amylase produced by different fungi are also largely used. Mold bran containing growth of
Aspergillus oryzae
is traditionally used for starch hydrolysis. Reference may be made to J. Ziffer and M. C. Losit. Biotechnology Letters 4, 573, 1982. Submerged fermentation process for the production of saccharifying &agr;-amylase was also developed and enzymes from
Mucor touxii, Mucor boulard, Rizopus delemer
etc. were used for starch saccharification.
However, the production of saccharifying &agr;-amylase by malting barley is a highly technical exercise, which depends on the use of selected varieties of cereals and on the malting technique, which was developed as an art over hundreds of year. On the other hand, bacterial and fungal enzymes have restricted use in food preparations. In the industrial production of fungal enzyme, frequent allergic outbreaks were reported time to time, possibly due to spreading of fungal spores in the environment.
OBJECTS OF THE INVENTION
The main object of the present invention is to provide a process for the preparation of &agr;-amylase useful for starch saccharification from a novel plant source
Tinospora cordifolia
Miers belonging to Menispermaceae group of plant.
Another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the enzyme preparation does not require presence of calcium ion for optimum activity.
Still another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the saccharifying enzyme has &agr;-glucosidase activity which hydrolyses maltose.
Yet another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the production of enzyme does not require any controlled environmental conditions like malting or growth of microorganism under defined physicochemical conditions.
One more object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the crude enzyme preparation contains mostly &agr;-amylase protein with little non-amylase contaminating protein.
One another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the enzyme preparation become homogeneous protein in gel electrophoresis in presence of sodium dodecyl sulphate by a single step process involving ion exchange chromatography.
Another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the enzyme preparation under suitable conditions could digest soluble starch until the degree of polymerization is between 2 and 3.
Still another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the enzyme preparation under suitable conditions could digest soluble starch to generate up to 20% free glucose.
Yet another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the starting material used as the source enzyme is a plant which has been known to be edible to humans for a longtime.
One more object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the plant used as starting material is not a vegetable or fodder having alternative demand.
One another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the plant used as starting material grows wild throughout the whole seasons.
Another object of the present invention is to provide a process for the preparation of &agr;-amylase wherein the plant used as starting material could grow from any part of the mature stem under proper growth supporting environments.
SUMMARY OF THE INVENTION
The present invention provides a process for the preparation of &agr;-amylase useful for starch saccharification from a novel plant source
Tinospora cordifolia
Miers belonging to Menispermaceae group of plant.
DETAILED DESCRIPTION OF THE INVENTION
Accordingly, the present invention provides a process for the preparation of &agr;-amylase useful for starch saccharification, which includes blending cut pieces of
Tinospora codifolia
Mi
Jana Mohanlal
Naskar Amal Kumar
Naskar Panchanan
Sengupta Subhabrata
Council for Scientific and Industrial Research
Dinesh Agarwal, P.C.
Lilling Herbert J.
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