Title of Invention

AN IMPROVED PROCESS OF OBTAINING HIGH CONTENTS OF BOUND-PHENOLIC ACID RICH DIETARY FIBRE

Abstract

The present invention relates to an improved process of obtaining bound-phenolic acid rich dietary fibre from cereal malts by step-wise increase of about 7-14°C in temperature and thereby using in situ amylases only. The main advantage of the present invention the use of in situ amylases to isolate high contents of bound-phenolic acid rich dietary fibre from cereal malts and it does not involve high temperature as seen in conventional methods.

Full Text

AN IMPROVED PROCESS FOR OBTAINING BOUND-PHENOLIC ACID RICH DIETARY FIBRE Field of the present invention The present invention relates to an improved process of obtaining boundphenolic acid rich dietary fibres. The present invention relates to an improved process of obtaining boundphenolic acid rich dietary fibres from cereal malts by step-wise increase of about 7-14°C in temperature and thereby using in situ amylases only. Phenolic acids such as ferulic and coumaric acids are covalently linked to mainly dietary fibre components such as arabinoxylans and pectins and influence their physicochemical properties in various food preparations. Besides acting as flavour compounds, phenolic acids are known to be possessing anticarcinogenic, antimutagenic, and antioxidant properties and hence their presence in dietary fibre components is very important for preparing health, geriatric and infant foods based on cereals. Phenolic acids are also present as free acids in very small amounts both in the cereals as well as malts. However, they undergo decarboxylation, in the process decreasing their utility value. Hence the bound phenolic acid content is very important for the positive health benefits of dietary fibre components. For phenolic acids' chemistry, biochemistry and biotechnology the following articles can be referred. • Journal of the Science of Food and Agriculture, Vol. 79 (1999) No. 3, pp 355- 490. • Non-starchy polysaccharide and bound phenolic acids from native and malted finger millet (Ragi, Eleusine coracana, Indaf- 15), M. V. S. S. T. Subba Rao and G. Muralikrishna, Food Chemistry, Vol. 72 (2001) 187-192. • Evaluation of the antioxidant properties of free and bound phenolic acids from native and malted finger millet (Ragi, Eleusine coracana, Indaf- 15), M. V. S. S. T. Subba Rao and G. Muralikrishna, J. Agric. Food. Chem., Feb 2002. Removal of starch by specific enzymatic treatments is a prerequisite step to prepare phenolic acid rich dietary fibre components from cereals and their malts. For preparing dietary fibres from cereals and their malts following reviewing articles can be referred. • Food Chemistry, Vol. 57 (1996) No. 1, pp 15-21. • Food Rev. Int., Vol. 13 (1997) pp 29-76. In the existing processes, enzymes from bacteria (Termamyl, thermostable enzyme) and fungi (Aspergillus niger, glucoamylase) are used to remove starch. Bacterial a-amylase is a thermostable enzyme and it degrades starch in to dextrins at boiling temperature (~98°C), which in turn are converted in to glucose by glucoamylase at 55°C. This procedure is a long drawn one, which involves high temperature treatment and enzyme cost. The main object of the present invention is to develop an improved process for obtaining bound-phenolic acid rich dietary fibre which obviates the drawbacks detailed above. Another object of the present invention is to develop an improved process of obtaining bound-phenolic acid rich dietary fibre from cereal malts. Yet another object of the present invention is to obtain bound-phenolic acid rich dietary fibre from cereal malts using in situ enzymes. Still another object of the present invention is to obtain bound-phenolic acid rich dietary fibre from cereal malts by step-wise increase in the temperature. In another object of the present invention is to develop a faster process of obtaining bound-phenolic acid rich dietary fibre from cereal malts. Yet another object of the present invention is to develop an economical process of obtaining bound-phenolic acid rich dietary fibre from cereal malts. Still another object of the present invention is to develop a faster process of obtaining bound-phenolic acid by retaining of all the bound phenolic acids present in the cereal malts unlike in the case of endosperm and bran, wherein only partial recovery of bound phenolic acid is achieved. Accordingly, the present invention relates to an improved process for obtaining bound-phenolic acid rich dietary fibres from cereal malts by step-wise increase in temperature by about 7-14°C for net temperature to be ranging between 36 and 84°C and using in situ amylases only, the said process comprising steps of: In an embodiment of the present invention, adding about 7-15 gm% (w/v) of cereal malt in about 40-60 mM of acetate buffer of pH ranging between 4.5-5.5 to obtain malt solution In another embodiment of the present invention, incubating malt solution at about 45°C for about 10-14 hours. Yet in another embodiment of the present invention, incubating malt solution of immediate above step further for about 10-14 hours at about 55°C. Still in another embodiment of the present invention, incubating malt solution of immediate above step further for about 10-14 hours at about 65°C. In another embodiment of the present invention, incubating malt solution of immediate above step for another about 10-14 hours at about 75°C to obtain a residue. Yet in another embodiment of the present invention, measuring starch levels after the each incubation. Still in another embodiment of the present invention, centrifuging the said residue of immediate above step. In another embodiment of the present invention, washing the centrifuged residue. Yet in another embodiment of the present invention, drying the washed residue with ethanol. Still in another embodiment of the present invention, measuring levels of bound-phenolic acid in the dried residue by conventional processes. In another embodiment of the present invention, obtaining bound-phenolic acid rich dietary fibres. Yet in another embodiment of the present invention, the said process eliminates the need for exogenous enzymes. Still in another embodiment of the present invention, the concentration of cereal malt in acetate buffer may be about 10 gm% (w/v). Yet in another embodiment of the present invention, the pH of the acetate buffer may be about 5.0. In another embodiment of the present invention, the concentration of the acetate buffer may be about 50mM. Still in another embodiment of the present invention, the cereal malts may be selected from a group comprising rice malt, maize malt, wheat malt, and ragi malt. Yet in another embodiment of the present invention, the said process eliminates starch from cereal malts comprising rice malt, maize malt, and wheat malt within about 21-39 hours. In another embodiment of the present invention, the said process eliminates starch at a maximum of about 65°C from cereal malts comprising rice malt, maize malt, and wheat malt. Still in another embodiment of the present invention, the said process eliminates starch from ragi malt within about 35-45 hours. Yet in another embodiment of the present invention, the said process eliminates starch at a maximum of about 75°C from ragi malt. In another embodiment of the present invention, the said process removes starch form cereal malt at a much faster rate as compared to conventional processes. Still in another embodiment of the present invention, the said process is economical as compared to conventional processes. In another embodiment of the present invention, centrifuging the residue at 5000 rpm for 1- 30 minutes. Still in another embodiment of the present invention, the concentration of ethanol may range between 40 and 100%. Yet in another embodiment of the present invention, the stepwise increase in temperature may help overcome the need of using exogenous enzymes. In another embodiment of the present invention, incubating the said mixture in an oven. Yet in another embodiment of the present invention, the starch levels may be measured by iodine test. Still in another embodiment of the present invention, washing the centrifuged residue in water. In the present invention, the improved process is faster than the conventional processes because in the conventional processes, the exogenous enzymes from bacteria and fungi are added and temperature is to raised to as high as about 100°C to obtain bound-phenolic acid rich dietary fibres from cereal malts. It is a very long drawn process with average results. The novelty in the present process is to exploit the malt/use of in situ enzymes to prepare fibre rich components without making use of bacterial amylase as well as glucoamylase. The steps of the improved process are as follows: 1. Weighing cereal malts (rice, maize, wheat and ragi) 1 g each and taking in 10 ml of acetate buffer (pH 5.0, 50 mM). 2. Incubation of the cereal malts at 45°C for 12 h. 3. Incubation of the cereal malts at 45°C for 12 h. 4. Incubation of the cereal malts at 45°C for 12 h. 5. Incubation of the cereal malts at 45°C for 12 h. 6. Testing the aliquots for starch at each time interval. 7. Centrifugation at 5000 rpm. 8. Washing the residue and drying with graded ethanol (50%, 70% and 90%) and ethanol. 9. Quantification of bound phenolic acids by standard procedure. The following examples are given by way of illustration of the present invention only and should not be construed to limit the scope of invention. Example 1 Rice malt (1 g) was dispersed in 10 ml of 0.05 M acetate buffer (pH 5.0) and kept in an oven at 45°C for 12 h. The temperature of the oven is raised by 10°C for every 12 hours (55°C - 12 h, 65°C - 12 h and 75°C - 12 h) and aliquots (1 ml) were withdrawn every 12 h and tested for the presence of starch by iodine reagent. At 65°C after 12 h (overall 36 h) of incubation the resultant malt gave a negative colour for starch indicating the action of in situ amylases of malt on the starch, producing dextrins. 8 Example 2 Maize malt (1 g) was dispersed in 10 ml of 0.05 M acetate buffer (pH 5.0) and kept in an oven at 45°C for 12 h. The temperature of the oven is raised by 10°C for every 12 hours (55°C - 12 h, 65°C - 12 h and 75°C - 12 h) and aliquots (1 ml) were withdrawn every 12 h and tested for the presence of starch by iodine reagent. At 65°C after 12 h (overall 36 h) of incubation the resultant malt gave a negative colour for starch indicating the action of in situ amylases of malt on the starch, producing dextrins. Example 3 Wheat malt (1 g) was dispersed in 10 ml of 0.05 M acetate buffer (pH 5.0) and kept in an oven at 45°C for 12 h. The temperature of the oven is raised by 10°C for every 12 hours (55°C - 12 h, 65°C - 12 h and 75°C - 12 h) and aliquots (1 ml) were withdrawn every 12 h and tested for the presence of starch by iodine reagent. At 65°C after 12 h (overall 36 h) of incubation the resultant malt gave a negative colour for starch indicating the action of in situ amylases of malt on the starch, producing dextrins. Example 4 Ragi malt (1 g) was dispersed in 10 ml of 0.05 M acetate buffer (pH 5.0) and kept in an oven at 45°C for 12 h. The temperature of the oven is raised by 10°C for every 12 hours (55°C - 12 h, 65°C - 12 h and 75°C - 12 h) and aliquots (1 ml) were withdrawn every 12 h and tested for the presence of starch by iodine reagent. At 75°C after 4 h (overall 40 h) of incubation the resultant malt gave a negative colour for starch indicating the action of in situ amylases of malt on the starch, producing dextrins. The main advantages of the present invention are: 1. The main advantage of the present invention the use of in situ amylases to isolate high contents of bound-phenolic acid rich dietary fibre from cereal malts. 2. Another advantage of the present invention is that is faster than the conventional processes. 3. Yet another advantage of the present invention is that it helps increase the nutrient value of the cereal malts. 4. Still another advantage of the present invention is that it does not involve high temperature to obtain bound-phenolic acid rich dietary fibre from cereal malts as seen in conventional methods. 5. Still another advantage of the present invention is that instant process is economical than the conventional processes, which involve use of commercially available bacterial and fungal enzymes. 10 6. Still another advantage of the present invention is that it involves shorter incubation periods as compared to conventional processes. This helps obtained higher nutritious cereal malt in much shorter duration. 7. Retaining of all the bound phenolic acids present in the cereal malts unlike in the case of endosperm and bran where in only partial recovery of bound phenolic acid is achieved. We Claim: 1. An improved process of obtaining high contents of bound-phenolic acid rich dietary fibre, the said process comprising steps of: (a) adding about 7-15 gm% (w/v) of cereal malt ranges from 40-60 mM of acetate buffer of pH ranging between 4.5-5.5 to obtain malt solution, (b) incubating malt solution at a temperature of 45°C for a period of 10-14 hours, (c) incubating malt solution of step (b) further for a period of 10-14 hours at a temperature of 55°C, (d) incubating malt solution of step (c) further for a period of 10-14 hours at a temperature of 65°C, (e) incubating malt solution of step (d) further for a period of 10-14 hours at a temperature of 75°C to obtain a residue, (f) measuring starch levels after the each incubation, (g) centrifuging the said residue of step (e), (h) washing the centrifuged residue, (i) drying the washed residue with ethanol, (j) measuring levels of bound-phenolic acid in the dried residue by conventional processes, and (k) obtaining bound-phenolic acid rich dietary fibres. 2. An improved process as claimed in claim 1, wherein concentration of cereal malt in acetate buffer is preferably 10gm %(w/v). 3. An improved process as claimed in claim 1, wherein pH of the acetate buffer is preferably 5.0. 4. An improved process as claimed in claim 1, wherein concentration of the acetate buffer is preferably 50mM. 5. An improved process as claimed in claim 1, wherein the cereal malts are selected from a group comprising rice malt, maize malt, wheat malt, and ragi malt. 6. An improved process as claimed in claim 1, wherein said process eliminates starch from cereal malts comprising rice malt, maize malt, and wheat malt for a period of 21-39 hours. 7. An improved process as claimed in claim 1, wherein said process eliminates starch at a maximum at a temperature of 65°C from cereal malts comprising rice malt, maize malt, and wheat malt. 8. An improved process as claimed in claim 1, wherein said process eliminates starch from ragi malt within about 35-45 hours. 9. An improved process as claimed in claim 1, wherein said process eliminates starch at a maximum of about 75°C from ragi malt. 10. An improved process as claimed in claim 1, wherein centrifuging the residue at 5000 rpm for time period ranging between 1-30 minutes. 11. An improved process as claimed in claim 1, wherein the concentration of ethanol is ranging between 40 to 100%. 12. An improved process as claimed in claim 1, wherein the stepwise increase in temperature helps overcome the need of using exogenous enzymes. 13. An improved process as claimed in claim 1, wherein incubating the said mixture in an oven. 14. An improved process as claimed in claim 1, wherein washing the centrifuged residue in water. 15. An improved process of obtaining high contents of bound-phenolic acid rich dietary fibre substantially as herein described with reference to the examples accompanying this specification.

Documents:

415-del-2002-Abstract-(26-02-2010).pdf

415-del-2002-abstract.pdf

415-del-2002-Claims-(26-02-2010).pdf

415-del-2002-claims.pdf

415-del-2002-Correspondence-Others-(26-02-2010).pdf

415-del-2002-correspondence-others.pdf

415-del-2002-Description (Complete)-(26-02-2010).pdf

415-del-2002-description (complete).pdf

415-del-2002-Form-1-(26-02-2010).pdf

415-del-2002-form-1.pdf

415-del-2002-form-18.pdf

415-del-2002-Form-2-(26-02-2010).pdf

415-del-2002-form-2.pdf

415-del-2002-Form-3-(26-02-2010).pdf

415-del-2002-form-3.pdf

415-del-2002-form-5.pdf

415-del-2002-Petition 137-(26-02-2010).pdf