KR101489601B1 - Method of Manufacturing Soluble β-Glucan with High Extraction Yield and Purity from Barley - Google Patents

Abstract

The present invention relates to a method for producing high purity and high yield water-soluble beta-glucan from barley. In the method of the present invention, barely powder is heated with hydrogen peroxide, so that the extraction yield is increased and a beta-glucanase is simultaneously inactivated. To prevent the additional decomposition of beta-glucan caused by hydrogen peroxide, catalase and phytic acid are added as hydrogen peroxide inhibitors. To remove residual polysaccharides and oligo saccharides, an alpha-amylase treatment is performed. By means of the method of the present invention, high purity and high yield water-soluble beta-glucan can be produced.

Description

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for producing high-purity beta-glucan from barley,

The present invention relates to a process for preparing high-purity beta-glucan from barley.

(1 → 3) and (1 → 4) -β-D-glucan (hereinafter referred to as "β-glucan") refers to a chain of non-ionic polysaccharides having β- (linear non-starch polysaccharide), which is distinguished from starch which is? -glucan. In the case of barley beta -glucan, it exists in a water-soluble or insoluble form. Unlike cellulose, which is composed of only β- (1 → 4) -conjugation, beta-glucan has an irregular structure of β- (1 → 3) Resulting in partial water solubility and more susceptibility to hydrolysis. Barley and bovine barley contain about 2-8% of total beta-glucan, of which the content of water-soluble beta-glucan is less than about 50%.

Extraction of beta-glucan is carried out by solid-liquid extraction, and acid or alkali is used as a solvent, but water extraction is the most common, and beta-glucan is swollen and gradually dissolved in an aqueous solution, Temperature, moisture content, moisture penetration and obstacles to the diffusion of dissolved materials. In addition, cereal beta-glucan is complex and tightly bound with starch, protein, and pentosan in the cell wall of barley and oat. Therefore, the extraction rate varies greatly depending on the extraction conditions and methods, and the characteristics of extracted beta-glucan It is different. As the extraction temperature increases, the amount of beta-glucan increases with increasing acidity and alkalinity. Depending on the extraction temperature, the 1 → 4 binding ratio, the size of the 1 → 4 binding block and the frequency of 1 → 3 binding are different .

The water soluble beta-glucan in barley has attracted attention as one of the representative water-soluble dietary fibers due to its known physiological functions such as lowering of cholesterol content in blood and improvement of blood lipid composition, and has better physiological activity of water-soluble beta-glucan than insoluble beta-glucan Are reported. However, the content of barley-soluble beta-glucan is relatively small, the molecular weight is high and the viscosity of the aqueous solution is high as compared with other grains. This hinders filtration in the production of beer using barley and lowers the digestibility of the barley to limit its use as a livestock feed. There is a limitation in industrial use such as formation of an undesirable precipitate. In order to solve these various problems and to control the molecular weight, studies are being carried out to lower molecular weight through physical and chemical methods. These low molecular weight methods can effectively reduce the molecular weight but can cause problems such as high treatment cost, low yield and long processing time, environmental pollution due to use of strong acid / base solution.

To overcome these disadvantages, researches have recently been conducted to lower molecular weight polysaccharides using oxidizing agents such as hydrogen peroxide, nitrite and ozone. The low molecular weight method by the oxidizing agent is a method in which the oxidizing agent generates a large amount of radicals by an external factor such as heat or an initiator, and the generated radical decomposes the molecular bonds of the polysaccharide into small molecules. Especially, in the case of hydrogen peroxide, the Gibbs free energy can be spontaneously decomposed to a negative value, so that an environmentally friendly treatment is possible. In addition, when the polysaccharides were oxidized and converted to low molecular weight, a large amount of treatment was possible in a shorter time than the conventional chemical method, and the yield of the product was higher than that of the other methods. However, studies on the oxidative low molecular weight of beta - glucan have been mainly focused on inhibiting the decrease of beta - glucan content by spontaneous oxidative low molecular weight in the liquid form beta - glucan product.

In order to increase the extraction rate of β-glucan by controlling the extraction conditions, it may be possible to control by the extraction temperature, the pH of the solvent, the type of solvent, the ionic strength of the solvent, the addition ratio of the solvent and the extraction time, The effects of temperature, pH, extraction temperature and pH have been investigated. However, studies on the content and purity of beta - glucan by oxidizing agent alone or in combination with oxidizing agent and heat treatment are insufficient. No study on condition optimization has been attempted.

In order to increase the yield of soluble beta-glucan from barley in the method of producing water-soluble beta-glucan from barley gills and the use thereof (Patent Publication No. 1995-0002867) in the prior art for increasing the extraction yield of beta-glucan from cereals However, the yield of β-glucan was low, and the ethanol treatment process was repeated, resulting in an increase in cost and time. In the method for producing a water-soluble dietary fiber (Patent Publication No. 1997-0064406), β-glucanase inactivating, liquefaction enzyme and glucose saccharifying enzyme treatment for producing high purity and high yield water-soluble beta-glucan from barley There were many unit processes to produce beta-glucan. In the method for extracting cereal beta-glucan (Patent Publication No. 1999-0065617), there is a process of finely grinding cereal grains and heat-treating beta-glucanase in order to increase the yield of beta-glucan . In order to promote the digestion and absorption of beta-glucan, a number of patents have been introduced in a method for producing low-molecular-weight beta-glucans (Patent Publication No. 2012-0021891), such as hydrogen peroxide and low molecular weight treatment of extracted beta-glucan .

Numerous papers and patent documents are referenced and cited throughout this specification. The disclosures of the cited papers and patent documents are incorporated herein by reference in their entirety to better understand the state of the art to which the present invention pertains and the content of the present invention.

Korean Patent Publication No. 1995-0002867 Korean Patent Publication No. 1997-0064406 Korean Patent Publication No. 1999-0065617 Korean Patent Publication No. 2012-0021891

Methods for increasing the extraction yield and purity of beta-glucan from conventional grains include a first step of removing microparticles or an embryo part to produce a fraction having a high beta-glucan content; A second step of heating to inactivate beta-glucanase, an enzyme that degrades beta-glucan; A third step of separating the solid from the solution by centrifugation and separation; A fourth step of decomposing the protein with a protease; A fifth step of decomposing the polysaccharide or oligosaccharide into monosaccharides with a starch hydrolyzing enzyme; A sixth step of separating the beta-glucan by precipitation and centrifugation; A seventh step of crystallizing beta -glucan in a separated aqueous solution by using alcohol or alcohol; And the eighth step of drying and pulverizing the crystallized beta-glucan. However, it is difficult to change the structure so that the beta-glucan can be easily extracted from the cereal. In order to solve this problem, it is necessary to further carry out the micro-pulverization process. Since the degradation efficiency of the protein is low, Is low. In addition, the inactivation of beta-glucanase and the heating process necessary for application of the protein hydrolyzing enzyme are repeatedly performed, and the process of crystallization of beta-glucan by alcohol is repeatedly performed, thereby increasing processing cost and time.

In order to overcome the disadvantages of the conventional process as described above, the inventors of the present invention have found that by increasing the extraction yield and deactivation of beta-glucanase simultaneously with hydrogen peroxide and heat treatment of the barley powder, Glucan was prepared by adding catalase and phytic acid in order to prevent degradation of the polysaccharide and to remove the residual polysaccharide and oligosaccharide by α-amylase treatment, thereby producing high purity and high yield of water-soluble beta-glucan The present invention has been completed.

Accordingly, it is an object of the present invention to provide a method for producing water-soluble beta-glucans of high purity and yield from barley.

The objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

According to one aspect of the present invention, the present invention provides a method for preparing beta-glucan from barley with high yield and high purity, comprising:

(a) a pulverizing step of pulverizing barley to obtain a barley powder;

(b) pre-processing step for homogenizing the powder mixture by mixing barley was added to ferrous sulfate (FeSO ₄₎ and distilled water to the obtained powder and barley;

(c) a step of adding heat to the homogenized barley powder suspension after adding hydrogen peroxide to the homogenized barley powder suspension;

(d) quenching the heat-treated barley powder suspension to room temperature and adding catalase and phytic acid;

(e) a starch hydrolysis step of treating the reaction solution which has not been reacted with the calcium chloride and the heat-resistant alpha-amylase to decompose the residual polysaccharide and oligosaccharide into monosaccharides;

(f) a precipitation step of centrifuging the hydrolyzed reaction solution to obtain a supernatant and adding alcohol thereto to crystallize beta-glucan; And

(g) a pulverizing step of separating, drying and pulverizing the crystallized and precipitated beta -glucan.

Hereinafter, the present invention will be described in detail with reference to the respective steps.

Step (a): a pulverizing step of pulverizing barley to obtain barley powder

Uncooked barley is pulverized into barley powder. The method of grinding barley is not particularly limited, but a barley grinder or the like can be used, and the size of barley powder to be pulverized is preferably 60-80 mesh.

Step (b): a pretreatment step of homogenizing the barley powder solution by adding ferrous sulfate (FeSO ₄ ) and distilled water to the obtained barley powder and mixing

Ferrous sulfate (FeSO ₄ ) and distilled water are added to the obtained barley powder and mixed to homogenize the barley powder solution. The ferrous sulfate (FeSO ₄ ) acts as a catalyst for the beta-glucan decomposition reaction by the Fenton oxidation reaction involving the following hydrogen peroxide. The concentration of the added ferrous sulfate (FeSO ₄ ) is not particularly limited, but is preferably added so as to have a concentration of 0.01-0.5 mM.

Step (c): a step of adding a hydrogen peroxide to the homogenized barley powder suspension followed by heating

Hydrogen peroxide is added to the homogenized barley powder suspension, followed by heat treatment. By simultaneously performing the hydrogen peroxide treatment and the heat treatment, the beta-glucan contained in the barley is decomposed and the beta-glucanase is inactivated. By treatment with hydrogen peroxide, beta-glucan can be separated and purified from barley at a high yield. The hydrogen peroxide is added so as to have a concentration of 0.2-0.8% (v / v), more preferably 0.509-0.514% (v / v). The heating is preferably carried out at a temperature of 90-150 DEG C for 5-35 minutes, more preferably at a temperature of 115-117 DEG C for 11-24 minutes.

Step (d): After the heat-treated barley powder suspension is cooled to room temperature, a reaction stopping step of adding catalase and phytic acid

After the hydrogen peroxide is added and the heat treated barley powder suspension is cooled to room temperature, catalase and phytic acid are added. The catalase and phytic acid decompose the hydrogen peroxide to stop further decomposition of the beta-glucan by the subsequent hydrogen peroxide. Preferably, the catalase is added to a concentration of 0.1-3 kU / mL, and the phytic acid is added to a concentration of 0.25-5 mM.

Step (e): a starch hydrolysis step of decomposing residual polysaccharides and oligosaccharides into monosaccharides by treating calcium chloride and a heat-resistant alpha-amylase to the quiescent reaction solution

The reaction solution in which the reaction is stopped is treated with calcium chloride and alpha-amylase to decompose the residual polysaccharide and oligosaccharide into monosaccharides. Preferably, the calcium chloride is added at a concentration of 6-8 mg / 100 mL, and the heat-resistant alpha-amylase is added at a concentration of 0.1-0.3 kU / 100 mL.

Step (f): The hydrolyzed reaction solution is centrifuged to obtain a supernatant, and alcohol is added thereto to precipitate beta-glucan

The hydrolyzed reaction solution is centrifuged to obtain a supernatant, and alcohol is added thereto to crystallize and precipitate the beta-glucan. The alcohol to be added is preferably a lower alcohol having from 1 to 4 carbon atoms, and most preferably ethanol. The alcohol is preferably added in an alcohol having 97-99% purity to a final concentration of 75-85%.

Step (g): a powdering step of separating, drying and pulverizing the crystallized and precipitated beta -glucan

The crystallized and precipitated beta -glucan is separated, dried and pulverized to powder. The drying is performed by vacuum drying at 55-65 ° C. According to one embodiment of the present invention, the finally obtained content of beta-glucan is 5.25-6.43 g per 100 g, the purity is 74.24-74.77%, and the solubility index is 92.63-94.78%.

The present invention relates to a process for producing high purity and high yield of water-soluble beta-glucans from barley. In the method of the present invention, in order to simultaneously increase the extraction yield and deactivate the beta-glucanase by treating the barley powder with hydrogen peroxide and heating, and to prevent further decomposition of beta-glucan by hydrogen peroxide, catalase And phytic acid are added, and alpha-amylase treatment is performed to remove residual polysaccharides and oligosaccharides. According to the method of the present invention, water-soluble beta-glucan can be produced with high purity and high yield.

FIG. 1 shows a typical property change according to the concentration of hydrogen peroxide and the heat treatment conditions in the heat treatment treatment process by adding hydrogen peroxide to the barley powder suspension.
Fig. 2 shows the reaction surface graph and contour graph of the total beta-glucan content of barley according to the treatment with hydrogen peroxide and heating.
FIG. 3 shows a reaction surface graph and a contour graph for the content of barley-soluble beta-glucan (.beta.-glucan) according to the treatment with hydrogen peroxide and heating.
FIG. 4 shows a reaction surface graph and a contour line graph of the ratio of the total beta-glucan content of the barley water-soluble beta-glucan according to the treatment with hydrogen peroxide and heating.
Fig. 5 is a process diagram showing the production process of total beta-glucan and water-soluble beta-glucan in barley by the combination treatment with hydrogen peroxide and heating.

Hereinafter, the present invention will be described in more detail with reference to Examples. It is to be understood by those skilled in the art that these embodiments are only for describing the present invention in more detail and that the scope of the present invention is not limited by these embodiments in accordance with the gist of the present invention .

Example

Example 1: Experimental design for optimization of extraction conditions of water-soluble beta-glucan

The barley harvested in 2011 was harvested from the National Institute of Agricultural Science and Technology, Rural Development Administration, and crushed to 60-80 mesh (Micro hammer cutter mill type-3, Culatti AG, Zurich, Swiss) And used as a sample. In this example, the central composite design (CCD) was used for the experimental design and the response surface methodology (RSM) was used to optimize the treatment conditions. In the central synthesis program, hydrogen peroxide concentration (0.2-0.8%, v / v; X ₁ ), heat treatment temperature (90-150 ° C, X ₂ ) and heat treatment time (5-35 min, X ₃ ) 0, and 1, and processed into 16 sections according to the central synthesis plan (Table 1). Further, as the dependent variable (Y _n) a total beta-was in the content of the glucan (Y _CSB), purity (Y _PSB) and ratio (Y _RSB) - the content of the glucan (Y _CTB) and purity (Y _PTB), water soluble beta , All tests were repeated three times and mean values were used for regression analysis.

Example 2: Combined treatment with hydrogen peroxide

5 g of barley powder, 0.1 mM of FeSO ₄ .7H ₂ O and 100 mL of distilled water were added to the heat treatment vessel, and the pH was adjusted to 4.8 ± 0.2 using 0.1 N NaOH and HCl. Hydrogen peroxide was added to the barley suspension and heat treated in a preheated oil bath. The hydrogen peroxide concentration, heat treatment temperature and time were treated by central synthetic scheme (Table 1). After the heat treatment was completed, the reaction mixture was rapidly cooled to room temperature. To suppress the Fenton-oxidation reaction, 50 μL of a catalase solution and 1.34 mL of a phytic acid solution were added and reacted at 25 ° C. for 1 hour to remove residual hydrogen peroxide The resulting hydroxy radical was removed. The changes in properties of barley powder according to hydrogen peroxide and heat treatment were observed, and representative results thereof are shown in Fig. (B) in which ferrous sulfate (FeSO ₄ ), which acts as a catalyst for catalyzing the decomposition reaction (Fenton-oxidation reaction) of organic compounds by hydrogen peroxide, differs from the untreated treatment (A) in which distilled water is added to barley powder (FeSO ₄ ) and hydrogen peroxide (C), bubbles were formed with color change. In the treatment (DF) treated with ferrous sulfate (FeSO ₄ ) and hydrogen peroxide at the concentration of 120 캜 for 35 min, the degree of browning increased with increasing hydrogen peroxide concentration.

Example 3: Extraction and purification of total beta-glucan and water-soluble beta-glucan

The total beta-glucan was extracted from the barley solution with hydrogen peroxide and heating. The volume was adjusted to 200 mL with distilled water, 1N NaOH was added, and the mixture was stirred for 1 hour. The mixture was centrifuged (3,500 rpm, 15 min) Respectively. The pH of the supernatant was adjusted to pH 6.5 by addition of 1N HCl and 7 mg of calcium chloride (CaCl ₂ ) and α-amylase (A3306, Sigma-Aldrich Co. LLC, MO, USA) 0.1 mL was added to the solution, which was then subjected to enzyme treatment in a water bath set at 95 ° C for 1 hour. After the enzyme treatment was completed, the solution was cooled to room temperature, adjusted to pH 4.5 with 1N HCl, centrifuged, and the supernatant was vigorously shaken with 99% ethanol so that the concentration of ethanol became 80% For 30 minutes to precipitate beta-glucan, which was centrifuged at 3,500 rpm for 30 minutes to obtain total beta-glucan.

The water-soluble beta-glucan was extracted by adding distilled water to the solution of hydrogen peroxide and heated barley, adjusted to a volume of 200 mL, and then extracted for 1 hour in a water bath set at 65 ° C. Then, CaCl2 and heat resistant alpha-amylase 7 mg and 0.1 mL were added, followed by enzymatic treatment for 1 hour in a water bath set at 95 ° C. After completion of the enzyme treatment, the mixture was cooled to room temperature, centrifuged, and 99% ethanol was added to the supernatant so that the concentration of ethanol became 80%. The mixture was shaken vigorously and allowed to stand in a refrigerator at 4 ° C for 12 hours to precipitate beta- This was centrifuged at 3,500 rpm for 30 minutes to obtain water-soluble beta-glucan.

The purity of extracted total beta-glucan and soluble beta-glucan was measured using a beta -D-glucan assay kit. That is, 0.2 mL of 50% ethanol and 4.0 mL of 20 mM sodium phosphate buffer (pH 6.5) were sequentially added to about 100 mg of dry-pulverized beta-glucan, stirred for 5 minutes, and left in boiling water for 3 minutes Then, 0.2 mL of lichanase (10 U) was added and the enzyme was treated at 50 ° C for 1 hour. To the enzyme-treated solution, 5.0 mL of 200 mM sodium acetate buffer (pH 4.0) was added to terminate the reaction, followed by centrifugation at 4,000 rpm for 10 minutes to obtain a supernatant. Add 0.1 mL of the supernatant to 3 test tubes, add 0.1 mL of 50 mM sodium acetate buffer (pH 6.0) to one test tube, add β-glucosidase (β -glucosidase (10 U) was added to the cells and the cells were incubated at 50 ° C for 10 minutes. 3.0 mL of GOPOD (glucose oxidase / peroxidase) was added to the culture and incubated at 50 ° C for 20 minutes. Then, the purity of beta-glucan was calculated by measuring the absorbance at 510 nm (spectrophotometer UV-1650 PC, Shimadzu, Tokyo, Japan).

The results of measuring the content of total beta-glucan in the barley extract according to the extraction conditions are shown in Table 2 and FIG. (P <0.05), which was 8.43-8.47% in the treatment condition (No. 8, 9) treated with hydrogen peroxide concentration of 0.5%, heat treatment temperature of 120 ℃ and heat treatment time of 20 minutes. On the other hand, total beta-glucan content decreased with 0.8% and 150 ℃ treatment temperature, and decreased to 1.23% at 35 ℃ treatment (No. 16) treatment with high concentration of hydrogen peroxide and heat treatment temperature. As a result of the analysis of the content of total beta-glucan, the concentration of hydrogen peroxide was significantly influenced by the concentration of F-value (102.08 ^*** ), the heat treatment temperature (66.03 ^*** ) and the time (13.38 ^{* **} ) also showed a great effect. The coefficient of determination (R ² ) of the regression equation through the response surface analysis was as high as 0.943, indicating that the model equation is suitable (Table 4). The reaction surface results were estimated to be 0.463% of hydrogen peroxide concentration, 7.54% at heat treatment temperature of 120 ℃ and 21 minutes of heat treatment, and the experimental value was 7.83% (p> 0.05).

The content of water-soluble beta-glucan in the barley extract according to the extraction conditions is shown in Table 2 and FIG. (P <0.05), which was 6.45-6.52% in the treatment condition (No. 8, 9) treated with hydrogen peroxide concentration of 0.5%, heat treatment temperature of 120 ℃ and heat treatment time of 20 minutes. On the other hand, total beta-glucan content decreased with 0.8% and 150 ℃ treatment temperature and 0.68% at 35 min treatment (No. 16). As a result of analysis of the content of water-soluble beta-glucan, the concentration of hydrogen peroxide was significantly influenced by the concentration of hydrogen peroxide (22.76 ^*** ) and the heat treatment time (11.16 ^*** ) 1.39 ^NS ) showed no effect. The coefficient of determination (R2) of the regression equation through the response surface analysis was 0.827, suggesting that the model equation is appropriate (Table 4). The reaction surface results were estimated to be 6.32% in the hydrogen peroxide concentration of 0.514%, the heat treatment temperature of 115 ℃ and the heat treatment time of 24 minutes, and the experimental value was 6.43%, which was not significant (p> 0.05).

Table 2 shows the results of measuring the total beta-glucan purity of the barley extract according to the extraction conditions. The highest purity (95%) was obtained in the treatment conditions (No. 15, 16) at the treatment temperature of 150 ℃ and the heat treatment time of 5 and 35 minutes, respectively. On the other hand, when treated with low hydrogen peroxide concentration and heat treatment temperature, the purity of total beta-glucan tended to decrease, and the lowest purity was 0.5%, 90 ° C and 38.24% at 5 minutes treatment (No. 1). As a result of the analysis of the purity of total beta-glucan, the concentration of hydrogen peroxide was significantly influenced by the concentration of hydrogen peroxide as shown in Table 3 (597.20 ^*** ), the heat treatment temperature (161.50 ^*** ) and the time (22.26 ^*** ) It was found that the effect was significant. The coefficient of determination (R ² ) of the regression equation through the response surface analysis was high as 0.987, indicating that the model equation is suitable (Table 4). At this time, the reaction surface results were estimated to be 98.68% in the hydrogen peroxide concentration of 1.146%, the heat treatment temperature of 190 ° C. and the heat treatment time of 20 minutes, and the experimental value was 98.59%, which was not significant (p> 0.05).

Table 2 shows the results of measuring the purity of the water-soluble beta-glucan in the barley extract according to the extraction conditions. (P <0.05), which was 98.26% and 98.01%, respectively, under conditions (No. 15, 16) treated with hydrogen peroxide concentration of 0.8%, heat treatment temperature of 150 ℃ and heat treatment time of 5 minutes and 35 minutes, respectively. On the other hand, when treated with low hydrogen peroxide concentration and heat treatment temperature, the purity of total beta-glucan tended to decrease, and the lowest purity was 0.5%, 90 ° C, and 41.48% at 5 minutes treatment (No. 1). As a result of the analysis of the purity of the water-soluble beta-glucan, the influence of the hydrogen peroxide concentration (182.22 ^*** ) and the heat treatment temperature (49.95 ^*** ) was greatly affected and the heat treatment time (3.16 ^* appear. The coefficient of determination (R ² ) of the regression equation through the response surface analysis was as high as 0.962, indicating that the model equation is suitable (Table 4). At this time, the reaction surface result was predicted to be 97.28 at the hydrogen peroxide concentration of 0.804%, the heat treatment temperature of 175 ° C and the heat treatment time of 20 minutes, and the experimental value was 96.59%, which was not significant (p> 0.05).

The solubility index (ratios of soluble β-glucan, RSB) to the total β-glucan content of the barley extracts according to the extraction conditions was shown in Table 2 and FIG. (P <0.05), which was 91.28-91.56% in the condition (No. 8, 9) treated with hydrogen peroxide concentration of 0.5%, heat treatment temperature of 120 ℃ and heat treatment time of 20 minutes. As a result of analysis of variance of solubility index, the concentration of hydrogen peroxide was significantly influenced by the concentration of hydrogen peroxide as shown in Table 3 (15.88 ^*** ), and also by the heat treatment temperature (6.03 ^*** ) and time (5.88 ^*** ) appear. The coefficient of determination (R ² ) of the regression equation through the response surface analysis was 0.812, suggesting that the model equation is suitable (Table 4). At this time, the reaction surface result was estimated to be 94.24% at the hydrogen peroxide concentration of 0.509%, the heat treatment temperature of 117 ° C and the heat treatment time of 11 minutes, and the experimental value was 94.78%, which was not significant (p> 0.05).

Example 4: Low-molecular characterization of water-soluble beta-glucan

The molecular weight distribution of water-soluble beta-glucan extracted from hydrogen peroxide and heated barley was determined by HPLC (Acme 9000 system, Younglin Ins., Anyang, Korea) on YMC Diol-300 size exclusion column (300 ㅧ 4.6 mm ID, (RI 750F, Younglin Ins., Anyang, Korea) with a flow rate of 0.5 ml / min, and a mobile phase HPLC grade water was loaded with a pore size 30 nm, YMC Co. Ltd., Kyoto, Japan. The molecular weight of the standard substance was 708,000 for P-800, 375,000 for P-400, 200,000 for P-200, and 200,000 for P-200. Pullulan standard kit (P-82, Showa-Denko, Tokyo, Japan) -100 was 107,000, P-50 was 47,100, P-10 was 9,600, and P-5 was 5,900. The molecular weight of the aqueous beta-glucan treated with hydrogen peroxide and heating was measured by substituting the elution time of the chromatogram analyzed under the same conditions after obtaining the regression equation from the standard chromatogram. The apparent viscosity of the water-soluble beta-glucan was determined by adding the water-soluble beta-glucan dry powder to distilled water at a concentration of 2.0% (w / v), heating the mixture at 95 ° C for 30 minutes, homogenizing, and then measuring the viscosity with a viscometer (RVT DV -II, Brookfield Co., Middleboro, MA, USA). No.02 was used for the spindle of the viscometer and it was measured three times at 25 rpm at 100 rpm. The re-dissolution rate of water-soluble β-glucan was prepared by using distilled water at a concentration of 1.0% (w / v), and then cooled and centrifuged (12,000 rpm, 10 minutes) The solubility was measured by measuring the weight of the insoluble solids.

As shown in Table 5, the molecular weight distribution of the barley water-soluble beta-glucan treated with hydrogen peroxide and heating was measured. As shown in Table 5, the molecular weight tended to decrease with increasing hydrogen peroxide concentration, heat treatment temperature and time. The molecular weight of No. 1 (0.2%, 90 ℃, 5 min) was 543063 Da, but decreased to 12460 Da at 16 (0.8%, 150 ℃, 35 min) As a result of measuring the viscosity change of the water-soluble β-glucan, the viscosity tended to decrease with increasing hydrogen peroxide concentration, heat treatment temperature and time, as shown in Table 5. The viscosity of 1% (0.2%, 90 ℃, 5 min) was 12.44 cP in the case of 2% water - soluble β - glucan aqueous solution but decreased to 1.27 cP at 16 (0.8%, 150 ℃, 35 min). As a result of measuring the change in the redissolution rate of water-soluble beta-glucan, the re-dissolution rate tended to increase with increasing hydrogen peroxide concentration, heat treatment temperature and time, as shown in Table 5. The redissolution rate of No. 1 (0.2%, 90 ℃, 5 min) was 46.30%, but it increased to 92.01% at 16 (0.8%, 150 ℃, 35 min)

Example 5 Prediction and Test of Optimum Extraction Conditions of Water-Soluble Beta-Glucan

Glucan content and purity, soluble beta - glucan content and purity and solubility index of extracts in order to optimize the extraction conditions to increase the yield and purity of water - soluble beta - glucan from barley through hydrogen peroxide and heat treatment. (Table 4). The results are shown in Table 4. In addition, the molecular weight, viscosity and redissolution rate of water-soluble beta-glucan were measured under optimum extraction conditions for each reaction variable. Although the purity of water - soluble beta - glucan could be greatly improved at high hydrogen peroxide concentration and heat treatment temperature, beta - glucan degradation proceeded at the same time, resulting in lower yield and lower molecular weight and viscosity. Therefore, the optimal extraction conditions for increasing the yield and purity of water-soluble beta-glucan are determined to be hydrogen peroxide concentration of 0.509-0.514%, heat treatment temperature of 115-117 ° C, and heat treatment time of 11-24 minutes, The purity was found to be 74.24-74.77% and the solubility index 92.63-94.78% for each 100 g of barley.

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the same is by way of illustration and example only and is not to be construed as limiting the scope of the present invention. Accordingly, the actual scope of the present invention will be defined by the appended claims and their equivalents.

Claims (8)

A process for producing beta-glucan in high yield and high purity from barley comprising the steps of:
(a) a pulverizing step of pulverizing barley to obtain a barley powder;
(b) pre-processing step for homogenizing the powder mixture by mixing barley was added to ferrous sulfate (FeSO ₄₎ and distilled water to the obtained powder and barley;
(c) a step of adding heat to the homogenized barley powder suspension after adding hydrogen peroxide to the homogenized barley powder suspension;
(d) quenching the heat-treated barley powder suspension to room temperature and adding catalase and phytic acid;
(e) a starch hydrolysis step of treating the reaction solution which has not been reacted with the calcium chloride and the heat-resistant alpha-amylase to decompose the residual polysaccharide and oligosaccharide into monosaccharides;
(f) a precipitation step of centrifuging the hydrolyzed reaction solution to obtain a supernatant and adding alcohol thereto to crystallize beta-glucan; And
(g) a pulverizing step of separating, drying and pulverizing the crystallized and precipitated beta -glucan.
The method according to claim 1, wherein the ferrous sulfate (FeSO ₄ ) is added in a concentration of 0.01-0.5 mM in the pretreatment step (b).
The method according to claim 1, wherein the hydrogen peroxide is added so as to have a concentration of 0.2-0.8% (v / v) in the step (c).
The method according to claim 1, wherein the heating step (c) is performed by heating at 90-150 캜 for 5-35 minutes.
2. The method according to claim 1, wherein the catalase is added in a concentration of 0.1-3 kU / mL in the reaction termination step (d), and the phytic acid is added in a concentration of 0.25-5 mM Lt; / RTI &gt;
The method according to claim 1, wherein the starch hydrolysis step (e) comprises adding the calcium chloride at a concentration of 6-8 mg / 100 mL and the heat-resistant alpha-amylase at a concentration of 0.1-0.3 kU / 100 mL Lt; / RTI &gt;
The method according to claim 1, wherein in the precipitation step (f), the alcohol is added to a final concentration of 75-85%.
The method according to claim 1, wherein the crystallized and precipitated beta -glucan in the pulverizing step (g) is vacuum-dried at 55-65 DEG C, pulverized and pulverized.

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