CN113519685B - Method for separating acid-soluble glutenin from wheat gluten by urea crystallization method - Google Patents
Method for separating acid-soluble glutenin from wheat gluten by urea crystallization method Download PDFInfo
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- CN113519685B CN113519685B CN202110702441.6A CN202110702441A CN113519685B CN 113519685 B CN113519685 B CN 113519685B CN 202110702441 A CN202110702441 A CN 202110702441A CN 113519685 B CN113519685 B CN 113519685B
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23J—PROTEIN COMPOSITIONS FOR FOODSTUFFS; WORKING-UP PROTEINS FOR FOODSTUFFS; PHOSPHATIDE COMPOSITIONS FOR FOODSTUFFS
- A23J1/00—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites
- A23J1/12—Obtaining protein compositions for foodstuffs; Bulk opening of eggs and separation of yolks from whites from cereals, wheat, bran, or molasses
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Abstract
The method for separating acid-soluble glutenin from wheat gluten by the urea crystallization method utilizes the characteristic that an acid-containing urea solution can disperse glutenin aggregates, and separates urea and glutenin in the solution by combining the characteristic that high-concentration urea is easy to crystallize. Wheat gluten obtained after alcohol extraction of wheat gluten is used as a raw material, urea solution with the pH of 2-5 and the concentration of 2-6 mol/L is used for stirring and dissolving the wheat gluten, and the weight ratio of the protein to the urea solution is 1: 2-1: 6. And concentrating the volume of the urea solution by a rotary evaporator to be half of the volume of the original solution. Standing the concentrated solution at the temperature of 2-10 ℃ for 12-48 h, and standing at the temperature of 15-30 ℃ for 18-36h to separate out urea crystals and protein. Taking out the solid urea at the upper part, and remaining the semi-solid acid-soluble glutenin at the lower part. Dialyzing to remove a small amount of acetic acid and urea, drying and crushing to obtain powdery glutenin.
Description
Technical Field
The invention relates to the field of food processing, in particular to a method for separating acid-soluble glutenin from wheat gluten by a urea crystallization method.
Background
The first method for extracting glutenin from wheat gluten is to extract wheat gluten with about 65% ethanol, dissolve prolamine in ethanol, and obtain the residual precipitate as glutenin. However, the protein is an aggregate, is insoluble in water, and even if it has an anti-retrogradation function, it is difficult to apply the protein as an additive to a food dough. The existing glutenin separation and identification methods comprise polyacrylamide gel electrophoresis (SDS-PAGE), high Performance Capillary Electrophoresis (HPCE), biuret colorimetry, gel chromatography and the like, wherein extracting agents mainly comprise Sodium Dodecyl Sulfate (SDS), dithiothreitol (DTT) and urea, and alcohol soluble protein, albumin and other monomer proteins are removed by acetone, isopropanol containing beta-mercaptoethanol and the like. The glutenin is still dissolved in the extractant and cannot be precipitated out. SDS and DTT in the extracting agent used in the prior literature can not be applied in the field of food, and the safety of the material is relatively high when the urea is used in the preparation process of the food additive. The method is based on that urea containing acetic acid with certain concentration can partially dissolve glutenin aggregates prepared by extracting alcohol-soluble protein with gluten powder, then a large amount of urea is crystallized and separated out under the conditions of controlled temperature and urea concentration, and then all urea and acetic acid are removed by a dialysis bag, so that acid-soluble glutenin is prepared.
Disclosure of Invention
The invention aims to overcome the defects in the prior art, and the acid-soluble glutenin in the wheat gluten is separated by utilizing a urea crystallization method, expensive equipment is not needed in the method, alcohol-soluble protein is not needed to be precipitated by acetone and isopropanol, the glutenin aggregate is dissolved by utilizing acid-containing urea with higher food safety, then the urea and the protein are separated by utilizing the characteristic that the urea in concentrated solution is easy to crystallize, and the residual acetic acid and the urea of the protein are removed by dialysis to prepare the glutenin monomer which can be rapidly dispersed in dilute acid, thereby being convenient for the application in the food field.
The invention is realized by the following technical scheme:
a method for separating acid-soluble glutenin from wheat gluten by a urea crystallization method is characterized by comprising the following steps: the method comprises the following steps:
mixing wheat gluten and 65% ethanol according to the mass ratio of 1: 20, and stirring and leaching in a 35 ℃ water bath kettle for 2-3h to dissolve alcohol soluble protein in the wheat gluten into the ethanol; centrifuging the leached solution for 5min at 3500r/min with a centrifuge, and collecting precipitate to obtain glutenin aggregate. The glutenin aggregate is dissolved by stirring with a urea solution with pH of 2-5 and concentration of 2-6 mol/L, and the weight ratio of the protein to the urea solution is 1: 2-1: 6. Concentrating the supernatant with rotary evaporator at 65 deg.C under 0.09MPa to obtain concentrated urea solution with volume one half of that of the original solution. Standing the concentrated solution at the temperature of 2-10 ℃ for 12-48 h, and standing at the temperature of 15-30 ℃ for 18-36h to separate urea crystals from protein. Taking out the solid urea at the upper part, and remaining the semi-solid acid-soluble glutenin at the lower part. Putting semi-solid acid-soluble glutenin into dialysis bag with dialysis molecular weight of 500, dialyzing at room temperature for 5-10 times, and removing small amount of acetic acid and urea. Removing semi-solid acid soluble glutenin by dialysis, drying at 40-45 deg.C for 24-48 hr, and pulverizing into powder.
The invention has the following technical effects:
the method utilizes the characteristic that disulfide bonds among glutenin molecules prepared from alcohol-extracted glutenin powder can be opened by urea containing acid so as to disperse glutenin aggregates, and further utilizes the characteristic that high-concentration urea is easy to crystallize to separate urea from glutenin in the solution. And (3) taking out the upper solid urea, and dialyzing the lower semi-solid glutenin to remove residual urea and acetic acid to prepare the water-dispersible glutenin for the food field. The preparation process is simple and cheap, the crystallized and precipitated solid urea can be reused, and the method is beneficial to environmental protection and is a green and environment-friendly method for preparing food-grade glutenin.
Drawings
FIG. 1 is a diagram of urea crystals in an acid-soluble glutenin-containing urea solution (urea at the upper part and glutenin at the lower part);
FIG. 2 is a diagram of the solid urea crystals taken out of FIG. 1;
fig. 3 is a diagram of semi-solid acid-soluble glutenins in the lower part of fig. 1.
Detailed Description
The present invention will be described in detail with reference to specific examples.
Example 1
Mixing 1000g of wheat gluten (dry weight) and 65% of ethanol according to the mass ratio of 1: 20, and stirring and leaching for 2 hours in a 35 ℃ water bath kettle to dissolve prolamin in the wheat gluten into the ethanol; centrifuging the leached solution at 3500r/min for 5min to obtain precipitate with wet weight of 1100g, which is glutenin aggregate. The glutenin aggregate is dissolved by stirring with a urea solution with pH of 2 and concentration of 2mol/L, and the weight ratio of protein to urea solution is 1: 2. Concentrating the supernatant with rotary evaporator at 65 deg.C under 0.09MPa to obtain concentrated urea solution with a volume of one-half of the original solution. Standing the concentrated solution at 2 deg.C for 12 hr, standing at 15 deg.C for 18 hr to separate urea crystal from protein. Taking out the solid urea at the upper part, and remaining the semi-solid acid-soluble glutenin at the lower part. Dialyzing with dialysis bag with molecular weight of 500, placing semisolid acid soluble glutenin into the dialysis bag, dialyzing at room temperature for 5 times, and removing small amount of acetic acid and urea. Dialyzed to obtain semi-solid acid-soluble glutenin with a wet weight of 34.46 g. Drying at 40 deg.C for 24 hr to constant weight, and pulverizing with pulverizer to obtain 16.54g powdered protein.
Example 2
Mixing 2000g of wheat gluten and 65% of ethanol according to the mass ratio of 1: 20, and stirring and leaching in a 35 ℃ water bath kettle for 2-3h to dissolve alcohol soluble protein in the wheat gluten into the ethanol; centrifuging the leached solution for 5min at 3500r/min with a centrifuge to obtain 2400g wet weight precipitate as glutenin aggregate. Stirring and dissolving the glutenin aggregate by using a urea solution with pH of 5 and concentration of 6mol/L, wherein the weight ratio of protein to the urea solution is 1: 6. Concentrating the supernatant with rotary evaporator at 65 deg.C under 0.09MPa to obtain concentrated urea solution with a volume of one-half of the original solution. Standing the concentrated solution at 10 deg.C for 48h, standing at 30 deg.C for 36h to separate urea crystal from protein. Taking out the solid urea at the upper part, and remaining the semi-solid acid-soluble glutenin at the lower part. Dialyzing with dialysis bag with molecular weight of 500, adding semisolid acid soluble glutenin into dialysis bag, dialyzing at room temperature for 10 times, and removing small amount of acetic acid and urea. Dialyzed to obtain semi-solid acid-soluble glutenin with wet weight of 75.81 g. Drying at 45 deg.C for 48 hr to constant weight, and pulverizing with pulverizer to obtain 32.59g powder protein.
Claims (4)
1. A method for separating acid-soluble glutenin from wheat gluten by a urea crystallization method is characterized by comprising the following steps: the method comprises the following steps:
mixing wheat gluten and 65% ethanol according to the mass ratio of 1: 20, and stirring and leaching in a 35 ℃ water bath kettle for 2-3h to dissolve alcohol soluble protein in the wheat gluten into the ethanol; centrifuging the leached solution for 5min at 3500r/min by using a centrifugal machine, taking a precipitate to obtain a glutenin aggregate, stirring and dissolving the glutenin aggregate by using an acid-containing urea solution, wherein the concentration of the urea solution is 2-6 mol/L, adjusting the pH value to 2-5 by using acetic acid, the weight ratio of the protein to the urea solution is 1: 2-1: 6, taking the supernatant, concentrating the supernatant by using a rotary evaporator, standing the concentrated supernatant at the temperature of 2-10 ℃ for 12-48 h, standing the concentrated supernatant at the temperature of 15-30 ℃ for 18-36h to separate urea crystals from the protein, taking out solid urea at the upper part, removing the residual acetic acid and urea at the lower part by using semi-solid acid-soluble glutenin, drying and crushing the mixture into powder protein after dialyzing to remove the residual acetic acid and urea.
2. The method for separating acid-soluble glutenin from wheat gluten by using the urea crystallization method as claimed in claim 1, wherein the urea solution is concentrated by using a rotary evaporator, the rotary evaporation temperature is 65 ℃, the vacuum degree is 0.09MPa, and the volume of the evaporated urea solution is one half of the volume of the original solution.
3. The method for separating acid-soluble glutenin from wheat gluten by urea crystallization as claimed in claim 1, wherein the semi-solid acid-soluble glutenin is placed in a dialysis bag with a dialysis molecular weight of 500, and dialyzed 5-10 times at room temperature to remove a small amount of acetic acid and urea.
4. The method for separating acid-soluble glutenin from wheat gluten by urea crystallization as claimed in claim 1, wherein the semi-solid acid-soluble glutenin is dried at 40-45 ℃ for 24-48 h.
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