CN107400690A - Rice protein preparation method - Google Patents

Abstract

The invention relates to a method for preparing rice protein, which comprises treating rice protein powder and water with combined enzymolysis of papain and neutral or alkaline protease. The preparation method of the invention can improve the solubility of rice protein, thereby obtaining high Protein yield of rice protein products.

Description

Method for preparing rice protein

technical field

The invention relates to a preparation method of rice protein.

Background technique

Rice protein is a high-quality vegetable protein, which is rich in essential amino acids, and its amino acid composition pattern is consistent with that of the World Health Organization (WHO) and the United Nations Food and Agriculture Organization (Food and Agriculture Organization of the United Nations, FAO). The recommended mode is similar, and it is easy to be digested and absorbed by the human body. In addition, rice protein is hypoallergenic and is very suitable as a protein supplement for special populations, especially patients. However, due to the special structure of glutenin, the main component of rice protein, there are a large number of sulfhydryl groups and disulfide bonds in the molecule and inside the molecule, which makes rice protein difficult to dissolve in water under neutral conditions, which affects its application in the food and health care products industries. Application, therefore, improving the solubility of rice protein is the key to solve the problem.

There are physical methods, chemical methods, and biological methods to improve protein solubility, among which enzymatic modification has the characteristics of strong specificity, mild conditions, and no toxic side effects, so it is widely used in the modification of the solubility of various plant proteins. Generally, proteases such as soybean protein, peanut protein, and rice bran protein are used to hydrolyze and break the peptide chain of protein molecules, improve the solubility of protein, and achieve the effect of promoting the solubility of insoluble vegetable protein powder.

Chinese patent CN1417344C, published in 2003, discloses a new process for extracting rice protein from rice grains, which is characterized in that the ratio by weight of the by-product rice grains of the monosodium glutamate factory and the compound enzyme is 1000: 1.5-4.5; The compound enzyme is composed of 0.05%-0.15% amylase, 0.025%-0.075% lipase, 0.025%-0.075% cellulase and 0.05%-0.15% protease; To 3, adjust the temperature to 35-40°C, add cellulase according to the proportion, react for 30 minutes, add 30% NaOH, adjust the pH to 5.0-5.5, add lipase, react for 30 minutes, then add NaOH to adjust the pH to neutral 7.0, add amylase and protease, react for 20 minutes, heat up to 60-70°C, react for 30 minutes, filter, add dilute lye with pH 9.5 to dissolve the filter residue, control the temperature below 60°C to obtain a protein solution, and then use The acid adjusts the pH to 5.5, and the protein precipitates to obtain concentrated rice protein with a content of 75%.

Another Chinese patent CN1257984C, published in 2006, discloses, for example, a preparation method of rice protein peptide, which is characterized in that rice dregs and water are mixed in a weight ratio of 1:4-6, the temperature is 55-60°C, and the temperature is kept for 30 minutes. Then, according to the activity of the enzyme preparation, it is 50,000U/mg, add respectively α-amylase and neutral protease of rice dregs weight 0.05%-0.1%, wherein, the weight ratio of α-amylase and neutral protease is 1: 1. Stir for 5-7 hours at pH 6.5-7.5 and temperature 50-60°C, filter the reactant, and spray-dry the resulting liquid to obtain a white powder product, in which the content of rice protein peptide is about 80%.

At present, it is still necessary to continue to develop better rice protein preparation methods to improve the problem of insoluble rice protein and increase protein yield.

Contents of the invention

In order to improve the problem of insoluble rice protein and increase the yield of protein, the present invention provides a method for preparing rice protein. The present invention uses a compound enzyme composed of papain and neutral or alkaline protease to enzymolyze rice protein powder, which can significantly Significantly improve the solubility of rice protein, increase the solubility of rice protein to more than 60%, and the protein yield of the obtained rice protein product reaches more than 90%.

In one aspect, the present invention provides a method for preparing rice protein that improves the solubility of rice protein powder to increase protein yield, characterized in that rice protein powder and water are mixed in an appropriate ratio and treated with an enzyme combination, wherein the enzyme combination It is a composite enzyme composed of papain and neutral protease, or a composite enzyme composed of papain and neutral protease, and the weight ratio of papain to neutral or alkaline protease is 1:1-3.

In a specific embodiment of the present invention, the weight ratio of papain to neutral protease, or papain to alkaline protease in the compound enzyme is 1:2.

According to the present invention, the enzyme combination further comprises a flavor enzyme to improve bitterness.

In a specific embodiment of the present invention, the flavor enzyme is a combination of Aspergillus oryzae and aminopeptidases.

In a specific embodiment of the present invention, the method of the present invention is characterized in that it includes the following steps: add water to the rice protein powder in an appropriate proportion and stir evenly, adjust the pH value of the obtained solution to medium to alkaline, add the enzyme combination for enzymolysis, and continue stirring After a period of time, the temperature is raised to inactivate the enzyme, and it is obtained by filtering, concentrating and drying.

In another aspect, the present invention provides a rice protein product prepared by the method of the present invention, wherein the solubility of rice protein in the rice protein product is greater than 60%, and the protein yield of rice protein is greater than 90%.

detailed description

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.

As used herein, the singular forms "a" and "the" include plural reference unless the context clearly dictates otherwise. Thus, for example, reference to "a sample" includes a plurality of such samples and equivalents thereof well known to those skilled in the art.

The word "papain (Papain)" used herein, or simply papain, papain, is a cysteine protease present in papaya fruit. Papain plays an important role in tenderizing meat. It is an endoprotease with protease and esterase activities.

The term "Neutral Protease" used herein refers to a class of enzymes whose optimum pH is between 6.0-7.5, molecular weight is 35000-40000, and isoelectric point is 8-9. Its action conditions are mild, and it is an endoprotease refined by advanced extraction technology after submerged fermentation of selected bacillus subtilis. It can decompose macromolecular proteins into polypeptides and amino acids, forming a unique hydrolysis flavor. Neutral protease is the first protease preparation discovered and widely used in industrial production. It can be used in the baked food industry, the synthesis of aspartame, and the removal of bitterness from protein hydrolyzate.

The term "Alkaline Protease" used in this paper is a kind of proteolytic enzyme produced by submerged fermentation, extraction and refining of a Bacillus subtilis variety cultivated by bacterial protoplast mutagenesis. Serine is a highly alkaline protease, which can hydrolyze the peptide chain of protein molecules under alkaline conditions to generate polypeptides or amino acids.

The word "Aspergillus oryzae" used in this paper is a kind of flavor enzyme, which includes two kinds of activities of endoprotease and exopeptidase, which can be used to remove the bitter peptide chain of low degree of hydrolysis product, and make it completely degraded to amino acids.

The term "aminopeptidases" as used herein refers to an exopeptidase that selectively cleaves amino acid residues from the N-terminus of a protein or peptide chain, allowing amino acids to be sequenced one by one from the N-terminus of the polypeptide chain. free out.

Because glutenin, the main component of rice protein, has a large number of sulfhydryl groups and disulfide bonds in the molecule and inside the molecule, it makes rice protein difficult to dissolve in water under neutral conditions. According to the present invention, it is characterized in that an enzyme combination is used, including a compound enzyme composed of papain and neutral protease, or a compound enzyme composed of papain and neutral protease, so as to improve the problem of insoluble rice protein and increase protein yield.

The invention provides a method for preparing rice protein by using a combination of special enzymes to improve the hydrolysis degree of rice protein and increase the protein yield. The method of the invention comprises mixing rice protein powder and water in an appropriate ratio, and treating with an enzyme combination, wherein the enzyme combination is a compound enzyme composed of papain and neutral protease, or a compound enzyme composed of papain and alkaline protease. However, after enzymatic treatment, it often has a bitter taste, so flavor enzymes are added to improve the bitter taste.

The ratio of papain to neutral or alkaline protease used in the present invention is preferably 1:1-3 by weight, preferably 1:2. Flavor enzymes can be added as needed to improve the bitter taste.

According to the present invention, the comparison of Table 2 and Table 3 in the following examples shows that the present invention utilizes the composite enzyme composed of papain and neutral protease, or the composite enzyme composed of papain and neutral protease, and its protein yield is as high as 90%. The above is far higher than using one of them alone, wherein the protein yields of papain, neutral protease, alkaline protease, Aspergillus oryzae or aminopeptidase alone are all less than 75%. While Aspergillus oryzae has the effect of improving bitter taste, it was unexpectedly found that the combination of Aspergillus oryzae and aminopeptidase has the best effect of improving bitter taste.

The method of the present invention can obtain rice protein powder by any known method steps, including but not limited to physical or chemical methods. For example, it can be obtained in powder form from rice through the processing steps of general protein extraction such as pulverization, purification, drying, etc., so as to carry out the enzymatic hydrolysis of the method of the present invention. According to an embodiment of the present invention, the rice protein powder is obtained by grinding the rice for preliminary treatment, adding water in an appropriate proportion and stirring evenly, for example, the weight ratio of material to water is 1:3-10, preferably 1:4, and the obtained solution is adjusted to Neutral or slightly alkaline, eg about pH 8.0. Enzymolysis is generally enzymatic hydrolysis at a relatively high temperature for a period of time, such as about 50-60°C, preferably 55°C, after continuous stirring, the temperature is raised to about a high temperature to kill the enzyme, such as about 85°C, and rice is obtained by filtering, concentrating, and drying protein products. The resulting product can be filtered, concentrated, dried, and further purified by conventional methods.

The following examples are provided to illustrate the invention and are not to be construed in any way as limiting the scope of the invention.

Example

Example 1

Grind the rice for preliminary treatment to obtain rice protein powder, add water in an appropriate proportion and stir evenly, for example, the weight ratio of material to water is 1:3-10, adjust the pH value of the obtained solution to slightly alkaline, for example about pH8.0, add the following table 1 Various enzyme combinations, enzymatic hydrolysis at a higher temperature for a period of time, such as about 50-60 ° C, continue to stir, then heat up to about 85 ° C to kill the enzyme, filter, concentrate, and dry to obtain a rice protein product for inspection.

Enzyme combination of each experimental group in table 1

group Enzyme Portfolio Content Experimental group 1 Complex protease A (papain + alkaline protease) + flavor enzyme A (Aspergillus oryzae) Experimental group 2 Compound Protease A (Papain + Alkaline Protease) + Flavor Enzyme B (Aspergillus oryzae + Aminopeptidase) Experimental group 3 Complex protease B (papain + neutral protease) + flavor enzyme A (Aspergillus oryzae) Experimental group 4 Compound protease B (papain + neutral protease) + flavor enzyme B (Aspergillus oryzae + aminopeptidase) Experimental group 5 blank

Example 2

The rice protein product of embodiment 1 gained is checked by following test method.

1. Solubility test method

A certain amount of raw material X is subjected to a hydrolysis test according to the test process. After the hydrolysis is completed, it is centrifuged, the supernatant is collected, and dried to obtain the weight Y after drying. The solubility is

2. Experimental method for the determination of amino nitrogen

Utilize the amphoteric effect of amino acid, add formaldehyde to fix the basicity of amino acid, make the carboxyl group show acidity, and quantify after titrating with sodium hydroxide standard solution. First, accurately draw 1ml of the sample solution to be tested, add 79ml of distilled water, start the magnetic stirrer, titrate to pH 8.2 with a sodium hydroxide standard solution with a concentration of 0.05mol/l, add 10ml of formaldehyde precisely, mix well, and then use a concentration of The 0.05mol/l sodium hydroxide standard solution continued to be titrated to pH9.2, and the volume of the sodium hydroxide standard solution consumed after adding formaldehyde was recorded. Next, take 80ml of distilled water, accurately add 10ml of formaldehyde, titrate to pH9.2 with a sodium hydroxide standard solution with a concentration of 0.05mol/l, record the volume of the sodium hydroxide standard solution consumed, and use it as an experimental blank. Calculated according to the following formula:

V1: Determination of the volume of sodium hydroxide standard solution consumed after adding formaldehyde to the sample solution to be tested (ml)

V2: the volume of sodium hydroxide standard solution consumed after adding formaldehyde to the actual blank (ml)

C: concentration of sodium hydroxide standard solution (mol/l)

W: The amount of the sample to be weighed

0.0141: The mass of amino nitrogen expressed in grams equivalent to 1ml sodium hydroxide standard solution

3. Protein quantitative test (Kjeldahl method)

The sample is heated and digested with concentrated sulfuric acid and a catalyst to hydrolyze the protein, in which carbon and hydrogen are oxidized into carbon dioxide and water to escape, while the organic nitrogen in the sample is converted into ammonia and combined with sulfuric acid to form ammonium sulfate, and then distilled with alkali to make the ammonia Distill it off, absorb it with boric acid and then titrate it with standard hydrochloric acid solution, and calculate the protein content according to the volume consumed by hydrochloric acid. First, accurately weigh 0.2-2 grams of the sample (liquid 1-2ml), carefully transfer it into a dry and clean digestion bottle, then add 0.5 grams of copper sulfate, 9.5 grams of potassium sulfate and 20ml of concentrated sulfuric acid, shake it gently and place it on the electric furnace Bring to a boil over low heat. After all the contents in the bottle are carbonized and the foam stops, increase the firepower and keep the liquid in the bottle slightly boiling until the liquid turns blue-green and transparent, then continue to heat and boil slightly for 1-2 hours (the length of time is determined by the protein content). Then, after the solution is cooled, add 200ml of pure water, move it into the distillation bottle, add 150ml of 40% NaOH solution, plug the bottle tightly, and insert the lower end of the condenser tube under the liquid level of the absorption bottle, which contains 50ml of 4% boric acid solution and Mix 2-3 drops of the indicator, heat and distill until all the ammonia is evaporated (the distillate should be 250ml). Titrate with 0.1mol/L HCl standard solution to gray as the end point, and make a blank at the same time. Calculated according to the following formula:

V1: The volume of hydrochloric acid standard solution consumed by the sample (ml)

V2: the volume of the actual blank hydrochloric acid standard solution (ml)

W: The amount of the sample to be weighed

0.014: The mass of amino nitrogen expressed in grams equivalent to 1ml of hydrochloric acid standard solution

C: concentration of hydrochloric acid standard solution (mol/l)

F: Factor for converting nitrogen into protein, F=6.25

The test results are summarized in Table 2. The protein yields of each experimental group were as high as 90% or more, and the solubility was significantly improved, as high as 60% or more. Among them, the experimental group 4 (composite protease B+flavor enzyme B) is particularly preferred, the degree of protein hydrolysis is as high as 98%, the solubility is more than 70%, and there is no bitter taste.

Table 2 Test results of each experimental group

test group AN% (g/100mg) Protein yield % Solubility flavor Experimental group 1 0.3397 96.5% 68.9% Light rice aroma, obvious bitterness Experimental group 2 0.3739 97.5% 72.0% light rice aroma, bitter taste Experimental group 3 0.3654 96.2% 63.8% Light rice aroma, a little bit of bitterness Experimental group 4 0.3839 98.6% 73.5% Light rice aroma, slightly sweet Experimental group 5 0.0093 0.4% 6.1% tasteless

With the same method and steps, a single enzyme preparation is used to carry out enzymolysis, and the results obtained through inspection are shown in Table 3, and the protein yield of each group is more than 40% to 75%. Obviously, the enzyme combination of the present invention obviously improves the solubility of rice protein powder. Improve the solubility of rice protein, up to 60%. Among them, the experimental group 4 (composite protease B+flavor enzyme B) is particularly preferred, the degree of protein hydrolysis is as high as 98%, the solubility is more than 70%, and there is no bitter taste.

The inspection result of each single enzyme preparation group of table 3

Group No use enzyme preparation AN% (g/100mg) Protein yield % flavor 1 papain 0.2450 69.7% light rice aroma, bitter taste 2 alkaline enzyme protease 0.2733 73.5% Light rice aroma, heavy bitterness 3 Dispase 0.2061 63.2% Light rice aroma, a little bit of bitterness 4 Aspergillus oryzae 0.1468 43.5% Light rice aroma, no bitterness

In summary, the compound enzyme combined with papain and neutral or alkaline protease can improve the solubility of rice protein and significantly increase the protein yield of rice protein (both as high as 90%). Among them, papain, neutral protease, and flavor enzyme B) are preferred, the degree of protein hydrolysis is as high as more than 98%, the solubility is more than 70%, and there is no bitter taste. With the same method and steps, a single enzyme preparation is used to carry out enzymolysis, and the results obtained through inspection are shown in Table 3, and the protein yield of each group is more than 40% to 75%. Obviously, the enzyme combination of the present invention can obviously improve the solubility of rice protein powder, improve the The solubility of rice protein is as high as 60%. Among them, the experimental group 4 (composite protease B+flavor enzyme B) is particularly preferred, the degree of protein hydrolysis is as high as 98%, the solubility is more than 70%, and there is no bitter taste.

The above embodiments are only exemplary, and do not constitute any limitation to the scope of the present invention. Those skilled in the art should understand that the details and forms of the technical solutions of the present invention can be modified or replaced without departing from the spirit and scope of the present invention, but these modifications and replacements all fall within the protection scope of the present invention.

Claims (10)

1. A preparation method for improving the solubility of rice protein powder to promote the rice protein of protein yield, characterized in that rice protein powder is mixed with water in an appropriate proportion, and processed with an enzyme combination, wherein the enzyme combination is papain and Chinese The composite enzyme composed of neutral protease, or the composite enzyme composed of papain and alkaline protease, the weight ratio of papain to neutral or alkaline protease is 1:1-3. 2. The method according to claim 1, characterized in that the weight ratio of papain and neutral protease, or papain and alkaline protease in the compound enzyme is 1:2. 3. The method according to claim 1 or 2, wherein the enzyme combination further comprises a flavor enzyme to improve bitterness. 4. The method according to claim 3, wherein the flavor enzyme is a combination of Aspergillus oryzae and aminopeptidases. 5. The method according to claim 1, characterized in that it comprises the following steps: adding water to the rice protein powder in an appropriate proportion and stirring evenly, adjusting the pH value of the obtained solution to medium to alkaline, adding the enzyme combination for enzymolysis, and continuing to stir for a period of time After heating to inactivate the enzyme, it can be obtained by filtering, concentrating and drying. 6. The method according to claim 1, characterized in that the enzyme combination comprises papain and neutral protease compound enzyme with a weight ratio of 1:2, and the flavor of Aspergillus oryzae and aminopeptidase (aminopeptidases) combination enzyme. 7. A rice protein product made by the method as claimed in claim 1. 8. A rice protein product made by the method as claimed in claim 3. 9. A rice protein product made by the method as claimed in claim 6. 10. The rice protein product according to any one of claims 7-9, characterized in that the solubility of rice protein in the rice protein product is greater than 60%, and the protein yield of rice protein is greater than 90%.