CN110713514A - Extraction method of wheat small molecule active peptide - Google Patents
Extraction method of wheat small molecule active peptide Download PDFInfo
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/36—Extraction; Separation; Purification by a combination of two or more processes of different types
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/30—Extraction; Separation; Purification by precipitation
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/14—Extraction; Separation; Purification
- C07K1/34—Extraction; Separation; Purification by filtration, ultrafiltration or reverse osmosis
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Medicinal Chemistry (AREA)
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- Proteomics, Peptides & Aminoacids (AREA)
- Analytical Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Water Supply & Treatment (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention discloses a method for extracting small-molecule active peptide of wheat, which comprises the following steps: pulverizing wheat gluten into powder by a pulverizer, adding a sodium hydroxide dilute solution for size mixing, adjusting the pH to 11, stirring at 50 ℃, centrifuging to obtain a supernatant, adding an acidic substance to adjust the pH value to 4.5, centrifuging to leave a protein precipitate, melting in water, heating at 100 ℃, cooling to 37 ℃, adding an alkaline substance to adjust the pH value to 7.0, stirring the liquid continuously, adding an enzyme for hydrolysis for 3 hours, adding an acidic substance to adjust the pH value to 4.0, deactivating the enzyme, centrifuging the solution in a centrifuge, separating and filtering the supernatant by an ultrafiltration membrane, deodorizing and decolorizing by activated carbon, and obtaining the wheat protein micromolecule polypeptide substance by utilizing a directional enzyme digestion specific small peptide separation technology. The invention separates and extracts the wheat protein micromolecule polypeptide substance by utilizing the wheat gluten powder with high protein content, reasonably utilizes resources and reduces the cost, the preparation method is convenient and fast, and the extraction rate and the quality of the micromolecule polypeptide substance are improved.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a method for extracting small-molecule active peptide of wheat.
Background
Wheat is a general name of wheat plants, is a monocotyledon, is a gramineous plant widely planted all over the world, has caryopsis as one of staple food of human beings, and can be used for making foods such as bread, steamed bread, biscuits, noodles and the like after being ground into flour; after fermentation, the fermented product can be made into beer, alcohol, liquor or biomass fuel. Wheat is rich in starch, protein, fat, minerals, calcium, iron, thiamine, riboflavin, nicotinic acid, vitamin A, vitamin C, etc. Wheat is one of the three major grains, is almost all eaten, and only about one sixth of the wheat is used as feed. The two river basin is the earliest region for cultivating wheat in the world, and China is one of the earliest countries for cultivating wheat in the world.
The wheat gluten is natural protein extracted from wheat, has protein content as high as 75-85%, is rich in nutrients, and has high viscosity, elasticity, extensibility, filming performance and fat absorbing performance. The wheat gluten is an excellent dough improver and is also a basic raw material of high-grade aquatic feeds. At present, wheat gluten is also used as an efficient green flour gluten fortifier in China, and is used for producing high gluten flour and bread special flour, and is an effective method for increasing the content of plant protein in food.
In recent years, the bioactive peptides are contained in the peptide chain of wheat protein, and are released in the process of protein degradation. Plays the role of metabolic regulation. The wheat protein has a glutamic acid content of about 37.5% of the total protein content, and the glutamic acid is glutamine in the protein in the form of glutamic acid, so that the wheat protein is very suitable for preparing glutamine-rich active peptides. The existing extraction method of wheat bioactive peptide has complex process, low extraction efficiency and high cost during extraction, and is not beneficial to industrial production, so that a method which is convenient and fast in process and high in extraction efficiency and quality of the prepared wheat bioactive peptide is needed.
Disclosure of Invention
The invention aims to overcome the defects and provides a method for extracting the wheat small-molecule active peptide.
In order to achieve the purpose, the invention adopts the following technical scheme:
a method for extracting wheat small molecule active peptide is characterized by comprising the following steps:
(1) crushing and drying the wheat protein raw material: pulverizing wheat protein raw material into powder by a pulverizer, and keeping the temperature for 2 h;
(2) wheat protein source solubilization: adding a wheat protein raw material into a sodium hydroxide dilute solution for size mixing treatment, adjusting the pH to 11, stirring at 50 ℃, stirring for 1.5h, centrifuging at the speed of 1500r/min for 10min, and taking supernatant for next-step separation;
(3) isolating the protein: adding acidic substance into the supernatant to adjust pH to 4.5, centrifuging at 1500r/min for 10min, removing supernatant, and collecting protein precipitate;
(4) enzymatic hydrolysis and enzyme deactivation: melting protein precipitate in water, heating in 100 deg.C water bath for 30min, cooling to 37 deg.C, adding alkaline substance to adjust pH to 7.0, stirring, adding enzyme, hydrolyzing for 3 hr, adding acidic substance to adjust pH to 4.0, inactivating enzyme, and cooling;
(5) filtering and separating: putting the solution into a centrifuge, centrifuging for 20min at 3000r/min, removing lower layer precipitate, collecting supernatant, and separating with ultrafiltration membrane to obtain filtrate;
(6) extraction of small molecular polypeptide substance of wheat protein: deodorizing and decolorizing the filtrate with active carbon, performing directional enzyme digestion on specific small peptide to obtain small molecular polypeptide substance of wheat protein, and sterilizing at high temperature.
According to the preparation method of the present invention, preferably, the wheat protein raw material used in the step (1) is wheat gluten.
According to the preparation method of the invention, preferably, the ratio of the sodium hydroxide dilute solution to the wheat gluten liquid in the step (1) is 1: 10.
According to the preparation method of the present invention, preferably, the acidic substance mentioned in the step (3) is citric acid.
According to the preparation method of the present invention, preferably, the alkaline substance in the step (4) is Na2SO3。
According to the preparation method of the present invention, preferably, the acidic substance mentioned in the step (4) is citric acid.
According to the preparation method of the present invention, preferably, the ultrafiltration membrane in the step (5) is a 300Dalton ultrafiltration membrane.
According to the preparation method of the present invention, preferably, the high-temperature sterilization temperature in the step (6) is 60 ℃ or higher.
The invention also provides the wheat small-molecule active peptide which is prepared by the method. The wheat small molecule active peptide prepared by the method comprises the following steps:
1. the realization process is simple, and convenient and fast industrial production is realized: the raw material of the method is wheat gluten, the protein content is high, the separation and extraction of the protein are convenient, and meanwhile, the efficiency of separating the protein by using the strong alkaline substance in the first step is high, so that the extraction working efficiency is improved.
2. The purpose of improving the quality of the wheat small molecule active peptide is realized: the invention improves the protein quality by filtering the protein extracted from wheat gluten through an ultrafiltration membrane of 300Dalton, and obtains the high-quality wheat protein micromolecule polypeptide substance by utilizing the specific small peptide separation technology of directional enzyme digestion.
3. The extraction efficiency is high: according to the invention, when strong alkaline substances are used for extraction, the setting environment pH value is increased to 11 when extraction is carried out, the temperature is 50 ℃, stirring is carried out for 1.5h, the liquid-material ratio is 1:10, the protein extraction efficiency is improved, and finally, the wheat protein small molecule polypeptide substance is efficiently and directionally separated by adopting a directional enzyme digestion specific small peptide separation technology.
In addition to the objects, features and advantages described above, other objects, features and advantages of the present invention are also provided. The present invention will be described in further detail below.
Drawings
In order to more clearly illustrate the technical solution of the embodiment of the present invention, the drawings needed to be used in the description of the embodiment will be briefly introduced below, and it is obvious that the drawings in the following description are only for more clearly illustrating the embodiment of the present invention or the technical solution in the prior art, and for those skilled in the art, other drawings can be obtained according to the drawings without any creative effort.
FIG. 1 is a schematic diagram of the temperature and protein extraction rate of the extraction method of wheat small molecule active peptide of the present invention.
FIG. 2 is a schematic diagram of the Ph value and the protein extraction rate of the extraction method of the wheat small molecule active peptide.
FIG. 3 is a schematic diagram of liquid-to-feed ratio and protein extraction rate of the extraction method of wheat small molecule active peptide.
FIG. 4 is a schematic diagram of the stirring time and the protein extraction rate of the extraction method of the wheat small molecule active peptide.
The specific implementation mode is as follows:
the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments.
A method for extracting wheat small molecule active peptide is characterized by comprising the following steps:
(1) crushing and drying the wheat protein raw material: pulverizing wheat protein raw material into powder by a pulverizer, and keeping the temperature for 2 h;
(2) wheat protein source solubilization: adding a wheat protein raw material into a sodium hydroxide dilute solution for size mixing treatment, adjusting the pH to 11, stirring at 50 ℃, stirring for 1.5h, centrifuging at the speed of 1500r/min for 10min, and taking supernatant for next-step separation;
(3) isolating the protein: adding acidic substance into the supernatant to adjust pH to 4.5, centrifuging at 1500r/min for 10min, removing supernatant, and collecting protein precipitate;
(4) enzymatic hydrolysis and enzyme deactivation: melting protein precipitate in water, heating in 100 deg.C water bath for 30min, cooling to 37 deg.C, adding alkaline substance to adjust pH to 7.0, stirring, adding enzyme, hydrolyzing for 3 hr, adding acidic substance to adjust pH to 4.0, inactivating enzyme, and cooling;
(5) filtering and separating: putting the solution into a centrifuge, centrifuging for 20min at 3000r/min, removing lower layer precipitate, collecting supernatant, and separating with ultrafiltration membrane to obtain filtrate;
(6) extraction of small molecular polypeptide substance of wheat protein: deodorizing and decolorizing the filtrate with active carbon, performing directional enzyme digestion on specific small peptide to obtain small molecular polypeptide substance of wheat protein, and sterilizing at high temperature.
In the invention, the wheat protein raw material used in the step (1) is wheat gluten; in the step (1), the ratio of the sodium hydroxide dilute solution to the wheat gluten liquid material is 1: 10; the acidic substance mentioned in the step (3) is citric acid; the alkaline substance in the step (4) is Na2SO3(ii) a The acidic substance mentioned in the step (4) is citric acid; the ultrafiltration membrane in the step (5) is a 300Dalton ultrafiltration membrane; the high-temperature sterilization temperature in the step (6) is above 60 ℃.
The extraction method of the wheat small-molecule active peptide is a preparation method which can facilitate the extraction mode of the wheat small-molecule active peptide and improve the extraction rate and the quality of the wheat small-molecule active peptide.
When the wheat protein raw material wheat gluten is prepared and extracted from a sodium hydroxide dilute solution, the extraction conditions are that the temperature is 50 ℃, the Ph value is 11.0, the liquid-material ratio is 10: 1, and the stirring time is 1.5 h. As shown in the graph of fig. 1, the protein extraction efficiency is not higher as the temperature is higher, but is highest at 50 ℃ and then sharply decreases; as shown in the Ph value and protein extraction rate diagram of fig. 2, the protein extraction efficiency is positively correlated with the Ph value, but the growth rate reaches the highest when the Ph value is 11.0, and the subsequent growth rate decreases gradually; as shown in the schematic diagram of fig. 3, the liquid-to-material ratio and the protein extraction rate are in positive correlation, but when the liquid-to-material ratio is 10: 1, the growth rate reaches the highest, and the subsequent growth rate decreases gently; as shown in fig. 4, the graph of the stirring time and the protein extraction rate shows that the increase rate of the protein extraction rate is highest when the stirring time is 1.5h, and then the increase rate tends to be gentle and the protein extraction rate begins to decrease after 2.0 h. In conclusion, when the protein is extracted from wheat protein raw material wheat gluten, the extraction conditions are that the temperature is 50 ℃, the Ph value is 11.0, the liquid-material ratio is 10: 1, and the stirring time is 1.5 h.
When extracting the wheat protein small molecule polypeptide substance, the specific small peptide separation technology of directional enzyme digestion is adopted to cut DNA fragments by using restriction endonuclease, a specific deoxynucleotide sequence is fixedly identified due to the specificity of the enzyme, and the corresponding DNA fragments of the wheat protein small molecule polypeptide substance are cut by using the specific enzyme, so that the purpose of directional cutting is achieved, and the extraction quality of the wheat protein small molecule polypeptide substance is improved.
The above description is only for the preferred embodiment of the present invention, but the scope of the present invention is not limited thereto, and any person skilled in the art should be considered to be within the technical scope of the present invention, and the technical solutions and the inventive concepts thereof according to the present invention should be equivalent or changed within the scope of the present invention.
Claims (8)
1. A method for extracting wheat small molecule active peptide is characterized by comprising the following steps:
(1) crushing and drying the wheat protein raw material: pulverizing wheat protein raw material into powder by a pulverizer, and keeping the temperature for 2 h;
(2) wheat protein source solubilization: adding a wheat protein raw material into a sodium hydroxide dilute solution for size mixing treatment, adjusting the pH to 11, stirring at 50 ℃, stirring for 1.5h, centrifuging at the speed of 1500r/min for 10min, and taking supernatant for next-step separation;
(3) isolating the protein: adding acidic substance into the supernatant to adjust pH to 4.5, centrifuging at 1500r/min for 10min, removing supernatant, and collecting protein precipitate;
(4) enzymatic hydrolysis and enzyme deactivation: melting protein precipitate in water, heating in 100 deg.C water bath for 30min, cooling to 37 deg.C, adding alkaline substance to adjust pH to 7.0, stirring, adding enzyme, hydrolyzing for 3 hr, adding acidic substance to adjust pH to 4.0, inactivating enzyme, and cooling;
(5) filtering and separating: putting the solution into a centrifuge, centrifuging for 20min at 3000r/min, removing lower layer precipitate, collecting supernatant, and separating with ultrafiltration membrane to obtain filtrate;
(6) extraction of small molecular polypeptide substance of wheat protein: deodorizing and decolorizing the filtrate with active carbon, performing directional enzyme digestion on specific small peptide to obtain small molecular polypeptide substance of wheat protein, and sterilizing at high temperature.
2. The method for extracting the wheat small molecule active peptide according to claim 1, wherein the method comprises the following steps: the wheat protein raw material used in the step (1) is wheat gluten.
3. The method for extracting the wheat small molecule active peptide according to claim 1, wherein the method comprises the following steps: in the step (1), the ratio of the sodium hydroxide dilute solution to the wheat gluten liquid material is 1: 10.
4. The method for extracting the wheat small molecule active peptide according to claim 1, wherein the method comprises the following steps: the acidic substance mentioned in the step (3) is citric acid.
5. The method for extracting the wheat small molecule active peptide according to claim 1, wherein the method comprises the following steps: the alkaline substance in the step (4) is Na2SO3。
6. The method for extracting the wheat small molecule active peptide according to claim 1, wherein the method comprises the following steps: the acidic substance mentioned in the step (4) is citric acid.
7. The method for extracting the wheat small molecule active peptide according to claim 1, wherein the method comprises the following steps: the ultrafiltration membrane in the step (5) is a 300Dalton ultrafiltration membrane.
8. The method for extracting the wheat small molecule active peptide according to claim 1, wherein the method comprises the following steps: the high-temperature sterilization temperature in the step (6) is above 60 ℃.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111670995A (en) * | 2020-06-08 | 2020-09-18 | 颜如玉医药科技有限公司 | Preparation method of wheat protein peptide |
| CN112042909A (en) * | 2020-08-31 | 2020-12-08 | 江苏师范大学 | Preparation method of oral liquid for dispelling effects of alcohol and protecting liver |
-
2019
- 2019-11-07 CN CN201911082161.9A patent/CN110713514A/en not_active Withdrawn
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111670995A (en) * | 2020-06-08 | 2020-09-18 | 颜如玉医药科技有限公司 | Preparation method of wheat protein peptide |
| CN111670995B (en) * | 2020-06-08 | 2021-05-18 | 颜如玉医药科技有限公司 | Preparation method of wheat protein peptide |
| CN112042909A (en) * | 2020-08-31 | 2020-12-08 | 江苏师范大学 | Preparation method of oral liquid for dispelling effects of alcohol and protecting liver |
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