CN108085356B - Method for industrially producing high-purity walnut peptide by taking low-temperature squeezed walnut meal as raw material - Google Patents
Method for industrially producing high-purity walnut peptide by taking low-temperature squeezed walnut meal as raw material Download PDFInfo
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- CN108085356B CN108085356B CN201711444692.9A CN201711444692A CN108085356B CN 108085356 B CN108085356 B CN 108085356B CN 201711444692 A CN201711444692 A CN 201711444692A CN 108085356 B CN108085356 B CN 108085356B
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Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P21/00—Preparation of peptides or proteins
- C12P21/06—Preparation of peptides or proteins produced by the hydrolysis of a peptide bond, e.g. hydrolysate products
-
- 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
- A23J3/00—Working-up of proteins for foodstuffs
- A23J3/14—Vegetable proteins
-
- 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
- A23J3/00—Working-up of proteins for foodstuffs
- A23J3/30—Working-up of proteins for foodstuffs by hydrolysis
- A23J3/32—Working-up of proteins for foodstuffs by hydrolysis using chemical agents
- A23J3/34—Working-up of proteins for foodstuffs by hydrolysis using chemical agents using enzymes
- A23J3/346—Working-up of proteins for foodstuffs by hydrolysis using chemical agents using enzymes of vegetable proteins
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/17—Amino acids, peptides or proteins
- A23L33/18—Peptides; Protein hydrolysates
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
Abstract
The invention discloses a method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut meal as a raw material. The invention takes cold-pressed walnut meal as a raw material, after dispersion and hydration, the walnut meal is subjected to enzymolysis through tannase, cellulase and medium-temperature amylase, then is denatured, and is subjected to enzymolysis step by step through alkaline protease and neutral protease, enzyme deactivation, centrifugation and plate-and-frame filtration to obtain the high-purity walnut peptide. The invention realizes the very economical industrialized preparation of the high-purity walnut peptide through the scientific optimization of the pretreatment, enzymolysis and refining processes, the protein content is more than 90 percent, the peptide content is more than 85 percent, the relative molecular weight is less than 2000Dal peptide and more than 85 percent, the relative molecular weight is less than 1000Dal peptide and more than 50 percent, the product has good quality and weak bitter taste, is easy to match with other food ingredients, has outstanding effects of strengthening brain, benefiting intelligence and enhancing immunity, and can be widely applied to various foods and health care products as a high-quality functional base material.
Description
Technical Field
The invention belongs to the technical field of high-added-value processing and utilization of agricultural and sideline products, and particularly relates to a method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut meal as a raw material.
Background
Walnut is one of four dry fruits in the world, and is popular with consumers at home and abroad due to the rich nutritional value. Every 100 g of walnut kernel contains 63-70% of fat (mainly unsaturated fatty acid, accounting for about 90% of the total weight), 15-29% of protein and 15% of saccharide; in addition, it also contains nutrient substances such as calcium, phosphorus, ferrum, zinc, magnesium, carotene, riboflavin, vitamin B6, vitamin E, juglone, and phospholipid. China is one of the original places of walnuts, has more than 2000 years of cultivation history, and the cultivation area and the yield of the Chinese walnuts are at the top of the world at present. However, the development and utilization degree and depth of walnut products are far from enough, and deep-processed products for other purposes are rare except for traditional stir-frying, can making, moon cake stuffing, walnut powder, walnut milk and walnut oil. The walnut oil is listed as one of 3 vegetable oils with higher international and domestic market demand in recent years, and the consumption proportion is continuously increased. Domestic walnut oil manufacturers reach dozens of families, a large amount of degreased walnut cakes are used as feed and fertilizer or discarded, and walnut resources are seriously wasted. Research shows that the protein content in the walnut meal is up to more than 50 percent, the walnut protein mainly comprises 4 types of proteins, namely albumin, globulin, prolamin and glutelin, which respectively account for 6.81 percent, 17.57 percent, 5.33 percent and 70.11 percent of the total amount of the walnut protein, the protein composition is close to the recommended standard of the Food and Agriculture Organization (FAO) and the World Health Organization (WHO) of the United nations, the content of 8 necessary amino acids is reasonable, the contents of glutamic acid, aspartic acid and arginine which have important functions on the physiological action of the human body are higher, and the walnut protein is a plant protein with great development potential.
The walnut protein has poor solubility due to high proportion of hydrophobic amino acids in the walnut meal glutelin and the globulin, and the utilization of the walnut protein in the food industry is greatly limited, so that the research on deep processing of the walnut meal protein has important social and economic significance. In recent years, researches show that bioactive peptides with various physiological functions can be obtained from edible protein raw materials by a scientific and reasonable bioengineering means, the bioactive peptides comprise bioactive peptides which are easy to digest and absorb, are antioxidant, eliminate fatigue, reduce blood sugar, reduce blood pressure, promote ethanol metabolism, enhance immunity and the like, belong to natural green functional food ingredients, have remarkable effects, no side effect and high safety, can be widely applied to foods (especially health-care foods) and medicines, have good market application prospects, and the development of walnut peptides is the most potential and promising direction in the fine and deep processing of walnut meal.
The natural walnut protein molecules are curled inside the protein molecules to be similar to a sphere due to the effects of hydrogen bonds, hydrophobic bonds, disulfide bonds and the like, and the peptide chains are compact in structure and have strong hydrophobicity and are difficult to hydrolyze by protease; in addition, considerable amount of starch, grease, pigment and the like exist in the walnut meal, which influences the deep processing and utilization of walnut protein. At present, the research on deep processing of walnut protein at home and abroad mainly focuses on how to carry out high-efficiency enzymolysis on the walnut protein to obtain high-yield low-molecular-weight walnut peptide. The outstanding difficult problems of walnut peptide preparation mainly comprise: (1) the product purity is low, namely the content of the walnut peptide protein is low, mainly a certain amount of starch and grease are still contained in the walnut meal, and the starch and the grease cannot be effectively removed in pretreatment before enzymolysis and separation and refining after the enzymolysis, so that the content of non-protein components of the final peptide product is high, and the quality of the product is influenced; (2) the walnut protein has compact structure and low enzymolysis efficiency, and the enzymolysis product has low content of low molecular weight peptide; (3) the walnut dregs contain walnut kernel peels, are black in color and bitter and astringent, and the color and flavor of the product can be influenced if the walnut dregs are not effectively treated; (4) the preparation process is complicated, scientific optimization is not performed, the cost is high, and industrial implementation is difficult. For example, patent CN103109971B discloses that walnut pulp obtained by cold pressing walnut is used as a raw material, and a walnut peptide with high arginine content is obtained by grinding, enzymolysis with compound protease, membrane filtration, concentration and drying, wherein the yield of active peptide is lower than 30%, the protein content of the product is low, and the molecular weight of the main peptide is less than or equal to 3000 kD; patent CN 102630804A discloses a method for preparing walnut peptide powder by using defatted walnut meal as raw material, adding water, mixing uniformly, homogenizing, performing enzymolysis with complex enzyme, inactivating enzyme, centrifuging, concentrating and drying, wherein the product prepared by the method has low protein content, wide peptide molecular weight range and low micromolecular peptide content; patent CN 103103244B discloses a preparation method of walnut antihypertensive peptide, which takes walnut dregs as raw materials, walnut protein is purified by an alkali-soluble acid-precipitation method, then proper denaturation of the walnut protein is treated by microwave and ultrasonic wave to improve enzymolysis efficiency, neutral protease is added for enzymolysis, then permeate liquid is collected by a 0.45um microfiltration membrane, an 8kD ultrafiltration membrane and a 1kD ultrafiltration membrane, B-cyclodextrin is added for embedding and vacuum freeze drying is carried out in the permeate liquid to obtain the walnut antihypertensive peptide.
Disclosure of Invention
In order to solve the difficulties and disadvantages of the prior art, the invention aims to provide a method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut meal as a raw material. The method comprises the steps of using low-temperature squeezed walnut pulp as a raw material, treating with tannase, cellulase and medium-temperature amylase, passivating enzyme by combining moderate heat treatment, changing protein structure, washing and purifying walnut protein, adding compound protease for enzymolysis step by step, inactivating enzyme, performing centrifugal separation and plate-frame filter pressing on enzymolysis products, concentrating filtrate, drying and preparing high-purity walnut peptide, and is compact and optimized in process and suitable for industrial production.
It is a further object of the invention to provide an application of said method.
The invention further aims to provide the high-purity walnut peptide prepared by the method. The peptide can be used as raw material of health food and common food, especially for brain strengthening, intelligence improving and immunity enhancing products.
The invention is realized by the following technical scheme: a method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut meal as a raw material comprises the following steps:
(1) dispersion and hydration of walnut meal: weighing cold-pressed walnut meal, and preparing walnut protein solution by using water; shearing, dispersing and hydrating the walnut protein solution uniformly;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 45-55 ℃, adding tannase, cellulase and medium-temperature amylase, and carrying out enzymolysis; the mass dosage of each enzyme is calculated by taking the mass of the walnut pulp as a reference, and the mass dosage is as follows: 0.1-0.2% of tannase, 0.1-1.0% of cellulase and 0.5-2.0% of medium-temperature amylase;
(3) protein denaturation, centrifugation, water washing: treating the walnut protein solution subjected to enzymolysis in the step (2) at 100-121 ℃ for 0.5-1.0 h, centrifuging to remove upper-layer liquid (a grease layer and an aqueous solution layer), collecting precipitate, adding water at the temperature of 50-60 ℃, uniformly stirring, centrifuging to remove the upper-layer liquid, and collecting precipitate;
(4) and (3) protease enzymolysis and enzyme deactivation step by step: adding water with the temperature of 50-60 ℃ into the precipitate obtained in the step (3), and uniformly stirring to obtain walnut protein slurry; adjusting the pH value of the walnut protein slurry to 7.5-8.5, firstly adding alkaline protease, stirring and hydrolyzing at 50-55 ℃ for 1-2 h, then adding neutral protease, stirring and hydrolyzing at 50-55 ℃ for 4-16 h; inactivating enzyme after enzymolysis; the mass dosage of each enzyme is calculated by taking the mass of the protein in the walnut protein slurry as a reference, and the mass dosage is as follows: 0.5 to 2 percent of alkaline protease and 0.5 to 2 percent of neutral protease;
(5) centrifugal separation and plate-and-frame filter pressing: and (3) centrifugally separating the enzyme-inactivated walnut protein enzymolysis product, carrying out plate-and-frame filter pressing on the obtained clear liquid, and taking the clear filtrate to obtain the high-purity walnut peptide.
The method for industrially producing the high-purity walnut peptide by taking the low-temperature squeezed walnut meal as the raw material further comprises the following steps:
(6) concentrating, drying and packaging: and (5) concentrating the clear filtrate obtained in the step (5), and performing spray drying on the concentrated solution to obtain a walnut peptide finished product.
The water in the present invention is preferably deionized water.
In the step (1):
the cold-pressed walnut meal refers to the walnut meal degreased by adopting a hydraulic cold pressing method.
The concentration of the walnut meal in the walnut protein solution is preferably 5-15% by mass percent.
The shearing condition is preferably shearing at the speed of 2000-3000 rpm for 10-30 min; more preferably, the shear is performed at a speed of 2000rpm to 3000rpm for 10 to 20 min.
In the step (2):
the temperature of the enzymolysis is preferably 45-50 ℃.
The enzymolysis time is preferably 30min to 90 min.
The enzymolysis is preferably carried out in a stirring state, and the stirring speed is preferably lower than 60rpm, and more preferably 18-36 rpm.
In the step (3):
the centrifugation condition is preferably centrifugation at 3000 rpm-6000 rpm for 10 min-20 min; more preferably, the centrifugation is carried out at 3000rpm to 4000rpm for 15min to 20 min.
The amount of the water is preferably such that the precipitate can be sufficiently dispersed, and is preferably 3 to 5 times the mass of the precipitate.
The temperature of the water is preferably 50-55 ℃.
In the step (4):
the amount of the water is preferably enough to fully disperse the precipitate, and is preferably 3-8 times of the mass of the precipitate; more preferably 6 to 8 times.
The temperature of the water is preferably 55 ℃.
The pH value is preferably 8-8.5.
The pH value is preferably adjusted by sodium hydroxide solution or hydrochloric acid solution.
The concentration of the sodium hydroxide solution is preferably 1mol-1～5mol.L-1。
The concentration of the hydrochloric acid solution is preferably 1mol-1～4mol.L-1。
The alkaline protease is preferably at least one of pancreatin, trypsin and bacillus licheniformis protease.
The neutral protease is preferably at least one of neufrasin neutral protease, papain, bromelain and ficin.
The enzymolysis is preferably carried out in a stirring state, and the stirring speed is preferably lower than 60rpm, and more preferably 18-36 rpm.
The enzyme deactivation condition is preferably that the enzyme is deactivated for 10-40 min at 85-95 ℃; more preferably, the enzyme is inactivated at 85-90 ℃ for 15-30 min.
In the step (5):
the centrifugal separation is to pump the walnut protein enzymolysis product after enzyme deactivation into a horizontal decanter centrifuge for centrifugation, discard the residue and take clear liquid.
The plate-frame filter pressing refers to the filter pressing by adopting a plate-frame diatomite filter, so that residual protein and macromolecular peptides in centrifugal clear liquid are further removed, and the components of the micromolecular peptides are enriched.
In the step (6):
the concentration degree is preferably to be 20-45% of the solid content.
The concentration is preferably performed by a double-effect or multi-effect falling film evaporator.
The drying is preferably carried out by a drying tower, and the operation parameters are preferably that the air inlet temperature is 170-200 ℃ and the air outlet temperature is 75-95 ℃.
The method is applied to the industrial production of the high-purity walnut peptide.
A high-purity walnut peptide is prepared by the method.
The high-purity walnut peptide has the protein content of more than 90 percent, the peptide content of more than 85 percent, the relative molecular weight of less than 2000Dal peptide accounts for more than 85 percent, and the relative molecular weight of less than 1000Dal peptide accounts for more than 50 percent.
The high-purity walnut peptide product has good quality and weak bitter taste, can be widely used as a raw material of health food and common food, and particularly can be used for preparing products for strengthening brain, benefiting intelligence and enhancing immunity.
Compared with the prior art, the invention has the following advantages and beneficial effects:
(1) because a certain amount of walnut shells remain in the walnut meal by the high-temperature hot pressing method and are not easy to remove, and the protein has serious excessive denaturation and deep color, the walnut meal by-product of the low-temperature cold pressing method is adopted as the raw material, the process of the low-temperature pressing method is mild, the protein denaturation degree is low, and a substrate guarantee is provided for the preparation of high-quality walnut peptide.
(2) Scientifically, a combined enzyme (tannase, cellulase and medium-temperature amylase) synergistic enzymolysis process is combined with protein denaturation, centrifugation and water washing processes, on one hand, walnut kernel skins, starch and fibers in walnut protein powder are efficiently removed, the purity and flavor of enzymolysis substrate walnut protein powder are improved, wherein the tannase can effectively remove the color and flavor influence generated by the walnut kernel skins, and the cellulase and the amylase can effectively degrade the fibers and starch in the walnut protein powder, so that the protein separation is conveniently neutralized in water washing and centrifugation; on the other hand, the proper denaturation treatment can effectively promote the separation of protein and impurities (mainly carbohydrate), and simultaneously, the compact structure of the walnut protein is opened, the hydrophobicity is reduced, the subsequent enzymolysis is facilitated, the yield of the walnut peptide product is improved, the color and the flavor of the product are improved, and the method is low in energy consumption, convenient to operate and easy to realize industrially.
(3) Because the content of glutamic acid, aspartic acid and arginine in the walnut protein is high, alkaline protease and neutral protease are adopted for enzymolysis step by step in the enzymolysis process to improve the enzymolysis efficiency and the walnut peptide yield, the pH value is adjusted by controlling the initial pH value and the final pH value, the action advantage of enzyme combination can be fully played, excessive acid liquor and alkali liquor are prevented from being introduced, and the production cost is reduced.
(4) After enzymolysis is finished, residual protein and macromolecular peptide in centrifugal clear liquid are removed through centrifugal separation and plate-and-frame filter pressing, and micromolecular peptide components are enriched.
The invention realizes the very economical industrialized preparation of the high-purity walnut peptide through the scientific optimization of the pretreatment, enzymolysis and refining processes, the protein content is more than 90 percent, the peptide content is more than 85 percent, the relative molecular weight is less than 2000Dal peptide and more than 85 percent, the relative molecular weight is less than 1000Dal peptide and more than 50 percent, the product has good quality and weak bitter taste, is easy to match with other food ingredients, has outstanding effects of strengthening brain, benefiting intelligence and enhancing immunity, and can be widely applied to various foods and health care products as a high-quality functional base material. The invention can realize high added value comprehensive utilization of walnut processing by-products, promote economic benefits of walnut deep processing enterprises and promote virtuous circle of walnut planting and processing.
Detailed Description
The present invention will be described in further detail with reference to examples, but the embodiments of the present invention are not limited thereto.
Example 1
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (1.0%) which are equivalent to the mass of walnut meal and are 0.15%, and carrying out enzymolysis for 60min under constant-temperature stirring (36 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring and hydrolyzing at 55 ℃ for 1.5h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for 8rpm, hydrolyzing, and heating to 90 ℃ to inactivate the enzyme for 30min after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Example 2
Dispersing and hydrating walnut pulp, namely preparing 5 mass percent walnut protein solution from low-temperature squeezed and degreased walnut pulp (the protein content is more than or equal to 55 percent and the grease content is less than or equal to 15 percent on a dry basis) by using deionized water, and dispersing and hydrating the walnut protein powder for 10min by using a stirrer at the rotating speed of 3000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 45 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (0.5 percent) which are equivalent to the mass of walnut meal, and carrying out enzymolysis for 90min at constant temperature and stirring (18 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 121 ℃ for 0.5h, centrifuging for 20min at 3000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 3 times of deionized water at 55 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.5, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 0.5 percent of the mass of the protein in the walnut protein slurry, stirring and hydrolyzing at 50 ℃ for 2 hours, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 0.5 percent of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for hydrolysis at 18rpm for 16 hours, and heating to 85 ℃ to inactivate the enzyme for 40 minutes after the enzymolysis is finished;
(5) centrifugal separation, centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 20%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 170 ℃ and the outlet temperature to be 75 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Example 3
(1) Dispersion and hydration of walnut meal: preparing walnut protein solution with the mass percent of 15% by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (2.0%) which are equivalent to the mass of walnut meal and are 0.2% of the mass of the walnut meal, and carrying out enzymolysis for 30min under constant-temperature stirring (25 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 5 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 8 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.5, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 2.0% of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for hydrolysis for 1h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 2.0% of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for hydrolysis at 25rpm for 4h, heating to 90 ℃ after the enzymolysis is finished, and inactivating the enzyme for 30 min;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid state of 45%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 190 ℃ and the outlet temperature to be 80 ℃, and packaging after drying to obtain the walnut peptide finished product.
Example 4
(1) Dispersion and hydration of walnut meal: preparing 12% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and moderate temperature amylase (1.5%) which are equivalent to the mass of walnut meal and are 0.15%, and carrying out enzymolysis for 60min under constant temperature stirring (36 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4.5 times of 50 ℃ deionized water, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 7 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0% of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for hydrolysis for 2h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 1.0% of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for hydrolysis at 36rpm for 8h, and heating to 90 ℃ to inactivate the enzyme for 30min after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 40%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 200 ℃ and the outlet temperature to be 90 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 1
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) enzymolysis with cellulase and medium-temperature amylase: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding cellulase (neutral, Beijing Xiasang Biotech development Co., Ltd.) with the mass equivalent to 0.5% of the walnut meal and 1.0% moderate temperature amylase (Huaxing bioengineering Co., Ltd. in Hunan century), and stirring at constant temperature (36rpm) for enzymolysis for 60 min;
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring and hydrolyzing at 55 ℃ for 1.5h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for 8rpm, hydrolyzing, and heating to 90 ℃ to inactivate the enzyme for 30min after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 2
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase and medium-temperature amylase enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (Huaxing bioengineering Co., Ltd., Hunan century) which is 0.15 percent of the mass of the walnut meal and medium temperature amylase (Huaxing bioengineering Co., Ltd., Hunan century) which is 1.0 percent of the mass of the walnut meal, and stirring at constant temperature (36rpm) for enzymolysis for 60 min;
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring and hydrolyzing at 55 ℃ for 1.5h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for 8rpm, hydrolyzing, and heating to 90 ℃ to inactivate the enzyme for 30min after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 3
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase and cellulase enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (Huaxing bioengineering Co., Ltd., Hunan century) which is 0.15 percent of the mass of the walnut meal and cellulase (neutral, Beijing Xiusheng Biotechnology development Co., Ltd.), stirring at constant temperature (36rpm) for enzymolysis for 60 min;
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring and hydrolyzing at 55 ℃ for 1.5h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for 8rpm, hydrolyzing, and heating to 90 ℃ to inactivate the enzyme for 30min after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 4
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase and medium-temperature amylase enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (Huaxing bioengineering Co., Ltd., Hunan century) which is 0.15 percent of the mass of the walnut meal and medium temperature amylase (Huaxing bioengineering Co., Ltd., Hunan century) which is 1.5 percent of the mass of the walnut meal, and stirring at constant temperature (36rpm) for enzymolysis for 60 min;
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring and hydrolyzing at 55 ℃ for 1.5h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for 8rpm, hydrolyzing, and heating to 90 ℃ to inactivate the enzyme for 30min after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 5
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and enzymatic hydrolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (Huaxing bioengineering Co., Ltd., Hunan century) which is 0.15 percent of the mass of the walnut meal and cellulase (neutral, Beijing Xiusheng Biotechnology development Co., Ltd.), stirring at constant temperature (36rpm) for enzymolysis for 60 min;
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring and hydrolyzing at 55 ℃ for 1.5h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for 8rpm, hydrolyzing, and heating to 90 ℃ to inactivate the enzyme for 30min after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 6
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (1.0%) which are equivalent to the mass of walnut meal and are 0.15%, and carrying out enzymolysis for 60min under constant-temperature stirring (36 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 90 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of 50 ℃ deionized water, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring and hydrolyzing at 55 ℃ for 1.5h, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is equivalent to 1.0 percent of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for 8rpm, hydrolyzing, and heating to 90 ℃ to inactivate the enzyme for 30min after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 7
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (1.0%) which are equivalent to the mass of walnut meal and are 0.15%, and carrying out enzymolysis for 60min under constant-temperature stirring (36 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 1.0% of the mass of protein in the walnut protein slurry, stirring at 55 ℃ for 9.5 hours, hydrolyzing, and heating to 90 ℃ for enzyme deactivation for 30 minutes after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 8
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (1.0%) which are equivalent to the mass of walnut meal and are 0.15%, and carrying out enzymolysis for 60min under constant-temperature stirring (36 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 7.0, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is 1.0% of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for hydrolysis at 36rpm for 9.5h, and heating to 90 ℃ after the enzymolysis is finished to inactivate the enzyme for 30 min;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 9
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (1.0%) which are equivalent to the mass of walnut meal and are 0.15%, and carrying out enzymolysis for 60min under constant-temperature stirring (36 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment Limited) which is equivalent to 2.0% of the mass of protein in the walnut protein slurry, stirring at 55 ℃ for 9.5 hours, hydrolyzing, and heating to 90 ℃ for enzyme deactivation for 30 minutes after the enzymolysis is finished;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 10
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (1.0%) which are equivalent to the mass of walnut meal and are 0.15%, and carrying out enzymolysis for 60min under constant-temperature stirring (36 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 7.0, adding neutral protease (neutral protease Neutrase, Nuoweixin (China) investment Limited) which is 2.0% of the mass of the protein in the walnut protein slurry, stirring at 55 ℃ for hydrolysis at 36rpm for 9.5h, and heating to 90 ℃ after the enzymolysis is finished to inactivate the enzyme for 30 min;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Comparative example 11
(1) Dispersion and hydration of walnut meal: preparing 10% walnut protein solution by using deionized water to squeeze the degreased walnut meal at low temperature (the protein content is more than or equal to 55% and the grease content is less than or equal to 15% in terms of dry basis), and dispersing and hydrating the walnut protein powder for 20min by using a stirrer at the rotating speed of 2000 rpm;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 50 ℃, adding tannase (China bioengineering, Inc. of Hunan century), cellulase (neutral, Beijing Xiusheng biotechnological development, Inc.) and medium temperature amylase (1.0%) which are equivalent to the mass of walnut meal and are 0.15%, and carrying out enzymolysis for 60min under constant-temperature stirring (36 rpm);
(3) protein denaturation, centrifugation, water washing: pumping the walnut protein solution treated in the step (2) into a tank, treating at 100 ℃ for 1.0h, centrifuging for 15min at the rotating speed of 4000r/min by using a centrifugal separator, removing fat and liquid, collecting precipitate, adding 4 times of deionized water at 50 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) enzymolysis and enzyme deactivation: adding 6 times of deionized water at 55 ℃ into the precipitate obtained in the step (3), uniformly stirring, adjusting the pH value of the walnut protein slurry to 8.0, adding alkaline protease (Alcalase2.4L, Nuoweixin (China) investment limited company) which is equivalent to 1.0% of the mass of protein in the walnut protein slurry and neutral protease (neutral protease Neutrase, Nuoweixin (China) investment limited company) which is 1.0% of the mass of protein in the walnut protein slurry, stirring and hydrolyzing at 55 ℃ for 9.5h, and heating to 90 ℃ after the enzymolysis is finished to inactivate the enzyme for 30 min;
(5) centrifugal separation and plate-and-frame filter pressing: pumping the walnut protein slurry after enzyme deactivation into a horizontal screw centrifuge for centrifugation, and performing filter pressing on the centrifuged clear liquid by a plate and frame type diatomite filter;
(6) concentrating, drying and packaging: and (4) concentrating the filtrate obtained in the step (5) to a solid form of 30%, then carrying out spray drying on the concentrated solution, controlling the inlet temperature to be 180 ℃ and the outlet temperature to be 85 ℃, finishing drying, and packaging to obtain the walnut peptide finished product.
Effects of the embodiment
The walnut peptide powder prepared in the examples 1 to 4 and the comparative examples 1 to 11 is detected as follows: measuring the color, moisture, ash content and protein content of the walnut peptide powder by methods specified in GB/T5492, GB/T5009.3, GB/T5009.4, GB/T5009.5 and GB/T5009.6 respectively; adding semen Juglandis peptide powder into 50 deg.C pure water to obtain 3% solution, tasting its taste and smell; analyzing the content of the walnut peptide powder peptide by adopting a method specified in GB/T22729-2008 at 6.3; the proportion of the walnut peptide powder with the relative molecular weight of less than 2000Dal and 1000Dal peptides is analyzed by adopting GB/T22729-2008 appendix A high-efficiency gel filtration chromatography, and the result is shown in tables 1-3.
TABLE 1 analysis and detection results of walnut peptide powder prepared in examples 1 to 4
Note: wherein, the walnut peptide powder prepared in the embodiments 1 to 4 is marked as 1#, 2#, 3#, 4# respectively.
TABLE 2 analysis and detection results of walnut peptide powder prepared in comparative examples 1-5
Note: wherein, the walnut peptide powder prepared by the comparative examples 1 to 5 is respectively marked as 5#, 6#, 7#, 8# and 9 #.
Table 3 analysis and detection results of walnut peptide powder prepared in comparative examples 6-11
Note: wherein, the walnut peptide powder prepared by the comparative examples 6 to 10 is respectively marked as 10#, 11#, 12#, 13#, 14#, and 15 #.
As can be seen from Table 1, the walnut peptides prepared by the methods of examples 1 to 4 of the present invention have protein mass percentages of 91.88-93.12% (all being equal to or greater than 90%), peptide contents of 86.95-88.21% (all being equal to or greater than 85%), relative molecular weights of less than 2000Dal peptides of all being equal to or greater than 85%, relative molecular weights of less than 1000Dal peptides of all being equal to or greater than 50%, and are high-quality high-purity protein peptide products.
As shown in tables 2 to 3, compared with the example 1, the comparative example 1 has no tannase, the product has dark yellow color and bitter taste, the peptide content and the relative molecular weight are less than 2000Dal and 1000Dal, and the addition of the tannase can improve the color and the flavor of the product. Comparative example 2 is not added with cellulase, comparative example 3 is not added with medium temperature amylase, the obtained product has dark yellow color and luster, the flavor is not different from that of example 1, the protein content, the peptide content and the relative molecular weight are less than 2000Dal and 1000Dal peptide ratio are reduced, the reduction of comparative example 3 is more obvious, and the addition of the cellulase and the medium temperature amylase can improve the protein content, the peptide content and the small molecular peptide content of the product. Comparative example 4 no cellulase is added, the dosage of the medium temperature amylase is increased to 1.5%, the obtained product has a dark yellow color, the flavor is not greatly different from that of comparative example 2, the protein content, the peptide content and the relative molecular weight are less than 2000Dal, and 1000Dal peptides account for the reduction of comparative example 1, and are not obviously different from comparative example 2; comparative example 5 no medium temperature amylase is added, the dosage of the cellulase is increased to 1.5%, the obtained product is light yellow and deepened, the flavor is not different from that of comparative example 3, the protein content, the peptide content and the relative molecular weight of peptides are less than 2000Dal and 1000Dal peptides account for the reduction of comparative example 1, and the difference from comparative example 3 is not obvious; therefore, the effect of singly adopting the cellulase and the medium-temperature amylase to remove the carbohydrate from the walnut protein substrate is not good when the cellulase and the medium-temperature amylase are combined, and the combination has obvious synergistic effect. Comparative example 6 only changes the walnut protein solution denaturation condition to be that the walnut protein solution is treated at the temperature of 90 ℃ for 1.0h, compared with example 1, the color, the flavor and the protein content are not obviously changed, but the peptide content and the peptide proportion of the relative molecular weight of less than 2000Dal and 1000Dal are obviously reduced, and the walnut protein denaturation degree in the cold-pressed walnut pulp has obvious influence on the characteristics of the enzymolysis products. Compared with the prior art, the neutral protease is not added in the comparative example 7, the alkaline protease is not added in the comparative example 8 (the initial enzymolysis pH is adjusted to be 7.0), the color, the flavor and the protein content are not obviously different from those in the example 1, but the peptide content is reduced, the relative molecular weight is less than 2000Dal, the ratio of 1000Dal peptides is obviously reduced, and the selection of the complex enzyme is favorable for improving the peptide content and the small molecular peptide content. Comparative example 9 has no neutral protease added, the dosage of the alkaline protease is increased to 2.0%, the color, the flavor and the protein content of the product are not greatly different from those of example 1, the peptide content and the relative molecular weight of peptides are less than 2000Dal and 1000Dal are reduced compared with comparative example 1, and the difference from comparative example 7 is not obvious; comparative example 10 without adding alkaline protease, the neutral protease dosage increased to 2.0% (initial enzymolysis pH adjusted to 7.0), product color, flavor, protein content and example 1 were not greatly different, peptide content and relative molecular weight less than 2000Dal, 1000Dal peptide account for comparative example 1 and was not obvious different from comparative example 8; therefore, the effect of singly adopting the alkaline protease and the neutral protease to prepare the walnut peptide by enzymolysis of the walnut protein is not good when the alkaline protease and the neutral protease are combined and used, and the combination has obvious synergistic effect. In the comparative example 11, the alkaline protease and the neutral protease are subjected to synchronous enzymolysis, the color, the flavor and the protein content are not obviously different from those in the example 1, but the peptide content is reduced, the peptide proportion of the peptides with the relative molecular weights of less than 2000Dal and 1000Dal is obviously reduced, the peptide content and the peptide proportion of the peptides with the relative molecular weights of less than 2000Dal and 1000Dal are close to those in the comparative example 7, so that the influence of the adding sequence of the complex enzyme on the physicochemical properties of the walnut peptides is obvious, and the synchronous enzymolysis synergistic effect of the alkaline protease and the neutral protease is not obvious.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Claims (7)
1. A method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut meal as a raw material is characterized by comprising the following steps:
(1) dispersion and hydration of walnut meal: weighing cold-pressed walnut meal, and preparing walnut protein solution by using water; shearing, dispersing and hydrating the walnut protein solution uniformly;
(2) tannase, cellulase and medium-temperature amylase for enzymolysis: heating the dispersed and hydrated walnut protein solution to 45-55 ℃, adding tannase, cellulase and medium-temperature amylase, and carrying out enzymolysis; the mass dosage of each enzyme is calculated by taking the mass of the walnut pulp as a reference, and the mass dosage is as follows: 0.1-0.2% of tannase, 0.1-1.0% of cellulase and 0.5-2.0% of medium-temperature amylase;
(3) protein denaturation, centrifugation, water washing: treating the walnut protein solution subjected to enzymolysis in the step (2) at 100-121 ℃ for 0.5-1.0 h, centrifuging to remove upper-layer liquid, collecting precipitate, adding water at 50-60 ℃, uniformly stirring, centrifuging to remove upper-layer liquid, and collecting precipitate;
(4) and (3) protease enzymolysis and enzyme deactivation step by step: adding water with the temperature of 50-60 ℃ into the precipitate obtained in the step (3), and uniformly stirring to obtain walnut protein slurry; adjusting the pH value of the walnut protein slurry to 7.5-8.5, firstly adding alkaline protease, stirring and hydrolyzing at 50-55 ℃ for 1-2 h, then adding neutral protease, stirring and hydrolyzing at 50-55 ℃ for 4-16 h; inactivating enzyme after enzymolysis; the mass dosage of each enzyme is calculated by taking the mass of the protein in the walnut protein slurry as a reference, and the mass dosage is as follows: 0.5 to 2 percent of alkaline protease and 0.5 to 2 percent of neutral protease;
(5) centrifugal separation and plate-and-frame filter pressing: centrifugally separating the enzyme-inactivated walnut protein enzymolysis product, carrying out plate-and-frame filter pressing on the obtained clear liquid, and taking the clear filtrate to obtain high-purity walnut peptide;
the alkaline protease in the step (4) is Novoverin Alcalase 2.4L;
the neutral protease in the step (4) is neufrasin neutral protease Neutrase.
2. The method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut pulp as a raw material according to claim 1, characterized by further comprising the following steps:
(6) concentrating, drying and packaging: and (5) concentrating the clear filtrate obtained in the step (5), and performing spray drying on the concentrated solution to obtain a walnut peptide finished product.
3. The method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut pulp as a raw material according to claim 1 or 2, which is characterized by comprising the following steps:
the concentration of the walnut meal in the walnut protein solution in the step (1) is 5-15% by mass percent;
the shearing condition in the step (1) is shearing at the speed of 2000-3000 rpm for 10-30 min.
4. The method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut pulp as a raw material according to claim 1 or 2, which is characterized by comprising the following steps:
the enzymolysis conditions in the step (2) are as follows: the temperature is 45-50 ℃, and the time is 30-90 min;
the enzymolysis in the step (2) and the step (4) is carried out in a stirring state, and the stirring speed is lower than 60 rpm.
5. The method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut pulp as a raw material according to claim 1 or 2, which is characterized by comprising the following steps:
the centrifugation condition in the step (3) is centrifugation at 3000 rpm-6000 rpm for 10 min-20 min;
the amount of the water used in the step (3) is 3-5 times of the mass of the precipitate;
the temperature of the water in the step (3) is 50-55 ℃;
the amount of the water used in the step (4) is 3-8 times of the mass of the precipitate;
the temperature of the water in the step (4) is 55 ℃;
the pH value in the step (4) is 8-8.5;
and (4) inactivating the enzyme at 85-95 ℃ for 10-40 min.
6. The method for industrially producing high-purity walnut peptide by using low-temperature squeezed walnut pulp as a raw material according to claim 2, characterized by comprising the following steps:
the concentration degree in the step (6) is to be concentrated until the solid content is 20-45%;
the drying in the step (6) is drying through a drying tower, and the operation parameters are as follows: the air inlet temperature is 170-200 ℃, and the air outlet temperature is 75-95 ℃.
7. Use of the method of any one of claims 1 to 6 for the industrial production of high purity walnut peptides.
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| CN109022525A (en) * | 2018-08-14 | 2018-12-18 | 吉林农业大学 | A kind of walnut active peptide and preparation method thereof improving memory function |
| CN109055472A (en) * | 2018-09-21 | 2018-12-21 | 滁州学院 | A kind of complex enzyme production method of walnut peptide |
| CN109527163A (en) * | 2018-11-21 | 2019-03-29 | 云南省林业科学院 | A kind of walnut polypeptide coffee and preparation method thereof |
| CN109845876A (en) * | 2018-12-28 | 2019-06-07 | 广州合诚实业有限公司 | Highly purified soybean oligopeptide and preparation method and application with unique solubility for high energy |
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| CN110819674A (en) * | 2019-11-14 | 2020-02-21 | 江南大学 | Method for preparing antihypertensive peptide by enzymolysis of walnut meal with compound protease |
| CN110710592A (en) * | 2019-11-16 | 2020-01-21 | 四川农业大学 | Method for improving antioxidant activity of walnut cake protein |
| CN112841393B (en) * | 2020-12-30 | 2022-11-15 | 广州合诚实业有限公司 | Walnut peptide stable to zinc ions as well as preparation method and application thereof |
| CN112868877A (en) * | 2021-01-22 | 2021-06-01 | 昆明生物制造研究院有限公司 | Method for preparing walnut modified protein by using extrusion puffing technology |
| CN112753925A (en) * | 2021-01-27 | 2021-05-07 | 昆明生物制造研究院有限公司 | Preparation method of walnut kernel skin solid beverage |
| CN113057248A (en) * | 2021-04-14 | 2021-07-02 | 王朝辉 | Extraction method of walnut peptide |
| CN113854398A (en) * | 2021-10-08 | 2021-12-31 | 长春大学 | Method for preparing digestion-free walnut protein by using walnut cake meal as raw material |
| CN116076699A (en) * | 2021-11-05 | 2023-05-09 | 安琪生物科技有限公司 | Plant raw material meal enzymolysis product and preparation method and application thereof |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2012149959A1 (en) * | 2011-05-03 | 2012-11-08 | Nestec S.A. | A hydrolysate of a protein substrate and a process for producing thereof |
| CN104293870A (en) * | 2014-09-25 | 2015-01-21 | 汾州裕源土特产品有限公司 | Preparation method of antioxidant walnut polypeptide health product |
| CN105002247A (en) * | 2015-07-30 | 2015-10-28 | 北京天肽生物科技有限公司 | Micromolecule walnut peptide and preparation method thereof |
| CN105063152A (en) * | 2015-09-10 | 2015-11-18 | 无限极（中国）有限公司 | Polypeptide raw material prepared by enzymolysis of walnuts and application thereof |
-
2017
- 2017-12-27 CN CN201711444692.9A patent/CN108085356B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2012149959A1 (en) * | 2011-05-03 | 2012-11-08 | Nestec S.A. | A hydrolysate of a protein substrate and a process for producing thereof |
| CN104293870A (en) * | 2014-09-25 | 2015-01-21 | 汾州裕源土特产品有限公司 | Preparation method of antioxidant walnut polypeptide health product |
| CN105002247A (en) * | 2015-07-30 | 2015-10-28 | 北京天肽生物科技有限公司 | Micromolecule walnut peptide and preparation method thereof |
| CN105063152A (en) * | 2015-09-10 | 2015-11-18 | 无限极（中国）有限公司 | Polypeptide raw material prepared by enzymolysis of walnuts and application thereof |
Non-Patent Citations (1)
| Title |
|---|
| 果汁中单宁脱除方法的研究进展;张磊 等;《食品科学》;20100201;第31卷(第3期);第312-315页 * |
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