A desensitizing wheat gluten peptide, and a preparation method and application thereof

Abstract

The application discloses a desensitization wheat gluten peptide and a preparation method and application thereof, and belongs to the field of food deep processing. The desensitization wheat gluten peptide is obtained by modifying and screening wheat gluten by means of joint methods including enzymolysis, ultrasonic, ultrafiltration segmentation and microwave. The wheat gluten peptide is fine and has uniform granularity, color and flavor, and can be directly used as a functional food, and has wide application value in the field of food processing industry and snacks. The application constructs a compound nutrient powder by using the wheat gluten peptide and starch, and the processing and sensory characteristics, nutritional property and antioxidant property of the compound nutrient powder are improved, and the allergenicity is reduced, so that the compound nutrient powder meets the international gluten-free (<10ppm) food standard and can be applied to food industry low-gluten cereal food base. The combined processing of various food processing shows a good synergistic effect, improves the comprehensive performance of the food system, and provides technical guidance and a good production model for preparing gluten-free food with good color, fragrance and flavor and health.

Description

Technical Field

[0001] This invention belongs to the field of food deep processing and also to the field of functional foods, and specifically relates to a desensitizing wheat gluten peptide, its preparation method and application. Background Technology

[0002] Wheat flour consists of 70-85% starch (including damaged starch) and 10-15% protein, of which 73-82% is gluten, composed of different varieties of gliadin and glutenin in varying proportions. Wheat flour is rich in nutrients and has many functional activities, such as the shaping function produced by the network formed by wheat gluten, and the unique aroma, color, and flavor qualities produced by the Maillard reaction under heat processing. Therefore, wheat flour is widely used in the production of various breads, cakes, biscuits, puffed foods, and fried doughs.

[0003] However, some individuals are sensitive to wheat gluten (a group of mixed proteins composed of gliadin and glutenin), which can easily lead to celiac disease, Becker's asthma, allergies, and even allergic mental illness. In particular, gliadin exhibits resistance to hydrolysis by gastrointestinal and brush membrane (BBM) enzymes, and its 13-mer α-gliadin peptide p31-43 can induce pro-inflammatory responses, potentially leading to celiac disease and innate immunity. Furthermore, products made from wheat flour (a protein-to-starch mixture) often have a high glycemic index (GI) due to the high digestibility of wheat starch, increasing the risk of diseases such as diabetes.

[0004] Regarding protein sensitization, enzymatic hydrolysis of proteins using specific enzymes yields peptide fragments with specific sequences (another common form of protein). Most enzymatically hydrolyzed peptides alter the original protein's spatial structure to varying degrees, losing the complex higher-order structures of protein subunits and multiple subunits, thereby changing the spatial structure and even the sequence of the sensitizing region of the original protein. Specific and non-specific screening can yield active peptides with certain functions and desensitizing properties. Ultrasonic treatment of proteins can generate cavitation through the penetration of high-frequency mechanical sound waves into the propagation medium. The collapse of these bubbles creates localized high pressure and temperature within the food matrix, inducing disulfide bond recombination or altering intermolecular forces, thus rearranging the protein's higher-order structure and changing its sensitization. For starch, processes such as controlling heat, moisture, retrogradation, enzymatic hydrolysis, and even modifying the starch structure can increase the proportion of amylose, increase the number of double helix structures, improve the B-type crystal form, and enhance crystallinity, making the starch structure more compact, improving its resistance to digestion, and slowing down the glycemic effect. Combination formulations can make it possible to create foods with unique advantages through the interaction of different components. For example, adding some polysaccharides can coat the protein structure, thereby masking its allergenic sites or areas; adding complex starches such as proteins, lipids, and polyphenols can change the binding sites of starch and degrading enzymes, thus reducing digestibility.

[0005] While food processing and compounding are essential steps in the general formulation of food products, effectively controlling food-induced allergic reactions and altering digestibility, formulated foods, as a more advanced stage of natural food ingredients, involve more complex production processes compared to raw materials and minimally processed products. Formulated foods combine multiple food base materials in specific proportions, utilizing specialized processing or cooking techniques to create food products with enhanced sensory qualities (color, aroma, and taste) and unique nutritional and functional advantages. However, the current understanding and mechanisms of simultaneously compounding multiple components to synergistically modify food base materials through combined processing are still unclear. It is difficult to integrate the advantages of multiple methods and apply theory to practice; most formulation and modification processes remain in the exploratory stage. Appropriately combining various processes and modifying the main formulation can provide feasibility for grain foods or their substitutes to fully achieve the combined advantages.

[0006] Therefore, drawing on the concept of formulated foods, and based on the main components of wheat flour, this approach involves deconstructing the entire system, combining multiple processing methods to synergistically modify the components, optimize and reconstruct the overall system, thereby eliminating the defects of the original system while preserving and enhancing the food's color, aroma, flavor, and nutritional functions. This method provides guidance and ideas for preparing the globally popular gluten-free, non-allergenic foods (i.e., foods with no (low) gluten or (low) gluten-free adverse reactions prepared through other non-grain ingredients and combined processing, with improved sensory quality and nutritional functions), and for enhancing the overall quality of food. Summary of the Invention

[0007] To overcome the shortcomings and deficiencies of existing materials and technologies, the primary objective of this invention is to provide a method for preparing desensitized wheat glutenin peptides. This invention utilizes a combination of multiple processing methods, including enzymatic hydrolysis, ultrasound, and ultrafiltration, to modify and screen wheat glutenin, thereby obtaining desensitized wheat glutenin peptides.

[0008] Another objective of this invention is to provide a desensitizing wheat gluten peptide prepared by the above-described preparation method.

[0009] Another objective of this invention is to provide a compound nutritional powder containing the above-mentioned desensitizing wheat gluten peptides.

[0010] Another object of the present invention is to provide the application of the above-mentioned desensitized wheat gluten peptides in the deep processing of food, especially in gluten-free products.

[0011] The objective of this invention is achieved through the following technical solution:

[0012] A method for preparing desensitizing wheat gluten peptides includes the following steps:

[0013] (1) Wheat gluten was dissolved in PBS phosphate buffer solution, and the mixture was subjected to ultrasonic cavitation and enzymatic hydrolysis to obtain mixed peptides;

[0014] (2) The mixed peptides obtained in step (1) are subjected to enzyme inactivation treatment, and then ultrafiltration is used to obtain desensitized wheat gluten peptides.

[0015] In step (1), the pH of the PBS phosphate buffer solution is 2.0-11.0, preferably 6.0; the concentration of the PBS phosphate buffer solution is 0.01-1.0M, preferably 0.2M.

[0016] In step (1), the ultrasonic power of ultrasonic cavitation is 100-600W, preferably 250W; the ultrasonic cavitation time is 25-60min, preferably 30min.

[0017] In step (1), the enzymatic hydrolysis can be performed using at least one of the following enzymes: Protamex complex protease, papain, alkaline protease, trypsin, neutral protease, etc.

[0018] Preferably, the enzymatic hydrolysis is performed using Protamex complex protease.

[0019] Preferably, the mass ratio of protease to wheat gluten in the enzymatic hydrolysis is (0.2-1.5):100, more preferably (0.5-1.0):100; and most preferably 0.81:100.

[0020] Preferably, the enzymatic hydrolysis time is 3-24 hours, more preferably 6 hours.

[0021] Preferably, the temperature of the enzymatic hydrolysis treatment is 25-60℃, more preferably 40℃.

[0022] In step (2), the specific steps of the enzyme inactivation treatment are as follows: inactivate the mixed peptides in a water bath at 80-100℃ for 10-90 min; further inactivate the mixed peptides in a water bath at 95℃ for 1 h.

[0023] Preferably, the mixed peptides can be separated by centrifugation after enzyme inactivation treatment, and then subjected to ultrafiltration to obtain the supernatant.

[0024] Preferably, the centrifugation separation is carried out at a temperature of 4℃-30℃ and a rotation speed of 2000-12000 r / min for 5-30 min. More preferably, it is carried out at 25℃ and 9500 r / min for 20 min.

[0025] In step (2), after ultrafiltration and segmentation, the wheat gluten peptides are obtained with a molecular weight preferably 30K or less, more preferably 8-30K or <8K; and even more preferably 8-30K.

[0026] Preferably, the prepared desensitized wheat gluten peptides can be freeze-dried.

[0027] This invention modifies wheat glutenin through a combination of ultrasonic physical modification and enzymatic modification, altering the active regions or corresponding epitopes and expression sequences of the original sensitizing sites. Simultaneously, ultrafiltration technology is used to screen wheat glutenin peptides by molecular weight to obtain peptides with high functional activity, ultimately yielding wheat glutenin peptides with excellent flavor, good nutrition, antioxidant properties, and low allergenicity.

[0028] The present invention also provides a desensitizing wheat gluten peptide prepared by the above preparation method.

[0029] The present invention also provides an application of the above-mentioned desensitizing wheat gluten peptide in the deep processing of food, especially in gluten-free products.

[0030] The present invention also provides a compound nutritional powder containing the above-mentioned desensitizing wheat gluten peptide, which is obtained by heat treatment after uniform mixing of desensitizing wheat gluten peptide and starch.

[0031] Preferably, the starch is a cereal starch, further including at least one of wheat starch and rice starch, with rice starch being preferred in terms of flavor and wheat starch being preferred in terms of nutritional function.

[0032] Preferably, the mass ratio of the desensitizing wheat gluten peptide to starch is 1:(0.2-3), more preferably 1:(1-2); and even more preferably 1:1.

[0033] The starch can be gelatinized, enzymatically hydrolyzed, freeze-dried, and then reacted with desensitized wheat gluten peptides.

[0034] The gelatinization process specifically involves dissolving starch in dilute acid or heat-treating it to gelatinize it into a starch solution.

[0035] Preferably, the dilute acid can be an acetate-sodium acetate buffer solution, more preferably a 0.2M acetate-sodium acetate buffer solution with pH 6.0.

[0036] Preferably, the starch solution concentration is 1-10% (w / v), more preferably 5% (w / v).

[0037] Preferably, the enzymatic hydrolysis of starch can be performed using amylase, more preferably α-thermal amylase (20000U / mL).

[0038] Preferably, the amount of α-high-temperature amylase is added to the system at 100-200 U / mL / g starch. More preferably, it is added to a 5% (w / v) starch solution at a ratio of 120 U / mL / g starch, that is, 1.25g starch to 7.5μL enzyme is added to the gelatinized starch solution.

[0039] Preferably, the amount of amylase added to the system is 100-200 U / mL / g starch, more preferably 120 U / mL / g starch.

[0040] Preferably, the enzymatic hydrolysis temperature is 60-85℃, more preferably 60-65℃ water bath enzymatic hydrolysis.

[0041] Preferably, the enzymatic hydrolysis time is 30-150 min, more preferably 40-120 min.

[0042] Preferably, the starch after enzymatic hydrolysis can be freeze-dried.

[0043] Preferably, the heat treatment can be performed by microwave heating or hydrothermal treatment.

[0044] Preferably, the mixing and homogenization can be achieved by stirring, grinding, or vortex oscillation.

[0045] Preferably, the microwave heating conditions are: water content 0-50% and not 0, relative humidity 60-90%, heating at 300-800W for 0-80 minutes and not 0%, more preferably: water content 10-50%, relative humidity 60-90%, heating at 360-600W for 5-70 minutes, and even more preferably: water content 15%, relative humidity 79%, heating at 600W for 15 minutes.

[0046] The heating is non-continuous, with a 30-90s interval between heatings of 3-8 minutes; further, with a 30-90s interval (preferably 30s) between heatings of 5 minutes.

[0047] Preferably, the hydrothermal treatment conditions are as follows: at a temperature of 30-95℃, the concentration of wheat glutenin peptide is 0.1-5% (w / v), the concentration of starch is 0.1-5% (w / v), and the mass ratio of wheat glutenin peptide to starch is 5:1-1:5, heated for 0-3 hours and not 0; more preferably, at a heating temperature of 90℃, the concentration of wheat glutenin peptide is 0.5% (w / v), the concentration of starch is 0.5% (w / v), and the mass ratio of wheat glutenin peptide to starch is 1:1, heated for 2 hours.

[0048] Preferably, the compound nutritional powder can be cooled, pulverized and sieved after heat treatment to form compound nutritional powder or molded into low (or gluten-free) compound nutritional basic model biscuits with the desired shape and structure.

[0049] Preferably, the compound nutritional powder may also have added excipients as needed. These excipients are conventional food processing ingredients or additives, including at least one of sugar, eggs, milk, oil, salt, grains, cocoa, food flavorings, food additives, and preservatives.

[0050] This invention utilizes thermally initiated Maillard saccharification to generate wheat gluten peptides and starch flavor, color, and antioxidant properties. The resulting compound nutritional powder has the characteristics of moderate color (light yellow to light brown), good flavor, antioxidant properties, anti-digestive properties, and low allergenicity.

[0051] This invention produces wheat gluten peptides with fine particles, uniform particle size and color, and good flavor through combined processing. The product is natural, safe, healthy, and nutritious, and can be directly used as a base for cereal foods or shortbread biscuits, with broad application value in the food processing industry and snack food fields. Allergic reactions and symptoms caused by wheat gluten, especially in baked goods, and hyperglycemia induced by the easy digestibility of starch, can harm human health. This invention produces different wheat gluten proteins and their hydrolysates through a combined ultrasonic and enzymatic processing method, which are then mixed with starch (from wheat and rice) or starch hydrolysates. The mixture is then microwaved or hydrothermally heated to prepare Maillard complex products for baked cereal-based or biscuit molds. Due to the high glutamic acid content and the generation of appropriate Maillard products during combined processing, the product exhibits a light yellow to light brown sensory color, a suitable baked cereal aroma, strong antioxidant capacity, moderate digestibility, and low immunogenicity (wheat gluten content meets multiple national standards, i.e., <10ppm). The combined processing of multiple food ingredients improves the overall performance of this food system, demonstrating a good synergistic effect. It provides technical guidance for the preparation of (low) gluten-free foods that are delicious, flavorful, and healthy, and offers a good example for production.

[0052] The present invention has the following advantages and effects compared with the prior art:

[0053] (1) The present invention modifies wheat gluten, which was originally directly processed, by a combination of physical ultrasound and enzymatic methods. By changing the allergenic sequence and allergenic structural region, the allergic reaction caused by allergens is reduced. At the same time, peptide screening is carried out, and the wheat gluten peptides obtained have good flavor and color and high antioxidant properties.

[0054] (2) The present invention uses modified wheat gluten peptides and staple starch as basic materials to construct a compound nutritional powder, which improves the processing characteristics, nutritional and sensory characteristics of the compound, and also improves the combined antioxidant properties of the compound. It can be applied to low (gluten-free) cereal food base in the food processing industry, which can enhance nutritional characteristics and extend shelf life.

[0055] (3) The present invention uses staple starch and degradation products as compounding agents to improve the plasticity, environmental resistance, energy supply and satiety of compound nutritional powder, so that the blood glucose value of wheat gluten peptide compound nutritional powder can be significantly improved, and the sensory quality is improved after compounding. The synergistic effect of wheat gluten peptide and starch further enhances the edible characteristics. Attached Figure Description

[0056] Figure 1 This is a schematic diagram of the preparation process of the present invention containing wheat gluten peptides and compound nutritional powder.

[0057] Figure 2The absorbance values ​​of the Maillard intermediate and final products of the composite nutritional powders of wheat gluten and different starch hydrolysates in Comparative Examples 1-4 of this invention are shown.

[0058] Figure 3 The potential and antioxidant properties (ability to eliminate ABTS free radicals) of wheat glutenin peptides prepared under different treatments in Comparative Examples 1, 5-6 of this invention are shown.

[0059] Figure 4 The image shows the physical product of the composite nutrient powder prepared in Comparative Examples 1, 5-6 of this invention.

[0060] Figure 5 The absorbance values ​​and corresponding antioxidant properties (ability to eliminate ABTS free radicals) of Maillard intermediates and final products of different treated protein-starch complex nutrients in Comparative Examples 1, 5-6 of this invention are shown.

[0061] Figure 6 The absorbance values ​​of Maillard intermediates and final products of wheat gluten peptides and different starch compound nutritional powders prepared by different heating methods (dry method and wet method) in Comparative Examples 7-10 and Examples 1-2 of this invention are shown.

[0062] Figure 7 The potential and antioxidant properties (ability to eliminate ABTS free radicals) of different ultrafiltration-screened peptide components in Comparative Example 7 and Examples 1-2 of this invention are shown.

[0063] Figure 8 This is a photograph of the composite nutrient powder prepared in Comparative Example 7 and Examples 1-2 of this invention.

[0064] Figure 9 The absorbance values ​​and corresponding antioxidant properties (ability to eliminate ABTS free radicals) of Maillard intermediate and final products of different peptide-starch composite nutritional powders in Comparative Examples 7 and Examples 1-2 of this invention are shown.

[0065] Figure 10 This is the molecular weight distribution of proteins and polypeptides after different treatments in Comparative Example 1 (protein 1), Comparative Examples 5-6 (protein 2-3), Comparative Example 7 (1 peptide), and Examples 1-2 (3 peptides, 2 peptides) of the present invention. Detailed Implementation

[0066] The present invention will be further described in detail below with reference to the embodiments and accompanying drawings, but the embodiments of the present invention are not limited thereto. Unless otherwise specified, all raw materials and reagents in the present invention are commercially available conventional raw materials and reagents. The amounts of each component in the embodiments are expressed in parts by mass and volume, in g and mL.

[0067] A schematic diagram of the preparation process of the wheat gluten peptides and compound nutritional powder of this invention is shown below. Figure 1 .

[0068] The flavor amino acids involved in the examples are described below:

[0069]

[0070] Comparative Example 1: Preparation of Untreated Wheat Glutenin and its Compound Nutritional Powder

[0071] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. After stirring at 40℃ for 6 h, Protamex complex protease was added. The undigested system was inactivated by enzyme in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was then freeze-dried to obtain dissolved wheat gluten.

[0072] (2) The dissolved wheat gluten (protein 1) and wheat (rice) starch obtained in step (1) are mixed at a mass ratio of 1:1 and ground slowly until uniform. Then, under the conditions of 15% moisture content and 79% relative humidity, the mixture is microwaved at 600W for 15 minutes (not continuous, with 30s intervals every 5 minutes) to obtain the compound nutritional powder.

[0073] Comparative Example 2: Preparation of Untreated Wheat Glutenin and its Compound Nutritional Powder

[0074] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. After stirring at 40℃ for 6 h, Protamex complex protease was added. The undigested system was inactivated by enzyme in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was then freeze-dried to obtain dissolved wheat gluten.

[0075] (2) The dissolved wheat gluten obtained in step (1) and wheat (rice) starch (starch 1) that has been gelatinized by α-high temperature amylase for 40 min are mixed at a mass ratio of 1:1 and ground slowly until uniform. Then, under the conditions of 15% moisture content and 79% relative humidity, the mixture is microwaved at 600W for 15 min (non-continuous, with 30s intervals every 5 min) to obtain the compound nutritional powder.

[0076] Comparative Example 3: Preparation of Untreated Wheat Glutenin and its Compound Nutritional Powder

[0077] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. After stirring at 40℃ for 6 h, Protamex complex protease was added. The undigested system was inactivated by enzyme in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was then freeze-dried to obtain dissolved wheat gluten.

[0078] (2) The dissolved wheat gluten obtained in step (1) and wheat (rice) starch (starch 2) that has been gelatinized by α-high temperature amylase for 80 min are mixed at a mass ratio of 1:1 and slowly ground evenly. Then, under the conditions of 15% moisture content and 79% relative humidity, the mixture is microwaved at 600W for 15 min (not continuous, every 5 min at 30 s intervals) to obtain the compound nutritional powder.

[0079] Comparative Example 4: Preparation of Untreated Wheat Glutenin and its Compound Nutritional Powder

[0080] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. After stirring at 40℃ for 6 h, Protamex complex protease was added. The undigested system was inactivated by enzyme in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was then freeze-dried to obtain dissolved wheat gluten.

[0081] (2) The dissolved wheat gluten obtained in step (1) and wheat (rice) starch (starch 3) that has been gelatinized by α-high temperature amylase for 120 min are mixed at a mass ratio of 1:1 and slowly ground evenly. Then, under the conditions of 15% moisture content and 79% relative humidity, the mixture is microwaved at 600W for 15 min (not continuous, every 5 min at 30 s intervals) to obtain the compound nutritional powder.

[0082] like Figure 2 As shown, the absorbance values ​​of Maillard products of the composite nutrient powders prepared in Comparative Examples 1-4 are compared. Maillard intermediates or final products can reflect color and flavor. It can be found that the composite nutrient powders prepared from untreated starch have relatively few intermediates and final products, but there is no substantial difference from the enzymatically hydrolyzed components. For starch, without significant changes in overall properties, the cost and time required for processing increase accordingly, and the shaping function may be lost. Therefore, untreated native starch is still used as the main material for preparing nutrient powders.

[0083] Comparative Example 5: Preparation of Wheat Glutenin and its Compound Nutritional Powder after Enzymatic Hydrolysis

[0084] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. Then, Protamex complex protease was added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected and freeze-dried to obtain wheat gluten mixed peptides.

[0085] (2) The wheat gluten mixed peptide (protein 2) obtained in step (1) and wheat (rice) starch are mixed at a mass ratio of 1:1 and slowly ground evenly. Then, under the conditions of 15% moisture content and 79% relative humidity, the mixture is microwaved at 600W for 15 minutes (not continuous, with 30s intervals every 5 minutes) to obtain compound nutritional powder.

[0086] Comparative Example 6: Preparation of Wheat Glutenin and its Compound Nutritional Powder after Ultrasonic and Enzymatic Treatment

[0087] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was then freeze-dried to obtain a mixed peptide of wheat gluten.

[0088] (2) The wheat gluten mixed peptide (protein 3) obtained in step (1) and wheat (rice) starch are mixed at a mass ratio of 1:1 and slowly ground evenly. Then, under the conditions of 15% moisture content and 79% relative humidity, the mixture is microwaved at 600W for 15 minutes (not continuous, with 30s intervals every 5 minutes) to obtain compound nutritional powder.

[0089] The wheat gluten proteins prepared in Comparative Examples 1 and 5-6 were subjected to amino acid composition analysis, antioxidant activity assay, and zeta potential assay. The results are shown in Table 1 and 2. Figure 3 As shown in Table 2, the color index, antioxidant activity, and absorbance values ​​of Maillard intermediates (294 nm) and final products (420 nm) of the composite nutrient powders prepared in Comparative Examples 1 and 5-6 were analyzed and scored. The results are also shown in Table 2 and 3. Figure 5 .

[0090] The antioxidant activity was determined as follows: The ABTS scavenging capacity of the protein / protein hydrolysate was determined according to the operating procedure of the T-AOC assay kit and detected using a microplate reader (Infinite 200 PRO, Tecan, Switzerland). The absorbance of the prepared ABTS working solution was diluted to 0.7 ± 0.05 (49-fold dilution) at 734 nm and then allowed to stand for 12–16 hours. 10 μL of a 5 mg / mL sample was added to 200 μL of the ABTS working solution, and the absorbance was measured at 734 nm. The absorbance was calculated using a standard curve prepared with different concentrations of Trolox (0.15, 0.3, 0.6, 0.9, 1.2, 1.5 mM). The final total antioxidant capacity was usually expressed as the total antioxidant capacity of water-soluble vitamins per milligram or gram of protein by weight, expressed in mmol / mg or mmol / g. The zeta potential of the samples was measured using a nano-scale zeta potential meter (Nano-ZS90, Malvern Instruments, UK).

[0091] Table 1. Amino acid composition of wheat gluten hydrolysates from different treatments

[0092]

[0093] As shown in Table 1, the amino acid composition of different treatments is not significantly different. The content of glutamic acid and glutamine, which have a savory flavor, is relatively high. The content of glycine, alanine, threonine, serine, and proline, which have a sweet flavor, still accounts for more than 25%. This indicates that ultrasonic or enzymatic treatment of wheat gluten does not cause loss of the nutritional components and flavor of wheat gluten.

[0094] Depend on Figure 3 It is evident that wheat gluten peptides prepared by enzymatic hydrolysis or combined ultrasonic hydrolysis of wheat glutenin exhibit higher antioxidant capacity. This is mainly because enzymatic hydrolysis or combined ultrasonic hydrolysis can alter the protein sequence to a certain extent (such as breaking), and change the spatial structure through stretching and loose rearrangement, thereby exposing the internal amino acid residues of the protein, enabling them to bind with free radicals and quench them. Simultaneously, the potential of wheat gluten peptides prepared by enzymatic hydrolysis or combined ultrasonic hydrolysis is below -20 mV, while the potential of untreated wheat glutenin is -2.97 mV. This indicates that enzymatic hydrolysis or combined ultrasonic hydrolysis can expose the polar amino acids of the protein, increase the effective surface charge, and induce electrostatic repulsion between molecules, which is beneficial for protein dispersion in water.

[0095] Table 2. Color of wheat gluten hydrolysate with different treatments and wheat (rice) starch compound nutritional powder

[0096]

[0097] Note: Different letters in each column indicate significant differences (P<0.05); L=lightness (0=black, 100=white), a=red or green (+a=red; -a=green), b=yellow or blue (+b=yellow; -b=blue), whiteness (W*), chroma (C*), color difference (△E*);

[0098] W* = 100-[(100-L) 2 +a 2 +b 2 ] 1 / 2 C* = (a 2 +b 2 ) 1 / 2 ; △E* = [(L-L0) 2 +(a-a0) 2 +(b-b0) 2 ] 1 / 2 .

[0099] The physical images of the composite nutrient powders prepared in Comparative Examples 1 and 5-6 are shown below. Figure 4 As shown; among them, the composite nutrient powder prepared by Comparative Example 1 and Comparative Example 5-6 is denoted as heated protein 1-3 wheat (rice) starch, while the composite without microwave heating treatment is denoted as protein 1-3 wheat (rice) starch.

[0100] like Figure 4 As shown in Table 2, it is evident that the unheated components are relatively white, while the heated protein / peptide-starch complex turns yellow, indicating the occurrence of the Maillard reaction. Combined with Table 2, this shows that the various indicators of the compound nutritional powder have changed after compounding, improving the sensory properties of the complex. Furthermore, no significant caramelization or carbonization occurred under these conditions, indicating that the conditions were suitable.

[0101] As shown in Table 2 and Figure 4The color indices differ slightly among the different treatments. Lower L* and W* values ​​in the treated groups indicate a deeper Maillard reaction, reflecting a higher level of browning pigment formation. The ultrasonic + enzymatic hydrolysis group showed a higher degree of Maillard reaction than the enzymatic hydrolysis group. The wheat-treated group had a higher C* value, indicating that the Maillard reaction level of the peptide and wheat starch compound nutritional powder was higher than that of the rice starch-containing nutritional powder. This means the treated wheat gluten peptide compound nutritional powder had a more pronounced color, providing an appealing color to the food. The enzymatic hydrolyzed peptide nutritional powder had the highest a* (positive) value, possibly due to the higher content of various Maillard intermediates. Relatively speaking, due to the superposition of a* and b* values, both enzymatic hydrolysis and ultrasonic + enzymatic hydrolysis resulted in a more pronounced darker color than the untreated group. The ultrasonic + enzymatic hydrolysis group might produce a tea-colored color, while the enzymatic hydrolysis group might produce a brownish-red color. For ΔE*, the ultrasonic + enzymatic hydrolysis group had the smallest color difference and relatively more uniform color. Although the enzymatic hydrolysis group had a high degree of Maillard reaction, its color uniformity was relatively poor. Enzyme-resistant starch increases b* (yellow), indicating that the ultrasonic + enzymatic hydrolysis treatment group contains a higher level of enzyme-resistant starch than the enzymatic hydrolysis treatment group. These results also demonstrate the combined inhibitory effect on starch gelatinization, thereby reducing the GI value (glycemic index) of food and benefiting human health.

[0102] Depend on Figure 5 It is evident that, compared with compound nutrient powder prepared from untreated wheat gluten, the compound nutrient powder prepared by enzymatic hydrolysis or ultrasonic combined enzymatic hydrolysis has significantly improved antioxidant properties.

[0103] Comparative Example 7: Preparation of wheat glutenin peptides with a molecular weight >30K and their compound nutritional powder

[0104] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0105] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation. Enzymatically hydrolyzed peptide segments with molecular weight >30K (1 peptide) are screened and freeze-dried into wheat gluten peptide powder.

[0106] (3) Mix the wheat gluten peptide powder and wheat (rice) starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a moisture content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0107] Comparative Example 8: Heating Process for Compound Nutritional Powder

[0108] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0109] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with molecular weight of 8-30K (2 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0110] (3) The wheat gluten peptide powder obtained in step (2) and wheat (rice) starch are mixed at a mass ratio of 1:1 (0.5% (w / v): 0.5% (w / v)) and ground slowly until uniform. Then, the mixture is heated at 90°C for 2 hours to obtain a compound nutrient powder.

[0111] Comparative Example 9: Heating Process for Compound Nutritional Powder

[0112] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0113] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation. Enzymatically hydrolyzed peptide segments with molecular weight <8K (3 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0114] (3) The wheat gluten peptide powder obtained in step (2) and wheat (rice) starch are mixed at a mass ratio of 1:1 (0.5% (w / v): 0.5% (w / v)) and ground slowly until uniform. Then, the mixture is heated at 90°C for 2 hours to obtain a compound nutrient powder.

[0115] Comparative Example 10: Heating Process for Compound Nutritional Powder

[0116] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0117] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation. Enzymatically hydrolyzed peptide segments with molecular weight >30K (1 peptide) are screened and freeze-dried into wheat gluten peptide powder.

[0118] (3) The wheat gluten peptide powder obtained in step (2) and wheat (rice) starch are mixed at a mass ratio of 1:1 (0.5% (w / v): 0.5% (w / v)) and ground slowly until uniform. Then, the mixture is heated at 90°C for 2 hours to obtain a compound nutrient powder.

[0119] like Figure 6 The absorbance values ​​of Maillard products of the composite nutritional powders prepared by Comparative Examples 8-10 (wet method), Comparative Example 7, and Examples 1-2 (dry method) were compared. Maillard intermediate or final products can reflect color and flavor. It can be found that the peptide starch complex prepared by the dry method has a higher degree of Maillard reaction overall. In contrast, the wet method is not only time-consuming and less efficient, but also detrimental to the final product shaping. Therefore, the dry method is chosen to prepare the composite nutritional powder.

[0120] Example 1: Preparation of wheat glutenin peptides with a molecular weight <8K and their compound nutritional powder

[0121] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0122] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation. Enzymatically hydrolyzed peptide segments with molecular weight <8K (3 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0123] (3) Mix the wheat gluten peptide powder and wheat (rice) starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a moisture content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0124] Example 2: Preparation of wheat glutenin peptides with a molecular weight of 8-30K and their compound nutritional powder

[0125] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0126] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with molecular weight of 8-30K (2 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0127] (3) Mix the wheat gluten peptide powder and wheat (rice) starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a moisture content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0128] The wheat gluten peptides prepared in Comparative Example 7, Example 1, and Example 2 were subjected to amino acid composition analysis, antioxidant activity assay, and zeta potential assay. The results are shown in Table 3 and 4. Figure 7 .

[0129] Table 3. Amino acid composition of wheat gluten hydrolysates at different molecular weight ranges

[0130]

[0131] As shown in Table 3, the amino acid composition varies across different molecular weight ranges. Among the dipeptides (8-30K), glycine (moderate sweetness), isoleucine (lowest) with a predominantly bitter taste, valine (lowest) with a mixed sweet and bitter taste, asparagine (moderate) with a predominantly sour taste, and glutamic acid (highest) with a umami taste. Dipeptides (8-30K) are the optimal choice for preparing wheat gluten peptide compound nutritional powder (base material) due to their superior flavor, good formability, and excellent nutritional function. However, the decrease in umami amino acids and the increase in bitter amino acids in tripeptides and monopeptides may hinder their further utilization.

[0132] At the same time, by Figure 7 It can be seen that the wheat gluten peptides of Examples 1 and 2 (<8K, 3 peptides; 8-30K, 2 peptides) have good antioxidant properties. At the same time, the wheat gluten peptides of Example 2 (8-30K, 2 peptides) have the lowest potential, which is beneficial to protein dispersion and dissolution. However, the wheat gluten peptides of Comparative Example 7 (>30K, 1 peptide) do not have outstanding characteristics in terms of both antioxidant properties and potential. Therefore, wheat gluten peptides with a molecular weight >30K (1 peptide) are not selected in this invention.

[0133] The color index, antioxidant activity, and absorbance values ​​of the compound nutrient powders prepared in Comparative Example 7, Example 1, and Example 2 were analyzed and scored. The results are shown in Table 4. Figure 9 .

[0134] Table 4. Color of wheat (rice) starch compound nutritional powder with different fractions of wheat gluten hydrolysate.

[0135]

[0136] Note: Different letters in each column indicate significant differences (P<0.05); L = lightness (0 = black, 100 = white), a = red or green (+a = red; a = green), b = yellow or blue (+b = yellow; -b = blue), whiteness (W*), chroma (C*), color difference (△E*);

[0137] W* = 100-[(100-L) 2 +a 2 +b 2 ] 1 / 2 C* = (a 2 +b 2 ) 1 / 2 ; △E* = [(L-L0) 2 +(a-a0) 2 +(b-b0) 2 ] 1 / 2 .

[0138] Comparative Example 7, physical images of the composite nutrient powders prepared in Examples 1-2 are shown below. Figure 8 As shown; wherein, the composite nutrient powder prepared in Comparative Example 7 and Examples 1-2 is referred to as heated 1-3 peptide wheat (rice) starch, while the composite without microwave heating treatment is referred to as 1-3 peptide wheat (rice) starch.

[0139] like Figure 8 As shown, it is intuitive to see that the unheated component is relatively white, but compared to... Figure 4 The middle section represents a further segmented protein / peptide complex. This segmented peptide-starch composite nutritional powder is darker in color. As the molecular weight of the segmented peptides decreases, the particle size becomes finer. The color change of the segmented peptide-starch composite complex after heating is more pronounced compared to... Figure 4 More noticeably, the color becomes yellower, even showing caramel and brown hues. Small areas of carbonization are even observed in the tripeptides, indicating that the Maillard reaction may occur with decreasing peptide molecular weight, leading to more reaction sites and accelerating the reaction. Combined with Table 4, this shows that the various indicators of the compounded nutritional powder changed significantly after compounding, improving the sensory properties of the compound. However, for <8K (tripeptides), especially tripeptide wheat starch compound nutritional powder, this treatment may be excessive. This provides further reference for subsequent selection.

[0140] As shown in Table 4 and Figure 8 In Examples 1 and 2, L*, W*, and C* are relatively larger than in Comparative Example 7. However, the products obtained from Examples 1 and 2 show a relatively darker color. This suggests that the Maillard reaction in Examples 1 and 2 may have led to the formation of larger particles, resulting in greater reflection and thus increased brightness (i.e., the absorption effect is less than the reflection and diffusion effects). However, the composite nutrient powder prepared in Examples 1 and 2 exhibits better color, smaller color difference (ΔE*), and more uniform color, while the composite nutrient powder prepared in Comparative Example 7 shows a larger color difference. Due to the superposition of a* and b* values, the comparative example appears brownish. Figure 9 As is known, the compound nutrient powders of Examples 1 and 2 have high absorbance values ​​and high antioxidant properties, while the compound nutrient powder of Comparative Example 7 has low absorbance, light color, and low antioxidant properties. (See Table 4 for details.) Figure 9 The results indicate that wheat gluten peptides with a molecular weight >30K (1 peptide) are not suitable for food processing applications, while those <8K, i.e., 3 peptide complexes, may also need to consider their acceptable color. Therefore, they are not selected for the preparation of compound nutritional powder in the technical solution of this invention.

[0141] The compound nutritional powders prepared by Comparative Examples 1, 5-7, Example 1, and Example 2 were tested for allergen gluten, and the specific results are shown in Table 5.

[0142] Table 5. Effects of different treatment methods on gluten content in compound nutritional powder

[0143]

[0144] In the gluten allergen test (Table 5), the gluten allergen levels in the ultrasonically and enzymatically hydrolyzed wheat gluten mixed peptide-wheat (rice) starch composite nutritional powder and the wheat gluten peptide (8-30K, 2-peptide; or, <8K, 3-peptide)-wheat (rice) starch composite nutritional powder were both below 10.00 ppm, far below the 10-20 mg / kg (ppm) standard for gluten-free foods in countries such as the Codex Alimentarius Commission, the United States, the European Union, Australia, New Zealand, and Argentina. The enzymatically hydrolyzed group had levels between 10-20 ppm, while the untreated group had levels above 20 ppm. In the ultrasonically + enzymatically hydrolyzed + ultrafiltration group, the difference between 3-peptides and 2-peptides was not significant, while the 1-peptide level was much higher than 20 ppm. This indicates that the combined-processed peptides or the ultrafiltration-reduced 2- and 3-peptides and their prepared nutritional powders can be used as gluten-free health foods. However, considering their overall properties, 2-peptides may be more suitable for the development of nutritional powders and gluten-free basic biscuit models. Untreated and 1-peptides still pose a potential risk of allergic reactions and are not suitable for promotion as nutritional health foods.

[0145] The molecular weight distribution of Comparative Example 1 (protein 1), Comparative Examples 5-6 (proteins 2-3), Comparative Example 7 (monopeptide), and Examples 1-2 (tripeptides, dipeptides) was determined, and the results are as follows: Figure 10 As shown.

[0146] Experimental methods: Molecular weight distribution

[0147] Proteins were dissolved in 200 mM sodium phosphate buffer (pH 6.8) at a concentration of 1 mg / mL. The solution was first filtered through a 0.22 μm filter membrane and then analyzed using a gel permeation high-performance liquid chromatography (SE-HPLC, Water 600, Milford, MA) column (7.8 mm inner diameter × 300 mm length, Tokyo, Japan). The detection wavelength was 214 nm. 20 μL of sample was injected at a flow rate of 1 mL / min, with 200 mM sodium phosphate buffer (pH 6.8). Standard curves (r) were plotted by the peak times (x) of protein standards of different molecular weights (Mw) (iron transporter (80000 Da), cytochrome C (12384 Da), synthetic peptides PVLGPVRGPH (1361 Da), glutathione (307 Da), and glutamate (147 Da)). 2=0.9904), thus obtaining the functional relationship y. Each sample was divided into different molecular weight fragment ranges according to the time range corresponding to the standard curve, namely >10 kDa, 5-10 kDa, 3-5 kDa, 1-3 kDa, and <1 kDa. The percentage content of different fragment ranges was calculated as follows:

[0148] ;

[0149] according to Figure 10 It is known that components with >10K in protein or peptide powder may contain more sensitizing sites and regions due to the retention of higher-order structures. Therefore, untreated proteins 1 and 2, and the larger fragments of peptide 1 (>30K) after ultrafiltration, pose a potential risk of allergic reactions and are unsuitable for promotion as nutritional health foods. For the co-treated protein 3, which contains more small-molecule peptides, further ultrafiltration of the fragmented peptide 2 (8-30K) and peptide 3 (<8K) is more suitable as a (low) gluten-free food base material under mainstream international standards where sensitizing gluten retention should be <20ppm, or even more stringent <10ppm. Furthermore, considering amino acid composition and other factors such as color and flavor, excessive small-molecule peptides <8K (i.e., peptide 3) result in a decline in flavor and sensory quality, making them relatively less desirable. The next preferred option is co-treated protein 3, and the next preferred option is co-treated ultrafiltration peptide 2 (8-30K).

[0150] Example 3: Preparation of wheat gluten peptides and their compound nutritional powder

[0151] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0152] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with molecular weight of 8-30K (2 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0153] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 5 minutes with a water content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0154] Example 4: Preparation of wheat gluten peptides and their compound nutritional powder

[0155] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0156] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with molecular weight of 8-30K (2 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0157] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 70 minutes (non-continuous, with 30s intervals every 5 minutes) with a moisture content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0158] Example 5: Preparation of wheat gluten peptides and their compound nutritional powder

[0159] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0160] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with molecular weight of 8-30K (2 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0161] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a water content of 50% and a relative humidity of 79% to obtain a compound nutrient powder.

[0162] Example 6: Preparation of wheat gluten peptides and their compound nutritional powder

[0163] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0164] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with molecular weight of 8-30K (2 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0165] (3) Mix the wheat gluten peptide powder and rice starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 75 minutes (non-continuous, with 30s intervals between every 5 minutes) with a moisture content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0166] Example 7: Preparation of wheat gluten peptides and their compound nutritional powder

[0167] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0168] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation. Enzymatically hydrolyzed peptide segments with molecular weight <8K (3 peptides) are screened and freeze-dried into wheat gluten peptide powder.

[0169] (3) Mix the wheat gluten peptide powder and rice starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 20 minutes (non-continuous, with 30s intervals every 5 minutes) with a water content of 15% and a relative humidity of 79% to obtain a compound nutritional powder.

[0170] Example 8: Preparation of wheat gluten peptides and their compound nutritional powder

[0171] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.27. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0172] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30K are screened and freeze-dried into wheat gluten peptide powder.

[0173] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a water content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0174] Example 9: Preparation of wheat gluten peptides and their compound nutritional powder

[0175] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:1.3. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0176] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30K are screened and freeze-dried into wheat gluten peptide powder.

[0177] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a water content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0178] Example 10: Preparation of wheat gluten peptides and their compound nutritional powder

[0179] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 3 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0180] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30K are screened and freeze-dried into wheat gluten peptide powder.

[0181] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a water content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0182] Example 11: Preparation of wheat gluten peptides and their compound nutritional powder

[0183] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 24 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0184] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30k are screened and freeze-dried into wheat gluten peptide powder.

[0185] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:2 and grind them evenly at a slow speed. Then, microwave the mixture at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a moisture content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0186] Example 12: Preparation of wheat gluten peptides and their compound nutritional powder

[0187] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 25℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0188] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30K are screened and freeze-dried into wheat gluten peptide powder.

[0189] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a water content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0190] Example 13: Preparation of wheat gluten peptides and their compound nutritional powder

[0191] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 60℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0192] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30k are screened and freeze-dried into wheat gluten peptide powder.

[0193] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a water content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0194] Example 14: Preparation of wheat gluten peptides and their compound nutritional powder

[0195] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0196] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30k are screened and freeze-dried into wheat gluten peptide powder.

[0197] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 600W for 15 minutes (non-continuous, with 30s intervals between every 5 minutes) with a water content of 15% and a relative humidity of 90% to obtain a compound nutrient powder.

[0198] Example 15: Preparation of wheat gluten peptides and their compound nutritional powder

[0199] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0200] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30k are screened and freeze-dried into wheat gluten peptide powder.

[0201] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 360W for 15 minutes (non-continuous, with 30s intervals every 5 minutes) with a water content of 15% and a relative humidity of 79% to obtain a compound nutrient powder.

[0202] Example 16: Preparation of wheat gluten peptides and their compound nutritional powder

[0203] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0204] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30k are screened and freeze-dried into wheat gluten peptide powder.

[0205] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 600W for 15 minutes (non-continuous, with 30s intervals every 5 minutes) with a water content of 10% and a relative humidity of 79% to obtain a compound nutrient powder.

[0206] Example 17: Preparation of wheat gluten peptides and their compound nutritional powder

[0207] (1) Wheat gluten was dissolved in 0.2M PBS phosphate buffer at pH 6.0 and stirred overnight. The mixture was then sonicated at 250W for 30 min. Protamex complex protease was then added to enzymatically hydrolyze the wheat gluten. The mass ratio of wheat gluten to Protamex complex protease was 100:0.81. The hydrolysis time was 6 h, and the hydrolysis temperature was 40℃. After hydrolysis, the enzyme was inactivated in a 95℃ water bath for 1 h, and then centrifuged at 9500 r / min for 20 min at 25℃. The supernatant was collected to obtain a mixed peptide of wheat gluten.

[0208] (2) The wheat gluten mixed peptides obtained in step (1) are subjected to ultrafiltration and segmentation, and enzymatically hydrolyzed peptide segments with a molecular weight of 8-30k are screened and freeze-dried into wheat gluten peptide powder.

[0209] (3) Mix the wheat gluten peptide powder and wheat starch obtained in step (2) at a mass ratio of 1:1 and grind them evenly at a slow speed. Then, microwave them at 600W for 15 minutes (non-continuous, with 90s intervals between every 5 minutes) with a water content of 10% and a relative humidity of 79% to obtain a compound nutrient powder.

[0210] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. A method for preparing desensitizing wheat glutenin peptides, characterized in that: Includes the following steps: (1) Wheat gluten was dissolved in PBS phosphate buffer solution, and the mixture was subjected to ultrasonic cavitation and enzymatic hydrolysis to obtain mixed peptides; (2) The mixed peptides obtained in step (1) were subjected to enzyme inactivation treatment, and then ultrafiltration was used to obtain desensitized wheat glutenin peptides. In step (2), after ultrafiltration and segmentation, subdivided wheat gluten peptides with a molecular weight of 30K and below are obtained.

2. The preparation method according to claim 1, characterized in that: In step (2), after ultrafiltration and segmentation, subdivided wheat gluten peptides are obtained with a molecular weight of 8-30K or <8K.

3. The preparation method according to claim 1, characterized in that: In step (1), the pH of the PBS phosphate buffer solution is 2.0-11.0, and the concentration of the PBS phosphate buffer solution is 0.01-1.0M; And / or, in step (1), the ultrasonic power of the ultrasonic cavitation is 100-600W, and the ultrasonic cavitation time is 25-60min; And / or, in step (1), the enzymatic hydrolysis is performed using at least one of Protamex complex protease, papain, alkaline protease, trypsin, and neutral protease. And / or, in step (1), the mass ratio of protease to wheat gluten in the enzymatic hydrolysis is (0.2-1.5):100; And / or, in step (1), the enzymatic hydrolysis treatment time is 3-24 hours; And / or, in step (1), the temperature of the enzymatic hydrolysis treatment is 25-60℃.

4. The preparation method according to claim 1, characterized in that: In step (2), the specific steps of the enzyme inactivation treatment are as follows: inactivate the mixed peptides in a water bath at 80-100℃ for 10-90 min; And / or, in step (2), the mixed peptides are centrifuged after enzyme inactivation treatment to obtain supernatant for ultrafiltration fractionation; And / or, in step (2), the desensitized wheat gluten peptides prepared are freeze-dried.

5. The preparation method according to claim 4, characterized in that: The centrifugation is carried out at a temperature of 4℃-30℃ and a rotation speed of 2000-12000 r / min for 5-30 min.

6. A desensitizing wheat glutenin peptide, characterized in that: It is prepared by the preparation method according to any one of claims 1 to 5.

7. The application of the desensitizing wheat gluten peptide according to claim 6 in the deep processing of food.

8. A compound nutritional powder, characterized in that: It is obtained by heat treatment after uniformly mixing and heating the desensitizing wheat gluten peptides as described in claim 1 and starch.

9. The compound nutritional powder according to claim 8, characterized in that: The starch is a cereal starch, and further includes at least one of wheat starch and rice starch; And / or, the mass ratio of the desensitized wheat gluten peptide to starch is 1:(0.2-3). And / or, the starch is gelatinized, enzymatically hydrolyzed, freeze-dried, and then compounded with desensitized wheat gluten peptides; And / or, the heat treatment is performed by microwave heating or hydrothermal treatment; And / or, the mixing and homogenization are carried out by stirring, grinding, or vortex oscillation; And / or, the composite nutrient powder is cooled after heat treatment, pulverized and sieved to form composite nutrient powder or molded to form composite nutrient basic model biscuits with the desired shape and structure. And / or, the compound nutritional powder may also have additional ingredients added as needed; the additional ingredients are conventional ingredients or additives used in food processing.

10. The compound nutritional powder according to claim 9, characterized in that: The enzymatic hydrolysis treatment uses amylase; And / or, the temperature of the enzymatic hydrolysis treatment is 60-85℃; And / or, the enzymatic hydrolysis time is 30-150 min; And / or, the starch after enzymatic hydrolysis is freeze-dried; And / or, the microwave heating conditions are: water content 0-50% and not 0, relative humidity 60-90%, heating at 300-800W for 0-80 minutes and not 0; And / or, the microwave heating is non-continuous heating, with a 30-90 second interval between heating intervals of 3-8 minutes; And / or, the hydrothermal treatment conditions are as follows: at a temperature of 30-95℃, the concentration of wheat glutenin peptides is 0.1-5% w / v, the concentration of starch is 0.1-5% w / v, and the mass ratio of wheat glutenin peptides to starch is 5:1-1:5, heated for 0-3 hours and not 0. And / or, the excipients include at least one of sugar, eggs, milk, oil, salt, cereals, cocoa, food flavorings, food additives, and preservatives.

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