A method for producing a corn oligopeptide

Abstract

The application provides a corn oligopeptide production method and belongs to the technical field of corn oligopeptide; the production method comprises pretreatment, enzyme carrier preparation, functional enzyme preparation and enzymolysis steps; the pretreatment step is that 50-90wt% of an ethanol solution is added to corn yellow powder for leaching treatment or 10-60wt% of a sodium bicarbonate solution is added for stirring treatment; after treatment, the pretreated corn yellow powder is obtained after washing and drying; the corn oligopeptide prepared by the method of the application effectively removes pigment and bitterness, and the yield and protein content of the corn oligopeptide are high.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of corn oligopeptide, and particularly relates to a production method of corn oligopeptide. BACKGROUND

[0002] Corn oligopeptide is a powder product with a relative molecular weight less than 1000 obtained by enzyme degradation and specific small peptide separation technology from corn protein extracted from natural food corn; corn oligopeptide has reliable safety and various biological activities, such as antioxidant, antihypertensive, immune enhancement, antifatigue, liver protection and the like; in recent years, with the gradual improvement of the understanding of corn peptide, the development of corn peptide food has gradually attracted people's attention, and the development of corn peptide in the United States, Europe and Japan and other countries and regions has been in the leading position, and the technical research has been relatively mature, especially Japan has been in the world's leading position in the field of corn peptide research.

[0003] The research on peptides in China started late, and the related research on corn peptides has just begun to step into the right track; at present, the methods for preparing corn peptides mainly include microbial fermentation method, enzymatic hydrolysis method and chemical synthesis method.

[0004] The chemical synthesis method generally refers to the protein hydrolysis method, and the protein hydrolysis method includes acid hydrolysis and alkali hydrolysis, that is, strong acid, strong base or strong acid and strong base are used to directly hydrolyze proteins to obtain a hydrolysis liquid; in general production, hydrochloric acid and sulfuric acid are used as strong acid, and barium hydroxide and sodium hydroxide are used as strong base, the hydrolysis speed is fast, and the degree is high; the hydrolysis process is difficult to control, and the acid and alkali reagents used will cause different degrees of damage to arginine, cystine and serine.

[0005] The main principle of the microbial fermentation method for preparing corn peptides is to use the microorganism enzymes produced in the growth process of microorganisms, and the method mainly relies on the enzyme system in the microorganism to decompose and process corn protein powder; this technology started late in China, so that the related research on the preparation of corn peptides by using the microbial fermentation method is less in China at present, especially there is almost no introduction of microbial strains, the microbial fermentation method is not mature, the conversion rate is low, the refining treatment is difficult, and the industrial production cannot be carried out.

[0006] The enzymatic hydrolysis method is to use single alkaline protease or complex enzymatic hydrolysis method to degrade corn prolamin as a substrate, so as to produce bioactive short peptides and multifunctional mixed peptides; the use of the enzymatic hydrolysis method for preparing corn oligopeptide has the disadvantages of difficult separation and low yield, and the pigment and bitter taste of corn oligopeptide cannot be effectively removed, which affects the taste and limits the application of corn peptides.

[0007] And the prior art adopts enzymatic method to prepare corn oligopeptide, on the one hand, due to the complex and close structure of corn protein, a single enzyme or a complex enzyme is difficult to fully act on the corn protein, which weakens the enzymatic effect and reduces the production efficiency; on the other hand, in the enzymatic process, in addition to generating product corn oligopeptide, a large amount of macromolecular protein, intermediate product not completely enzymolyzed and other impurities are also generated, which brings great difficulty to subsequent purification and separation, thereby reducing the yield and purity of the product; furthermore, the enzymatic condition is harsh, and the requirements for temperature and pH value and the like are relatively strict, which is easy to affect the activity of the enzyme, and further affect the enzymatic effect. SUMMARY

[0008] In order to solve the technical problems existing in the prior art, the present application provides a production method of corn oligopeptide, which adopts enzymatic method, has high enzymatic efficiency, effectively removes the pigment and bitter taste of corn oligopeptide, and has high product yield and purity.

[0009] In view of the above technical problems, the present application adopts the following technical solutions:

[0010] A production method of corn oligopeptide, comprising pretreatment, preparation of enzyme carrier, preparation of functional enzyme and enzymolysis steps;

[0011] 1. Pretreatment

[0012] The pretreatment liquid is added to the corn yellow powder for treatment, after the treatment is completed, the pretreated corn yellow powder is obtained after washing and drying;

[0013] The mass-volume ratio of the corn yellow powder to the pretreatment liquid is 100g:800-1000mL;

[0014] The addition of the pretreatment liquid for treatment is the addition of 50-90wt% ethanol solution for leaching treatment or the addition of 10-60wt% sodium bicarbonate solution for stirring treatment;

[0015] The leaching treatment is leaching for 3.0-3.5h at 70-73℃ and 100-120rpm;

[0016] The stirring temperature of the stirring treatment is 45℃, and the stirring time is 18min;

[0017] When the 10-60wt% sodium bicarbonate solution is added for stirring treatment, after the treatment is completed, the pH value can be adjusted to 4.7, and the lysocell amylase is added, and the stirring is continued for 1.0h, and the lysocell amylase is 1.2wt% of the mass of the corn yellow powder.

[0018] 2. Preparation of enzyme carrier

[0019] (1) Activation

[0020] The diatomite is placed in a 6-8 times mass hydrochloric acid solution, the temperature is raised to 35-40℃, and the soaking is kept for 3.6-4.2 hours; after the soaking is completed, the diatomite is filtered and washed, and then dried at 78-82℃ for 8-12 hours; after the temperature is naturally recovered to room temperature, the diatomite is put into a muffle furnace, the temperature is raised to 350-380℃ at a rate of 2.0-4.0℃ / min, and the calcination is kept for 4.7-5.3 hours; after the calcination is completed, the temperature is naturally reduced to room temperature, and the activated diatomite is obtained;

[0021] The particle size of the diatomite is 210-240 nm;

[0022] The mass concentration of the hydrochloric acid solution is 20-25%;

[0023] (2) primary treatment

[0024] The hydroxylamine hydrochloride is put into deionized water and stirred to obtain a hydroxylamine hydrochloride solution; the chitosan is put into an acetic acid solution and stirred to obtain a chitosan solution; the activated diatomite is placed in deionized water, the temperature is raised to 37-43℃, the hydroxylamine hydrochloride solution and the chitosan solution are slowly added, the adding rate is controlled to be 1.8-2.2 g / min, the stirring speed is kept at 230-250 rpm during the adding, after the adding is completed, the temperature is continuously raised to 68-72℃, and the soaking and stirring are kept for 2.3-2.6 hours; after being washed and dried, the primary enzyme carrier is obtained;

[0025] The mass ratio of the hydroxylamine hydrochloride to the deionized water is 6.5-7.5:100;

[0026] The mass ratio of the chitosan to the acetic acid solution is 4.6-5.3:100;

[0027] The mass concentration of the acetic acid solution is 3.0-5.0%;

[0028] The mass ratio of the activated diatomite, the deionized water, the hydroxylamine hydrochloride solution and the chitosan solution is 18-22:200:34-38:33-37;

[0029] (3) secondary treatment

[0030] The primary carrier is put into N,N-dimethylformamide, and after being stirred uniformly, succinic anhydride and triethylamine are added, the temperature is raised to 72-78℃, and the reaction is kept for 3.8-4.3 hours; after the reaction is completed, the enzyme carrier is obtained by filtering, washing and drying;

[0031] The mass ratio of the primary carrier, the N,N-dimethylformamide, the succinic anhydride and the triethylamine is 9.5-10.5:100:3.4-3.8:0.37-0.42.

[0032] 3. preparation of a functional enzyme

[0033] Mix the enzyme carrier with the phosphate buffer, stir uniformly, add the enzyme, carry out oscillation at room temperature, the oscillation speed is 130-150 rpm, the oscillation time is 3.8-4.2 h, then add the cinnamyl aldehyde solution, continue to oscillate the reaction for 1.8-2.2 h, after the reaction is completed, after centrifugal washing, natural drying, the functional enzyme is prepared;

[0034] The mass-volume ratio of the enzyme carrier, the phosphate buffer, the enzyme and the cinnamyl aldehyde solution is 2.8-3.2 g:48-53 mL:0.24-0.26 g:16.0-17.0 g;

[0035] The enzyme is one or both of Alcase2.4L protease and Flavourzyme1000L enzyme, and the mass ratio of the Alcase2.4L protease and the Flavourzyme1000L enzyme is 2:1;

[0036] The mass concentration of the cinnamyl aldehyde solution is 1.3-1.7%;

[0037] The molar concentration of the phosphate buffer is 0.04-0.06 ml / L, and the pH value is 7.8-8.2.

[0038] 4. Enzymolysis

[0039] Add deionized water to the pretreated corn yellow powder, adjust the pH to 7.5-8.0, add the functional enzyme, react at 53-56 DEG C for 7.5-8.5 h, after the reaction is completed, the supernatant and solid are obtained by suction filtration, 0.8-1.2 wt% activated carbon is added to the supernatant, stirring treatment is carried out for 1.0-1.3 h, after the stirring is completed, vacuum drying is carried out after rotary evaporation, and the corn oligopeptide product is obtained;

[0040] The mass ratio of the pretreated corn yellow powder, the deionized water and the functional enzyme is 100:900-1000:22-40.

[0041] Compared with the prior art, the present application has the following beneficial effects:

[0042] 1. In the process for preparing corn oligopeptide, the corn yellow powder is pretreated to remove pigment and oil, and then subjected to enzymatic hydrolysis using functional enzyme. In the preparation method of the functional enzyme, diatomite is used as the enzyme carrier matrix. The diatomite is treated with dilute acid solution to remove the impurity components on the surface, increase the specific surface area and activity of the diatomite, and improve the stability of the diatomite after calcination. In the one-step treatment, hydroxylamine hydrochloride and diatomite are introduced to introduce amino groups on the surface of the diatomite. The chitosan molecules can further enhance the hydrophilic property and biocompatibility of the diatomite carrier. The primary enzyme carrier is reacted with succinic anhydride to introduce carboxyl groups on the surface of the enzyme carrier, further increasing the hydrophilic property and reactivity of the carrier. In the preparation process of the functional enzyme, cinnamaldehyde solution is added, which can react with the enzyme molecules and the amino groups on the surface of the enzyme carrier, enhancing the binding force between the enzyme and the enzyme carrier, so that the enzyme molecules are more firmly fixed on the enzyme carrier, ensuring the stability of the functional enzyme and reducing the probability of enzyme inactivation and denaturation. Therefore, the enzyme activity can be maintained for a long time during the enzymatic hydrolysis. The combination with the activated carbon adsorption step effectively removes the pigment and bitterness of the product, improves the reusability of the enzyme, and increases the contact area with the substrate, improves the enzymatic hydrolysis efficiency, promotes the enzymatic hydrolysis reaction, and thus improves the product yield and protein content.

[0043] 2. The corn oligopeptide product obtained by the method of the present application has a yield of 61.2-63.8% and a protein content of 91.0-93.4 wt%. DETAILED DESCRIPTION

[0044] In order to more clearly understand the technical features, objectives and effects of the present application, the specific embodiments of the present application will be described.

[0045] Example 1

[0046] 1. Pretreatment

[0047] 100 g of corn yellow powder was added to 1000 mL of 50 wt% ethanol solution, and extracted at 70℃ and 100 rpm for 3.5 h. After extraction, the anhydrous ethanol was removed by suction filtration, and then washed and dried to obtain pretreated corn yellow powder.

[0048] 2. Preparation of enzyme carrier

[0049] (1) Activation

[0050] The diatomite was placed in 7 times the mass of 23 wt% hydrochloric acid solution, the temperature was raised to 38℃, and soaked for 4.0 h. After soaking, it was filtered and washed, and then dried at 80℃ for 10 h. After natural recovery to room temperature, it was put into a muffle furnace, and the temperature was raised to 370℃ at a rate of 3.0℃ / min, and calcined for 5.0 h. After calcination, the temperature was naturally lowered to room temperature to obtain activated diatomite.

[0051] The particle size of the diatomite is 220 nm;

[0052] (2) Primary treatment

[0053] 7.0 g of hydroxylamine hydrochloride was put into 100 g of deionized water and stirred uniformly to obtain a hydroxylamine hydrochloride solution; 5.0 g of chitosan was put into 100 g of 4.0 wt% acetic acid solution and stirred uniformly to obtain a chitosan solution; 20 g of activated diatomite was placed in 200 g of deionized water, the temperature was raised to 40℃, 36 g of hydroxylamine hydrochloride solution and 35 g of chitosan solution were slowly added at a rate of 2.0 g / min, and stirring was maintained at a speed of 240 rpm at the same time; after the addition was completed, the temperature was continuously raised to 70℃, and the temperature was maintained for 2.5 h with stirring; after washing and drying, a primary enzyme carrier was obtained;

[0054] (3) Secondary treatment

[0055] 10.0 g of the primary carrier was put into 100 g of N, N-dimethylformamide, and after uniform stirring, 3.6 g of succinic anhydride and 0.40 g of triethylamine were added, the temperature was raised to 75℃, and the reaction was maintained for 4.0 h; after the reaction was completed, the enzyme carrier was obtained by filtration, washing and drying.

[0056] 3. Preparation of functional enzyme

[0057] 3.0 g of the enzyme carrier was mixed with 50 mL of phosphate buffer solution, stirred uniformly, 0.25 g of enzyme was added, and oscillation was carried out at room temperature at a speed of 140 rpm for 4.0 h; then 16.5 g of 1.5 wt% cinnamaldehyde solution was added, and the oscillation reaction was continued for 2.0 h; after the reaction was completed, the functional enzyme was obtained by centrifugal washing and natural drying;

[0058] The enzyme is Alcase2.4L protease;

[0059] The molar concentration of the phosphate buffer solution is 0.05 ml / L, and the pH value is 8.

[0060] 4. Enzymatic hydrolysis

[0061] 100 g of pretreated corn meal was added with 900 g of deionized water, the pH was adjusted to 7.5, 23 g of functional enzyme was added, and the reaction was carried out at 55℃ for 8.0 h; after the reaction was completed, the clear liquid and solid material were obtained by suction filtration; 1 wt% activated carbon was added to the clear liquid, and stirring treatment was carried out for 1.0 h; after the stirring was completed, vacuum drying was carried out after rotary evaporation to obtain a corn oligopeptide product.

[0062] The corn oligopeptide product obtained by the method of Example 1 has a yield of 61.2%, and the protein content is 91.6 wt%.

[0063] Example 2

[0064] 1. Pretreatment

[0065] 1000 mL of sodium bicarbonate solution was added to 100 g of corn meal, and the mixture was stirred at 40°C for 20 min. After washing and drying, pretreated corn meal was obtained.

[0066] The concentration of the sodium bicarbonate solution was 10%.

[0067] 2. Preparation of enzyme carrier

[0068] (1) Activation

[0069] The diatomite was placed in 6 times the mass of 20 wt% hydrochloric acid solution, and the temperature was raised to 35°C for 3.6 h of soaking. After soaking, the filtrate was washed and dried at 78°C for 12 h. After natural recovery to room temperature, it was put into a muffle furnace, and the temperature was raised to 350°C at a rate of 2.0°C / min, and the temperature was maintained for 4.7 h of calcination. After calcination, the temperature was naturally lowered to room temperature, and activated diatomite was obtained.

[0070] The particle size of the diatomite was 210 nm.

[0071] (2) First treatment

[0072] 6.5 g of hydroxylamine hydrochloride was added to 100 g of deionized water and stirred to obtain a hydroxylamine hydrochloride solution. 4.6 g of chitosan was added to 100 g of 3.0 wt% acetic acid solution and stirred to obtain a chitosan solution. 18 g of activated diatomite was placed in 200 g of deionized water, and the temperature was raised to 37°C. 34 g of hydroxylamine hydrochloride solution and 33 g of chitosan solution were slowly added at a rate of 1.8 g / min while maintaining a stirring speed of 230 rpm. After the addition was completed, the temperature was raised to 68°C and stirred for 2.3 h. After washing and drying, a primary enzyme carrier was obtained.

[0073] (3) Second treatment

[0074] 9.5 g of the primary carrier was placed in 100 g of N,N-dimethylformamide, and 3.4 g of succinic anhydride and 0.37 g of triethylamine were added after stirring. The temperature was raised to 72°C, and the reaction was maintained for 3.8 h. After the reaction was completed, the filtrate was washed and dried to obtain an enzyme carrier.

[0075] 3. Preparation of functional enzyme

[0076] Mix 2.8 g enzyme carrier with 48 mL phosphate buffer, stir evenly, add 0.24 g enzyme, oscillate at room temperature, oscillation speed is 130 rpm, oscillation time is 3.8 h, then add 16.0 g 1.3 wt% cinnamaldehyde solution, continue to oscillate for 1.8 h, after the reaction is completed, centrifugal washing is carried out, and natural drying is carried out, to obtain functional enzyme;

[0077] The enzyme is Alcase2.4L protease;

[0078] The molar concentration of the phosphate buffer is 0.06 ml / L, and the pH value is 7.8.

[0079] 4. Enzymolysis

[0080] Add 900 g deionized water to 100 g pretreated corn yellow powder, adjust the pH to 7.8, add 25 g functional enzyme, and react at 53℃ for 8.5 h. After the reaction is completed, the supernatant and solid are obtained by suction filtration. Add 1.2 wt% activated carbon to the supernatant, stir for 1.2 h, and then perform rotary evaporation and vacuum drying to obtain a corn oligopeptide product.

[0081] The corn oligopeptide product obtained by the method of Example 2 has a yield of 63.8% and a protein content of 91.0 wt%.

[0082] Example 3

[0083] 1. Pretreatment

[0084] Add 800 mL sodium bicarbonate solution to 100 g corn yellow powder, stir at 45℃ for 18 min, wash and dry, adjust the pH to 4.7, add 1.2 g rizole amylase, continue to stir for 1.0 h, and then perform suction filtration, washing and drying to obtain pretreated corn yellow powder;

[0085] The concentration of the sodium bicarbonate solution is 60%.

[0086] 2. Preparation of enzyme carrier

[0087] (1) Activation

[0088] Put diatomite into 8 times mass of 25 wt% hydrochloric acid solution, raise the temperature to 40℃, and soak for 4.2 h. After soaking, filter out and wash, dry at 82℃ for 8 h, naturally restore to room temperature, then put into a muffle furnace, raise the temperature to 380℃ at a rate of 4.0℃ / min, and keep the temperature for 5.3 h. After calcination, naturally reduce to room temperature to obtain activated diatomite;

[0089] The particle size of the diatomite is 240 nm;

[0090] (2) First treatment

[0091] Put 7.5 g of hydroxylamine hydrochloride into 100 g of deionized water, stir until uniform, to obtain a hydroxylamine hydrochloride solution; put 5.3 g of chitosan into 100 g of 5.0 wt% acetic acid solution, stir until uniform, to obtain a chitosan solution; put 22 g of activated diatomite into 200 g of deionized water, raise the temperature to 43℃, slowly add 38 g of hydroxylamine hydrochloride solution and 37 g of chitosan solution, control the addition rate to be 2.2 g / min, keep the stirring speed at 250 rpm during the addition, after the addition is completed, continue to raise the temperature to 72℃, keep stirring for 2.6 h, after washing and drying, obtain a primary enzyme carrier;

[0092] (3) Secondary treatment

[0093] Put 10.5 g of the primary carrier into 100 g of N,N-dimethylformamide, stir until uniform, then add 3.8 g of succinic anhydride and 0.42 g of triethylamine, raise the temperature to 78℃, keep the reaction for 4.3 h, after the reaction is completed, filter, wash and dry, to obtain an enzyme carrier.

[0094] 3. Preparation of functional enzyme

[0095] Mix 3.2 g of the enzyme carrier with 53 mL of phosphate buffer, stir until uniform, add 0.26 g of enzyme, perform oscillation at room temperature, the oscillation speed is 150 rpm, the oscillation time is 4.2 h, then add 17.0 g of 1.7 wt% cinnamaldehyde solution, continue to perform oscillation reaction for 2.2 h, after the reaction is completed, perform centrifugal washing, and then naturally dry, to obtain a functional enzyme;

[0096] The enzyme is Alcase2.4L protease;

[0097] The molar concentration of the phosphate buffer is 0.04 ml / L, and the pH value is 8.2.

[0098] 4. Enzymatic hydrolysis

[0099] Add 1000 g of deionized water to 100 g of pretreated corn meal, adjust the pH to 8.0, add 22 g of the functional enzyme, and perform reaction at 53℃ for 8.5 h, after the reaction is completed, perform suction filtration to obtain a clear liquid and a solid, add 1.2 wt% activated carbon to the clear liquid, perform stirring treatment for 1.2 h, after the stirring is completed, perform rotary evaporation, and then perform vacuum drying, to obtain a corn oligopeptide product.

[0100] The corn oligopeptide product obtained by the method of Example 3 has a yield of 63.2%, and a protein content of 92.5 wt%.

[0101] Example 4

[0102] 1. Pretreatment

[0103] To 100 g of corn meal, 800 mL of 90 wt% ethanol solution was added, and the mixture was extracted at 73 °C for 3.0 h at 120 rpm. After the extraction, the mixture was filtered to remove the ethanol, and the pretreated corn meal was obtained after washing and drying.

[0104] 2. Preparation of enzyme carrier

[0105] (1) Activation

[0106] The diatomite was placed in 8 times the mass of 22 wt% hydrochloric acid solution, and the temperature was raised to 37 °C for 3.8 h. After the soaking, the mixture was filtered, washed, and dried at 80 °C for 12 h. The mixture was then placed in a muffle furnace, and the temperature was raised to 360 °C at a rate of 3.5 °C / min. The mixture was calcined for 4.8 h, and then the temperature was lowered to room temperature. The activated diatomite was obtained.

[0107] The diatomite had a particle size of 230 nm.

[0108] (2) First treatment

[0109] The hydroxylamine hydrochloride solution was prepared by adding 7.2 g of hydroxylamine hydrochloride to 100 g of deionized water and stirring until the mixture was uniform. The chitosan solution was prepared by adding 4.8 g of chitosan to 100 g of 4.5 wt% acetic acid solution and stirring until the mixture was uniform. The activated diatomite was placed in 200 g of deionized water, and the temperature was raised to 42 °C. Then, 37 g of the hydroxylamine hydrochloride solution and 34 g of the chitosan solution were slowly added to the mixture at a rate of 2.1 g / min while stirring at 245 rpm. After the addition was completed, the temperature was raised to 69 °C, and the mixture was stirred for 2.4 h. The primary enzyme carrier was obtained after washing and drying.

[0110] (3) Second treatment

[0111] The primary carrier was placed in 100 g of N,N-dimethylformamide, and the mixture was stirred until the mixture was uniform. Then, 3.7 g of succinic anhydride and 0.38 g of triethylamine were added, and the temperature was raised to 76 °C. The mixture was reacted for 4.2 h, and then the enzyme carrier was obtained after filtering, washing, and drying.

[0112] 3. Preparation of functional enzyme

[0113] The enzyme carrier was mixed with 50 mL of phosphate buffer solution, and the mixture was stirred until the mixture was uniform. Then, 0.26 g of enzyme was added, and the mixture was shaken at room temperature at a speed of 138 rpm for 3.8 h. Then, 16.8 g of 1.4 wt% cinnamaldehyde solution was added, and the mixture was shaken for an additional 2.1 h. The functional enzyme was obtained after centrifugation, washing, and natural drying.

[0114] The enzymes are Alcase 2.4L protease and Flavourzyme 1000L enzyme, and the mass ratio of the Alcase 2.4L protease and the Flavourzyme 1000L enzyme is 2:1.

[0115] The molar concentration of the phosphate buffer is 0.05ml / L, and the pH value is 8.

[0116] 4. Enzymolysis

[0117] 1000g of deionized water is added to 100g of pretreated corn meal, the pH value is adjusted to 8.0, 40g of functional enzyme is added, and the reaction is carried out at 56℃ for 7.5h. After the reaction is completed, the filtrate and solid are obtained by suction filtration, 0.8wt% of activated carbon is added to the filtrate, and stirring treatment is carried out for 1.3h. After the stirring is completed, rotary evaporation is carried out, and vacuum drying is carried out, to obtain a corn oligopeptide product.

[0118] The corn oligopeptide product obtained by the method of Example 4 has a yield of 62.5% and a protein content of 93.4wt%.

[0119] The repeated use performance of the functional enzyme prepared in Example 4 is tested. Specifically, after the enzymolysis reaction is completed, the filtrate and solid are obtained by suction filtration, the solid is washed with a phosphate buffer, the molar concentration of the phosphate buffer is 0.05ml / L, and the pH value is 8. After being washed for 3 times, drying is carried out at 40℃ until the constant weight is reached. Then, the same operation steps in the enzymolysis are carried out to obtain a corn oligopeptide product. The above operation is one cycle, and the corn oligopeptide product obtained by testing is cycled for 20 times. The yield of the corn oligopeptide product is 58.7%, and the protein content is 88.6%.

[0120] Comparative Example 1

[0121] On the basis of Example 4, the pretreatment step is omitted. In the enzymolysis step, the pretreated corn meal is replaced with corn meal that is not treated in any way.

[0122] The remaining operations are the same.

[0123] The corn oligopeptide product obtained by the method of Comparative Example 1 has a yield of 35.8% and a protein content of 68.5wt%.

[0124] Comparative Example 2

[0125] On the basis of Example 4, the steps of preparing the enzyme carrier and preparing the functional enzyme are omitted.

[0126] The enzymolysis step is that 1000g of deionized water is added to 100g of pretreated corn meal, the pH is adjusted to 8.0, 3.0g of enzyme is added, and the reaction is carried out at 56 DEG C for 7.5h; after the reaction is completed, the supernatant is obtained by suction filtration, 0.8wt% of activated carbon is added to the supernatant, stirring treatment is carried out for 1.3h, after the stirring is completed, rotary evaporation is carried out, and vacuum drying is carried out to obtain a corn oligopeptide product;

[0127] The enzyme is Alcase2.4L protease and Flavourzyme1000L enzyme, and the mass ratio of the Alcase2.4L protease and the Flavourzyme1000L enzyme is 2:1;

[0128] The remaining operations are the same.

[0129] The corn oligopeptide product obtained by the method of the comparative example 2 has a yield of 42.3% and a protein content of 73.6wt%.

[0130] In the process of preparing the corn oligopeptide, the corn meal is pretreated to remove pigments and oils, and then enzymolysis is carried out by using a functional enzyme; in the preparation method of the functional enzyme, diatomite is used as an enzyme carrier matrix, the diatomite is treated by using a dilute acid solution to remove impurities on the surface, increase the specific surface area and the activity of the diatomite, and the stability of the diatomite is improved after calcination; in the first treatment step, the amino group is introduced on the surface of the diatomite by introducing hydroxylamine hydrochloride and diatomite, and the chitosan molecules can also enhance the hydrophilic property and biocompatibility of the diatomite carrier; the primary enzyme carrier is reacted with succinic anhydride, and then the carboxyl group is introduced on the surface of the enzyme carrier, and the hydrophilic property and the reactivity of the carrier are further increased; in the preparation process of the functional enzyme, the cinnamyl aldehyde solution is added, which can react with the enzyme molecules and the amino group on the surface of the enzyme carrier, and the binding force between the enzyme and the enzyme carrier is enhanced, so that the enzyme molecules are more firmly fixed on the enzyme carrier, the stability of the functional enzyme is ensured, the probability of enzyme inactivation and denaturation is reduced, the enzyme activity can be maintained for a long time during the enzymolysis process, the activated carbon adsorption step is combined, the pigments and bitterness of the product are effectively removed, the reusability of the enzyme is improved, the contact area with the substrate is increased, the enzymolysis efficiency is improved, the enzymolysis reaction is promoted, and the product yield and the protein content are improved.

[0131] The comparative example 1 omits the pretreatment step for the corn gluten meal, which cannot effectively remove the pigment and oil of the corn gluten meal, and the structure of the corn protein is still relatively tight, which is not conducive to the subsequent binding and action of the enzyme and the protein molecule, thereby leading to incomplete enzymolysis, so that the corn protein cannot be converted into oligopeptide, which exists in the form of a large molecule protein in the solid matter, finally reducing the product yield and protein content; the comparative example 2 does not load the enzyme, in the enzymolysis step, the contact area of the enzyme and the substrate is small, which affects the progress of the enzymolysis reaction, and the reaction process is unstable, finally reducing the yield of the corn oligopeptide and the protein content.

[0132] Unless otherwise specified, the proportions described in the present application are mass proportions, and the percentages described are mass percentages.

[0133] Finally, it should be noted that: the above only describes the preferred embodiments of the present application, and is not used to limit the present application, although the present application has been described in detail with reference to the foregoing embodiments, for those skilled in the art, the technical solutions recorded in the foregoing embodiments can still be modified, or some technical features can be replaced equivalently. Any modification, equivalent replacement, improvement, etc. made within the spirit and principles of the present application shall be included in the protection scope of the present application.

Claims

1. A method for producing corn oligopeptides, characterized in that, This includes pretreatment, preparation of enzyme carriers, preparation of functional enzymes, and enzymatic hydrolysis steps; The pretreatment step is to add a pretreatment solution to the corn yellow powder to obtain pretreated corn yellow powder. In the pretreatment step, the mass-to-volume ratio of corn yellow powder to pretreatment liquid is 100g:800-1000mL; The pretreatment process involves either extraction with 50-90 wt% ethanol solution or stirring with 10-60 wt% sodium bicarbonate solution. The extraction process is carried out at 70-73℃ and 100-120 rpm for 3.0-3.5 hours. The stirring process was carried out at a temperature of 45°C for 18 minutes. When adding 10-60 wt% sodium bicarbonate solution and stirring, after the treatment is completed, the pH can be adjusted to 4.7, and rifasulfanilase can be added, followed by stirring for another 1.0 h. The rifasulfanilase is 1.2 wt% of the corn yellow flour. The preparation of the enzyme carrier includes activation, primary treatment and secondary treatment steps; The first processing step is as follows: hydroxylamine hydrochloride is added to deionized water and stirred evenly to obtain a hydroxylamine hydrochloride solution; chitosan is added to acetic acid solution and stirred evenly to obtain a chitosan solution; activated diatomaceous earth is placed in deionized water, the temperature is raised to 37-43℃, and the hydroxylamine hydrochloride solution and chitosan solution are slowly added, controlling the addition rate at 1.8-2.2 g / min, while maintaining a stirring speed of 230-250 rpm. After the addition is completed, the temperature is further raised to 68-72℃, and the mixture is kept warm and stirred for 2.3-2.6 h. After washing and drying, the primary enzyme carrier is obtained. The secondary processing step is as follows: the primary carrier is placed in N,N-dimethylformamide, stirred evenly, succinic anhydride and triethylamine are added, the temperature is raised to 72-78℃, and the reaction is maintained for 3.8-4.3h. After the reaction is completed, the carrier is filtered, washed and dried to obtain the enzyme carrier. The steps for preparing the functional enzyme are as follows: mix the enzyme carrier with phosphate buffer, stir evenly, add the enzyme, and shake at room temperature at a speed of 130-150 rpm for 3.8-4.2 h. Then add cinnamaldehyde solution and continue shaking for 1.8-2.2 h. After the reaction is completed, the enzyme is washed by centrifugation and then dried naturally to obtain the functional enzyme. The enzyme is one or both of Alcase 2.4L protease and Flavorzyme 1000L enzyme, and the mass ratio of Alcase 2.4L protease to Flavorzyme 1000L enzyme is 2:

1. The enzymatic hydrolysis step is as follows: deionized water is added to pretreated corn yellow powder to adjust the pH to 7.5-8.0, functional enzymes are added, and the reaction is carried out at 53-56℃ for 7.5-8.5 hours. After the reaction is completed, the clear liquid and solids are obtained by suction filtration. 0.8-1.2wt% activated carbon is added to the clear liquid and stirred for 1.0-1.3 hours. After stirring is completed, the product is vacuum dried after rotary evaporation to obtain the corn oligopeptide product. The mass ratio of the pretreated corn yellow powder, deionized water, and functional enzyme is 100:900-1000:22-40.

2. The method for producing corn oligopeptides according to claim 1, characterized in that, The activation step is as follows: diatomaceous earth is placed in a hydrochloric acid solution with a mass of 6-8 times its weight, the temperature is raised to 35-40℃, and it is kept at this temperature for 3.6-4.2 hours. After soaking, it is filtered out, washed, and dried at 78-82℃ for 8-12 hours. After naturally returning to room temperature, it is then placed in a muffle furnace and heated to 350-380℃ at a rate of 2.0-4.0℃ / min. It is kept at this temperature for calcination for 4.7-5.3 hours. After calcination, it is allowed to naturally cool to room temperature to obtain activated diatomaceous earth. The particle size of the diatomaceous earth is 210-240 nm; The mass concentration of the hydrochloric acid solution is 20-25%.

3. The method for producing corn oligopeptides according to claim 1, characterized in that, The mass ratio of hydroxylamine hydrochloride to deionized water is 6.5-7.5:100; The mass ratio of chitosan to acetic acid solution is 4.6-5.3:100; The acetic acid solution has a mass concentration of 3.0-5.0%; The mass ratio of the activated diatomaceous earth, deionized water, hydroxylamine hydrochloride solution, and chitosan solution is 18-22:200:34-38:33-37.

4. The method for producing corn oligopeptides according to claim 1, characterized in that, In the secondary processing step, the mass ratio of the primary carrier, N,N-dimethylformamide, succinic anhydride, and triethylamine is 9.5-10.5:100:3.4-3.8:0.37-0.

42.

5. The method for producing corn oligopeptides according to claim 1, characterized in that, The mass-to-volume ratio of the enzyme carrier, phosphate buffer, enzyme, and cinnamaldehyde solution is 2.8-3.2 g: 48-53 mL: 0.24-0.26 g: 16.0-17.0 g; The cinnamaldehyde solution has a mass concentration of 1.3-1.7%; The phosphate buffer solution has a molar concentration of 0.04-0.06 ml / L and a pH value of 7.8-8.2.