Preparation methods and applications of yeast protein-polysaccharide complexes

Abstract

This invention relates to the field of biotechnology, and particularly to a method for preparing and applying yeast protein-polysaccharide complexes. The invention provides a method for preparing yeast protein-polysaccharide complexes, which involves compounding yeast protein with polysaccharides while enzymatically hydrolyzing the protein. This rapidly improves the functional properties of the yeast protein while encapsulating some bitterness and unpleasant flavors. The yeast protein-polysaccharide complexes prepared by this invention through enzymatic hydrolysis and compounding processes are less prone to sedimentation. The yeast protein prepared by this method exhibits a foaming power of 150.00% and an emulsifying property of 67 μm. 2 / g, with a water-holding capacity of 700%, a delicate taste and mild aroma, and a production process that does not involve organic solvents, making it clean and environmentally friendly. It is suitable for use in food systems such as solid beverages and artificial meat.

Description

Technical Field

[0001] This invention relates to the field of biotechnology, and in particular to a method for preparing and applying yeast protein-polysaccharide complexes. Background Technology

[0002] Yeast protein is a microbial protein extracted from yeast powder. Yeast protein contains all nine essential amino acids for the human body (including histidine, essential for children's growth and development). Except for sulfur-containing amino acids, which are slightly lower than the FAO / WHO (2007) recommended essential amino acid score (AAS), the AAS of other amino acids are all above 100. The total essential amino acids account for 47.58% of the total amino acids, and the ratio of essential to non-essential amino acids reaches 0.91, meeting the FAO / WHO standards of 40% and 0.6, respectively, which is higher than that of soy protein and close to that of whey protein. However, ordinary yeast protein has disadvantages such as lack of prominent functional properties, easy sedimentation, and a gritty texture. Its large molecular structure easily aggregates, making it difficult to dissolve in water, which greatly limits its application in the food industry.

[0003] To enhance the practicality of yeast proteins, methods such as glycosylation, high-pressure homogenization, ultrasonication, acid-base extraction, and enzymatic modification are commonly used to modify them. While moderate modification can improve some functional properties of yeast proteins, the effects are still not significant, and most methods have problems such as specific equipment requirements, low recovery rates, and the tendency to produce bitterness and develop a salty or umami flavor.

[0004] CN111903967A describes a method for preparing protein-polysaccharide complexes using spray drying technology. This method accelerates the dispersion of protein suspensions by dry mixing of proteins, polysaccharides, and inorganic salts, and adjusts the pH of the protein-polysaccharide-inorganic salt suspension using heating and high-speed shearing processes. The protein and polysaccharide complexes are bound together via chemical covalent bonds, avoiding complex separation under neutral conditions. The addition of inorganic salts accelerates the complex preparation rate. The complex of this invention consists of two polysaccharides bound to one protein molecule, providing better protection for the protein and resulting in improved heat resistance. However, this method uses a 5:1 to 1:5 ratio of proteoglycans, which, while enhancing the functional properties of yeast proteins through a Maillard reaction, significantly reduces protein purity.

[0005] CN101429226A This invention utilizes different types of glycosyl donors mixed with rice protein in solution for Maillard grafting coupling reaction. The products obtained by the Maillard reaction of rice protein with different types of sugars exhibit superior solubility, emulsifying activity, emulsion stability, and foaming properties compared to the unsweetened protein control. This method integrates protein and polysaccharides through a wet Maillard reaction to enhance the functional properties of rice protein. However, this method uses a pH value between 11 and 12, requires a large amount of alkali solution, consumes significant energy, and this pH range easily induces bitterness in the protein.

[0006] CN115677818A (Angel) inactivates high-protein yeast broth, separates the heavy phase to obtain yeast milk, then enzymatically hydrolyzes the yeast milk with a compound enzyme and ethyl acetate, inactivates it, centrifuges to obtain the heavy phase, and yields low-yeast-flavor yeast protein. The yeast protein obtained by this method has a reduced yeast odor, but a stronger gritty texture and less delicate mouthfeel.

[0007] CN112868884A describes a method that uses microwave heating to induce a Maillard reaction in a mixture of whey protein and lactose, thereby enhancing the emulsifying and antioxidant activity of whey protein isolate. This method utilizes rapid, internally heated molecular vibrations instead of the traditional temperature gradient-based heat conduction method, increasing the reaction rate and significantly shortening the reaction time required to obtain the desired product, thus improving production efficiency. While this method primarily accelerates the Maillard reaction through microwaves, enabling whey protein to bind with lactose and enhancing its functional properties, microwave heating is currently not feasible for large-scale industrial production and still has certain limitations.

[0008] In summary, existing yeast protein products have poor functional properties, especially emulsifying and water-holding properties; they also leave a residue in the mouth and have a pungent feeling in the throat; and they tend to precipitate quickly after dissolving in water, which greatly limits their application in the food industry. Summary of the Invention

[0009] In view of this, the present invention provides a method for preparing and applying a yeast protein-polysaccharide complex. The method comprises the following steps: A) dissolving polysaccharide and protein evenly in pure water and mixing them in a specific ratio, wherein the total amount of polysaccharide added shall not exceed 5% of the protein content; B) adjusting the pH of the uniformly mixed protein-polysaccharide solution to 6.5–7.0 and preheating it to 50–60°C; C) adding protease to the mixed solution to decompose some large-particle aggregates of protein, with an enzymatic hydrolysis time of 2–6 hours; D) spray drying the suspension, with an inlet air temperature not lower than 170°C and an outlet air temperature not lower than 90°C, to obtain a protein-polysaccharide complex powder. Experiments have shown that the preparation method of the present invention rapidly improves the functional properties of yeast protein while also removing some bitterness and unpleasant flavors. The yeast protein and polysaccharide complex prepared by this invention through enzymatic hydrolysis and compounding process is not easy to settle. The yeast protein prepared by this method has a foaming power of 150.00%, an emulsifying property of 67 m2 / g, and a water holding capacity of 700%. It has a delicate taste and light aroma. The production process does not involve organic solvents, is clean and environmentally friendly, and is suitable for use in food systems such as solid beverages and artificial meat.

[0010] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0011] The present invention provides a method for preparing a yeast protein and polysaccharide complex, comprising: preparing an aqueous solution containing yeast protein, mixing it with polysaccharide, enzymatically hydrolyzing it, inactivating the enzyme, drying it, and obtaining the yeast protein and polysaccharide complex;

[0012] On a dry basis, the yeast protein has a protein content of ≥60%, a polysaccharide content of ≤20%, and a nucleic acid content of ≤5%; the mass ratio of yeast protein to polysaccharide includes 100:(1~5).

[0013] Preferably, the mass ratio of yeast protein to polysaccharide is 100:(1.5~2.5).

[0014] In some specific embodiments of the present invention, the mass concentration of the yeast protein in the aqueous solution is 8-15%.

[0015] In some specific embodiments of the present invention, the mass concentration of the yeast protein in the aqueous solution includes 10-12%.

[0016] In some specific embodiments of the present invention, the polysaccharide includes one or more combinations of carrageenan, konjac gum, xanthan gum, pectin, seaweed polysaccharide, or glucan.

[0017] In some specific embodiments of the present invention, the enzymes used in the enzymatic hydrolysis include one or more combinations of alkaline protease, neutral protease, papain, transglutaminase, bromelain, or flavor protease.

[0018] In some specific embodiments of the present invention, the pH value of the solution during enzymatic hydrolysis is 6.0-7.5; the temperature of enzymatic hydrolysis is 45-65°C; the time of enzymatic hydrolysis is 2-6 h; preferably, a pH adjuster is used for the pH value; the pH adjuster includes hydrochloric acid or citric acid; preferably, the temperature of enzymatic hydrolysis is 50-60°C; the time of enzymatic hydrolysis is 2.5-3.5 h.

[0019] In some specific embodiments of the present invention, the amount of enzyme added for enzymatic hydrolysis includes 180.15~1200 U / g of the yeast protein.

[0020] In some specific embodiments of the present invention, the drying includes spray drying; preferably, the inlet air temperature of the spray drying is not lower than 170°C and the outlet air temperature is not lower than 90°C.

[0021] Preferably, the enzyme inactivation conditions include inactivating the enzyme at 90°C for 30 minutes.

[0022] The present invention also provides a yeast protein and polysaccharide complex prepared by the aforementioned method.

[0023] In some specific embodiments of the present invention, on a dry basis, the yeast protein and polysaccharide complex has a protein content ≥80% and free amino nitrogen <5%; the yeast protein and polysaccharide complex has a foaming power ≥150.00% and an emulsifying property ≥67 m. 2 / g, water holding capacity ≥700%.

[0024] The present invention also provides the application of the yeast protein and polysaccharide complex in food processing.

[0025] In some specific embodiments of the present invention, the food includes solid beverages or artificial meat.

[0026] This invention provides the following beneficial effects:

[0027] This invention provides a method for preparing yeast protein and polysaccharide complexes. The method involves compounding yeast protein and polysaccharides while simultaneously using a protease for enzymatic hydrolysis. This rapidly enhances the functional properties of the yeast protein while encapsulating some bitterness and unpleasant flavors. The yeast protein and polysaccharide complexes prepared by this invention through enzymatic hydrolysis and compounding processes are less prone to sedimentation. The yeast protein prepared by this method exhibits a foaming power of 150.00% and an emulsifying property of 67 μm. 2 / g, with a water-holding capacity of 700%, a delicate taste and mild aroma, and a production process that does not involve organic solvents, making it clean and environmentally friendly. It is suitable for use in food systems such as solid beverages and artificial meat. Detailed Implementation

[0028] This invention discloses a method for preparing yeast protein-polysaccharide complexes and their applications. Those skilled in the art can refer to this document and appropriately modify the process parameters to achieve the desired results. It is particularly important to note that all similar substitutions and modifications are obvious to those skilled in the art and are considered to be included within the scope of this invention. The methods and applications of this invention have been described through preferred embodiments. Those skilled in the art can clearly modify or appropriately change and combine the methods and applications described herein without departing from the content, spirit, and scope of this invention to realize and apply the technology of this invention.

[0029] This invention improves the functional properties of yeast protein by enzymatic hydrolysis and polysaccharide combination. While enzymatic hydrolysis opens up the protein macromolecular chain, polysaccharides are added to combine with the protein. This not only exposes more active sites in the protein and enhances its functional properties, but also greatly avoids the activation of bitterness and off-flavor in the protein during the enzymatic hydrolysis process. It is also relatively easy to implement in production.

[0030] This invention relates to a method for preparing a yeast protein-polysaccharide complex. The characteristics of this protein are as follows: protein content is above 80% (dry basis), polysaccharide content is ≤4%, and free amino nitrogen is less than 5%. However, the key feature is its foaming power of 150.00% and emulsifying properties of 67 m. 2 / g, with a water-holding capacity of 700%, and a delicate taste and light aroma.

[0031] Specifically, the preparation method of yeast protein-polysaccharide complex includes the following steps:

[0032] Yeast protein and polysaccharide were prepared into solutions, and protease was added to moderately hydrolyze the protein. After adjusting the pH, the mixture was spray-dried.

[0033] The yeast protein used in this invention is characterized by having a protein content of more than 60%, a polysaccharide content of less than 20%, and a nucleic acid content of less than 5%.

[0034] Preparation: Prepare a solution of yeast protein with water with a mass concentration of 8-15%, preferably 10-12%, and prepare a homogeneous solution of polysaccharide with water.

[0035] Mixing: Add polysaccharides to the protein solution at an addition amount of 1% to 5%, preferably 1.5% to 2.5%. The polysaccharide molecular chains all have a large number of ionic bonds or hydrogen bonds, which can stably bind with proteins or water molecules in the solution. They are one or more of carrageenan, konjac gum, xanthan gum, pectin, seaweed polysaccharides, and dextran.

[0036] Enzymatic hydrolysis: Modified yeast protein is treated with enzymes. The pH of the above mixed solution is adjusted to 6.0-7.0 using hydrochloric acid or citric acid, preferably citric acid. The temperature is 45-65℃, preferably 50-60℃, and the time is 2-6 h, preferably 2.5-4 h. The enzyme is one or a mixture of alkaline protease, neutral protease, papain, transglutaminase, and flavor protease.

[0037] Spray drying: The above suspension is spray dried with an inlet air temperature of not less than 170°C and an outlet air temperature of not less than 90°C to obtain a protein polysaccharide complex powder.

[0038] The reagents and instruments used in this invention are shown in the table below:

[0039] Table 1. Reagents and Instruments

[0040]

[0041] To address the shortcomings of existing technologies, this invention solves the following technical problems: (1) it solves the problems of gritty texture and off-flavor in traditional yeast proteins; (2) it solves the problems of unsatisfactory functional properties of existing yeast proteins, such as poor emulsification, water retention, and dispersion stability. Furthermore, this invention maintains a high protein content while improving the functional properties of the protein. The conditions of this invention are milder, avoiding the use of large amounts of acids and alkalis, and effectively improving the texture of the protein. This invention is not limited by equipment or materials and is simple and easy to implement.

[0042] Unless otherwise specified, the raw materials and reagents used in the preparation method and application of the yeast protein and polysaccharide complex provided by this invention can all be purchased from the market.

[0043] The present invention will be further illustrated below with reference to the embodiments:

[0044] Example 1

[0045] 1. Preparation of yeast protein aqueous solution:

[0046] 1 kg of yeast protein powder was added to purified water to prepare a 9 kg solution, resulting in a yeast protein aqueous solution with a mass concentration of 11.1%. Citric acid was added to adjust the pH to 6.5.

[0047] 2. Add polysaccharides

[0048] Prepare a 1 kg solution by adding 20 g xanthan gum to pure water, and after fully dispersing and dissolving, mix it evenly with the yeast protein solution.

[0049] 3. Enzymatic hydrolysis

[0050] After adjusting the pH to 6.5, add 2 g of flavor protease (enzyme activity 180,000 U / g) and hydrolyze at 50℃ for 3 h. After hydrolysis, inactivate the enzyme at 90℃ for 30 min.

[0051] 4. Spray drying

[0052] Yeast protein and polysaccharide complex was obtained by spray drying, with the inlet air temperature not lower than 170℃ and the outlet air temperature not lower than 90℃, thus producing protein-polysaccharide complex powder.

[0053] Example 2

[0054] 1. Preparation of yeast protein aqueous solution:

[0055] Add 1 kg of yeast protein powder to purified water to prepare a 10 kg solution, obtaining a yeast protein aqueous solution with a mass concentration of 10%. Add citric acid to adjust the pH to 7.5.

[0056] 2. Add polysaccharides

[0057] After 10g xanthan gum and 10g carrageenan are fully dispersed and dissolved, they are mixed evenly with yeast protein solution (polysaccharide addition is 0.2%, w / w).

[0058] 3. Enzymatic hydrolysis

[0059] After adjusting the pH to 7.5, add 2 g of basic protein (enzyme activity 180,000 U / g) and hydrolyze at 55℃ for 3 h. After hydrolysis, inactivate the enzyme at 90℃ for 30 min.

[0060] 4. Spray drying

[0061] Yeast protein and polysaccharide complex was obtained by spray drying, with the inlet air temperature not lower than 170℃ and the outlet air temperature not lower than 90℃, thus producing protein-polysaccharide complex powder.

[0062] Example 3

[0063] 1. Preparation of yeast protein aqueous solution:

[0064] Add 1 kg of yeast protein powder to purified water to prepare a 10 kg solution, obtaining a yeast protein aqueous solution with a mass concentration of 10%. Add citric acid to adjust the pH to 7.0.

[0065] 2. Add polysaccharides

[0066] After fully dispersing and dissolving 10 g xanthan gum and 15 g high-methoxyl pectin, mix them evenly with yeast protein solution (polysaccharide addition is 0.25%, w / w).

[0067] 3. Enzymatic hydrolysis

[0068] After adjusting the pH to 7.0, add 2 g of basic protein (enzyme activity 180,000 U / g) and hydrolyze at 50℃ for 3.5 h. After hydrolysis, inactivate the enzyme at 90℃ for 30 min.

[0069] 4. Spray drying

[0070] Yeast protein and polysaccharide complex was obtained by spray drying, with the inlet air temperature not lower than 170℃ and the outlet air temperature not lower than 90℃, thus producing protein-polysaccharide complex powder.

[0071] Example 4

[0072] 1. Preparation of yeast protein aqueous solution:

[0073] Add 1 kg of yeast protein powder to purified water to prepare a 10 kg solution, obtaining a yeast protein aqueous solution with a mass concentration of 10%. Add citric acid to adjust the pH to 7.5.

[0074] 2. Add polysaccharides

[0075] After fully dispersing and dissolving 15 g of carrageenan and 10 g of pectin, mix them evenly with the yeast protein solution (polysaccharide addition is 0.25%, w / w).

[0076] 3. Enzymatic hydrolysis

[0077] After adjusting the pH to 7.5, add 2 g of papain (enzyme activity of 600,000 U / g) and hydrolyze at 50℃ for 3 h. After hydrolysis, inactivate the enzyme at 90℃ for 30 min.

[0078] 4. Spray drying

[0079] Yeast protein and polysaccharide complex was obtained by spray drying, with the inlet air temperature not lower than 170℃ and the outlet air temperature not lower than 90℃, thus producing protein-polysaccharide complex powder.

[0080] Example 5

[0081] 1. Preparation of yeast protein aqueous solution:

[0082] Add 1 kg of yeast protein powder to purified water to prepare a 10 kg solution, obtaining a yeast protein aqueous solution with a mass concentration of 10%. Add citric acid to adjust the pH to 7.5.

[0083] 2. Add polysaccharides

[0084] After fully dispersing and dissolving 10 g of yeast β-glucan and 15 g of pectin, mix them evenly with the yeast protein solution (polysaccharide addition is 0.25%, w / w).

[0085] 3. Enzymatic hydrolysis

[0086] After adjusting the pH to 7.5, add 0.5 g of transglutaminase (enzyme activity 300 U / g) and 1 g of alkaline protease (180,000 U / g) and hydrolyze at 60℃ for 3 h. After hydrolysis, inactivate the enzyme at 90℃ for 30 min.

[0087] 4. Spray drying

[0088] Yeast protein and polysaccharide complex was obtained by spray drying, with the inlet air temperature not lower than 170℃ and the outlet air temperature not lower than 90℃, thus producing protein-polysaccharide complex powder.

[0089] Example 6

[0090] 1. Preparation of yeast protein aqueous solution:

[0091] Add 1 kg of yeast protein powder to purified water to prepare a 10 kg solution, obtaining a yeast protein aqueous solution with a mass concentration of 10%. Add citric acid to adjust the pH to 6.0.

[0092] 2. Add polysaccharides

[0093] After fully dispersing and dissolving 5g of yeast β-glucan, 15g of carrageenan and 5g of xanthan gum, mix them evenly with the yeast protein solution (polysaccharide addition is 0.25%, w / w).

[0094] 3. Enzymatic hydrolysis

[0095] After adjusting the pH to 6.0, add 1 g of papain (600,000 U / g) and 1.5 g of flavor protease (180,000 U / g) and hydrolyze at 50°C for 2.5 h. After hydrolysis, inactivate the enzyme at 90°C for 30 min.

[0096] 4. Spray drying

[0097] Yeast protein and polysaccharide complex was obtained by spray drying, with the inlet air temperature not lower than 170℃ and the outlet air temperature not lower than 90℃, thus producing protein-polysaccharide complex powder.

[0098] Example 7

[0099] 1. Preparation of yeast protein aqueous solution:

[0100] Add 1 kg of yeast protein powder to purified water to prepare a 10 kg solution, obtaining a yeast protein aqueous solution with a mass concentration of 10%. Add citric acid to adjust the pH to 6.5.

[0101] 2. Add polysaccharides

[0102] After fully dispersing and dissolving 5 g of konjac gum, 10 g of pectin and 10 g of xanthan gum, mix them evenly with the yeast protein solution (polysaccharide addition is 0.25%, w / w).

[0103] 3. Enzymatic hydrolysis

[0104] After adjusting the pH to 7.5, add 1 g of transglutaminase (enzyme activity 300 U / g) and 2 g of alkaline protease (180,000 U / g) and hydrolyze at 50℃ for 3 h. After hydrolysis, inactivate the enzyme at 90℃ for 30 min.

[0105] 4. Spray drying

[0106] Yeast protein and polysaccharide complex was obtained by spray drying, with the inlet air temperature not lower than 170℃ and the outlet air temperature not lower than 90℃, thus producing protein-polysaccharide complex powder.

[0107] Example 8

[0108] Replace xanthan gum with low-methoxyl pectin and flavor protease with acidic protease, and perform the remaining operations as in Example 1.

[0109] Example 9

[0110] Replace xanthan gum with seaweed polysaccharide, replace flavor protease with neutral protease, and perform the remaining operations as in Example 1.

[0111] Example 10

[0112] Replace xanthan gum with seaweed polysaccharide and flavor protease with bromelain (enzyme activity of 100,000 U / g), and perform the remaining operations as in Example 1.

[0113] Comparative Example 1: Enzymatic hydrolysis without polysaccharides

[0114] 1 kg of yeast protein powder was added to purified water to prepare a 10 kg solution. Citric acid was added to adjust the pH to 7.5, and 2 g of alkaline protease (enzyme activity of 180,000 U / g) was added for enzymatic hydrolysis at 50°C for 3.5 h. After hydrolysis, the enzyme was inactivated at 90°C for 30 min, and the yeast protein-polysaccharide complex was obtained by spray drying.

[0115] Comparative Example 2: Enzymatic hydrolysis without polysaccharides

[0116] 1 kg of yeast protein powder was added to purified water to prepare a 10 kg solution. Citric acid was added to adjust the pH to 7.5, and 2 g of alkaline protease (enzyme activity of 180,000 U / g) was added for enzymatic hydrolysis at 50°C for 5 h. After enzymatic hydrolysis, the enzyme was inactivated at 90°C for 30 min, and the yeast protein-polysaccharide complex was obtained by spray drying.

[0117] Comparative Example 3: Enzymatic hydrolysis + 1% polysaccharide

[0118] 1 kg of yeast protein powder was added to purified water to prepare a 9 kg solution. Citric acid was added to adjust the pH to 6.5. 10 g of xanthan gum was added to purified water to prepare a 1 kg solution. After thorough dispersion and dissolution, the xanthan gum solution was mixed evenly with the yeast protein solution. After adjusting the pH to 6.5, 2 g of flavor protease (enzyme activity of 180,000 U / g) was added and enzymatically hydrolyzed at 50℃ for 3 h. After enzymatic hydrolysis, the enzyme was inactivated at 90℃ for 30 min, and the yeast protein-polysaccharide complex was obtained by spray drying.

[0119] Comparative Example 4: Enzymatic hydrolysis + 5% polysaccharide

[0120] 1 kg of yeast protein powder was added to purified water to prepare a 9 kg solution. Citric acid was added to adjust the pH to 6.5. 20 g of xanthan gum and 30 g of high-methoxyl pectin were added to purified water to prepare a 1 kg solution. After thorough dispersion and dissolution, the xanthan gum and high-methoxyl pectin were mixed evenly with the yeast protein solution. After adjusting the pH to 6.5, 2 g of alkaline protease (enzyme activity of 180,000 U / g) was added and enzymatically hydrolyzed at 50°C for 3 h. After enzymatic hydrolysis, the enzyme was inactivated at 90°C for 30 min. The yeast protein and polysaccharide complex was obtained by spray drying.

[0121] Comparative Example 5: Polysaccharide without enzymatic hydrolysis

[0122] 1 kg of yeast protein powder was added to purified water to prepare a 10 kg solution. Citric acid was added to adjust the pH to 7.5. 20 g of xanthan gum and 30 g of pectin were fully dispersed and dissolved, and then mixed evenly with the yeast protein solution. The yeast protein and polysaccharide complex was obtained by spray drying.

[0123] Comparative Example 6: Enzymatic hydrolysis + monosaccharide

[0124] 1 kg of yeast protein powder was added to purified water to prepare a 9 kg solution. Citric acid was added to adjust the pH to 6.5. 20 g of fructose was added to purified water to prepare a 1 kg solution. After thorough dispersion and dissolution, the fructose solution was mixed evenly with the yeast protein solution. After adjusting the pH to 6.5, 2 g of flavor protease (enzyme activity of 180,000 U / g) was added and enzymatically hydrolyzed at 50 °C for 3 h. After enzymatic hydrolysis, the enzyme was inactivated at 90 °C for 30 min, and the yeast protein-polysaccharide complex was obtained by spray drying.

[0125] Comparative Example 7: Enzymatic hydrolysis + disaccharide

[0126] 1 kg of yeast protein powder was added to purified water to prepare a 9 kg solution. Citric acid was added to adjust the pH to 6.5. 20 g of lactose was added to purified water to prepare a 1 kg solution. After thorough dissolution, the lactose solution was mixed evenly with the yeast protein solution. After adjusting the pH to 6.5, 2 g of flavor protease (enzyme activity of 180,000 U / g) was added and enzymatically hydrolyzed at 50 °C for 3 h. After enzymatic hydrolysis, the enzyme was inactivated at 90 °C for 30 min. The yeast protein and polysaccharide complex was obtained by spray drying.

[0127] Comparative Example 8: pH value was 5.5

[0128] 1 kg of yeast protein powder was added to purified water to prepare a 9 kg solution. Citric acid was added to adjust the pH to 5.5. 20 g of lactose was added to purified water to prepare a 1 kg solution. After thorough dispersion and dissolution, the lactose solution was mixed evenly with the yeast protein solution. After adjusting the pH to 5.5, 2 g of flavor protease (enzyme activity of 180,000 U / g) was added and enzymatically hydrolyzed at 50 °C for 3 h. After enzymatic hydrolysis, the enzyme was inactivated at 90 °C for 30 min, and the yeast protein-polysaccharide complex was obtained by spray drying.

[0129] Comparative Example 9: pH value was 8.0

[0130] 1 kg of yeast protein powder was added to purified water to prepare a 9 kg solution. Citric acid was added to adjust the pH to 8.0. 20 g of lactose was added to purified water to prepare a 1 kg solution. After thorough dissolution, the lactose solution was mixed evenly with the yeast protein solution. After adjusting the pH to 8.0, 2 g of flavor protease (enzyme activity of 180,000 U / g) was added and enzymatically hydrolyzed at 50 °C for 3 h. After enzymatic hydrolysis, the enzyme was inactivated at 90 °C for 30 min. The yeast protein and polysaccharide complex was obtained by spray drying.

[0131] Example 1: Functional determination of yeast protein-polysaccharide complex

[0132] 1. Determination of foaming ability and foaming stability

[0133] Yeast protein and polysaccharide complexes obtained from Preparation Examples 1-10 and Comparative Examples 1-9 were used to prepare a 0.6% (w / w) yeast protein and polysaccharide complex dispersion using pH 7.0, 0.05 mol / L phosphate buffer. 20 mL of the dispersion was placed in a 100 mL graduated cylinder and homogenized (10000 r / min, 1 min). The foam volume V0 was read immediately and allowed to stand at room temperature for 30 min. The foam volume V30 was then read.

[0134]

[0135] 2. Determination of emulsifying properties and emulsion stability

[0136] Yeast protein and polysaccharide complexes obtained in Preparation Examples 1-10 and Comparative Examples 1-9 were prepared as 2 mg / mL yeast protein and polysaccharide complex dispersions and mixed with soybean oil at a ratio of 3:1 (v / v). The mixtures were then homogenized at 21000 r / min for 2 min. Immediately after homogenization and 10 min after homogenization, 50 μL of the emulsion was aspirated from the bottom of the container. The emulsion was then diluted to 5 mL with 0.1% SDS solution (0.01 mol / L, pH 7.0), mixed thoroughly, and the absorbance was measured at 500 nm using a UV-Vis spectrophotometer. The absorbance value A0 was recorded. After standing for 10 min, the absorbance was measured again, and the absorbance value A10 was recorded. Emulsifiable index (EAI) and emulsion stability (ESI) were calculated using the following formulas:

[0137]

[0138] Where DF represents the dilution factor (100); φ represents the optical path of the cuvette (1 cm); θ represents the oil phase volume fraction (0.25); and C represents the initial protein concentration (g / mL).

[0139] 3. Water retention

[0140] Take 3 g of the yeast protein and polysaccharide complex obtained from Preparation Examples 1-10 and Comparative Examples 1-9, mix with 25 mL of deionized water, and stir for 1 minute every 10 minutes for a total of 6 times. Then centrifuge at 3000 r / min at room temperature for 25 minutes, remove the supernatant, weigh, and calculate the water holding capacity using the following formula:

[0141]

[0142] The results are shown in Table 2. Examples 1 to 10 exhibited excellent performance in foaming ability, foaming stability, emulsification, emulsification stability, and water retention. The foaming ability was mostly above 130%, the foaming stability ranged from 78.2% to 92.3%, and the emulsification was consistently above 69.24 m. 2 / g, the emulsification stability of all samples exceeded 1200 minutes, and the water retention was all above 682.5%. Comparative Examples 1 to 9 were relatively weaker in these properties, especially Comparative Examples 1, 2, 6, and 7 without added polysaccharides, which showed poor emulsification and water retention. This indicates that the formulations or processes in the examples are superior and can significantly improve the foaming, emulsification, and water retention properties of the materials.

[0143] Table 2. Determination of foaming properties, emulsifying properties and water-holding capacity

[0144]

[0145] Note: a, b, c, d, e, f represent the differences between groups in each column of data, p < 0.05.

[0146] Example 2: Sensory Evaluation

[0147] Ten people were randomly selected to provide sensory evaluations.

[0148] Table 3. Sensory Evaluation Scoring Criteria

[0149]

[0150] The results are shown in Table 4. Considering color, odor, taste, and texture, the yeast protein and polysaccharide complexes of Examples 1-10 are superior to those of Comparative Examples 1-9. There are no significant differences in appearance, color, and texture between the examples and the comparative examples. The main differences are in taste and odor. The samples from the examples have a more delicate and smooth texture. In the comparative examples, the yeast protein that was only enzymatically hydrolyzed (Comparative Examples 1 and 2) could not mask the unpleasant odor produced by small molecules during the enzymatic hydrolysis process. Over-hydrolysis also affected the protein flavor to some extent. Adding a small amount of polysaccharide (Comparative Example 3) could not achieve the desired smooth texture, while adding too much (Comparative Example 4) would make the sample too viscous and have poor palatability. Adding polysaccharide alone (Comparative Example 5) could not completely remove the gritty texture of the yeast protein. Adding monosaccharides and disaccharides (Comparative Examples 6 and 7) did not significantly improve the taste of the yeast protein. In addition, an excessively high pH value (Comparative Example 9) would also cause a slightly astringent taste in the solution. However, using both methods in combination can optimize the functional properties of the yeast protein while ensuring its excellent sensory properties.

[0151] Table 4. Sensory Evaluation Table for 5% Solution

[0152]

[0153] Note: a, b, c, d, e, f represent the differences between groups in each column of data, p < 0.05.

[0154] Example 3: Determination of protein and free amino nitrogen content in protein-polysaccharide complex powder

[0155] Detection of protein and free amino nitrogen content

[0156] The protein content determination method refers to Method I of GB5009.5-2016, Kjeldahl method.

[0157] The free amino nitrogen content was determined using the ninhydrin method.

[0158] I. Reagent Preparation

[0159] 1. Standard amino acid solution

[0160] Accurately weigh a certain amount of glycine (e.g., 0.1 g), dissolve it in distilled water, and dilute to a specific volume (e.g., 100 mL) to prepare a 1 mg / mL standard solution. If necessary, this standard solution can be diluted to prepare a series of solutions of different concentrations for the purpose of constructing a standard curve.

[0161] 2. Ninhydrin solution

[0162] Weigh an appropriate amount of ninhydrin (usually about 2 g) and dissolve it in an appropriate amount of glacial acetic acid (e.g., 100 mL) to prepare the ninhydrin reagent. This solution is usually blue and should be sealed and stored in a brown bottle in the dark.

[0163] II. Construction of Standard Curve

[0164] 1. Sampling

[0165] Take 1 mL of standard amino acid solutions of different concentrations (e.g., 0.1 mg / mL, 0.2 mg / mL, 0.4 mg / mL, 0.6 mg / mL, 0.8 mg / mL, 1.0 mg / mL, etc.) into a series of test tubes, and take 1 mL of distilled water as a blank control.

[0166] 2. Color reaction

[0167] Add 2 mL of ninhydrin solution and 2 mL of citrate-sodium citrate buffer solution (pH 5.6–6.0) to each test tube, and gently mix. Place the test tubes in a boiling water bath and heat for 15–20 minutes. During this process, the solution will change color, reacting with free amino groups to form a blue compound.

[0168] 3. Cooling and colorimetry

[0169] After heating, remove the test tubes and quickly cool them to room temperature. Then transfer the solutions from each test tube to cuvettes and measure the absorbance at 570 nm using a spectrophotometer. Plot a standard curve with the concentration of the standard amino acid solution on the x-axis and the corresponding absorbance values ​​on the y-axis.

[0170] III. Sample Determination

[0171] 1. Sample processing

[0172] Take an appropriate amount of the protein solution to be tested and dilute it according to its approximate free amino nitrogen content (if the content is high, appropriate dilution is required to ensure that the test results are within the linear range of the standard curve). Generally, take 2 to 3 mL of the diluted sample solution in a test tube.

[0173] 2. Colorimetric reaction and colorimetry

[0174] Add the same volume of ninhydrin solution and buffer solution (e.g., 2 mL ninhydrin solution and 2 mL buffer solution) to the sample solution test tube as used for preparing the standard curve. Shake well and heat in a boiling water bath for the same time (15-20 minutes). After cooling, measure the absorbance at a wavelength of 570 nm.

[0175] IV. Result Calculation

[0176] The free amino nitrogen content (expressed as nitrogen, in mg / mL) can be calculated using the following formula:

[0177] Free amino nitrogen content (mg / mL) = Free amino acid concentration (mg / mL) × Amino acid nitrogen content ratio factor.

[0178] For the standard curve prepared by the glycine standard solution, since the molecular weight of glycine is 75.07 and the nitrogen content is 14.01 / 75.07≈0.1866, when calculating the free amino nitrogen content in the sample, the glycine concentration obtained from the lookup is multiplied by 0.1866 to obtain the content in terms of nitrogen.

[0179] The results are shown in Table 5. Adding excessive polysaccharides (greater than 5%) will reduce the protein content of the sample, while excessive enzymatic hydrolysis will increase the free amino nitrogen content of the sample, which will easily trigger the bitterness and off-flavor of the protein and affect the overall nutritional composition and sensory properties of the protein.

[0180] Table 5. Protein and Free Amino Nitrogen Content in Samples

[0181]

[0182] Note: a, b, c, d, e, f represent the differences between groups in each column of data, p < 0.05.

[0183] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for preparing a yeast protein-polysaccharide complex, characterized in that, include: An aqueous solution containing yeast protein was prepared, mixed with polysaccharide, enzymatically hydrolyzed, enzyme inactivated, and dried to obtain the yeast protein-polysaccharide complex; the mass ratio of yeast protein to polysaccharide was 100:(2~2.5). The mass concentration of the yeast protein in the aqueous solution is 8-15%; The polysaccharide is one or more of the following: carrageenan, konjac gum, xanthan gum, pectin, seaweed polysaccharide, or glucan; The enzymes used in the enzymatic hydrolysis are one or more combinations of alkaline protease, neutral protease, papain, transglutaminase, bromelain, or flavor protease. The pH of the solution during enzymatic hydrolysis is 6.0~7.

5.

2. The preparation method according to claim 1, characterized in that, The enzymatic hydrolysis temperature is 45~65℃; the enzymatic hydrolysis time is 2~6 h.

3. The preparation method according to claim 2, characterized in that, The amount of enzyme added for enzymatic hydrolysis is 180.15~1200 U / g of the yeast protein.

4. The preparation method according to claim 1, characterized in that, The drying process includes spray drying; the inlet air temperature of the spray drying is not lower than 170°C and the outlet air temperature is not lower than 90°C; the enzyme inactivation conditions include inactivation at 90°C for 30 minutes.

5. The yeast protein and polysaccharide complex prepared by the preparation method according to any one of claims 1 to 4.

6. The yeast protein and polysaccharide complex as described in claim 5, characterized in that, On a dry basis, the yeast protein and polysaccharide complex has a protein content ≥80% and free amino nitrogen <5%; the yeast protein and polysaccharide complex has a foaming power ≥150.00% and an emulsifying property ≥67 m. 2 / g, water holding capacity ≥700%.

7. The application of the yeast protein and polysaccharide complex as described in claim 6 in food processing.

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