Highly soluble defatted egg yolk powder and preparation method thereof

Abstract

This invention provides a highly soluble defatted egg yolk powder and its preparation method, comprising the following steps: adding water to defatted egg yolk powder to obtain a defatted egg yolk powder suspension; ultrasonic pretreatment (U); enzymatic hydrolysis to obtain a modified defatted egg yolk powder solution (E); enzyme inactivation and sterilization (P); ultrasonic post-treatment (U); and sequentially performing vacuum concentration, drying, grinding, and sieving to obtain highly soluble defatted egg yolk powder. By utilizing a combined ultrasonic-enzymatic-sterilization-ultrasonic treatment (UE-PU), the defatted egg yolk powder is modified to possess characteristics such as rapid dissolution, high solubility, and good dispersion stability. Emulsifying activity, emulsifying stability, foaming properties, and foam stability are also significantly improved. Furthermore, the product exhibits good stability in aqueous solution under relatively wide environmental conditions (temperature, pH value, salt concentration). Therefore, the highly soluble defatted egg yolk powder produced using the process of this invention can serve as a high-protein, low-fat, and low-cholesterol nutritional ingredient with broad application prospects in the food industry.

Description

Technical Field

[0001] This invention belongs to the field of egg processing, specifically a highly soluble defatted egg yolk powder and its preparation method. Background Technology

[0002] Defatted egg yolk powder is a byproduct of extracting egg yolk oil or lecithin from egg yolk powder. Produced using green integrated extraction technology, it is low in fat and cholesterol, and high in protein. Therefore, it is more suitable than regular egg yolk powder as a high-protein nutritional ingredient in various formulated foods, such as baked goods like cakes, egg tarts, egg yolk pastries, and bread, as well as nutritional breakfast powders, specialty milk powders, and specialty coffees and beverages.

[0003] However, after defatting, the spatial structure of egg yolk powder changes, exposing hydrophobic groups and increasing surface hydrophobicity. The greater the degree of defatting, the higher the protein content in the defatted egg yolk powder. Simultaneously, the greater the change in protein structure, the worse the solubility and dispersibility of the defatted egg yolk powder, significantly hindering its application in food. Therefore, improving its solubility and dispersibility is a current research focus. Conventional methods include enzymatic hydrolysis, a primary method for protein modification in industry, characterized by low cost and large-scale production. Different enzymes have different cleavage sites on substrates, resulting in varying hydrolytic abilities for the same substrate. A certain degree of enzymatic hydrolysis can significantly reduce the protein particle size in the material, improving protein solubility and imparting a certain emulsifying ability. Tang Shitao et al. used desugared egg yolk liquid as raw material, selected Bacillus subtilis protease, and employed ultrasonic combined with protease hydrolysis technology for treatment, followed by spray drying to produce egg yolk powder, significantly improving the water dispersibility, solubility, and stability coefficient of the egg yolk powder. However, excessive hydrolysis breaks down proteins into small peptides, causing them to lose many of their original properties. Therefore, selecting the appropriate type of protease and controlling the degree of hydrolysis is crucial, directly affecting the product's solubility, emulsifying properties, and other functional characteristics. Furthermore, in actual production, defatted egg yolk powder requires sterilization to improve product safety (meeting national standards for microbiological indicators), but heat sterilization can lead to heat aggregation, resulting in reduced solubility. Summary of the Invention

[0004] Technical Problem to be Solved: To address the aforementioned technical problems, the objective of this invention is to provide a highly soluble defatted egg yolk powder and its preparation method. Using defatted egg yolk powder, a byproduct of egg yolk oil or phospholipid extraction, as raw material, the invention employs a combined ultrasonic-enzymatic hydrolysis-sterilization-ultrasonic treatment (UE-PU) process—essentially a protein modification technique—to improve the solubility and dispersibility of the defatted egg yolk powder. This produces a highly soluble and dispersible defatted egg yolk powder and solves the problem of reduced solubility caused by heat aggregation after the sterilization process in the production flow.

[0005] Technical solution: A method for preparing highly soluble defatted egg yolk powder, comprising the following steps, in parts by weight:

[0006] (1) Grind 1 part of defatted egg yolk powder through a 60-mesh sieve, add 9-15 parts of water and stir evenly to obtain a defatted egg yolk powder suspension.

[0007] (2) Ultrasonic pretreatment: The defatted egg yolk powder suspension was subjected to ultrasonic pretreatment, and the pH value was adjusted to 10-11 before or after ultrasonic pretreatment.

[0008] (3) Enzymatic hydrolysis: Add alkaline protease to carry out enzymatic hydrolysis. After the hydrolysis is completed, adjust the pH to neutral to obtain a modified defatted egg yolk powder solution.

[0009] (4) Enzyme inactivation and sterilization: The modified defatted egg yolk powder solution was heat-treated at a temperature of 70-95℃;

[0010] (5) Ultrasonic post-treatment: Ultrasonic post-treatment is performed on the defatted egg yolk powder solution after enzyme inactivation and sterilization, and then cooled to below 50°C; or step (4) enzyme inactivation and sterilization and ultrasonic post-treatment are performed simultaneously, and then cooled to below 50°C.

[0011] (6) Vacuum concentration, drying, and grinding are performed sequentially to obtain high-solubility defatted egg yolk powder through a sieve of 60 mesh or higher.

[0012] Furthermore, the defatted egg yolk powder in step (1) includes ethanol defatted egg yolk powder and supercritical CO2 defatted egg yolk powder.

[0013] Furthermore, the ultrasonic treatment conditions in step (2) are: ultrasonic power of 100-400W and ultrasonic time of 5-30min. Furthermore, the enzymatic hydrolysis conditions in step (3) are: the amount of alkaline protease added is 0.5-1.5wt.% of the defatted egg yolk powder, the enzymatic hydrolysis temperature is 50-55℃, and the enzymatic hydrolysis time is 0.5-2.5h.

[0014] Furthermore, in step (5), the ultrasonic treatment power is 200-500W and the time is 5-20min.

[0015] Furthermore, in step (6), the vacuum concentration temperature is ≤50℃, and the drying methods include vacuum freeze-drying and spray drying. The highly soluble defatted egg yolk powder prepared by the above method...

[0016] Beneficial effects:

[0017] 1. The highly soluble defatted egg yolk powder produced by the process of this invention has the characteristics of fast dissolution speed, high solubility, and good dispersion stability. Its emulsifying activity, emulsifying stability, foaming property, and foam stability are also greatly improved. Moreover, the product has good stability in aqueous solution under a wide range of environmental conditions (temperature, pH value, salt concentration).

[0018] 2. In this invention, an alkaline protease with an optimal pH value of strongly alkaline is selected for enzymatic hydrolysis. The strongly alkaline pH value increases the intramolecular electrostatic interactions in the polypeptide chain, causing the protein to partially unfold and refold, forming a more hydrophilic morphology in the solution. Combined with ultrasonic treatment, more enzyme binding sites are further exposed. Furthermore, the combined use with an alkaline protease at this pH value significantly improves the protease's hydrolytic ability on defatted egg yolk powder, achieving a multi-dimensional modification effect on defatted egg yolk powder.

[0019] 3. In this invention, ultrasound is used as a pretreatment for enzymatic hydrolysis and a posttreatment for sterilization. Ultrasound loosens the particle structure of egg yolk powder, and the pretreatment with ultrasound can expose more enzyme binding sites, making the enzymatic hydrolysis reaction more complete. The posttreatment with ultrasound can redisperse the clumps of egg yolk powder formed by heat aggregation, thereby giving the defatted egg yolk powder higher solubility.

[0020] 4. The highly soluble defatted egg yolk powder produced by the process of this invention is a nutritional ingredient that is high in protein, low in fat, and low in cholesterol, and has broad application prospects in the food industry. Attached Figure Description

[0021] Figure 1 The images show the appearance of defatted egg yolk powder, where A is homemade ethanol defatted egg yolk powder and B is supercritical CO2 defatted egg yolk powder (commercially available).

[0022] Figure 2 The values ​​represent the solubility of defatted egg yolk powder under different enzymatic hydrolysis conditions in Example 1, where A represents different enzymatic hydrolysis times; B represents different amounts of alkaline protease added; and C represents different enzymatic hydrolysis pH. Different lowercase letters (a, b, c) indicate significant differences (p < 0.05).

[0023] Figure 3 This is a comparison chart of the solubility of the control group, the ethanol defatted egg yolk powder enzymatic hydrolysis treatment group (E), and the ethanol defatted egg yolk powder ultrasonic combined enzymatic hydrolysis treatment group (UE) in Example 2.

[0024] Figure 4 This is a comparison chart of the solubility of supercritical CO2 defatted egg yolk powder (control group), supercritical CO2 defatted egg yolk powder enzymatic hydrolysis group (E), and ultrasonic combined enzymatic hydrolysis group (UE) in Example 3.

[0025] Figure 5This is a comparison chart of the solubility of egg yolk powder in the E control group, UE control group, EP control group, UE-P control group, E-PU control group and UE-PU group in Example 4;

[0026] Figure 6 The graph shows the bactericidal effect test results of the E control group, UE control group, and UE-P group in Example 4;

[0027] Figure 7 The graph shows the enzyme inactivation effect test results for groups E, UE-P, UE-PU, and UE-PU extension group in Example 4;

[0028] Figure 8 The particle size variation diagram shows the ethanol defatted egg yolk powder prepared in Example 4 and the egg yolk powder at different modification stages in Example 4.

[0029] Figure 9 The graph shows the solubility variation of the UE-PU highly soluble defatted egg yolk powder prepared in Example 4 under different environments, where A represents different NaCl concentrations, B represents different pH conditions, and C represents different temperature conditions.

[0030] Figure 10 Photographs showing the changes in dispersion stability of UE-PU highly soluble defatted egg yolk powder prepared in Example 4 under different environments, where A represents different NaCl concentrations, B represents different pH conditions, and C represents different temperature conditions.

[0031] Figure 11 This is a comparison chart of the solubility of egg yolk powder in the E control group, UE control group, EP group, UE-P group, E-PU group and UE-PU group in Example 5;

[0032] Figure 12 Particle size variation diagram of supercritical CO2 defatted egg yolk powder (commercially available) and egg yolk powder at different modification stages;

[0033] Figure 13 The graph shows the solubility comparison of supercritical CO2 defatted egg yolk powder treated with combined ultrasonication and enzymatic hydrolysis in Example 6 and Comparative Example 1 under different sterilization process temperatures and after sterilization + ultrasonication treatment. In Example 6, A represents enzymatic hydrolysis using the alkaline protease of the present invention, and B represents enzymatic hydrolysis using subtilisin in Comparative Example 1. Detailed Implementation

[0034] This invention proposes a highly soluble defatted egg yolk powder and its preparation method. To make the objectives, technical solutions, and effects of this invention clearer and more explicit, the following will provide a more detailed description of the invention with reference to specific embodiments. It should be understood that the specific embodiments described herein are only for explaining the invention and are not intended to limit the invention.

[0035] Determination of nutritional content of defatted egg yolk powder:

[0036] The preparation method of ethanol defatted egg yolk powder is as follows:

[0037] S1: Weigh 100g of commercially available egg yolk powder into a 1L beaker, add 500mL of anhydrous ethanol, and stir until the egg yolk powder is fully dissolved;

[0038] S2: Stir at 50°C for 1.5 hours using a magnetic stirrer, then filter to separate the egg yolk powder and ethanol;

[0039] S3: Mix the egg yolk powder obtained in step S2 with 500 mL of anhydrous ethanol until homogeneous, and repeat step S2.

[0040] S4: Place the egg yolk powder obtained in step S3 in a 50℃ oven and dry it for 5 hours. Stir the sample every 1 hour to obtain ethanol defatted egg yolk powder, and store it in a desiccator at room temperature.

[0041] The composition of commercially available egg yolk powder, ethanol-defatted egg yolk powder, and supercritical CO2-defatted egg yolk powder was analyzed: Determination of fat content: Referring to GB 5009.6-2016, the fat content of egg yolk powder and defatted egg yolk powder was determined using Soxhlet extraction: 2g of sample was weighed, accurate to 0.001g. The receiving flask was dried to constant weight. The sample was placed in a filter paper tube and then placed in a Soxhlet extractor. Extraction was performed using petroleum ether with a boiling range of 30-60℃, continuously refluxed in a water bath for 10 hours. After extraction, the remaining solvent was evaporated in a water bath, and the receiving flask was dried to constant weight. The fat content of the sample was calculated.

[0042] Protein content determination: The protein content of defatted egg yolk powder was determined using an automatic Kjeldahl nitrogen analyzer, in accordance with GB 5009.5-2016.

[0043] Moisture content determination: Referring to GB 5009.3-2016, the moisture content of defatted egg yolk powder and egg yolk powder was determined by direct drying method;

[0044] Ash content determination: The ash content of egg yolk powder and defatted egg yolk powder was determined according to GB 5009.4-2016;

[0045] Cholesterol content determination: Refer to GB5009.128-2016, and use colorimetric method to determine the cholesterol content of defatted egg yolk powder and egg yolk powder.

[0046] The results are shown in Table 1 below:

[0047] Table 1

[0048]

[0049] Experimental Results and Analysis:

[0050] Compared to egg yolk powder, defatted egg yolk powder with ethanol has a reduced fat content of 11.4%, an increased protein content of 53.37%, and a reduced cholesterol content of 288.6 mg / 100g. Supercritical CO2 defatted egg yolk powder has a fat content of 3%, a protein content of 75%, and a cholesterol content of 76.8 mg / 100g, with cholesterol only 4.3% of that of regular egg yolk powder. Defatted egg yolk powder, as a high-protein, low-fat, and low-cholesterol nutritional protein powder, undoubtedly has broad application prospects in the food industry if its poor solubility and dispersibility issues are resolved. The defatted egg yolk powder and supercritical CO2 defatted egg yolk powder (commercially available) prepared by this invention have the following appearance: Figure 1 As shown.

[0051] Example 1: Optimization of Enzymatic Hydrolysis Process Conditions

[0052] 1. Preparation methods of enzymatically degreased egg yolk powder under different enzymatic hydrolysis times:

[0053] (1) Grind the defatted egg yolk powder with ethanol through a 60-mesh sieve and mix it with water at a ratio of 1:9 to obtain a defatted egg yolk powder suspension.

[0054] (2) Enzymatic hydrolysis: The pH of the defatted egg yolk powder suspension was adjusted to 11 with 2 mol / L sodium hydroxide solution. Alkaline protease was added at a concentration of 1.0 wt.% of the defatted egg yolk powder. The hydrolysis temperature was 50℃ and the hydrolysis reaction times were 0.5 h, 1.0 h, 1.5 h, 2.0 h, and 2.5 h, respectively. After the hydrolysis was completed, the pH was adjusted to 7.0 with 2 mol / L hydrochloric acid to obtain the modified defatted egg yolk powder solution.

[0055] (3) The defatted egg yolk powder solution obtained in step (2) is subjected to vacuum freeze concentration, drying, and grinding through a sieve of 60 mesh or higher to obtain enzymatically defatted egg yolk powder.

[0056] 2. Preparation methods of enzymatically degreased egg yolk powder under different amounts of alkaline protease:

[0057] (1) Grind the defatted egg yolk powder with ethanol through a 60-mesh sieve and mix it with water at a ratio of 1:9 to obtain a defatted egg yolk powder suspension.

[0058] (2) Enzymatic hydrolysis: The pH of the defatted egg yolk powder suspension was adjusted to 11 with 2 mol / L sodium hydroxide solution. Alkaline protease was added at concentrations of 0.5 wt.%, 0.75 wt.%, 1.0 wt.%, 1.25 wt.%, and 1.5 wt.% of the defatted egg yolk powder, respectively. The hydrolysis temperature was 50℃ and the hydrolysis reaction time was 1.5 h. After the hydrolysis was completed, the pH was adjusted to 7.0 with 2 mol / L hydrochloric acid to obtain the modified defatted egg yolk powder solution.

[0059] (3) The defatted egg yolk powder solution obtained in step (2) is vacuum concentrated, dried and ground through a sieve of 60 mesh or higher to obtain enzymatically defatted egg yolk powder.

[0060] 3. Methods for enzymatically degreasing egg yolk powder under different pH conditions:

[0061] (1) Grind the defatted egg yolk powder with ethanol through a 60-mesh sieve and mix it with water at a ratio of 1:9 to obtain a defatted egg yolk powder suspension.

[0062] (2) Enzymatic hydrolysis: The pH of the defatted egg yolk powder suspension was adjusted to 10, 10.25, 10.5, 10.75 and 11.0 respectively with 2 mol / L sodium hydroxide solution. Alkaline protease was added at a concentration of 1.0 wt.% of the defatted egg yolk powder. The hydrolysis temperature was 50℃ and the hydrolysis reaction time was 1.5 h. After the hydrolysis was completed, the pH was adjusted to 7.0 with 2 mol / L hydrochloric acid to obtain the modified defatted egg yolk powder solution.

[0063] (3) The defatted egg yolk powder solution obtained in step (2) is subjected to vacuum freeze concentration, drying, and grinding through a sieve of 60 mesh or higher to obtain enzymatically defatted egg yolk powder.

[0064] Determining the solubility of the enzymatically degreased egg yolk powder in Example 1:

[0065] Weigh 1g of sample at room temperature and dissolve it in 30mL of water in a 50mL centrifuge tube. Vortex to mix, then shake on a shaker for 10 minutes to ensure complete dissolution. Place the centrifuge tube in a centrifuge and centrifuge at 5000 rpm for 15 minutes to precipitate the insoluble matter. Discard the supernatant, rinse the precipitate with a small amount of water into a pre-weighed evaporating dish, and dry in an oven at 105℃ until constant weight. The solubility is calculated using the following formula:

[0066] Solubility (g / 100g) = (m1 - m2) × 100 / m

[0067] Where m, m1, and m2 represent the mass of the sample before drying, the mass of the evaporating dish and the sample after drying, and the mass of the evaporating dish before drying, respectively.

[0068] The results are shown in Table 2 below:

[0069] Table 2

[0070]

[0071] Experimental Results and Analysis:

[0072] As shown in Table 2 and Figure 2 As shown, where, as Figure 2As shown in Figure A, the single-factor optimization experiment of enzymatic hydrolysis time showed that after adding enzyme to the defatted egg yolk powder suspension to initiate the reaction, the solubility of the defatted egg yolk powder increased with increasing hydrolysis time, but the rate of increase slowed down with further time. The main component of defatted egg yolk powder is egg yolk protein; enzymatic hydrolysis can cleave protein peptide bonds, reducing the protein molecular weight, which increases solubility. Between 1.5h and 2.5h of enzymatic hydrolysis, the change in the solubility of defatted egg yolk powder was not significant; therefore, 1.5h was selected as the subsequent hydrolysis time. Figure 2 As shown in Figure B, the single-factor optimization experiment of alkaline protease content showed that the solubility of defatted egg yolk powder first increased and then decreased with increasing alkaline protease content, reaching its highest value at an enzyme content of 1.0 wt.%, with no significant difference between 1.0-1.5 wt.%. Figure 2 As shown in Figure C, the single-factor optimization experiment of enzymatic hydrolysis pH indicates that the solubility of defatted egg yolk powder increases with increasing enzymatic hydrolysis pH, and this trend is more pronounced at higher pH levels. Previous experiments revealed that the solubility of defatted egg yolk powder is extremely sensitive to pH changes. Adjusting the pH of the defatted egg yolk powder to a strongly alkaline state and then back to 7.0 resulted in an increase in solubility. Under alkaline conditions, pH shift and the enzymatic hydrolysis process synergistically increase solubility. pH 11 is the optimal pH value for enzymatic hydrolysis. Based on the current optimal solubility, pH 11 is selected as the optimal enzymatic hydrolysis condition for subsequent experiments.

[0073] Example 2: Optimization of Ultrasonic Conditions

[0074] 1. Ultrasonic + enzymatic hydrolysis method for defatted egg yolk powder under different ultrasonic powers:

[0075] (1) Grind the defatted egg yolk powder with ethanol through a 60-mesh sieve, mix it with water at a ratio of 1:9 to obtain a defatted egg yolk powder suspension, and adjust the pH of the defatted egg yolk powder suspension to 11 with 2 mol / L sodium hydroxide solution.

[0076] (2) Ultrasonic pretreatment: The defatted egg yolk powder suspension was ultrasonically treated with a reaction frequency of 20kHz, an ultrasonic mode of 3 seconds on time and 3 seconds off time, and ultrasonic power of 0W, 100W, 200W, 300W and 400W respectively, and ultrasonic treatment for 10min.

[0077] (3) Enzymatic hydrolysis: Add alkaline protease at a concentration of 1.0 wt.% of defatted egg yolk powder, at a hydrolysis temperature of 50°C, for a hydrolysis reaction time of 1.5 h. After hydrolysis, adjust the pH to 7.0 with 2 mol / L hydrochloric acid to obtain a modified defatted egg yolk powder solution.

[0078] (4) The defatted egg yolk powder solution obtained in step (3) is subjected to vacuum freeze concentration, drying, and grinding through a sieve of 60 mesh or higher to obtain highly soluble defatted egg yolk powder.

[0079] 2. Ultrasonic + enzymatic hydrolysis method for defatted egg yolk powder at different ultrasonic times:

[0080] (1) Grind the defatted egg yolk powder with ethanol through a 60-mesh sieve, mix it with water at a ratio of 1:9 to obtain a defatted egg yolk powder suspension, and adjust the pH of the defatted egg yolk powder suspension to 11 with 2 mol / L sodium hydroxide solution.

[0081] (2) Ultrasonic pretreatment: The defatted egg yolk powder suspension was ultrasonically treated with a reaction frequency of 20kHz, an ultrasonic mode of 3 seconds on and 3 seconds off, an ultrasonic power of 100W, and ultrasonic treatment times of 0min, 5min, 10min, 20min and 30min respectively.

[0082] (3) Enzymatic hydrolysis: Add alkaline protease at a concentration of 1.0 wt.% of defatted egg yolk powder, at a hydrolysis temperature of 50°C, for a hydrolysis reaction time of 1.5 h. After hydrolysis, adjust the pH to 7.0 with 2 mol / L hydrochloric acid to obtain a modified defatted egg yolk powder solution.

[0083] (4) The defatted egg yolk powder solution obtained in step (3) is subjected to vacuum freeze concentration, drying, and grinding through a sieve of 60 mesh or higher to obtain highly soluble defatted egg yolk powder.

[0084] Performance Test 1:

[0085] The solubility of the highly soluble defatted egg yolk powders in Examples 5 and 6 was determined, and the results are shown in Table 3 below:

[0086] Table 3

[0087]

[0088] Experimental Results and Analysis:

[0089] As shown in Table 3, solubility increases with increasing ultrasonic power, and initially increases then decreases with increasing ultrasonic time. The optimal ultrasonic power is 300W, and the optimal ultrasonic treatment time is 20 minutes. Ultrasonic pretreatment can effectively improve the solubility of defatted egg yolk powder. After determining the optimal enzymatic modification conditions, ultrasonic treatment of the defatted egg yolk powder suspension before enzymatic hydrolysis can further improve the solubility of defatted egg yolk powder.

[0090] Performance Test 2:

[0091] Comparison of the solubility effects of enzymatic hydrolysis and ultrasonic combined enzymatic hydrolysis on defatted egg yolk powder with ethanol:

[0092] Control group: The self-made ethanol-blended egg yolk powder mentioned above;

[0093] Ethanol defatted egg yolk powder enzymatic hydrolysis group (E): Same as Example 1.3, except that the enzymatic hydrolysis pH in step (2) is 11;

[0094] Ethanol defatted egg yolk powder ultrasonic combined with enzymatic hydrolysis group (UE): Same as Example 2.2, wherein the ultrasonic power in step (2) is 300W and the ultrasonic time is 20min.

[0095] The solubility of the three groups of treated defatted egg yolk powder was measured, and the results are as follows: Figure 3 As shown.

[0096] Experimental Results and Analysis:

[0097] according to Figure 3 It was found that after ultrasonic pretreatment, the solubility of the ultrasonic combined with enzymatic hydrolysis (UE) group was significantly higher than that of the enzymatic hydrolysis (E) group. This is because the ultrasonic cavitation effect makes the protein structure in the defatted egg yolk powder loose, exposing some hidden enzyme cleavage sites, making the enzymatic cleavage more complete, thereby reducing the average particle size of the defatted egg yolk powder and increasing the solubility.

[0098] Example 3: Comparison of the effects of enzymatic hydrolysis and ultrasonic combined enzymatic hydrolysis on supercritical CO2 defatted egg yolk powder:

[0099] Control group: Supercritical CO2 defatted egg yolk powder (commercially available);

[0100] Enzymatic hydrolysis (E): Same as in Example 1.3, except that in step (1), the self-made ethanol defatted egg yolk powder is replaced with supercritical CO2 defatted egg yolk powder, and in step (2), the enzymatic hydrolysis pH is 11;

[0101] Ultrasonic combined enzymatic hydrolysis (UE): Same as in Example 2.2, except that in step (1), supercritical CO2 defatted egg yolk powder is used instead of the ethanol defatted egg yolk powder prepared in Example 2, and in step (2), the ultrasonic power is 300W and the ultrasonic time is 20min.

[0102] The solubility of the modified defatted egg yolk powder in Example 3 was determined, and the results are as follows: Figure 4 .

[0103] Experimental Results and Analysis:

[0104] like Figure 4 As shown, ultrasonic combined with enzymatic hydrolysis (UE) is superior to enzymatic hydrolysis (E), but due to the stronger hydrophobicity of supercritical CO2 defatted egg yolk powder, its solubility is significantly lower than that of ethanol defatted egg yolk powder.

[0105] Example 4

[0106] Preparation method of UE-PU group highly soluble defatted egg yolk powder:

[0107] (1) Grind the defatted egg yolk powder with ethanol through a 60-mesh sieve and mix it with water at a ratio of 1:10 to obtain a defatted egg yolk powder suspension.

[0108] (2) Ultrasonic pretreatment: The defatted egg yolk powder suspension was ultrasonically treated with a reaction frequency of 20kHz, an ultrasonic mode of 3 seconds on and 3 seconds off, an ultrasonic power of 300W, and an ultrasonic treatment time of 20min.

[0109] (3) Enzymatic hydrolysis: The pH of the defatted egg yolk powder suspension was adjusted to 11 with 2 mol / L sodium hydroxide solution. Alkaline protease was added at a concentration of 1.0 wt.% of the defatted egg yolk powder. The hydrolysis temperature was 50℃ and the hydrolysis reaction time was 1.5 h. After the hydrolysis was completed, the pH was adjusted to 7.0 with 2 mol / L hydrochloric acid to obtain the modified defatted egg yolk powder solution.

[0110] (4) Enzyme inactivation and sterilization: The modified defatted egg yolk powder solution was subjected to enzyme inactivation and sterilization at a temperature of 72℃ for 15 minutes;

[0111] (5) Post-ultrasound processing: The ultrasonic probe is immersed 1.5 cm below the liquid surface, the ultrasonic power is 300W, the ultrasonic time is 15 min, and it is allowed to cool naturally to room temperature.

[0112] (6) The defatted egg yolk powder solution obtained in step (5) is subjected to vacuum concentration, freeze drying and grinding through a sieve of 60 mesh or higher to obtain highly soluble defatted egg yolk powder.

[0113] Different control groups were set up, as follows:

[0114] E control group: Unlike the UE-PU group, steps (2), (4), and (5) are omitted;

[0115] UE control group: Unlike the UE-PU group, steps (4) and (5) are omitted;

[0116] EP control group: Unlike the UE-PU group, steps (2) and (5) are omitted;

[0117] UE-P control group: Unlike the UE-PU group, step (5) is omitted;

[0118] E-PU control group: Unlike the UE-PU group, step (2) is omitted;

[0119] Performance Test 1: Solubility

[0120] The solubility of defatted egg yolk powder in Example 4 and different control groups was determined, and the results are as follows: Figure 5 As shown.

[0121] Experimental Results and Analysis:

[0122] like Figure 5 As shown, the solubility of ethanol-fattened egg yolk powder decreased significantly after the sterilization process (-P). Sterilization led to a decrease in solubility, and the solubilities of the ultrasonic combined enzymatic hydrolysis (UE) and enzymatic hydrolysis (E) groups became similar. This may be due to heat accumulation caused by heating. To solve the problem of decreased solubility caused by sterilization, an ultrasonic post-treatment step was introduced to further disperse the defatted egg yolk powder. After sterilization, a second ultrasonic post-treatment (-PU) was performed, and the solubility was significantly improved. Ultrasonic post-treatment plays an important role in this modification process. Therefore, the optimal process steps for modified ethanol-fattened egg yolk powder were finally determined to be: ultrasonic pre-treatment - enzymatic hydrolysis - enzyme inactivation and sterilization - ultrasonic post-treatment, i.e., UE-PU.

[0123] Performance Test 2: Sterilization Effect

[0124] The following groups are set up respectively:

[0125] E control group: Unlike the UE-PU group, steps (2), (4), and (5) are omitted;

[0126] UE control group: Unlike the UE-PU group, steps (4) and (5) are omitted;

[0127] UE-P group: Unlike UE-PU group, step (5) is omitted.

[0128] The total bacterial count of the three groups of treated defatted egg yolk powders was determined, and the results are as follows: Figure 6 As shown.

[0129] Experimental Results and Analysis:

[0130] according to Figure 6 After sterilization at 75℃ for 15 minutes (UE-P group), the total bacterial count in the modified defatted egg yolk powder was 8.15 x 10⁻⁶. 2 CFU / g was lower than 1.08 x 10⁻⁶ when the sample was not sterilized and only subjected to sonication (UE control group). 4 CFU / g, far below the national standard detection limit of 5.0x10. 4 CFU / g.

[0131] Performance Test 3: Enzyme Inactivation Effect

[0132] The following groups are set up respectively:

[0133] Group E: Unlike the UE-PU group, steps (2), (4), and (5) are omitted;

[0134] UE-P group: Unlike UE-PU group, step (5) is omitted;

[0135] UE-PU group;

[0136] UE-PU extension group: Based on the UE-PU group, continue the reaction for 5 hours.

[0137] The degree of hydrolysis of the four groups of treated defatted egg yolk powders was measured. The change in the degree of hydrolysis reflects the enzyme inactivation effect. The results are as follows: Figure 7 As shown.

[0138] Experimental Results and Analysis:

[0139] according to Figure 7 The degree of hydrolysis of defatted egg yolk powder after heat treatment at 75℃ for 15 min (UE-P group) with different modifications was compared. After enzymatic hydrolysis of defatted egg yolk powder, enzyme inactivation and sterilization were performed, resulting in a degree of hydrolysis of 2.37%. Ultrasonication combined with enzymatic hydrolysis significantly improved the degree of hydrolysis of defatted egg yolk powder. In the UE-PU extended group (ultrasonication + enzymatic hydrolysis + sterilization + ultrasonication followed by 5 h of further reaction), the degree of hydrolysis did not change significantly after two ultrasonic extensions of the enzymatic hydrolysis time, proving that the sterilization step had inactivated the enzyme.

[0140] Performance Test 4: Emulsification and emulsion stability, foaming and foaming stability; particle size distribution and ZETA potential were used to determine the functional properties of the self-made ethanol defatted egg yolk powder, the UE-PU group of highly soluble defatted egg yolk powder prepared in Example 4, and the E control group, UE control group, EP control group, and UE-P control group.

[0141] Emulsifying activity (EAI) and emulsifying stability (ESI) determination: Prepare a 1% concentration sample solution (10 mg / mL). Mix 15 mL of the solution with 5 mL of oil using a homogenizer at 13000 rpm for 2 min. Take 50 μL of the emulsion and mix it with 5 mL of 0.1% SDS. Measure the absorbance (A0) at 500 nm. Incubate at room temperature for 10 min and measure the absorbance again (A1). EAI and ESI are calculated using the following formula:

[0142]

[0143]

[0144] In the formula, DF, C, and θ represent the dilution coefficient 101, protein concentration 5.6, and oil volume fraction 0.25, respectively; ΔA is the difference between A0 and A1; and ΔT = 10 min.

[0145] Foaming property (FA) and foam stability (FS) determination: Prepare a 1% concentration sample solution (10 mg / mL). Mix 15 mL of the solution using a dispersion homogenizer at 13000 rpm for 2 min. Immediately pour the foam into a 50 mL graduated cylinder. Foaming property is characterized by comparing the foam volume at 0 min (V0) with the initial liquid volume of the sample (15 mL). Foam stability is determined by comparing the foam volume at 60 min with the initial foam volume. FA and FS are calculated using the following formulas:

[0146]

[0147]

[0148] In the formula, V0 is the foam volume at 0 min; V 60 The foam volume at 60 minutes.

[0149] Particle size distribution and ZETA potential determination: Prepare a 0.2% concentration sample solution (2 mg / mL); use a laser particle size analyzer and a Marven Zeta ZEN3700 electrochemical workstation to analyze particle size distribution, average particle size, and ZETA potential.

[0150] The results are shown in Tables 4 and 5 below:

[0151] Table 4

[0152]

[0153] Table 5

[0154] type D[4,3] Zeta potential Comparison The above-mentioned homemade ethanol defatted egg yolk powder 57.56±0.34 -6.25±0.45 E Enzymatic hydrolysis 50.21±2.43 -7.35±0.46 UE Ultrasonic pretreatment-enzymatic hydrolysis 34.94±1.45 -7.38±0.37 UE-P Ultrasonic pretreatment - enzymatic hydrolysis - sterilization 40.19±3.00 -6.57±0.13 UE-PU Ultrasonic pretreatment - enzymatic hydrolysis - sterilization - ultrasonic posttreatment 22.25±1.19 -8.18±0.13

[0155] Experimental Results and Analysis:

[0156] As shown in Tables 4 and 5, the functional properties of ethanol-treated defatted egg yolk powder were significantly improved after ultrasonic combined with enzymatic hydrolysis modification, with marked increases in emulsifying activity, emulsifying stability, foaming properties, and foam stability. All indicators in the ultrasonic combined with enzymatic hydrolysis (UE) group were higher than those in the ultrasonic combined with enzymatic hydrolysis sterilization group (UE-P). The enzyme inactivation and sterilization steps (P) impaired the functional properties of the defatted egg yolk powder to some extent. Meanwhile, all indicators in the UE-PU group were higher than those in the UE-P group, indicating that ultrasonic post-treatment is beneficial for improving the functional properties of defatted egg yolk powder. Particle size distribution changes are shown in [Table 4]. Figure 8 .

[0157] Performance Test 5: Environmental Stability Test of Highly Soluble Defatted Egg Yolk Powder

[0158] The solubility and dispersion stability of the highly soluble defatted egg yolk powder in the UE-PU group prepared in Example 4 were determined under different application conditions:

[0159] (1) Under different concentrations of NaCl solution conditions

[0160] The highly soluble defatted egg yolk powder prepared in the UE-PU group of Example 4 was dissolved in 0%, 0.5%, 1%, 2%, and 4% NaCl solutions, respectively. The pH was adjusted to 7, and the solution was kept at 20°C and shaken for 10 min. The solubility was measured, and the solution was allowed to stand at room temperature for 24 h to observe the dispersion stability and record the results.

[0161] (2) Under different pH conditions

[0162] The highly soluble defatted egg yolk powder prepared in the UE-PU group in Example 4 was dissolved in deionized water, and the pH was adjusted to 4, 5, 6, 7 and 8 respectively. The mixture was kept at 20°C and shaken for 10 min to measure the solubility. The mixture was then left to stand at room temperature for 24 h to observe the dispersion stability and to take pictures and record the results.

[0163] (3) Conditions under different temperatures

[0164] The highly soluble defatted egg yolk powder prepared in the UE-PU group in Example 4 was dissolved in deionized water, the pH was adjusted to 7, and the mixture was kept at 20°C and shaken for 10 min to determine the solubility. The mixture was then allowed to stand at 4°C, 20°C, 40°C, 60°C, and 65°C for 24 h to observe the dispersion stability and to record the results by taking pictures.

[0165] See results Figure 9 , 11 The solubility of highly soluble defatted egg yolk powder gradually decreases in high-salt environments. When the salt content is below 0.5%, the solubility is greater than 90g / 100g, and when the salt content is below 2%, the solubility is greater than 80g / 100g. When the pH is greater than 7, the solubility is greater than 90g / 100g. At pH 4-6, the solubility is lower. At pH 6-7, stratification occurs after 24 hours of standing. The solubility of highly soluble defatted egg yolk powder remains above 90g / 100g within the temperature range of 4-60℃, exhibiting good temperature applicability and dispersion stability. When the temperature is above 65℃, the solubility decreases significantly, and stratification occurs after 24 hours of standing.

[0166] Example 5

[0167] This embodiment uses supercritical CO2 defatted egg yolk powder instead of the self-made ethanol defatted egg yolk powder. The modification preparation method is completely the same as in Example 4, and it is also divided into E control group, UE control group, EP group, UE-P group, E-PU group and UE-PU group.

[0168] Performance Test 1: Solubility

[0169] The solubility of the highly soluble defatted egg yolk powder prepared in Example 5 was determined, and the results are as follows: Figure 11 As shown.

[0170] Experimental Results and Analysis:

[0171] Similar to Example 4, the solubility of supercritical CO2 defatted egg yolk powder modified by ultrasonic combined enzymatic hydrolysis (UE) was better than that modified by single enzymatic hydrolysis (E); after the sterilization process (-P), the solubility decreased significantly. The solubility change pattern of supercritical CO2 defatted egg yolk powder during the modification process was consistent with that of ethanol defatted egg yolk powder; the ultrasonic post-treatment after sterilization is also applicable to the modification of both ethanol defatted egg yolk powder and supercritical CO2 defatted egg yolk powder.

[0172] Performance Test 2: Emulsification and emulsion stability, foaming and foaming stability; particle size distribution and ZETA potential were used to determine the functional properties of supercritical CO2 defatted egg yolk powder (commercially available), and the UE, UE-P, and UE-PU groups of highly soluble defatted egg yolk powder prepared in Example 5. The results are shown in Tables 6 and 7 below:

[0173] Table 6

[0174]

[0175] Table 7

[0176]

[0177] Experimental Results and Analysis:

[0178] As shown in Tables 6 and 7, after ultrasonic combined with enzymatic modification, the emulsifying activity (EAI), emulsifying stability (ESI), and foaming properties (FA) of the critical CO2 defatted egg yolk powder were basically consistent with those of the ethanol-modified defatted egg yolk powder. The changes in particle size distribution are shown in Tables 6 and 7. Figure 12 .

[0179] Example 6

[0180] Using the defatted egg yolk powder raw material from Example 5 and the ultrasonic combined enzymatic hydrolysis process (UE), the enzymes were inactivated and sterilized by water baths at 72°C, 85°C, and 100°C, respectively, with a heat treatment time of 15 minutes for each. The conditions for ultrasonic post-treatment after sterilization were completely consistent with those for ultrasonic post-treatment in Examples 4 and 5.

[0181] Comparative Example 1

[0182] The difference between this comparative example and Example 6 is that the enzymatic hydrolysis conditions in step (3) are as follows: based on defatted egg yolk powder suspension, the amount of subtilisin added is 40u / g, the enzymatic hydrolysis temperature is 45℃, and the enzymatic hydrolysis time is 4h.

[0183] The solubility of the modified defatted egg yolk powder of Example 6 and Comparative Example 1 after sterilization and after sterilization + ultrasonic treatment was determined, and the results are as follows: Figure 13 As shown.

[0184] Experimental Results and Analysis:

[0185] Example 6 shows that the solubility of modified defatted egg yolk powder increases with increasing sterilization temperature; after ultrasonic post-treatment, the solubility of modified defatted egg yolk powder is significantly improved, such as... Figure 13 As shown in Figure A.

[0186] Comparative Example 1 describes the enzymatic hydrolysis of supercritical CO2 defatted egg yolk powder using the conditions described by Tang Shitao et al. from Huazhong Agricultural University in several publications. Figure 13 As shown in B, in the comparative example, the solubility of defatted egg yolk powder modified with subtilisin after heat sterilization and sterilization + ultrasonic treatment was far lower than that of the alkaline protease modified in this study. Figure 13 A). After sterilization at 72℃ and ultrasonic treatment, the solubility of the modified product (alkaline protease) according to this patent application is 40.92 g / 100 g, while the comparative example is 20.67 g / 100 g.

Claims

1. A method for preparing highly soluble defatted egg yolk powder, characterized in that, Includes the following steps, in parts by weight: (1) Grind 1 part of defatted egg yolk powder through a 60-mesh sieve, add 9-15 parts of water and stir evenly to obtain a defatted egg yolk powder suspension. (2) Ultrasonic pretreatment: The defatted egg yolk powder suspension was subjected to ultrasonic pretreatment, and the pH value was adjusted to 10-11 before or after ultrasonic pretreatment. (3) Enzymatic hydrolysis: Add alkaline protease to carry out enzymatic hydrolysis. After the hydrolysis is completed, adjust the pH to neutral to obtain a modified defatted egg yolk powder solution. (4) Enzyme inactivation and sterilization: The modified defatted egg yolk powder solution was heat-treated at a temperature of 70-95℃; (5) Ultrasonic post-treatment: Ultrasonic post-treatment is performed on the defatted egg yolk powder solution after enzyme inactivation and sterilization, and then cooled to below 50°C; or step (4) enzyme inactivation and sterilization and ultrasonic post-treatment are performed simultaneously, and then cooled to below 50°C. (6) Vacuum concentration, drying, and grinding are performed sequentially to obtain high-solubility defatted egg yolk powder through a sieve of 60 mesh or higher.

2. The method for preparing a highly soluble defatted egg yolk powder according to claim 1, characterized in that, The defatted egg yolk powder in step (1) includes ethanol defatted egg yolk powder and supercritical CO2 defatted egg yolk powder.

3. The method for preparing a highly soluble defatted egg yolk powder according to claim 1, characterized in that, The ultrasonic treatment conditions in step (2) are: ultrasonic power of 100-400W and ultrasonic time of 5-30min.

4. The method for preparing a highly soluble defatted egg yolk powder according to claim 1, characterized in that, The enzymatic hydrolysis conditions in step (3) are as follows: the amount of alkaline protease added is 0.5-1.5 wt.% of the defatted egg yolk powder, the hydrolysis temperature is 50-55℃, and the hydrolysis time is 0.5-2.5h.

5. The method for preparing a highly soluble defatted egg yolk powder according to claim 1, characterized in that, In step (5), the ultrasonic treatment power is 200-500W and the time is 5-20min.

6. The method for preparing a highly soluble defatted egg yolk powder according to claim 1, characterized in that, In step (6), the temperature of vacuum concentration is ≤50℃, and the drying methods include vacuum freeze drying and spray drying.

7. The highly soluble defatted egg yolk powder prepared by the preparation method according to any one of claims 1-6.

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