Method for enhancing the stability of protein peptide emulsion by synergistic modification of glycosylation-calcium bridge and preparation of nutritional emulsion thereof

Abstract

The application belongs to the technical field of nutritional formula food processing, and discloses a method for enhancing the stability of protein peptide emulsion by glycosylation-calcium bridge synergistic modification, that is, glycosylation reaction of protein peptide and carbohydrate, then adding calcium chloride before emulsification, and inducing moderate cross-linking of glycosylated protein peptide by calcium bridge under room temperature or heating condition, thereby regulating the self-assembly behavior of protein peptide and the emulsion formation process, so as to improve the thermal stability and ion tolerance of the protein peptide emulsion. The application also discloses a protein peptide nutritional emulsion prepared based on the synergistic modification technology, wherein 100 mL of the nutritional emulsion comprises 3-8 g of protein peptide, 2-12 g of oil, 8-30 g of carbohydrate, 0.25-2 mmol of calcium chloride (i.e. 10-80 mg of calcium element), 0.05-0.5 g of compound vitamin and 0.02-1 g of compound mineral. The obtained nutritional emulsion has significantly better long-term stability after high-temperature sterilization. The method can be widely applied to the manufacturing of high-stability protein base and full-nutrition emulsion and other special-diet emulsion foods; and the protein peptide nutritional emulsion can meet the high demand of protein peptide absorption of people.

Description

Technical Field

[0001] This invention belongs to the field of nutritional formula food processing technology, specifically relating to a method for enhancing the stability of protein peptide emulsions through glycosylation-calcium bridging synergistic modification and the preparation of nutritional emulsions. Background Technology

[0002] In recent years, protein nutritional emulsions have gained popularity due to their ready-to-eat and convenient characteristics, and are widely used in many health-related fields such as clinical nutrition, elderly health, infant nutrition, and sports and fitness, resulting in huge market demand. Among them, hydrolyzed proteins rich in short peptides have greater advantages in terms of digestion and absorption, bioactivity, and reduced allergenicity, which can meet the higher protein absorption needs of these populations. Therefore, expanding the application of protein peptides in nutritional emulsions has profound significance.

[0003] However, high-temperature sterilization and mineral metal ion-mediated protein flocculation are key factors that compromise the stability of nutritional emulsions. This is especially true for divalent metal ions (such as Ca). 2+ Mg 2+ Salts easily induce high cross-linking and aggregation of proteins through salt bridges (such as -COO-Ca-OOC-), forming large aggregates or even gels. This has long played a negative role in disrupting the stability of protein nutritional emulsions. Furthermore, Ca... 2+ The destructive effect is strongest. Although the industry avoids the destructive effects of Ca by adding calcium-bound salts (such as calcium phosphate, nano-calcium carbonate, calcium citrate, etc.). 2+ The adverse effects of other metal ions (such as Na) + K + Mg 2+ and Fe 3+ The destructive effects of certain substances (such as calcium chelates) are still unavoidable. Furthermore, their low calcium absorption rate easily leads to kidney stones. Conversely, calcium chelate peptides, as a novel calcium supplement, can improve calcium absorption. Moreover, the industry typically adds specific minerals (such as calcium- and iron-fortified products) after obtaining the primary emulsion, and then performs high-temperature sterilization. This process may exacerbate oil droplet flocculation through protein-ion interactions, thus negatively impacting emulsion stability. Therefore, improvements are necessary.

[0004] Glycosylation modification can improve the thermal stability and ionic tolerance of protein emulsions, thereby effectively resisting calcium ions. 2+ The flocculation and aggregation induced by 121°C heat sterilization are commonly used in the protein emulsion industry, but the long-term stability of nutritional emulsions prepared by single glycosylation modification is still poor, making it difficult to meet commercial requirements. Therefore, further improvement measures are needed. Summary of the Invention

[0005] The purpose of this invention is to overcome the shortcomings and deficiencies of existing technologies and provide a method for enhancing the stability of protein peptide nutritional emulsions through glycosylation-calcium bridging synergistic modification. This invention is based on the fact that glycosylation can effectively inhibit the growth of metal ions (such as Ca). 2+ Leveraging the characteristics of heat-induced protein peptide aggregation, this product incorporates CaCl2 and preheating before emulsion preparation. This calcium bridge promotes moderate cross-linking between protein peptides, regulating their self-assembly behavior, interfacial behavior, and emulsion formation process. This enhances interfacial layer stability and inhibits oil droplet flocculation. This protein peptide nutritional emulsion significantly reduces the adverse effects of divalent metal ions on product stability and greatly improves long-term product stability, providing a feasible solution for technological innovation in the nutritional emulsion industry.

[0006] The objective of this invention is achieved through the following technical solution: Each 100mL of protein peptide emulsion contains 3-8g of protein peptides, 2-12g of fats, 8-30g of carbohydrates, 0.25-2mmol of calcium chloride (i.e., 10-80mg of elemental calcium), with the remainder being water. It also includes the following steps: S1: Add the weighed protein peptides and carbohydrates to distilled water and stir thoroughly until fully hydrated. Stir at 400-1000 rpm for 2-6 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 8.0~11.0, and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a constant, the above protein peptide / carbohydrate aqueous solution undergoes a low-degree glycosylation reaction under ultrasonic and water bath heating conditions. The glycosylation reaction conditions are: reaction temperature 60~90℃, ultrasonic power 400~800W, reaction time 1~20min, and stirring speed 200~600rpm. The degree of glycosylation of the obtained glycosylated protein peptide is 10-25%. The reaction is quickly terminated by using a cold water bath and an ice water bath after the reaction. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 6.5~7.5, and calcium chloride solution is added for calcium heat pretreatment. The pretreatment temperature is 20~95℃, the treatment time is 5~120min, and the stirring speed is 400~800rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed fats, complex minerals and complex vitamins to the protein / carbohydrate reaction solution that has been pretreated with calcium heat, and perform homogenization and emulsification. The homogenization speed is 8000~15000 rpm and the homogenization time is 2~45 min to obtain a protein / carbohydrate / fat ternary crude emulsion. S6: The above-mentioned ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 60~150MPa, and the number of homogenizations is 2~6 times to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 115~125℃ for 10~25 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain a protein peptide emulsion with better stability.

[0007] The calcium chloride mentioned is used as a model of divalent metal ions in minerals, and magnesium chloride can also be used as a substitute in practice.

[0008] The second objective of this invention is to prepare a protein peptide nutritional emulsion using the above-mentioned glycosylation-calcium bridging synergistic modification technology, comprising the following characteristics: A protein peptide nutritional emulsion with high thermal stability and ion tolerance, in addition to the above-mentioned components, may also contain 0.05~0.5g of compound vitamins and 0.02~1g of compound minerals per 100mL of protein peptide nutritional emulsion.

[0009] The protein peptides are mainly derived from one or a mixture of several of the following: hydrolyzed milk protein, hydrolyzed soy protein, hydrolyzed cereal protein, hydrolyzed tuber protein, and collagen peptides. (The hydrolyzed milk protein mainly includes hydrolyzed whey protein and casein peptides; the hydrolyzed soy protein mainly includes hydrolyzed soybean protein and hydrolyzed pea protein; the hydrolyzed cereal protein mainly includes hydrolyzed wheat protein, hydrolyzed rice protein, hydrolyzed quinoa protein, rice bran protein peptides, and corn protein peptides; the hydrolyzed tuber protein mainly includes potato protein peptides and yam peptides.) The oils are mainly derived from one or a mixture of several of the following: soybean oil, corn oil, rapeseed oil, peanut oil, sesame oil, tea oil, sunflower seed oil, flaxseed oil, olive oil, and medium-chain fatty acid esters.

[0010] The carbohydrates are mainly derived from one or a mixture of several of the following: glucose, fructose, xylose, lactose, sucrose, maltose, galactomannan, xylooligosaccharides, fructooligosaccharides, maltodextrin, polydextrose, and maltodextrin.

[0011] Compared with the prior art, the present invention has the following advantages: (1) The glycosylation-calcium bridge synergistic modification provided by the present invention can significantly weaken the destructive effect of divalent metal ions on the stability of emulsions, and the resulting protein peptide nutritional emulsion has excellent ion tolerance and long-term stability.

[0012] (2) The above-mentioned protein peptide nutritional emulsion can meet the high demand of specific populations for protein and polypeptide absorption, and indirectly realize the application of calcium chelating peptides in nutritional emulsions, which is expected to improve the bioavailability of the calcium contained therein, and is conducive to promoting the synthesis of bones and muscles of the eaters.

[0013] (3) The protein peptide nutritional emulsion preparation method described in this invention is simple and convenient, and has a certain degree of universality. It can be applied to protein peptide matrix products from many sources, promoting the application of protein peptides in nutritional emulsions; it can become a new idea and strategy for upgrading nutritional emulsion processing technology. Attached Figure Description

[0014] Figure 1 The images show the appearance of the protein peptide nutritional emulsions obtained in Examples 1, 2, 1, and 2. Figure 2 The changes in backscattered light intensity during storage of the nutrient emulsions obtained in Examples 1, 2, and 1 are shown. Figure 3 The changes in average particle size of the nutrient emulsions obtained in Examples 1, 2, and 1 during storage; Figure 4 The images show the microstructure (transmission electron microscopy images) of the protein peptides obtained in Examples 1, 2, and 1. Detailed Implementation

[0015] To enable those skilled in the art to better understand the technical solution of the present invention, the present invention will be further described below in conjunction with the accompanying drawings and specific embodiments.

[0016] Example 1 A protein peptide nutritional emulsion, the raw materials of which include 4% (m / v) hydrolyzed whey protein, 4% soybean oil, 24% xylose, 0.06% calcium ions (derived from CaCl2), 0.2% complex vitamins and 0.15% complex minerals, with the balance being water.

[0017] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (hydrolyzed whey protein) and carbohydrates (xylose) to distilled water, stir thoroughly until fully hydrated, stir at 600 rpm for 4 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 10.0 and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a constant, the above protein peptide / carbohydrate aqueous solution was subjected to a low degree of glycosylation under ultrasonic and water bath heating conditions. The glycosylation reaction conditions were: reaction temperature 80℃, ultrasonic power 500W, reaction time 2min, and stirring speed 600rpm. The degree of glycosylation of the obtained glycosylated protein peptide was 20%. The reaction was quickly terminated by using a cold water bath and an ice water bath after the reaction. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 7.0, and calcium chloride solution is added for calcium heat pretreatment. The pretreatment temperature is 25℃ (room temperature), the treatment time is 60 min, and the stirring speed is 600 rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed oil (soybean oil), compound minerals and compound vitamins to the protein / carbohydrate reaction solution with added calcium chloride, and perform homogenization and emulsification at a speed of 10,000 rpm for 2 minutes to obtain a protein / carbohydrate / oil ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 125MPa, and the homogenization is performed 3 times to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 121°C for 15 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0018] Example 2 A protein peptide nutritional emulsion, the raw materials of which include 4% (m / v) hydrolyzed whey protein, 4% soybean oil, 24% xylose, 0.06% calcium ions (derived from CaCl2), 0.2% complex vitamins and 0.15% complex minerals, with the balance being water.

[0019] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (hydrolyzed whey protein) and carbohydrates (xylose) to distilled water, stir thoroughly until fully hydrated, stir at 600 rpm for 4 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 10.0 and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a constant, the above protein peptide / carbohydrate aqueous solution was subjected to a low degree of glycosylation under ultrasonic and water bath heating conditions. The glycosylation reaction conditions were: reaction temperature 80℃, ultrasonic power 500W, reaction time 2min, and stirring speed 600rpm. The degree of glycosylation of the obtained glycosylated protein peptide was 20%. The reaction was quickly terminated by using a cold water bath and an ice water bath after the reaction. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 7.0, and calcium chloride solution is added for calcium heat pretreatment. The pretreatment temperature is 95℃, the treatment time is 8min, and the stirring speed is 600rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed oil (soybean oil), compound minerals and compound vitamins to the protein / carbohydrate reaction solution with added calcium chloride, and perform homogenization and emulsification at a speed of 10,000 rpm for 2 minutes to obtain a protein / carbohydrate / oil ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 125MPa, and the homogenization is performed 3 times to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 121°C for 15 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0020] Example 3 A protein peptide nutritional emulsion, the raw materials of which include 3% (m / v) hydrolyzed whey protein, 4% soybean oil, 24% xylose, 0.08% calcium ions (derived from CaCl2), 0.2% complex vitamins and 0.15% complex minerals, with the balance being water.

[0021] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (hydrolyzed whey protein) and carbohydrates (xylose) to distilled water, stir thoroughly until fully hydrated, stir at 600 rpm for 4 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 10.0 and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a constant, the above protein peptide / carbohydrate aqueous solution was subjected to a low degree of glycosylation under ultrasonic and water bath heating conditions. The glycosylation reaction conditions were: reaction temperature 80℃, ultrasonic power 500W, reaction time 2min, and stirring speed 600rpm. The degree of glycosylation of the obtained glycosylated protein peptide was 20%. The reaction was quickly terminated by using a cold water bath and an ice water bath after the reaction. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 7.0, and calcium chloride solution is added for calcium heat pretreatment. The pretreatment temperature is 40℃, the treatment time is 30min, and the stirring speed is 600rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed oil (soybean oil), compound minerals and compound vitamins to the protein / carbohydrate reaction solution with added calcium chloride, and perform homogenization and emulsification at a speed of 10,000 rpm for 2 minutes to obtain a protein / carbohydrate / oil ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 125MPa, and the homogenization is performed 3 times to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 121°C for 15 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0022] Example 4 A protein peptide nutritional emulsion, the raw materials of which include 8% (m / v) hydrolyzed whey protein, 4% soybean oil, 24% xylose, 0.01% calcium ions (derived from CaCl2), 0.2% complex vitamins and 0.15% complex minerals, with the balance being water.

[0023] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (hydrolyzed whey protein) and carbohydrates (xylose) to distilled water, stir thoroughly until fully hydrated, stir at 600 rpm for 4 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 10.0 and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a constant, the above protein peptide / carbohydrate aqueous solution undergoes a low-degree glycosylation reaction under ultrasonic and water bath heating conditions. The glycosylation reaction conditions are: reaction temperature 80℃, ultrasonic power 500W, reaction time 2min, stirring speed 600rpm. After the reaction, the reaction is quickly terminated by using a cold water bath and an ice water bath. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 7.0, and calcium chloride solution is added for calcium heat pretreatment. The pretreatment temperature is 25℃ (room temperature), the treatment time is 60 min, and the stirring speed is 600 rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed oil (soybean oil), compound minerals and compound vitamins to the protein / carbohydrate reaction solution with added calcium chloride, and perform homogenization and emulsification at a speed of 10,000 rpm for 2 minutes to obtain a protein / carbohydrate / oil ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 125MPa, and the homogenization is performed 3 times to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 121°C for 15 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0024] Example 5 A protein peptide nutritional emulsion, the raw materials of which include 6% (m / v) casein peptides, 5% corn oil, 12% maltodextrin, 4% glucose, 0.08% calcium ions (derived from CaCl2), 0.3% complex vitamins and 0.5% complex minerals, with the balance being water.

[0025] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (casein peptides) and carbohydrates (maltodextrin, glucose) to distilled water, stir thoroughly until fully hydrated, stir at 600 rpm for 4 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 11.0 and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a fixed level, the above protein peptide / carbohydrate aqueous solution undergoes a low-degree glycosylation reaction under ultrasonic and water bath heating conditions. The glycosylation reaction conditions are: reaction temperature 90℃, ultrasonic power 600W, reaction time 5min, stirring speed 600rpm. After the reaction, the reaction is quickly terminated by using a cold water bath and an ice water bath. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 6.8, and magnesium chloride solution is added for heating pretreatment. The pretreatment temperature is 80℃, the treatment time is 15min, and the stirring speed is 800rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed oil (corn oil), compound minerals and compound vitamins to the protein / carbohydrate reaction solution with added magnesium chloride, and perform homogenization and emulsification at a speed of 12000 rpm for 5 minutes to obtain a protein / carbohydrate / oil ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 150MPa, and the homogenization is performed 4 times to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 125°C for 10 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0026] Example 6 A protein peptide nutritional emulsion, the raw materials of which include 3% (m / v) hydrolyzed soybean protein, 2% yam peptide, 6% low erucic acid rapeseed oil, 10% maltodextrin, 2% sucrose, 2% oligomaltose, 0.024% magnesium ions (derived from MgCl2), and the balance being water.

[0027] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (hydrolyzed soy protein, yam peptides) and carbohydrates (maltodextrin, sucrose, oligomaltose) to distilled water, stir thoroughly until fully hydrated, stir at 800 rpm for 2 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 9.0 and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a fixed level, the above protein peptide / carbohydrate aqueous solution undergoes a low-degree glycosylation reaction under ultrasonic and water bath heating conditions. The glycosylation reaction conditions are: reaction temperature 70℃, ultrasonic power 800W, reaction time 15min, stirring speed 400rpm. After the reaction, the reaction is quickly terminated by using a cold water bath and an ice water bath. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 6.8, and calcium chloride solution is added for calcium heat pretreatment. The pretreatment temperature is 40℃, the treatment time is 30min, and the stirring speed is 600rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed oil (low erucic acid rapeseed oil) to the protein / carbohydrate reaction solution with added calcium chloride, and perform homogenization and emulsification at a speed of 15000 rpm for 10 min to obtain a protein / carbohydrate / oil ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 100MPa, and the homogenization is performed twice to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 115°C for 20 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0028] Example 7 A protein peptide nutritional emulsion, the raw materials of which include 2% (m / v) hydrolyzed pea protein, 1% collagen peptide, 3% medium-chain triglycerides, 2% flaxseed oil, 12% glucose, 1% polydextrose, 0.04% calcium ions (derived from CaCl2), 0.1% complex vitamins and 0.1% complex minerals, with the balance being water.

[0029] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (hydrolyzed pea protein, collagen peptides) and carbohydrates (glucose, polydextrose) to distilled water, stir thoroughly until fully hydrated, stir at 500 rpm for 4 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 10.0 and bring the volume to the corresponding level; S3: After adjusting the pH and making up the volume, the above protein peptide / carbohydrate aqueous solution undergoes a low degree of glycosylation under ultrasonic and water bath heating conditions. The glycosylation reaction conditions are: reaction temperature 60℃, ultrasonic power 800W, reaction time 20min, stirring speed 600rpm. After the reaction, the reaction is quickly terminated by cold water bath and ice water bath. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 7.0, and calcium chloride solution is added for calcium heat pretreatment. The pretreatment temperature is 50℃, the treatment time is 30min, and the stirring speed is 600rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed oils (medium-chain triglycerides, flaxseed oil), complex minerals and complex vitamins to the protein / carbohydrate reaction solution with added calcium chloride, and perform homogenization emulsification at a speed of 12000 rpm for 5 minutes to obtain a protein / carbohydrate / oil ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 120MPa, and the homogenization is performed 3 times to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 121°C for 15 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0030] Comparative Example 1 A protein peptide nutritional emulsion, the raw materials of which include 4% (m / v) hydrolyzed whey protein, 4% soybean oil, 24% xylose, 0.06% calcium ions (derived from CaCl2), 0.2% complex vitamins and 0.15% complex minerals, with the balance being water.

[0031] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (hydrolyzed whey protein) and carbohydrates (xylose) to distilled water, stir thoroughly until fully hydrated, stir at 600 rpm for 4 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 10.0 and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a constant, the above protein peptide / carbohydrate aqueous solution was subjected to a low degree of glycosylation under ultrasonic and water bath heating conditions. The glycosylation reaction conditions were: reaction temperature 80℃, ultrasonic power 500W, reaction time 2min, and stirring speed 600rpm. The degree of glycosylation of the obtained glycosylated protein peptide was 20%. The reaction was quickly terminated by using a cold water bath and an ice water bath after the reaction. S4: After the glycosylation reaction is complete, adjust the pH of the obtained protein peptide / carbohydrate reaction solution to 7.0; S5: Add the weighed fats, complex minerals and complex vitamins to the above protein / carbohydrate reaction solution and perform homogenization emulsification at a speed of 10,000 rpm for 2 minutes to obtain a protein / carbohydrate / fat ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 125MPa, and the homogenization is performed 3 times to obtain the ternary fine emulsion. S7: After homogenization, add calcium chloride solution to the above ternary fine emulsion and mix thoroughly. Then fill and autoclave at 121°C for 15 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0032] Comparative Example 2 A protein peptide nutritional emulsion, the raw materials of which include 4% (m / v) hydrolyzed whey protein, 4% soybean oil, 24% xylose, 0.06% calcium ions (derived from CaCl2), 0.2% complex vitamins and 0.15% complex minerals, with the balance being water.

[0033] The preparation method of the above-mentioned protein peptide nutritional emulsion is as follows: S1: Add the weighed protein peptides (hydrolyzed whey protein) and carbohydrates (xylose) to distilled water, stir thoroughly until fully hydrated, stir at 600 rpm for 4 hours to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 7.0, add calcium chloride solution, and perform calcium heat pretreatment. The pretreatment temperature is 25℃ (room temperature), the treatment time is 60 min, and the stirring speed is 600 rpm to promote moderate cross-linking between calcium ions and protein peptides. S5: Add the weighed oil (soybean oil), compound minerals and compound vitamins to the protein / carbohydrate reaction solution with added calcium chloride, and perform homogenization and emulsification at a speed of 10,000 rpm for 2 minutes to obtain a protein / carbohydrate / oil ternary crude emulsion. S6: The above ternary crude emulsion is subjected to high-pressure homogenization. The high-pressure homogenization equipment is a high-pressure micro-jet homogenizer, the homogenization pressure is 125MPa, and the homogenization is performed 3 times to obtain the ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and subjected to high-pressure steam sterilization at a temperature of 115°C for 10 minutes. S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide nutritional emulsion.

[0034]

[0035] The test results of the seven embodiments are quite similar. Among them, Embodiment 1 and Embodiment 2 are representative, so they are used as the main objects of analysis.

[0036] Comparing the protein peptide nutritional emulsions prepared in Examples 1 and 2 with those prepared in Comparative Examples 1 or 2, the appearance images after high-temperature and high-pressure sterilization show that, compared with the nutritional emulsion prepared by glycosylation and the addition of calcium chloride after emulsification (Comparative Example 1) or the nutritional emulsion prepared by inducing calcium ion crosslinking with protein peptides at room temperature before emulsification without glycosylation (Comparative Example 2), the nutritional emulsion prepared by glycosylation combined with room temperature induction of calcium ion crosslinking with protein peptides before emulsification (Example 1) and the nutritional emulsion prepared by glycosylation combined with heating induction of calcium ion crosslinking with protein peptides before emulsification (Example 2) exhibited better stability. Neither of them showed obvious clumping or scaling, while scaling occurred at the liquid surface of Comparative Example 1, and obvious clumping occurred in Comparative Example 2. Figure 1 The results indicate that glycosylation-calcium bridging synergistic modification (Example 1 or Example 2) significantly enhances the thermal stability and sensory quality of protein peptide nutritional emulsions; while when glycosylation modification or calcium bridging crosslinking acts independently on protein peptides (Comparative Example 1 or Comparative Example 2), the stability of the nutritional emulsion is poor.

[0037] To further investigate the long-term stability of the comparative emulsion products, a 75-day storage test was conducted on the products obtained in Example 1, Example 2, and Comparative Example 1. The results of the backscattered light intensity show that, compared to Comparative Example 1, the nutritional emulsions obtained in Example 1 and Example 2 exhibited a lower phase separation rate and enhanced homogeneity during storage. Figure 2 Furthermore, compared to Comparative Example 1, the average particle size of the nutritional emulsions obtained in Examples 1 and 2 was significantly reduced during storage, and the increase in average particle size with prolonged storage time was also greatly slowed down. Moreover, the average particle size of Example 1 was lower than that of Example 2. Figure 3 The above results collectively demonstrate that the glycosylation-calcium bridge synergistic modification technique can significantly weaken the calcium content of calcium bridges. 2+ It reduces the adverse effects on the stability of nutritional emulsions, improves the thermal stability and ionic tolerance of emulsion products, and endows them with better long-term stability and shelf life.

[0038] To clarify the mechanism by which glycosylation-calcium bridging synergistic modification enhances the stability of protein peptide nutritional emulsions, the protein peptides were further characterized structurally. Transmission electron microscopy images showed that Comparative Example 1 formed large, irregular aggregates, while Example 1 formed smaller nanoparticles, and Example 2 formed a looser network structure. Figure 4 Furthermore, compared to Comparative Example 1, these two ordered self-assembled structures exhibit higher hydrophobicity and molecular flexibility, which typically have a positive impact on emulsion stability. The related results indicate that glycosylation-calcium bridging synergistic modification can mediate the formation of ordered assemblies from protein peptides, providing strong evidence for their enhanced emulsion stability.

[0039] In addition, Examples 3-7 showed similar effects to Examples 1 and 2, with no scaling or clumping observed. However, the average particle sizes differed significantly, being 862.79, 248.62, 344.26, 276.51, and 301.90 nm, respectively, all significantly lower than Comparative Example 1 (1358.50 nm). This further demonstrates that glycosylation-calcium bridging synergistic modification can significantly enhance the stability of protein peptide nutritional emulsions and can be widely applied to the manufacture of protein peptide nutritional emulsions from different sources. Furthermore, based on the results of Examples 1-7, the effect of this synergistic modification in enhancing emulsion stability mainly depends on the calcium ion concentration and the calcium crosslinking temperature. When the calcium ion concentration is below 0.04% (10 mM), the average particle size of the protein peptide nutritional emulsion is below 350 nm, while even at a concentration as high as 0.08% (20 mM), it remains within the ≤1000 nm range. When the calcium ion concentration is high, a lower calcium crosslinking temperature is preferable. On the other hand, the concentration and stability of the protein peptides also affect the synergistic modification effect; lower concentrations and better stability of the protein peptides result in better effects. It is important to note that the above effects are achieved through prolonged high-temperature steam sterilization (which has a very strong destructive effect on emulsion stability). In actual industrial manufacturing, ultra-high temperature (UHT) sterilization, which has a weaker destructive effect on emulsion stability, is usually used. Under these conditions, the stability of protein nutritional emulsions is continuously and significantly improved, and they can meet the corresponding product quality standards.

[0040] The above embodiments are merely specific implementations of the present invention, and their descriptions are quite specific and detailed. However, they should not be construed as limiting the patent scope of the present invention to these embodiments. It should be noted that several modifications and improvements can be made without departing from the concept of the present invention, and these obvious substitutions all fall within the protection scope of the present invention.

Claims

1. A method for enhancing the stability of protein peptide emulsions through glycosylation-calcium bridging synergistic modification, characterized in that, Each 100mL protein peptide emulsion contains 3-8g protein peptides, 2-12g oils, 8-30g carbohydrates, 0.25-2mmol calcium chloride or magnesium chloride, with the remainder being water, and includes the following steps: S1: Add the weighed protein peptides and carbohydrates to distilled water and stir thoroughly until fully hydrated to obtain a protein peptide / carbohydrate aqueous solution. S2: Adjust the pH of the above protein peptide / carbohydrate aqueous solution to 8.0~11.0, and bring the volume to the corresponding level; S3: After adjusting the pH and bringing the volume to a constant, the above protein peptide / carbohydrate aqueous solution undergoes a low-degree glycosylation reaction under ultrasonic and water bath heating conditions. The reaction is then quickly terminated using a cold water bath and an ice water bath. S4: After the glycosylation reaction is completed, the pH of the obtained protein peptide / carbohydrate reaction solution is adjusted to 6.5~7.5, and calcium chloride solution is added for calcium heat pretreatment to promote moderate cross-linking between calcium ions and protein peptides; S5: Add the weighed oil to the protein / carbohydrate reaction solution that has been pretreated with calcium heat, and perform homogenization and emulsification to obtain a ternary crude emulsion of protein / carbohydrate / oil. S6: The above ternary crude emulsion is subjected to high-pressure homogenization to obtain a ternary fine emulsion. S7: After homogenization, the above ternary fine emulsion is filled and sterilized; S8: After sterilization, remove the above ternary fine emulsion and cool it to room temperature to obtain the protein peptide emulsion.

2. The method for enhancing the stability of protein peptide emulsions through glycosylation-calcium bridging synergistic modification according to claim 1, characterized in that, In step S1, the stirring speed is 400~1000 rpm and the stirring time is 2~6 h; in step S3, the glycosylation reaction conditions are a reaction temperature of 60~90℃, an ultrasonic power of 400~800W, a reaction time of 1~20 min, and a stirring speed of 200~600 rpm, and the degree of glycosylation of the obtained glycosylated protein peptides is 10~25%.

3. The method for enhancing the stability of protein peptide emulsions through glycosylation-calcium bridging synergistic modification according to claim 1, characterized in that, In step S4, the calcium heat pretreatment temperature is 20~95℃, the treatment time is 5~120min, and the stirring speed is 400~800rpm; in step S5, the homogenization speed of the homogenized emulsion is 8000~15000rpm, and the homogenization time is 2~45min.

4. The method for enhancing the stability of protein peptide emulsions through glycosylation-calcium bridging synergistic modification according to claim 1, characterized in that, In step S6, the high-pressure homogenizing equipment is a high-pressure micro-jet homogenizer, with a homogenization pressure of 60~150MPa and a homogenization frequency of 2~6 times; in step S7, the sterilization refers to high-pressure steam sterilization, with a temperature of 115~125℃ and a time of 10~25min.

5. A protein peptide nutritional emulsion prepared based on the glycosylation-calcium bridge co-modification according to any one of claims 1 to 4, characterized in that, In addition to the components described in claim 1, 0.05-0.5g of complex vitamins and 0.02-1g of complex minerals may also be added to each 100mL of protein peptide nutritional emulsion.

6. The protein peptide nutritional emulsion according to claim 5, characterized in that, The protein peptides are mainly derived from one or a mixture of several of hydrolyzed milk protein, hydrolyzed soy protein, hydrolyzed cereal protein, hydrolyzed tuber protein, and collagen peptides. The hydrolyzed milk protein mainly includes hydrolyzed whey protein and casein peptide; the hydrolyzed legume protein mainly includes hydrolyzed soybean protein and hydrolyzed pea protein; the hydrolyzed cereal protein mainly includes hydrolyzed wheat protein, hydrolyzed rice protein, hydrolyzed quinoa protein, rice bran protein peptide, and corn protein peptide; the hydrolyzed tuber protein mainly includes potato protein peptide and yam peptide.

7. The protein peptide nutritional emulsion according to claim 5, characterized in that, The oils are mainly derived from one or a mixture of several of the following: soybean oil, corn oil, rapeseed oil, peanut oil, sesame oil, tea oil, sunflower seed oil, flaxseed oil, olive oil, and medium-chain fatty acid esters.

8. The protein peptide nutritional emulsion according to claim 5, characterized in that, The carbohydrates are mainly derived from one or a mixture of several of the following: glucose, fructose, xylose, lactose, sucrose, maltose, galactomannan, xylooligosaccharides, fructooligosaccharides, maltodextrin, polydextrose, and maltodextrin.

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