Method for preparing proanthocyanidin microcapsules by complex coacervation self-assembly

Abstract

This invention discloses a method for preparing proanthocyanidin microcapsules through complex coagulation self-assembly, belonging to the field of microcapsule preparation technology. It aims to solve problems such as the easy degradation and inactivation of proanthocyanidins, the high-temperature degradation of activity in existing preparation methods, and cumbersome processes. The method uses proanthocyanidins as the core material and sodium alginate and whey protein as the wall material. The product is obtained through a four-step process: wall material solution preparation, core material solution preparation, complex coagulation self-assembly reaction, and proanthocyanidin encapsulation and solidification. The wall material solution consists of a 1-3% (w / w) sodium alginate solution and a 1.0-5.0% (whey protein) solution. The core material solution is prepared by dissolving proanthocyanidins in anhydrous ethanol and ultrasonically dispersing them. The pH of the complex coagulation reaction is adjusted to 3.5-7.5, and the reaction is carried out with stirring at 30-50°C. Finally, the microcapsules are obtained by calcium chloride cross-linking solidification and vacuum freeze-drying. The preparation conditions of this invention are mild, without high-temperature and long-term treatment, and the core material encapsulation rate reaches 85% to 95%. The product has strong antioxidant activity and good stability. Moreover, the process steps are few, no organic solvents or additional excipients are required, the raw materials are readily available, and the production cost is low. It can also be applied to the encapsulation of other polyphenols and flavonoid active ingredients, and has broad application prospects in the fields of medicine, food, and cosmetics.

Description

Technical Field

[0001] This invention belongs to the field of proanthocyanidin microcapsule preparation technology, specifically relating to a method for preparing proanthocyanidin microcapsules through complex coagulation self-assembly. Background Technology

[0002] Proanthocyanidins are an important component of flavonoids, a polymer mixture formed by the carbon-carbon condensation of flavan-3-ol units such as catechins and epicatechins. They possess a variety of biological activities, including strong antioxidant activity, free radical scavenging, anti-inflammatory effects, immunomodulation, cardiovascular protection, obesity prevention, and anti-tumor activity. Their antioxidant capacity surpasses that of vitamin C, vitamin E, and β-carotene, and they are internationally recognized as highly effective natural antioxidants, showing broad application prospects in medicine, food preservation, nutrition, and cosmetics. However, during application, proanthocyanidins are susceptible to oxidative hydrolysis or ring-opening degradation due to environmental influences, leading to a decrease in physiological activity. Therefore, methods for improving the physiological activity of proanthocyanidins have considerable economic value and practical significance.

[0003] Existing methods for preparing proanthocyanidin microcapsules include, for example, the publication number CN109259234A, published on January 25, 2019, entitled "A proanthocyanidin microcapsule with resistant starch as the wall material and its preparation method". The method disclosed is as follows: using resistant starch as the wall material, proanthocyanidins extracted from lychee peel are mixed with resistant starch, emulsifiers and protectants are added, and after homogenization, spray drying is performed to obtain microcapsules. The main disadvantages of this method are: (1) This method uses spray drying to prepare microcapsules, with an inlet air temperature as high as 180-200℃ and an outlet air temperature of 80-90℃. High-temperature treatment can easily destroy the thermosensitive bioactivity of proanthocyanidins and affect the bioactivity of microcapsule products; (2) This method requires the pre-preparation of resistant starch wall material, which involves multiple complex processes such as enzymatic hydrolysis of α-amylase and glucoamylase, microwave treatment, and aging treatment. The wall material preparation process is cumbersome and the production cycle is long, which increases the production cost and the difficulty of technical control; (3) This method requires the addition of monoglycerides as emulsifiers, collagen and glycerol as protectants and other excipients, which increases the complexity of the formulation and the cost of raw materials. In addition, the spray drying process has high energy consumption, which is not conducive to energy conservation and environmental protection. Summary of the Invention

[0004] The purpose of this invention is to address the problems of insufficient stability, easy degradation and inactivation, lack of efficient encapsulation carriers and functional application products of proanthocyanidins in existing applications, and to provide a method for preparing proanthocyanidin microcapsules by complex coagulation. This method has the advantages of fewer operation steps, mild conditions, high encapsulation rate, low environmental pollution, low production energy consumption and cost, good product stability, strong antioxidant activity, and excellent sustained-release performance. Furthermore, it can be applied to facial masks to realize the high-value utilization of proanthocyanidins in the cosmetic field.

[0005] The mechanism of this invention is as follows: Proanthocyanidins are prepared using a complex coagulation method to self-assemble microcapsules, with sodium alginate and whey protein as wall materials. By adjusting the pH to 4.5, negatively charged sodium alginate and positively charged whey protein undergo electrostatic self-assembly on the surface of proanthocyanidin droplets, forming a dense interfacial film. Subsequently, Ca²⁺ is introduced, which specifically cross-links with sodium alginate to construct an "egg-box" three-dimensional gel network, achieving effective encapsulation of proanthocyanidins. Simultaneously, hydrogen bonds and hydrophobic interactions are formed between the phenolic hydroxyl groups of proanthocyanidins and the wall materials, further enhancing structural stability. This method is mild, avoiding the destruction of proanthocyanidin activity caused by prolonged high-temperature treatment, has high encapsulation efficiency, requires no organic solvents, is safe and environmentally friendly, and achieves high-value utilization of proanthocyanidins.

[0006] The technical solution to achieve the invention's objective is: a method for preparing proanthocyanidin microcapsules using a complex coagulation method. Using proanthocyanidins as the core material and sodium alginate and whey protein as wall materials, the method involves a simple process in a complex coagulation reaction system, including wall material solution preparation, core material solution preparation, complex coagulation self-assembly reaction, and proanthocyanidin encapsulation and solidification, to obtain the proanthocyanidin microcapsules. The specific steps of the method are as follows: (1) Preparation of wall material solution First, weigh 0.5–1.5 g of sodium alginate and 0.5–2.5 g of whey protein and dissolve them separately in 50 mL of distilled water. Then, stir magnetically in a water bath at 40–60 °C for 30–60 min until completely dissolved to obtain sodium alginate solution and whey protein solution, respectively. The mass concentration of the sodium alginate solution is 1–3%, and the mass concentration of the whey protein solution is 1.0–5.0%.

[0007] (2) Core material solution preparation After completing step (1), weigh 0.25-6g of proanthocyanidins, dissolve them in 5-15mL of anhydrous ethanol, and then ultrasonically disperse them for 10-20min under ultrasonic power of 100-200W and temperature of 20-30℃ to obtain the core material solution.

[0008] (3) Complex condensation autonomous reaction After step (2) is completed, the core material solution is added to the whey protein solution and stirred evenly according to the mass ratio of core material to wall material of 2:1 to 1:4. Then, the sodium alginate solution is slowly added dropwise. Next, the pH value of the system is adjusted to 3.5 to 7.5 with 10% acetic acid solution. Then, the mixture is stirred at 30 to 50°C and at a stirring speed of 300 to 600 r / min for 30 to 60 min. Finally, the mixture is cooled to room temperature to obtain the mixture.

[0009] (4) Proanthocyanidin encapsulation and solidification After step (3) is completed, the mixture is first added dropwise to 150-250 mL of calcium chloride solution with a mass concentration of 1%-3% at a rate of 1-3 mL / min using a peristaltic pump. Then, while adding the mixture, it is stirred at 200-400 r / min. After solidification for 2-4 h, the mixture is then separated by filtration. The solid is washed three times with distilled water to obtain wet microcapsules. Finally, the wet microcapsules are placed flat in a vacuum freeze dryer and freeze-dried at -70℃ for 48 h to obtain proanthocyanidin microcapsules.

[0010] After adopting the above technical solution, the present invention mainly has the following effects: (1) In this invention, sodium alginate and whey protein are combined as wall material, and proanthocyanidins are encapsulated by complex coagulation method. The encapsulation rate of proanthocyanidins in the core material can reach 85% to 95%. Water is used as solvent for wall material and only a small amount of ethanol is used to dissolve core material. No other emulsifiers, protective agents or other excipients are used, which is beneficial to environmental protection.

[0011] (2) The preparation conditions of the present invention using the complex coagulation self-assembly method are optimal at 40℃ and the reaction time is only 45min. This avoids the destruction of proanthocyanidin bioactivity by high temperature and long-term treatment, reduces energy consumption in the production process, and helps to reduce production costs. The preparation method is simple, green and environmentally friendly.

[0012] (3) This invention uses sodium alginate and whey protein as wall materials, which are widely available and readily available. They can also be widely used in the microencapsulation of other unstable active ingredients such as polyphenols and flavonoids, providing technical support for their application in the pharmaceutical, food, and cosmetic fields. The purpose of this invention is to address the problems of insufficient stability, easy degradation and inactivation, lack of efficient encapsulation carriers and functional application products in the existing application of proanthocyanidins, and to provide a method for preparing proanthocyanidin microcapsules by complex coagulation. This method has the characteristics of fewer operation steps, mild conditions, high encapsulation rate, low environmental pollution, low production energy consumption and cost, good product stability, strong antioxidant activity, and excellent sustained-release performance. Furthermore, it can be applied to facial masks to realize the high-value utilization of proanthocyanidins in the cosmetic field. Detailed Implementation

[0013] The present invention will be further described below with reference to specific embodiments: Example

[0014] The specific steps of a method for preparing proanthocyanidin microcapsules through complex coagulation self-assembly are as follows: (1) Preparation of wall material solution First, weigh 1g of sodium alginate and 1.5g of whey protein and dissolve them separately in 50mL of distilled water. Then, stir magnetically for 40min in a 50℃ water bath until completely dissolved to obtain sodium alginate solution and whey protein solution, respectively. The mass concentration of the sodium alginate solution is 2%, and the mass concentration of the whey protein solution is 3%.

[0015] (2) Core material solution preparation After completing step (1), weigh 1.25g of proanthocyanidins, dissolve them in 10mL of anhydrous ethanol, and then ultrasonically disperse them for 15min under ultrasonic power of 150W and temperature of 25℃ to obtain the core material solution.

[0016] (3) Complex condensation self-assembly reaction After step (2) is completed, the core material solution is added to the above whey protein solution and stirred evenly according to the mass ratio of core material to wall material of 1:2. Then, the above sodium alginate solution is slowly added dropwise. Next, the pH value of the system is adjusted to 4.5 with 10% acetic acid solution. Then, the mixture is stirred at 40°C and at a stirring speed of 400r / min for 45min. Finally, the mixture is cooled to room temperature to obtain the mixture.

[0017] (4) Proanthocyanidin encapsulation and solidification After step (3) is completed, the mixture is first added dropwise to 200 mL of 2% calcium chloride solution at a rate of 1.5 mL / min using a peristaltic pump. Then, the mixture is added dropwise while stirring at 300 r / min. After solidification for 3 h, the mixture is then separated by filtration. The solid is washed three times with distilled water to obtain wet microcapsules. Finally, the wet microcapsules are placed flat in a vacuum freeze dryer and freeze-dried at -70℃ for 48 h to obtain proanthocyanidin microcapsules. Example

[0018] A method for preparing proanthocyanidin microcapsules through complex condensation self-assembly, the same as in Example 1, wherein: In step (1), the concentration of sodium alginate is 1.5%, the concentration of whey protein is 2.0%, the dissolution temperature is 45℃, the dissolution time is 50min, and the stirring speed is 350r / min.

[0019] In step (2), the amount of proanthocyanidins used was 1.75g, the amount of anhydrous ethanol used was 8mL, the ultrasonic time was 12min, the core-to-wall ratio was 1:1, the pH value was 3.5, the reaction temperature was 35℃, the reaction time was 40min, and the stirring speed was 350r / min.

[0020] In step (3), the core-to-wall ratio is 1:1, the pH value is 3.5, the reaction temperature is 35℃, the reaction time is 40min, and the stirring speed is 350r / min.

[0021] In step (4), the calcium chloride concentration is 2.5%, the curing time is 2.5 h, the curing stirring speed is 250 r / min, and the peristaltic pump dripping speed is 2.0 mL / min. Example

[0022] A method for preparing proanthocyanidin microcapsules through complex condensation self-assembly, the same as in Example 1, wherein: In step (1), the concentration of sodium alginate is 2.5%, the concentration of whey protein is 4.0%, the dissolution temperature is 55℃, the dissolution time is 30min, and the stirring speed is 500r / min.

[0023] In step (2), the amount of proanthocyanidins used is 1.08g, the amount of anhydrous ethanol used is 12mL, and the ultrasonic time is 18min.

[0024] In step (3), the core-to-wall ratio is 1:3, the pH value is 5.5, the reaction temperature is 45℃, the reaction time is 55min, and the stirring speed is 450r / min.

[0025] In step (4), the calcium chloride concentration is 3.0%, the curing time is 3.5h, the curing stirring speed is 350r / min, and the peristaltic pump dripping speed is 1.0mL / min.

[0026] Experimental results The same method was used to investigate the effect of pH value on proanthocyanidin encapsulation efficiency in the complex condensation reaction system. 1. Comparison of encapsulation efficiency of proanthocyanidin microcapsules in Examples 1-3 Table 1. Comparison of encapsulation efficiency of proanthocyanidin microcapsules in Examples 1-3 2. The reaction temperature was 40℃, the reaction time was 45 min, the calcium chloride concentration was 2.0%, and the curing time was 3 h. The effect of different pH values ​​on the encapsulation rate of proanthocyanidins was investigated.

[0027] Table 2. Comparison of the effect of different pH values ​​on proanthocyanidin encapsulation efficiency in complex coagulation reaction systems 3. The reaction temperature was 40℃, the reaction time was 45 min, the calcium chloride concentration was 2.0%, and the curing time was 3 h. The effect of different pH values ​​on the encapsulation rate of proanthocyanidins was investigated.

[0028] Table 3. Comparison of the effect of different sodium alginate concentrations on proanthocyanidin encapsulation efficiency in complex condensation reaction systems 4. The reaction temperature was 40℃, the reaction time was 45 min, the calcium chloride concentration was 2.0%, and the curing time was 3 h. The effect of different pH values ​​on the encapsulation rate of proanthocyanidins was investigated.

[0029] Table 4. Comparison of the effect of different whey protein concentrations on proanthocyanidin encapsulation efficiency in complex coagulation reaction systems 5. The reaction temperature was 40℃, the reaction time was 45 min, the calcium chloride concentration was 2.0%, and the curing time was 3 h. The effect of different pH values ​​on the encapsulation rate of proanthocyanidins was investigated.

[0030] Table 5. Comparison of proanthocyanidin encapsulation efficiency with different core-wall ratios in complex condensation reaction systems 6. The reaction temperature was 40℃, the reaction time was 45 min, the calcium chloride concentration was 2.0%, and the curing time was 3 h. The effect of different proanthocyanidin concentrations on the DPPH free radical scavenging rate was investigated.

[0031] Table 6. Comparison of DPPH free radical scavenging rates between proanthocyanidins and proanthocyanidin microcapsules Sodium alginate concentration (%) Whey protein concentration (%) pH value Core-to-wall ratio Proanthocyanidin concentration (mg / mL) Proanthocyanidins Proanthocyanidin microcapsules 2 3 4.5 2:1 0.02 20.69 23.69 2 3 4.5 1:1 0.04 35.68 38.68 2 3 4.5 1:2 0.06 43.88 47.88 2 3 4.5 1:3 0.08 54.12 57.12 2 3 4.5 1:4 0.10 67.23 70.23 7. The reaction temperature was 40℃, the reaction time was 45 min, the calcium chloride concentration was 2.0%, and the curing time was 3 h. The effect of different proanthocyanidin concentrations on the ABTS free radical scavenging rate was investigated.

[0032] Table 7. Comparison of ABTS free radical scavenging rates between proanthocyanidins and proanthocyanidin microcapsules Sodium alginate concentration (%) Whey protein concentration (%) pH value Core-to-wall ratio Proanthocyanidin concentration (mg / mL) Proanthocyanidins Proanthocyanidin microcapsules 2 3 4.5 2:1 0.02 20.57 21.57 2 3 4.5 1:1 0.04 31.29 34.29 2 3 4.5 1:2 0.06 54.68 56.28 2 3 4.5 1:3 0.08 73.73 74.12 2 3 4.5 1:4 0.10 82.68 86.63 The above experiments show that the present invention uses a complex coagulation method to prepare proanthocyanidin microcapsules. When the sodium alginate concentration is 2%, the whey protein concentration is 3%, the core-to-wall ratio is 1:2, the pH value is 4.5, the reaction temperature is 40℃, and the reaction time is 45 min, the proanthocyanidin microcapsule product can be obtained after solidification and freeze-drying. This method has a high encapsulation efficiency of up to 91.32%, and achieves DPPH and ABTS free radical scavenging rates of 70.23% and 86.63%, respectively, while exhibiting good physical stability. Furthermore, the preparation conditions are mild, the operation is simple, and no organic solvents are required, avoiding environmental pollution and corrosion of production equipment, thus contributing to environmental protection and reducing production costs.

Claims

1. A method for preparing proanthocyanidin microcapsules through complex coagulation self-assembly, characterized in that, Using proanthocyanidins as the core material and sodium alginate and whey protein as the wall material, the product was prepared through a process including wall material solution preparation, core material solution preparation, complex coagulation self-assembly reaction, and proanthocyanidin encapsulation and solidification. The specific steps are as follows: (1) Preparation of wall material solution: Weigh 0.5-1.5g sodium alginate and 0.5-2.5g whey protein and dissolve them in 50mL of distilled water. Stir magnetically for 30-60min under water bath conditions of 40-60℃ until completely dissolved to obtain sodium alginate solution with a mass concentration of 1-3% and whey protein solution with a mass concentration of 1.0-5.0% respectively. (2) Preparation of core material solution: Weigh 0.25-6g of proanthocyanidins, dissolve them in 5-15mL of anhydrous ethanol, and ultrasonically disperse them for 10-20min under ultrasonic power of 100-200W and temperature of 20-30℃ to obtain core material solution; (3) Complex coagulation self-assembly reaction: The core material solution is added to the whey protein solution and stirred evenly according to the mass ratio of core material to wall material of 2:1 to 1:

4. Sodium alginate solution is slowly added dropwise, and the pH value of the system is adjusted to 3.5 to 7.5 with 10% acetic acid solution. The reaction is carried out at 30 to 50°C and 300 to 600 r / min for 30 to 60 min. The mixture is then cooled to room temperature to obtain the mixture. (4) Proanthocyanidin encapsulation and solidification: The mixture was added dropwise to 150-250 mL of 1%-3% calcium chloride solution at a rate of 1-3 mL / min using a peristaltic pump while stirring at 200-400 r / min. After solidification for 2-4 h, the mixture was filtered and separated. The solid was washed three times with distilled water to obtain wet microcapsules. The wet microcapsules were then freeze-dried under vacuum at -70℃ for 48 h to obtain proanthocyanidin microcapsules.

2. The method for preparing proanthocyanidin microcapsules by complex coagulation self-assembly according to claim 1, characterized in that, In step (1), the mass concentration of sodium alginate solution is 2%, the mass concentration of whey protein solution is 3%, the water bath temperature is 50℃, and the magnetic stirring time is 40min.

3. The method for preparing proanthocyanidin microcapsules by complex coagulation self-assembly according to claim 1, characterized in that, In step (2), the amount of proanthocyanidins used is 1.25g, the amount of anhydrous ethanol used is 10mL, the ultrasonic power is 150W, the ultrasonic temperature is 25℃, and the ultrasonic dispersion time is 15min.

4. The method for preparing proanthocyanidin microcapsules by complex coagulation self-assembly according to claim 1, characterized in that, In step (3), the mass ratio of core material to wall material is 1:2, the pH value of the system is adjusted to 4.5, the reaction temperature is 40℃, the stirring speed is 400r / min, and the stirring reaction time is 45min.

5. The method for preparing proanthocyanidin microcapsules by complex coagulation self-assembly according to claim 1, characterized in that, In step (4), the volume of the calcium chloride solution is 200 mL, the mass concentration is 2%, the peristaltic pump drip rate is 1.5 mL / min, the stirring speed is 300 r / min, and the curing time is 3 h.

6. The proanthocyanidin microcapsules prepared by the method for preparing proanthocyanidin microcapsules by complex condensation self-assembly according to any one of claims 1-5 are characterized in that, The microcapsules have a proanthocyanidin encapsulation rate of 85%–95% and exhibit good scavenging ability against DPPH and ABTS free radicals.

7. The application of the proanthocyanidin microcapsules according to claim 6 in the fields of cosmetics, pharmaceuticals, or food preservation.