High-stability sodium caseinate-based quercetin nano-particles, nano quercetin micro-capsule powder and preparation method of nano quercetin nano-particles and nano quercetin micro-capsule powder

Abstract

The invention discloses high-stability sodium caseinate-based quercetin nanoparticles, nano quercetin microcapsule powder and a preparation method of the high-stability sodium caseinate-based quercetin nanoparticles and the nano quercetin microcapsule powder. According to the method, the characteristic that sodium caseinate nano-micelles are fully dissociated under the conditions of high temperature and alkaline pH, so that quercetin crystal powder is introduced, and then a protein-quercetin nano-composite is rapidly formed is utilized, and the efficient nano-embedding effect on quercetin is achieved; the pH is adjusted back to weak acidity, corresponding nano-composites are induced to be further assembled to form nano-particles with compact structures, and the nano-particles can be further subjected to spray drying to obtain the nano quercetin microcapsule powder. According to the quercetin nano-embedding method provided by the invention, the encapsulation efficiency and the loading capacity are relatively high, and the water solubility and the stability of the quercetin are also remarkably improved; the obtained nano quercetin micro-capsule powder has the advantages of good resolubility, high bioavailability of quercetin and the like, so that the nano quercetin micro-capsule powder is expected to have wide application prospects in the fields of nutritional and healthy foods, medicines, feeds and the like.

Description

Technical Field

[0001] The invention belongs to the technical field of polyphenol nano-encapsulation and delivery, and particularly relates to high-stability sodium caseinate-based quercetin nanoparticles and nano-quercetin microcapsule powder and a preparation method thereof. Background Art

[0002] Numerous epidemiological studies have shown that a healthy consumption of fruits and vegetables can effectively prevent chronic diseases such as cardiovascular disease, cancer, stroke, diabetes, Alzheimer's disease, and cataracts, and contributes to maintaining good health. The health-promoting effects of fruits and vegetables are partly attributed to their rich content of flavonoids. Quercetin, one of the most important flavonoid polyphenols, possesses antioxidant, anti-inflammatory, anticancer, antiviral, and anti-cardiovascular effects. However, as a polyphenol, like many other polyphenols, quercetin suffers from poor dispersibility in aqueous environments, low stability, and low bioavailability, which limits its application in food.

[0003] To overcome these shortcomings, many researchers are actively exploring more efficient delivery systems to improve the absorption rate and stability of quercetin. Research on protein-encapsulated delivery of bioactive substances has been a hot topic in recent years. However, due to quercetin's poor chemical stability and sensitivity to alkaline environments, the current methods for introducing quercetin into protein systems are mostly antisolvent and covalent binding methods, but both methods have certain limitations. For example, the antisolvent method often involves the use of organic reagents to dissolve quercetin first when introducing quercetin. The use of organic solvents will limit its application in the food industry, and this method often uses alcohol-soluble proteins, which has certain requirements for the type of protein. The covalent binding method also requires the use of organic solvents when introducing quercetin to provide a quercetin solution dissolved in an organic reagent. Therefore, it is not easy to safely and easily introduce quercetin into proteins without using organic reagents to dissolve quercetin. Among the technologies for encapsulating bioactive substances in proteins, nanosizing is the most effective strategy and technology for improving the bioavailability of quercetin. National invention patent CN118045062A discloses a ternary protein-polysaccharide composite nanoparticle loaded with quercetin. This method uses zein to encapsulate quercetin as the core, and gelatin and carboxymethyl starch to form the shell, creating composite nanoparticles with a core-shell structure for delivering quercetin. The resulting nanoparticles are said to significantly improve the thermal stability, pH stability, photostability, and storage stability of the drug delivery system. However, this method is complex and involves multiple steps, including the preparation of an ethanolic solution of zein and quercetin, the preparation of a gelatin solution, the preparation of a dispersion of gelatin-modified zein nanoparticles loaded with quercetin, the preparation of a carboxymethyl starch solution, and the preparation of a dispersion of gelatin-carboxymethyl starch dual-modified zein nanoparticles loaded with quercetin. National invention patent CN118923859A proposes a method for preparing a high-content quercetin nano-microcapsule emulsion. The emulsion is obtained by high-pressure homogenization using a 70%-77% quercetin ester oil solution, 6%-8% sodium caseinate, 15%-22% isomaltooligosaccharide, 0.3%-0.7% dipotassium hydrogen phosphate, and 0.5%-1% sodium ascorbate. This method is said to not use organic solvents, making the entire process environmentally friendly and safe. It can achieve high quercetin content, reaching 5%-35%, and the solution system is stable, with little precipitation and stratification. Animal experiments have demonstrated that the quercetin nano-microcapsule emulsion prepared by this method has superior lung protective effects (anti-inflammatory, antioxidant, and mucus-reducing) compared to free quercetin powder. The patent does not characterize the stability and bioaccessibility of the nano-microcapsule emulsion. Judging from the existing patent results, compared with free quercetin, nano-encapsulation technology significantly improves the solubility, stability and bioaccessibility of quercetin.

[0004] In summary, the current technologies for delivering quercetin still have many limitations: (1) the preparation system is complex, requiring a variety of raw materials and equipment, and the preparation process is cumbersome; (2) the introduction method is complex (such as glycosylation process, esterification process), involving organic reagents, resulting in safety risks or high requirements for technical conditions, which may not be applicable to the food industry; (3) the loading efficiency of quercetin is poor or the unique health functions of quercetin such as antioxidant properties are ignored. Therefore, this patent uses sodium caseinate, the sodium salt form of casein, the main protein component commonly found in milk in daily life, as an encapsulation material, and utilizes the principle that pH alkaline shift will affect the structure of sodium caseinate, to propose a method for nano-encapsulation of quercetin by inducing protein self-assembly through pH shift and heating treatment. When the pH is far from the isoelectric point of sodium caseinate, the protein molecules unfold or even dissociate into subunits, and the hydrophobic groups inside the protein molecules are exposed to the hydrophilic environment. When quercetin is introduced under this state, quercetin can efficiently and quickly bind to the hydrophobic groups of the protein molecules. After the pH is adjusted back to near neutral, the alkaline-dissociated protein begins to fold back, and quercetin is encapsulated inside the subunits or bound to the surface of the protein molecules. The auxiliary effect of heat treatment can make the protein molecules unfold more completely and fully under alkaline conditions, exposing more hydrophobic groups, thereby improving the loading efficiency of quercetin and improving the thermal stability of the complex.

[0005] This method aims to modify sodium caseinate in a greener and safer manner using a simple, low-energy process, while also rationally introducing quercetin to prepare quercetin nanoparticles and nanoquercetin microcapsules with high loading capacity, high stability, and high bioavailability. This method is suitable for large-scale industrial production and processing, and has promising applications in pigments, feed, food, pharmaceuticals, and other fields. Summary of the Invention

[0006] The present invention aims to provide highly stable sodium caseinate-based quercetin nanoparticles and nanoquercetin microcapsule powder, and methods for their preparation, to address the shortcomings and problems of existing technologies. The nanoparticle preparation method overcomes the traditional view that quercetin is more susceptible to oxidation under high temperature and alkaline conditions. Instead, it utilizes the characteristics of "hot alkaline" conditions, which facilitate the introduction of quercetin crystals, and the rapid nanocomposite formation of sodium caseinate with quercetin, to achieve efficient nanoencapsulation of quercetin. The nanoparticle solution can also be spray-dried to produce the nanoquercetin microcapsule powder.

[0007] The first object of the present invention is to provide a method for preparing sodium caseinate-based quercetin nanoparticles, which has a simple preparation system, requires only quercetin and sodium caseinate as raw materials, and does not require large-scale experimental equipment. This method does not require the assistance of additional sugars or lipids and can be achieved only by pH regulation. The pH shift is achieved by adjusting the pH of the protein solution so that the pH of the liquid environment is away from the isoelectric point of the protein. As a result, the protein structure unfolds, exposing the hydrophobic active groups buried inside, providing quercetin with the opportunity to bind to it. When the pH is adjusted back to around neutral, the protein will refold back to a state similar to its natural structure, and quercetin will therefore be embedded in the interior of the protein.

[0008] A second objective of the present invention is to provide quercetin nanoparticles with high quercetin loading and high stability. These nanoparticles have a quercetin encapsulation efficiency of up to 98.01%, a loading of 24.50 mg / g protein, an average particle size of less than 200 nm, and a bioaccessibility of over 30%.

[0009] A third objective of the present invention is to provide a method for producing a protein-based quercetin nanocapsule powder that is highly bioaccessible, simple to introduce, requires no organic reagents, poses no safety risks, and is suitable for production in the food industry. This method involves the addition of only hydrochloric acid, sodium hydroxide, and carbohydrates, without the addition of any organic reagents, making it suitable for large-scale food industry production.

[0010] The purpose of the present invention is achieved through the following technical solutions:

[0011] The present invention fully utilizes the characteristic that "the nanomicelles of sodium caseinate fully dissociate under high temperature and alkaline pH conditions, thereby introducing quercetin crystal powder and rapidly forming a protein-quercetin nanocomposite", thereby achieving efficient nano-encapsulation of quercetin; by adjusting the pH to weak acidity, the corresponding nanocomposite is induced to further assemble into compact nanoparticles, which can be further spray-dried to obtain nano-quercetin microcapsule powder.

[0012] A method for preparing highly stable sodium caseinate-based quercetin nanoparticles comprises the following steps:

[0013] 1) dispersing sodium caseinate in deionized water and hydrating to obtain a protein solution;

[0014] 2) Adjust the pH of the protein solution to alkaline, keep stirring and maintain alkalinity;

[0015] 3) heating the alkaline sodium caseinate solution;

[0016] 4) adding quercetin powder to the dissociated protein solution and maintaining the alkaline state for a period of time to obtain a complex solution;

[0017] 5) Adjust the pH value of the complex solution to weak acidity and keep stirring until the solution is stable;

[0018] 6) Centrifuging the complex solution to remove unbound free quercetin and obtain sodium caseinate-based quercetin nanoparticles.

[0019] Furthermore, the concentration of the sodium caseinate solution in step 1) is 0.1 wt% to 3.0 wt%; preferably, the concentration of the sodium caseinate solution is 1.0 wt% to 2.0 wt%.

[0020] Furthermore, the alkaline pH value of step 2) is 9.0 to 12.0; preferably, the alkaline pH value is 10.0 to 11.0. In the present invention, the quercetin nanoparticles obtained under alkaline pH 11 conditions have the highest encapsulation efficiency and loading capacity.

[0021] Furthermore, the temperature of the heating treatment in step 3) is 80-100° C., and then the mixture is quickly cooled to room temperature with ice water.

[0022] Furthermore, in step 4), the mass ratio of the added quercetin powder to the mass ratio of the protein in the protein solution should be 1% to 5%; preferably, the mass ratio of the added quercetin powder to the mass ratio of the protein in the protein solution should be 2% to 4%.

[0023] Furthermore, in step 5), the pH range of the complex solution adjusted to a weakly acidic state is 5.0 to 7.0; preferably, the pH range of the complex solution adjusted to a weakly acidic state is 5.5 to 6.5.

[0024] Furthermore, in step 6), the centrifugal speed of the complex solution is 3000-6000 g, and the centrifugal time is 10-30 min.

[0025] A method for preparing nano-quercetin microcapsule powder comprises the following steps:

[0026] The high-stability sodium caseinate-based quercetin nanoparticles are added to a spray drying aid and spray-dried to obtain nano-quercetin microcapsule powder.

[0027] Furthermore, the spray drying aid is at least one of maltodextrin, sodium alginate and octenylsuccinate modified starch; and the added mass ratio of the spray drying aid to the protein in the high-stability sodium caseinate-based quercetin nanoparticles is 1:1 to 1:2.

[0028] The method is used to prepare sodium caseinate-based quercetin nanoparticles and nanoquercetin microcapsule powder, which have a simple preparation system, require only quercetin and sodium caseinate as raw materials, do not require large-scale experimental equipment, have a simple quercetin introduction method, do not require the addition of organic reagents, and pose no safety risks. The nanoparticles are suitable for food industry production and have high quercetin loading, high chemical stability, and high bioaccessibility.

[0029] The principle of the present invention is as follows: Quercetin is a flavonoid polyphenol compound with multiple phenolic hydroxyl groups, which makes it extremely insoluble in water, but it has multiple hydrophobic sites and can combine with hydrophobic substances. Casein is a common protein in milk. It has external hydrophilic groups and internal hydrophobic groups. In an alkaline pH environment far away from the isoelectric point, it will dissociate and the subunits will unfold due to the electrostatic repulsion, exposing the internal hydrophobic groups and hydrophobic sites. At this time, when quercetin powder is added, quercetin can quickly combine with the hydrophobic sites of casein. In the process of returning the complex solution to a weak acidic state, the casein folds back to the same state as the natural structure, and quercetin is therefore embedded in the interior of the casein.

[0030] Compared with the prior art, the present invention has the following beneficial effects:

[0031] (1) The present invention utilizes the structural characteristics of proteins and the properties of quercetin to propose for the first time a method for pH alkaline shift-induced protein assembly nano-encapsulation of quercetin, encapsulating quercetin inside the denatured and folded protein structure, greatly improving the water solubility and stability of quercetin.

[0032] (2) The present invention directly adds quercetin in the form of solid powder, has no special requirements for the ambient temperature, and does not require the addition of organic solvents or chaotropic agents. A highly stable and high-concentration quercetin protein solution can be obtained only by pH control and heating treatment. The production process and materials meet food safety requirements.

[0033] (3) The present invention uses heat treatment to assist pH shift in modifying sodium caseinate, so that the structure of sodium caseinate unfolds more thoroughly, exposing more hydrophobic active sites for quercetin to be combined with. The method is simple, does not require the addition of organic solvents or other additives, meets the requirements of food industry production, is safe and harmless, and fully realizes the potential of sodium caseinate to combine with active substances.

[0034] (4) The sodium caseinate-based quercetin nanoparticles and nanoquercetin microcapsule powder prepared by the method of the present invention can improve the water dispersibility, thermal stability, light stability, storage stability, antioxidant capacity and bioavailability of quercetin. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0036] Figure 1 This is the appearance of the sodium caseinate-based quercetin nanoparticle solutions pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, p12SC-Q6) obtained by treatment with different alkalinity in Example 1.

[0037] Figure 2 This is a diagram showing the particle size of sodium caseinate-based quercetin nanoparticles pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, p12SC-Q6) obtained by treatment with different alkalinity in Example 1.

[0038] Figure 3 1 is a particle size distribution diagram of sodium caseinate-based quercetin nanoparticles pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, p12SC-Q6) obtained by treatment with different alkalinity in Example 1.

[0039] Figure 4 The thermal stability of sodium caseinate-based quercetin nanoparticles pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, p12SC-Q6) obtained by treatment with different alkalinity in Example 1.

[0040] Figure 5 This is a diagram showing the particle size of sodium caseinate quercetin nanoparticles (PH10SC-Q, PH15SC-Q, PH20SC-Q, PH25SC-Q) obtained by heating for different times to assist pH shift in Example 2.

[0041] Figure 6 The thermal stability of sodium caseinate quercetin nanoparticles (PH10SC-Q, PH15SC-Q, PH20SC-Q, PH25SC-Q) obtained by heating for different times to assist pH shift in Example 2.

[0042] Figure 7 The appearance and reconstituted solution of the four nano-quercetin microcapsule powders (QC, Q-MD, QT, and Q-OSAS) prepared in Example 3 are shown.

[0043] Figure 8 The redissolution solubility of the four nano-quercetin microcapsule powders (QC, Q-MD, QT, and Q-OSAS) prepared in Example 3.

[0044] Figure 9 The bioaccessibility of the four nano-quercetin microcapsule powders (QC, Q-MD, QT, and Q-OSAS) prepared in Example 3 after simulated in vitro digestion. DETAILED DESCRIPTION

[0045] The present invention is further described in detail below through specific examples.

[0046] It should be understood that these examples are intended to illustrate the present invention and are not intended to limit the scope of the present invention. In addition, for the numerical ranges in the present invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Each smaller range between the intermediate value within any stated value or stated range and any other stated value or intermediate value within the range is also included in the present invention. The upper and lower limits of these smaller ranges may be independently included or excluded within the scope.

[0047] In the following examples, experimental methods without specific conditions are generally performed under conventional conditions or according to the conditions recommended by the manufacturer. Unless otherwise specified, the materials and reagents used in these examples were obtained from commercial sources.

[0048] Example 1

[0049] A high-stability sodium caseinate-based quercetin nanoparticle and a preparation method thereof, comprising the following steps:

[0050] (1) Dissolve 2 g of sodium caseinate powder in 100 ml of distilled water to prepare a 20 mg / mL protein solution. Stir magnetically for 2 h and then hydrate overnight in a 4°C refrigerator. The next day, remove the solution from the refrigerator, return it to room temperature, and centrifuge it (5000 g, 20 min) to remove insoluble impurities.

[0051] (2) The pH of the protein solution was adjusted to 9.0, 10.0, 11.0, and 12.0 with 2 M NaOH and magnetic stirring was maintained for 90 min. Then, 0.05 g of quercetin was added to the dissociated sodium caseinate solution to achieve a quercetin concentration of 0.5 mg / mL.

[0052] (3) After magnetic stirring for 30 seconds in the dark, the pH of the solution was adjusted to 6.0 with 2M HCl and magnetic stirring was continued for 1 hour to allow the solution to stabilize. The resulting solution was centrifuged at 5000g for 20 minutes to remove unbound quercetin. The supernatant was the sodium caseinate-based quercetin nanoparticles pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, and p12SC-Q6) prepared under different alkalinity treatments.

[0053] In this example, sodium caseinate was used as the material to encapsulate quercetin. The main purpose was to explore the differences in appearance, encapsulation efficiency and loading capacity, particle size and distribution, and stability of sodium caseinate-based quercetin nanoparticles prepared under different alkalinity treatments.

[0054] Performance testing:

[0055] The appearance of sodium caseinate-based quercetin nanoparticle solutions pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, p12SC-Q6) obtained by treatment with different alkalinity is shown in the figure below. Figure 1 shown.

[0056] Depend on Figure 1 As can be seen, the sodium caseinate-based quercetin nanoparticle solutions obtained after treatment with different alkalinity levels all appeared as stable, transparent liquids, indicating the formation of nanoscale particles. At pH 11 and 12, the complex color was darker, orange-brown. Quercetin has low solubility at pH 9 and 10, resulting in a yellow solution color. At pH 11, the composite nanoparticle solution obtained was the darkest, indicating that sodium caseinate bound the most quercetin. This may be because quercetin is sensitive to alkaline environments. A pH 12 environment causes quercetin to degrade rapidly, resulting in a reduced binding rate with sodium caseinate.

[0057] The quercetin encapsulation efficiency and loading capacity of sodium caseinate-based quercetin nanoparticles pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, p12SC-Q6) obtained by treatment with different alkalinity are shown in Table 1.

[0058] Table 1 Quercetin encapsulation efficiency and loading amount of sodium caseinate-based quercetin nanoparticles obtained by different alkalinity treatments

[0059]

[0060] As shown in Table 1, when the alkalinity was pH 9, pH 10, pH 11 and pH 12, the encapsulation efficiency of the prepared sodium caseinate-based quercetin nanoparticles for quercetin was 34.62%, 55.66%, 86.49% and 75.86%, respectively, and the loading amounts were 8.66 mg / g protein, 13.91 mg / g protein, 21.62 mg / g protein and 18.97 mg / g protein, respectively.

[0061] The particle sizes of sodium caseinate-based quercetin nanoparticles pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, p12SC-Q6) obtained by treatment with different alkalinity are as follows: Figure 2 As shown, the particle size distribution is Figure 3 shown.

[0062] Depend on Figure 2 and Figure 3 It can be seen that the average diameter of sodium caseinate-based quercetin nanoparticles obtained by different alkalinity treatments decreased significantly with the increase of pH of alkaline treatment, from 219.7 nm to 152.7 nm. This may be because the farther the pH value is from the isoelectric point of sodium caseinate, the stronger the electrostatic repulsion in the solution, resulting in greater changes in the SC structure and the formation of more compact and smaller particles during the reorganization and folding process.

[0063] The thermal stability of sodium caseinate-based quercetin nanoparticles pSC-Q6 (p9SC-Q6, p10SC-Q6, p11SC-Q6, p12SC-Q6) obtained by treatment with different alkalinity is shown in Figure 2. Figure 4 shown.

[0064] Depend on Figure 4 As can be seen, after heating at 65°C for 6 hours, the quercetin retention rates in the four sodium caseinate-based quercetin nanoparticles prepared in this example, obtained by treating with different alkalinity levels, were all above 35%, while the free quercetin content remained less than 20%. This indicates that the sodium caseinate-based quercetin nanoparticles prepared in this example have good thermal stability.

[0065] Example 2

[0066] A high-stability sodium caseinate-based quercetin nanoparticle and a preparation method thereof, comprising the following steps:

[0067] (1) Dissolve 2 g of sodium caseinate powder in 100 ml of distilled water to prepare a 20 mg / mL protein solution. Stir magnetically for 2 h and then hydrate overnight in a 4°C refrigerator. The next day, remove the solution from the refrigerator, return it to room temperature, and centrifuge it (5000 g, 20 min) to remove insoluble impurities.

[0068] (2) Adjust the pH of the protein solution to 11.0 with 2 M NaOH and maintain magnetic stirring for 90 min. Then, heat the dissociated sodium caseinate solution in a 95°C water bath for 10 min, 15 min, 20 min, and 25 min, respectively. After removal, quickly cool to room temperature with ice water.

[0069] (3) Add 0.05 g of quercetin to the sodium caseinate solution dissociated and heated in step (2) to achieve a quercetin concentration of 0.5 mg / mL.

[0070] (4) After magnetic stirring for 30 seconds in the dark, the pH value of the solution was adjusted to 6.0 with 2M HCl and magnetic stirring was continued for 1 hour to allow the solution to stabilize. The resulting solution was centrifuged at 5000g for 20 minutes to remove unbound quercetin. The supernatant was the sodium caseinate quercetin nanoparticles obtained by treating with different heating times to assist pH shift, and they were named PH10SC-Q, PH15SC-Q, PH20SC-Q, and PH25SC-Q according to the heating time. The sodium caseinate quercetin nanoparticles obtained without heating treatment were named PCSC and served as a control.

[0071] This example carried out a heating treatment based on Example 1, the main purpose of which was to explore the effect of heating-assisted pH shift on the quercetin encapsulation effect of modified sodium caseinate.

[0072] Performance testing:

[0073] The encapsulation efficiency and loading capacity of sodium caseinate-based quercetin nanoparticles (PH10SC-Q, PH15SC-Q, PH20SC-Q, PH25SC-Q) obtained by treating with different heating times to assist pH shift are shown in Table 2.

[0074] Table 2 Quercetin encapsulation efficiency and loading amount of sodium caseinate-based quercetin nanoparticles obtained by different heating time treatments to assist pH shift

[0075]

[0076] As can be seen from Table 2, the sodium caseinate-based quercetin nanoparticles obtained by treating with different heating times to assist pH shift have high encapsulation efficiency and loading capacity. When the heating time is 10 min, 15 min, 20 min and 25 min, the encapsulation efficiency of the prepared sodium caseinate-based quercetin nanoparticles for quercetin is 83.92%, 89.78%, 86.83% and 78.28%, respectively, and the loading capacity is 20.98 mg / g protein, 22.45 mg / g protein, 21.71 mg / g protein and 19.57 mg / g protein, respectively.

[0077] The particle sizes of sodium caseinate-based quercetin nanoparticles (PH10SC-Q, PH15SC-Q, PH20SC-Q, PH25SC-Q) obtained by pH shift assisted by heating time are as follows: Figure 5 shown.

[0078] Depend on Figure 5It can be seen that as the heating time increases, the average diameter of the prepared sodium caseinate quercetin nanoparticles shows a trend of first decreasing and then increasing. This indicates that the input of heat can promote the further collapse of the protein structure caused by pH shift, and the protein molecules are dissociated into smaller dispersions, which then fold back into more compact protein particles. At this time, further heating will cause the originally cleaved polypeptide chains to form aggregates again under the influence of thermal energy, and the particle size will increase significantly. The particle sizes of sodium caseinate quercetin nanoparticles PCSC, PH10SC-Q, PH15SC-Q, PH20SC-Q, and PH25SC-Q prepared without heat treatment and with different heating time treatments are 156.34nm, 166.8nm, 155.82nm, 167.8nm, and 172.7nm, respectively (the particle size data results are the average of three measurements).

[0079] The thermal stability of sodium caseinate quercetin nanoparticles (PH10SC-Q, PH15SC-Q, PH20SC-Q, PH25SC-Q) obtained by pH shift assisted by heating time is shown in Figure 2. Figure 6 shown.

[0080] Depend on Figure 6 As can be seen, after heating at 65°C for 6 hours, the quercetin retention rates in the five sodium caseinate-based quercetin nanoparticles prepared in this example, obtained by heating for different times to assist pH shift, were all above 60%, demonstrating a better quercetin protection effect than the nanoparticles obtained by pH shift alone. This indicates that appropriate heating-assisted pH shift can produce sodium caseinate-based quercetin nanoparticles with improved thermal stability.

[0081] Example 3

[0082] A nano-quercetin microcapsule powder and a preparation method thereof, comprising the following steps:

[0083] (1) Dissolve 2 g of sodium caseinate powder in 100 ml of distilled water to prepare a 20 mg / mL protein solution. Stir magnetically for 2 h and then hydrate overnight in a 4°C refrigerator. The next day, remove the solution from the refrigerator, return it to room temperature, and centrifuge it (5000 g, 20 min) to remove insoluble impurities.

[0084] (2) Adjust the pH of the protein solution to 11.0 with 2M NaOH and maintain magnetic stirring for 90 minutes. Then, heat the dissociated sodium caseinate solution in a 95°C water bath for 15 minutes and quickly cool it to room temperature with ice water.

[0085] (3) Add 0.05 g of quercetin to the sodium caseinate solution dissociated and heated in step (2) to achieve a quercetin concentration of 0.5 mg / mL.

[0086] (4) After magnetic stirring for 30 seconds in the dark, the pH of the solution was adjusted to 6.0 with 2M HCl and magnetic stirring was continued for 1 hour to allow the solution to stabilize. The resulting solution was centrifuged at 5000 g for 20 minutes to remove unbound quercetin. The supernatant was the sodium caseinate-based quercetin nanoparticles obtained by heat-assisted pH shift.

[0087] (5) The sodium caseinate-based quercetin nanoparticles obtained in step (4) were evenly divided into four portions, and four groups of spray-dried dispersions were prepared under magnetic stirring conditions:

[0088] a) No additives are added, recorded as QC;

[0089] b) adding a certain amount of maltodextrin (MD) to the sodium caseinate-based quercetin nanoparticle solution prepared above so that the ratio of maltodextrin mass to protein mass in the solution is 1:1. Stirring is continued for 30 minutes to obtain a maltodextrin-protein-based quercetin complex solution, which is recorded as Q-MD;

[0090] c) adding a certain amount of sodium alginate (T) to the sodium caseinate-based quercetin nanoparticle solution prepared above so that the ratio of the mass of sodium alginate to the mass of protein in the solution is 1:1. Stirring is continued for 30 minutes to obtain a sodium alginate-protein-based quercetin complex solution, which is recorded as QT;

[0091] d) adding a certain amount of octenyl succinate starch (OSAS) to the sodium caseinate-based quercetin nanoparticle solution prepared above so that the ratio of the octenyl succinate starch mass to the protein mass in the solution is 1:1. Stirring is continued for 30 minutes to obtain an octenyl succinate starch-protein-based quercetin complex solution, which is recorded as Q-OSAS;

[0092] (6) The dispersions were dried using a Büchi B290 spray dryer. The specific conditions were: nozzle diameter 0.7 mm, inlet temperature 160°C, feed rate 3.5 mL / min, and airflow rate 667 L / h. The powders obtained from the four dispersions were also designated as QC, Q-MD, QT, and Q-OSAS. The powders were collected, sealed in brown bottles, and stored in a desiccator containing phosphorus pentoxide in the dark.

[0093] The main purpose of this embodiment is to achieve the powder production of sodium caseinate-based quercetin nanoparticles to obtain quercetin powder with stable particle properties, excellent solubility and high bioaccessibility.

[0094] Performance testing:

[0095] The appearance and reconstitution solutions of the four prepared nanoquercetin microcapsules (QC, Q-MD, QT, Q-OSAS) are as follows: Figure 7 shown.

[0096] Depend on Figure 7 As can be seen, the powder appears golden or orange-yellow overall, preserving the yellow color of quercetin. Sample Q-MD, prepared using maltodextrin as a spray-drying aid, exhibits the darkest color and the highest quercetin content. The powder is uniform and fine, and exhibits good resolubility. The reconstituted solutions all appear as clear, transparent orange-yellow solutions, demonstrating that the quercetin powder prepared by spray drying has good water solubility and can form a uniform and stable system.

[0097] The re-dissolution solubility of the four prepared nano-quercetin microcapsule powders (QC, Q-MD, QT, Q-OSAS) is as follows: Figure 8 shown.

[0098] Depend on Figure 8 As can be seen, QC has the lowest solubility. In contrast, the solubility of microcapsules prepared with the addition of carbohydrates ranges from 70% to 90%, indicating that the use of carbohydrates significantly improves the solubility of the powder. The difference in solubility of nano-microcapsules prepared with different wall materials is related to the type of carbohydrate added. Among them, the microcapsules prepared with MD as an additive showed the highest solubility, exceeding 90%, while the solubility of microcapsules prepared with the other two carbohydrates as additives was slightly worse, but still higher than when no carbohydrates were added.

[0099] The bioaccessibility of the four prepared nanoquercetin microcapsules (QC, Q-MD, QT, and Q-OSAS) after in vitro simulated digestion is shown in Figure 2. Figure 9 shown.

[0100] Bioaccessibility generally refers to the ability of bioactive substances to transfer from the food matrix to mixed micelles during digestion. Figure 9The bioaccessibility of free quercetin is only 3.57%. However, the bioaccessibility of quercetin encapsulated with sodium caseinate is significantly improved after microencapsulation. The bioaccessibility of QC reaches 22.20%, approximately 6.2 times that of free quercetin. The addition of a carbohydrate adjuvant further enhances the bioaccessibility of quercetin in the microcapsules. The bioaccessibility of quercetin in Q-MD, QT, and Q-OSAS microcapsules reaches 43.23%, 40.89%, and 51.82%, respectively, approximately 12.1, 11.5, and 14.5 times that of free quercetin. This suggests that sodium caseinate-based quercetin nanoparticles, after spray drying to form powder particles, exhibit high stability and sustained-release properties, potentially enabling colonic delivery and release. Therefore, spray drying is an effective method for preparing water-soluble quercetin microcapsules.

[0101] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be considered as equivalent replacement methods and are included in the scope of protection of the present invention.

Claims

1. A method for preparing high-stability sodium caseinate-based quercetin nanoparticles, characterized in that: The method utilizes the characteristics of sodium caseinate nanomicelles that fully dissociate under high temperature and alkaline pH conditions to introduce quercetin crystal powder and then quickly form a protein-quercetin nanocomposite, thereby achieving efficient nano-encapsulation of quercetin; by adjusting the pH back to weak acidity, the corresponding nanocomposite is induced to further assemble to form sodium caseinate-based quercetin nanoparticles with a compact structure; The specific steps are as follows: 1) dispersing sodium caseinate in deionized water and hydrating to obtain a protein solution; 2) Adjust the pH of the protein solution to alkaline, keep stirring and maintain alkalinity; 3) heating the alkaline sodium caseinate solution; 4) adding quercetin powder to the dissociated protein solution and maintaining the alkaline state for a period of time to obtain a complex solution; 5) Adjust the pH value of the complex solution to weak acidity and keep stirring until the solution is stable; 6) Centrifuging the complex solution to remove unbound free quercetin and obtain sodium caseinate-based quercetin nanoparticles.

2. The preparation method according to claim 1, characterized in that The concentration of the protein solution in step 1) is 0.1 wt% to 3.0 wt%.

3. The preparation method according to claim 1, characterized in that In step 2), the pH value of the alkaline treatment is 9.0 to 12.

0.

4. The preparation method according to claim 1, characterized in that In step 3), the temperature of the heating treatment is 80-100° C., and then the mixture is rapidly cooled to room temperature with ice water.

5. The preparation method according to claim 1, characterized in that In step 4), the mass ratio of the added quercetin crystal powder to the mass ratio of the protein in the protein solution should be 1% to 5%; and the time for maintaining the alkaline state is 10 seconds to 30 minutes.

6. The preparation method according to claim 1, characterized in that In step 5), the pH of the complex solution is adjusted back to a weakly acidic state within a range of 5.0 to 7.

0.

7. High-stability sodium caseinate-based quercetin nanoparticles prepared by the preparation method according to any one of claims 1 to 6.

8. A method for preparing nano-quercetin microcapsule powder, characterized in that: The method comprises the following steps: adding the high-stability sodium caseinate-based quercetin nanoparticles according to claim 6 to a spray drying adjuvant and performing spray drying to obtain nano-quercetin microcapsule powder.

9. The preparation method according to claim 8, characterized in that: The spray drying aid is at least one of maltodextrin, sodium alginate and octenylsuccinate modified starch; the added mass ratio of the spray drying aid to the protein in the high-stability sodium caseinate-based quercetin nanoparticles is 1:1 to 1:

2.

10. A nano-quercetin microcapsule powder prepared by the preparation method according to claim 8 or 9, characterized in that: The corresponding embedded quercetin has good water solubility, thermal stability and bioavailability.

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