Water-soluble fucoxanthin and preparation method thereof

Abstract

The present invention relates to a water-soluble fucoxanthin and a preparation method thereof. The water-soluble fucoxanthin is a water-soluble particle formed by self-assembly of fucoxanthin, fatty acids, bovine serum albumin, and water-soluble chitosan. The preparation method comprises the following steps: (1) preparing water-soluble chitosan; (2) preparing a fucoxanthin-fatty acid mixture; (3) preparing alkaline electrolyte water; (4) preparing a fucoxanthin-protein complex by electrolysis water coupling ultrasound technology; (5) preparing a chitosan-protein-fatty acid-fucoxanthin complex; and (6) obtaining water-soluble fucoxanthin. The present invention adopts alkaline electrolysis water coupling ultrasound technology and non-covalent self-assembly technology to prepare water-soluble fucoxanthin with good stability and high absorption rate. The preparation process is simple, the embedding rate is high, the safety is good, the production cost is low, the fucoxanthin is suitable for industrial production, and the fucoxanthin can be widely used in food systems as a nutritional enhancer.

Description

Technical Field

[0001] The present invention relates to the technical field of food processing, and in particular to water-soluble fucoxanthin and a preparation method thereof. Background Art

[0002] Fucoxanthin is a marine carotenoid extracted from seaweed and microalgae, accounting for over 10% of the total natural carotenoid production. Numerous studies have demonstrated that fucoxanthin exhibits diverse functional properties, including antioxidant, anti-inflammatory, anti-obesity, and cardiovascular disease prevention, and holds broad application prospects in the food industry. However, as a lipid-soluble nutritional active substance, fucoxanthin suffers from shortcomings such as poor water solubility, instability under light / heat and acid-base conditions, and limited bioavailability. Therefore, simultaneously improving fucoxanthin's solubility, processing stability, and bioavailability remains a pressing technical challenge in developing fucoxanthin-based nutritional supplements or functional foods.

[0003] Currently, researchers at home and abroad have begun utilizing microemulsions, nanoemulsions, Pickering emulsions, and nanocomposite carriers to encapsulate carotenoids. Dai et al. prepared an oil-in-water emulsion loaded with fucoxanthin using medium-chain triglycerides as the oil phase and 1-α-phosphatidylcholine as the emulsifier (Dai & Kim, 2016). Ma et al. employed direct microchannel emulsification technology to investigate the formulation and stability characteristics of a monodisperse oil-in-water emulsion encapsulating fucoxanthin using medium-chain triglycerides, whey protein isolate, and Tween 20 as raw materials (Ma et al., 2020). Li et al. prepared caseinate-stabilized zein particles encapsulating fucoxanthin using a simple antisolvent precipitation method at neutral pH (Li et al., 2018). Current fucoxanthin delivery systems typically employ high-energy methods and chemical reagents to produce relatively small particles, large loading capacity, and good stability. However, these methods still have some defects: (1) There are safety risks, as strong acids and alkalis and organic solvent residues are added to the products; (2) Absorption is difficult, as the delivery vector is released too quickly or incompletely in the intestine, causing intestinal absorption disorders; (3) Production efficiency is low, and there is a large loss of fucoxanthin during the production process.

[0004] Natural macromolecules are green, environmentally friendly, and widely available, and have been widely used in delivery vehicle wall materials in recent years. Zhu et al. constructed and characterized a whey protein complex loaded with fucoxanthin using bovine serum albumin, β-lactoglobulin, and α-lactoglobulin (Zhu, Sun, Wang, Xu, & Wang, 2017). Wang et al. encapsulated fucoxanthin in carboxymethyl konjac glucomannan-coated gliadin nanoparticles, which improved the stability of fucoxanthin against both photo- and thermal degradation and exhibited excellent colon-targeting properties (Wang, Wei, & Xue, 2022). However, fucoxanthin is heat-sensitive and easily oxidized, and its structure and activity are somewhat damaged at high temperatures. Furthermore, natural macromolecules such as proteins and polysaccharides are good microbial culture media, making them highly susceptible to microbial growth and spoilage. High-temperature sterilization or the addition of preservatives is often required to extend the shelf life of food. It is worth noting that alkaline electrolyzed water is produced by placing electrolyte-added water in an electrolysis device and subjecting it to an electric field, which modifies the water's pH, available chlorine concentration, redox potential, and other parameters. Alkaline electrolyzed water has a high redox potential, exhibits certain antibacterial properties, and is highly permeable, safe, and low-cost. As a type of electro-functional water, electrolyzed water remains essentially water. After exposure to air, light, oxygen, and organic matter, its pH and ORP levels remain unchanged, and all parameters gradually approach those of the original water. Studies have shown that the application of alkaline electrolyzed water in konjac gel and Chongqing noodles has great potential. Li et al. combined alkaline electrolyzed water with ultrasonic treatment to improve the functional properties and digestibility of Antarctic krill protein. Furthermore, chitosan is a natural aminocationic polysaccharide biopolymer that not only possesses a variety of activities such as biocompatibility, biodegradability, and non-toxicity, but also has a wide range of applications in the field of nutrient delivery due to its unique properties. However, chitosan's inherent insolubility at neutral or high pH limits its application under these pH conditions. It is generally believed that the Maillard reaction occurs between sugars and proteins, but it is noteworthy that the presence of free amino groups in chitosan makes it a candidate for reaction with the carbonyl groups of reducing sugars, enabling it to participate in the Maillard reaction and increasing its pH solubility range. Summary of the Invention

[0005] In response to the difficulties in industrial application of fucoxanthin due to its poor solubility in water, poor stability and low absorption rate, the purpose of the present invention is to disclose a method for preparing fucoxanthin microparticles with good water solubility, strong stability and high absorption rate through the innovative use of alkaline electrolysis water coupled with ultrasound technology and non-covalent self-assembly technology. This processing method is low-cost, and the prepared water-soluble fucoxanthin microparticles are highly safe and can be widely used in food systems.

[0006] The present invention adopts alkaline electrolyzed water coupled ultrasonic self-assembly technology to prepare water-soluble fucoxanthin microparticles, which not only overcomes the defects of traditional nanoemulsions and does not require the addition of additional antibacterial agents and stabilizers, but also avoids the use of chemical reagents such as organic solvents, strong acids / alkalis, surfactants, etc. that have food safety risks. There are no toxic and harmful substances remaining, which fully guarantees the safety of the product. At the same time, during the production process, no unit operations such as high-pressure homogenization and high-speed shearing are required, the production cost is low, and it is suitable for industrial production.

[0007] Alkaline electrolyzed water has a small molecular cluster structure with low surface tension, strong permeability, and high redox potential. It can effectively inhibit the oxidative destruction of carotenoids during the self-assembly process and has a certain antibacterial effect. It is an ideal reaction medium. As an electro-functional water, the essence of electrolyzed water is still water. After being exposed to the air and contacted with light, oxygen, and organic matter, its pH value and ORP have no residual properties, and various indicators gradually approach those of the original water. Conventional alkaline water (such as NaOH aqueous solution) has larger molecular clusters and does not have reducing properties, which makes carotenoids dissolved in conventional alkaline water very susceptible to oxidative destruction. In addition, when restoring the pH of conventional alkaline water to neutral, a large amount of strong acid reagents are required, which does not meet the requirements of green and environmentally friendly production.

[0008] Compared with other nanoparticles, the water-soluble fucoxanthin microparticles prepared by the present invention do not need to be sterilized at high temperature, and the microparticle powder is also very stable, which is convenient for storage and transportation; compared with protein carriers, the introduction of glycosylated modified chitosan can delay the release of fucoxanthin in the gastrointestinal tract, and coupled with its good absorption-promoting effect, it has a wide range of uses in the food and biomedicine fields.

[0009] The specific plan is as follows:

[0010] A method for preparing water-soluble fucoxanthin, characterized in that the specific preparation method comprises the following steps:

[0011] (1) Preparation of water-soluble chitosan powder: chitosan, acetic acid solution, and reducing sugar are mixed and subjected to Maillard reaction. After the reaction, the pH is adjusted and the mixture is dried to obtain the water-soluble chitosan powder;

[0012] (2) Preparation of fucoxanthin-fatty acid mixture: fucoxanthin, ethanol and fatty acid are uniformly mixed to dissolve the fucoxanthin to obtain the fucoxanthin-fatty acid mixture;

[0013] (3) Preparation of alkaline electrolyte water: using carbonate as electrolyte, ionizing the aqueous solution of the electrolyte, and preparing alkaline electrolyte water at the anode;

[0014] (4) Preparation of a fucoxanthin-protein complex aqueous solution by electrolysis water coupled with ultrasonic technology: protein powder is mixed with the alkaline electrolyte water to obtain a protein solution; the fucoxanthin-fatty acid mixture is then added to the protein solution to obtain a mixed solution; the mixed solution is then ultrasonically treated to allow the fucoxanthin, the fatty acid, and the protein in the mixed solution to self-assemble; after the reaction, the mixture is left open at room temperature to obtain the fucoxanthin-protein complex aqueous solution;

[0015] (5) Preparation of chitosan-protein-fatty acid-fucoxanthin complex aqueous solution: the water-soluble chitosan powder prepared in step (1) is mixed with the fucoxanthin-protein complex aqueous solution prepared in step (4), and the reaction is carried out to combine the chitosan with the protein shell of the complex through non-covalent self-assembly to obtain the chitosan-protein-fatty acid-fucoxanthin complex aqueous solution;

[0016] (6) Obtaining water-soluble fucoxanthin dry powder: drying the chitosan-protein-fatty acid-fucoxanthin complex aqueous solution to obtain the water-soluble fucoxanthin dry powder.

[0017] Furthermore, the reducing sugar in step (1) includes at least one of glucose, fructose, galactose, xylose or mannose;

[0018] Optionally, chitosan, acetic acid solution, and reducing sugar are mixed so that the mass ratio of chitosan to reducing sugar in the solution is (1-2):(1-3), and the pH of the reaction system after mixing is 5.0-6.0;

[0019] Optionally, the Maillard reaction is carried out at a temperature of 60-70° C. and for a time of 48 to 96 hours.

[0020] Furthermore, the fatty acid in step (2) includes at least one of myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid or docosahexaenoic acid; the mass ratio of the fucoxanthin to the fatty acid is 1:(3-5); and the fatty acid in the water-soluble fucoxanthin has a significant absorption-promoting effect on the fucoxanthin.

[0021] Furthermore, the ionization in step (3) is carried out at a voltage of 220 V for 15-30 minutes, and the pH value of the obtained alkaline electrolyte water is 10.0-12.0, and the redox potential is -750--650 mV;

[0022] Preferably, the pH value is 10.5 to 11.5, and the redox potential is -710 to -700 mV;

[0023] More preferably, the pH value is 11 and the redox potential is -706 mV.

[0024] Furthermore, the mass ratio of the fucoxanthin to the protein powder in the mixed solution in step (4) is (0.5-1.5):(15-25);

[0025] Preferably, the mass ratio of the fucoxanthin to the protein powder in the mixed solution is (0.8-1.2):(18-22);

[0026] More preferably, the mass ratio of the fucoxanthin to the protein powder in the mixed solution is 1:20;

[0027] Optionally, the protein concentration is 8-12% by mass, and the ultrasonic treatment is performed at a power of 200-400 W for 1-5 minutes to obtain the chitosan-protein-fatty acid-fucoxanthin complex aqueous solution, wherein the fucoxanthin encapsulation rate is ≥98%;

[0028] Optionally, after the reaction, the mixture is left open at room temperature to adjust the pH value of the system to 7-8, thereby obtaining a clear aqueous solution of the fucoxanthin-protein complex.

[0029] Furthermore, in step (5), the concentration of the chitosan in the system after mixing is 0.1 to 4.0% by mass;

[0030] Preferably, the concentration of chitosan in the system is 0.5 to 2.5% by mass;

[0031] Optionally, the reaction is carried out at room temperature for 20-40 minutes to combine chitosan with the protein shell of the complex through non-covalent self-assembly to obtain an aqueous solution of the chitosan-protein-fatty acid-fucoxanthin complex with nano-scale particle size.

[0032] Furthermore, the drying treatment in step (6) refers to spray drying, or freeze drying after concentration under reduced pressure.

[0033] The present invention also protects a water-soluble fucoxanthin dry powder prepared by the above method.

[0034] Furthermore, the particle size of the water-soluble fucoxanthin dry powder after redissolving in deionized water is 100 to 300 nm, and the Zeta potential of the water-soluble fucoxanthin aqueous solution is -15.0 to -45.0 mV.

[0035] Furthermore, after the water-soluble fucoxanthin is heated at 80-120° C. for 10-30 minutes, the retention rate of the fucoxanthin is ≥90%;

[0036] Optionally, after the water-soluble fucoxanthin is subjected to simulated gastric digestion, the retention rate of the fucoxanthin is ≥89.0%;

[0037] Optionally, the fucoxanthin is subjected to simulated intestinal digestion, and the retention rate of the fucoxanthin is ≥75%;

[0038] The above scheme shows that the water-soluble fucoxanthin prepared by the present invention has strong digestion stability.

[0039] Furthermore, the bioaccessibility of the water-soluble fucoxanthin dry powder is increased by 10.6 to 25.2 times compared to the raw material fucoxanthin.

[0040] Furthermore, the water-soluble fucoxanthin has a sustained-release effect in the gastrointestinal tract.

[0041] Compared with the prior art, the present invention has the following beneficial effects:

[0042] (1) The water-soluble fucoxanthin is prepared using alkaline electrolyzed water-ultrasonic coupling technology. Compared with existing preparation technologies, the alkaline electrolyzed water-ultrasonic coupling technology avoids the use of organic reagents, surfactants, and strong acids / bases, and has the advantages of being green and safe. At the same time, due to the strong reduction potential of alkaline electrolyzed water, it can effectively prevent fucoxanthin from being oxidized and degraded during the preparation process. The coupling of alkaline electrolyzed water with ultrasound greatly improves the fucoxanthin encapsulation rate. The fucoxanthin encapsulation rate in the chitosan-protein-fatty acid-fucoxanthin complex aqueous solution is ≥98%.

[0043] (2) Using water-soluble chitosan as the particle shell can achieve digestion-controlled release of fucoxanthin. The present invention first uses a simple and convenient Maillard modification technology to increase the pH solubility range of chitosan. Then, non-covalent self-assembly technology is used to coat the surface of fucoxanthin particles with chitosan, which can effectively improve the stability of fucoxanthin during gastrointestinal digestion and achieve the effect of intestinal targeted sustained release.

[0044] (3) Compared with other delivery systems, the present invention selectively introduces fatty acid components that promote intestinal absorption of fucoxanthin, which can improve the body's absorption of fucoxanthin, thereby fully exerting the nutritional function of fucoxanthin.

[0045] (4) The present invention provides a method for preparing water-soluble fucoxanthin, which has a simple technical route, does not require high-pressure homogenization and high-speed shearing processing, has the advantage of low production cost, and is suitable for industrial production. DETAILED DESCRIPTION

[0046] The preferred embodiments of the present invention will be described in more detail below. Although the preferred embodiments of the present invention are described below, it should be understood that the present invention can be implemented in various forms and should not be limited by the embodiments set forth herein. In the examples, if specific techniques or conditions are not specified, the techniques or conditions described in the literature in this area or the product specifications are used. Reagents or instruments used that do not specify the manufacturer are conventional products that can be obtained commercially. In the following examples, if not otherwise specified, "%" refers to weight percentage and parts refers to parts by weight.

[0047] The test methods used below include:

[0048] The digestive stability, controlled release, and bioaccessibility of water-soluble fucoxanthin were analyzed using the standardized in vitro food digestion method of Brodkorb et al. (see Brodkorb et al., Nature Protocols, 2019, 14:991-1014 for detailed methods).

[0049] Example 1

[0050] Chitosan powder was dissolved in acetic acid solution (acetic acid concentration was 1% by volume), stirred evenly to fully dissolve, the chitosan concentration was 0.5% by mass, and xylose was added at a mass ratio of 2:1. After mixing evenly, the mixture was subjected to Maillard reaction at 65°C and pH 5.0 for 96 hours. After the reaction was completed, the pH was adjusted to neutral and the mixture was spray-dried to obtain water-soluble chitosan powder. Fucoxanthin powder was dissolved in a small amount of ethanol solution, and then oleic acid was added at a mass ratio of 1:3 and stirred evenly to obtain a fucoxanthin fatty acid dispersion solution; 10.0g of bovine serum albumin powder was dissolved in alkaline electrolyte water (pH 11.0, redox potential -706mV), and fully stirred to dissolve to obtain an alkaline protein electrolyte aqueous solution (protein concentration was 10% by mass); then the fucoxanthin-fatty acid mixture was added to the protein solution so that the mass ratio of fucoxanthin to protein was 1:20; ultrasonic treatment was performed at a power of 300W for 3 minutes to allow the fucoxanthin, fatty acid, and protein to undergo uniform self-assembly; and the reaction system was left open at room temperature to allow the pH to naturally return to 7.8 to obtain a clear fucoxanthin-protein complex aqueous solution. The prepared partially water-soluble chitosan powder was added to a fucoxanthin-protein complex aqueous solution, so that the fucoxanthin, fatty acid, protein, and water-soluble chitosan were mixed in a mass ratio of 1:3:20:1, wherein the concentration of chitosan was 0.5% by mass. The mixture was allowed to stand for 30 minutes, and the chitosan was combined with the protein shell of the complex through non-covalent self-assembly to obtain a chitosan-protein-fatty acid-fucoxanthin complex aqueous solution (the fucoxanthin encapsulation efficiency was 98.3%); and the mixture was spray-dried to obtain a water-soluble fucoxanthin dry powder.

[0051] The physicochemical properties and absorption effect of the water-soluble fucoxanthin powder were further investigated: after redissolving it in deionized water, a clear and transparent solution was formed with a particle size of 100.0 nm and a surface charge of -45.0 mV. The retention rate of the water-soluble fucoxanthin microparticles after heating at 100°C for 15 minutes was 90.3%. The digestive stability, controlled release effect, and bioaccessibility of the water-soluble fucoxanthin were analyzed using the standardized method of Brodkorb et al. for in vitro simulated food digestion (Brodkorb et al., Nature Protocols, 2019, 14:991-1014). The results showed that the retention rate of fucoxanthin after simulated gastric digestion was 89.4%, and the retention rate after simulated intestinal digestion was 75.3%. After simulated gastrointestinal digestion, the release rate of fucoxanthin was 96.8%, and the bioaccessibility was increased by 10.6 times.

[0052] Example 2

[0053] Chitosan powder was dissolved in acetic acid solution (acetic acid concentration was 1% by volume), stirred evenly to fully dissolve, the chitosan concentration was 0.5% by mass, and glucose was added at a mass ratio of 1:1. After mixing evenly, Maillard reaction was carried out at 65°C and pH 5.5 for 72 hours. After the reaction was completed, the pH was adjusted to neutral and spray-dried to obtain water-soluble chitosan powder. Fucoxanthin powder was dissolved in a small amount of ethanol solution, and then linoleic acid was added at a mass ratio of 1:3 and stirred evenly to obtain a fucoxanthin fatty acid dispersion solution; 10.0g of bovine serum albumin powder was dissolved in alkaline electrolyte water (pH 11.0, redox potential -706mV), and fully stirred to dissolve to obtain an alkaline protein electrolyte aqueous solution (protein concentration was 10% by mass); then the fucoxanthin-fatty acid mixture was added to the protein solution so that the mass ratio of fucoxanthin to protein was 1:20; ultrasonic treatment was performed at a power of 300W for 3 minutes to allow the fucoxanthin, fatty acid, and protein to undergo uniform self-assembly; and the reaction system was left open at room temperature to allow the pH to naturally return to 7.5 to obtain a clear fucoxanthin-protein complex aqueous solution. The prepared partially water-soluble chitosan powder was added to a fucoxanthin-protein complex aqueous solution, so that the fucoxanthin, fatty acid, protein, and water-soluble chitosan were mixed in a mass ratio of 1:3:20:2, wherein the concentration of chitosan was 1.0% by mass. The mixture was allowed to stand for 30 minutes, and the chitosan was combined with the protein shell of the complex through non-covalent self-assembly to obtain a chitosan-protein-fatty acid-fucoxanthin complex aqueous solution (the fucoxanthin embedding efficiency was 98.4%); after reduced pressure concentration, the mixture was freeze-dried to obtain a water-soluble fucoxanthin dry powder.

[0054] The physicochemical properties and absorption efficiency of the water-soluble fucoxanthin powder were further investigated. After reconstitution in deionized water, the powder formed a clear, transparent solution with a particle size of 153 nm and a surface charge of -37.0 mV. The water-soluble fucoxanthin microparticles retained 93.6% after heating at 100°C for 15 minutes. The digestive stability, controlled release, and bioaccessibility of the water-soluble fucoxanthin were analyzed using the standardized in vitro simulated food digestion method of Brodkorb et al. (Brodkorb et al., Nature Protocols, 2019, 14:991-1014). The results showed that the retention of fucoxanthin after simulated gastric digestion was 89.6%, and the retention after simulated intestinal digestion reached 78.4%. After simulated gastrointestinal digestion, the release rate of fucoxanthin was 97.1%, and bioaccessibility was increased by 13.2-fold.

[0055] Example 3

[0056] Chitosan powder was dissolved in acetic acid solution (acetic acid concentration was 1% by volume), stirred evenly to fully dissolve, the concentration of chitosan was 0.5% by mass, and fructose was added at a mass ratio of 1:2. After mixing evenly, the mixture was subjected to Maillard reaction at 65°C and pH 5.5 for 72 hours. After the reaction was completed, the pH was adjusted to neutral and spray-dried to obtain water-soluble chitosan powder. The fucoxanthin powder was dissolved in a small amount of ethanol solution, and then myristic acid was added at a mass ratio of 1:4 and stirred evenly to obtain a fucoxanthin fatty acid dispersion solution; 10.0g of bovine serum albumin powder was dissolved in alkaline electrolyte water (pH 11.0, redox potential -706mV), and fully stirred to dissolve to obtain an alkaline protein electrolyte aqueous solution (protein concentration was 10% by mass); the fucoxanthin-fatty acid mixture was then added to the protein solution so that the mass ratio of fucoxanthin to protein was 1:20; ultrasonic treatment was performed at a power of 300W for 3 minutes to allow the fucoxanthin, fatty acid, and protein to undergo uniform self-assembly; the reaction system was left open at room temperature to allow the pH to naturally return to 7.0, thereby obtaining a clear fucoxanthin-protein complex aqueous solution. The prepared partially water-soluble chitosan powder was added to a fucoxanthin-protein complex aqueous solution, so that the fucoxanthin, fatty acid, protein, and water-soluble chitosan were mixed in a mass ratio of 1:4:20:3, wherein the concentration of chitosan was 1.5% by mass. The mixture was allowed to stand for 30 minutes, and the chitosan was combined with the protein shell of the complex through non-covalent self-assembly to obtain a chitosan-protein-fatty acid-fucoxanthin complex aqueous solution (the fucoxanthin embedding efficiency was 98.3%); and the mixture was spray-dried to obtain a water-soluble fucoxanthin dry powder.

[0057] The physicochemical properties and absorption effect of the water-soluble fucoxanthin powder were further investigated: after redissolving it in deionized water, a clear and transparent solution was formed with a particle size of 189.0 nm and a surface charge of -30 mV. The retention rate of the water-soluble fucoxanthin microparticles after heating at 100°C for 15 minutes was 91.5%. The digestive stability, controlled release effect, and bioaccessibility of the water-soluble fucoxanthin were analyzed using the standardized method of Brodkorb et al. for in vitro simulated food digestion (Brodkorb et al., Nature Protocols, 2019, 14:991-1014). The results showed that the retention rate of fucoxanthin after simulated gastric digestion was 91.5%, and the retention rate after simulated intestinal digestion could reach 84.8%. After simulated gastrointestinal digestion, the release rate of fucoxanthin was 97.9%, and the bioaccessibility was increased by 16.5 times.

[0058] Example 4

[0059] Chitosan powder was dissolved in acetic acid solution (acetic acid concentration was 1% by volume), stirred evenly to fully dissolve it, the concentration of chitosan was 0.5% by mass, and then galactose was added at a mass ratio of 1:3. After mixing evenly, the Maillard reaction was carried out at 65°C and pH 6.0 for 48 hours. After the reaction was completed, the pH was adjusted to neutral and spray-dried to obtain water-soluble chitosan powder. The fucoxanthin powder was dissolved in a small amount of ethanol solution, and then palmitic acid was added at a mass ratio of 1:4 and stirred evenly to obtain a fucoxanthin fatty acid dispersion solution; 10.0g of bovine serum albumin powder was dissolved in alkaline electrolyte water (pH 11.0, redox potential -706mV), and fully stirred to dissolve to obtain an alkaline protein electrolyte aqueous solution (protein concentration was 10% by mass); the fucoxanthin-fatty acid mixture was then added to the protein solution to make the mass ratio of fucoxanthin to protein 1:20; ultrasonic treatment was performed at a power of 300W for 3 minutes to allow the fucoxanthin, fatty acid, and protein to undergo uniform self-assembly; the reaction system was left open at room temperature to allow the pH to naturally return to 8.0, thereby obtaining a clear fucoxanthin-protein complex aqueous solution. The prepared water-soluble chitosan powder was added to a fucoxanthin-protein complex aqueous solution, and the fucoxanthin, fatty acid, protein, and water-soluble chitosan were mixed in a mass ratio of 1:4:20:4, wherein the concentration of chitosan was 2.0% by mass. The mixture was allowed to stand for 30 minutes, and the chitosan was combined with the protein shell of the complex through non-covalent self-assembly to obtain a chitosan-protein-fatty acid-fucoxanthin complex aqueous solution (fucoxanthin embedding efficiency was 98.5%); after reduced pressure concentration, the mixture was freeze-dried to obtain a water-soluble fucoxanthin dry powder.

[0060] The physicochemical properties and absorption effect of the water-soluble fucoxanthin powder were further investigated: after redissolving it in deionized water, a clear and transparent solution was formed with a particle size of 254 nm and a surface charge of -27 mV. The water-soluble fucoxanthin microparticles retained 94.0% after being heated at 100°C for 15 minutes. The digestive stability, controlled release effect, and bioaccessibility of the water-soluble fucoxanthin were analyzed using the standardized method of Brodkorb et al. for in vitro simulated food digestion (Brodkorb et al., Nature Protocols, 2019, 14:991-1014). The results showed that the retention rate of fucoxanthin after simulated gastric digestion was 93.8%, and the retention rate after simulated intestinal digestion was 89.9%. After simulated gastrointestinal digestion, the release rate of fucoxanthin was 98.3%, and the bioaccessibility was increased by 21.6 times.

[0061] Example 5

[0062] Chitosan powder was dissolved in acetic acid solution (acetic acid concentration was 1% by volume), stirred evenly to fully dissolve, the concentration of chitosan was 0.5% by mass, and mannose was added at a mass ratio of 1:2. After mixing evenly, the mixture was subjected to Maillard reaction at 65°C and pH 6.0 for 48 hours. After the reaction was completed, the pH was adjusted to neutral and spray-dried to obtain water-soluble chitosan powder. The fucoxanthin powder was dissolved in a small amount of ethanol solution, and then stearic acid was added at a mass ratio of 1:5 and stirred evenly to obtain a fucoxanthin fatty acid dispersion solution; 10.0g of bovine serum albumin powder was dissolved in alkaline electrolyte water (pH 11.0, redox potential -706mV), and fully stirred to dissolve to obtain an alkaline protein electrolyte aqueous solution (protein concentration was 10% by mass); the fucoxanthin-fatty acid mixture was then added to the protein solution so that the mass ratio of fucoxanthin to protein was 1:20; ultrasonic treatment was performed at a power of 300W for 3 minutes to allow the fucoxanthin, fatty acid, and protein to undergo uniform self-assembly; the reaction system was left open at room temperature to allow the pH to naturally return to 7.3, thereby obtaining a clear fucoxanthin-protein complex aqueous solution. The prepared partially water-soluble chitosan powder was added to a fucoxanthin-protein complex aqueous solution, so that the fucoxanthin, fatty acid, protein, and water-soluble chitosan were mixed in a mass ratio of 1:5:20:5, wherein the concentration of chitosan was 2.5% by mass. The mixture was allowed to stand for 30 minutes, and the chitosan was combined with the protein shell of the complex through non-covalent self-assembly to obtain a chitosan-protein-fatty acid-fucoxanthin complex aqueous solution (the fucoxanthin encapsulation efficiency was 98.6%); and the mixture was spray-dried to obtain a water-soluble fucoxanthin dry powder.

[0063] The physicochemical properties and absorption effect of the water-soluble fucoxanthin powder were further investigated: after redissolving it in deionized water, a clear and transparent solution was formed with a particle size of 300 nm and a surface charge of -15 mV. After heating the water-soluble fucoxanthin microparticles at 100°C for 15 minutes, the retention rate was 92.6%. The digestive stability, controlled release effect, and bioaccessibility of the water-soluble fucoxanthin were analyzed using the standardized method of Brodkorb et al. for in vitro simulated food digestion (Brodkorb et al., Nature Protocols, 2019, 14:991-1014). The results showed that the retention rate of fucoxanthin after simulated gastric digestion was 90.9%, and the retention rate after simulated intestinal digestion was 87.1%. After simulated gastrointestinal digestion, the release rate of fucoxanthin was 97.9%, and the bioaccessibility was increased by 25.2 times.

[0064] Example 6

[0065] Chitosan powder was dissolved in acetic acid solution (acetic acid concentration was 1.5% by volume), stirred evenly to fully dissolve, the concentration of chitosan was 0.5% by mass, and then mannose was added at a mass ratio of 1:2. After mixing evenly, the mixture was subjected to Maillard reaction at 70°C and pH 6.0 for 48 hours. After the reaction was completed, the pH was adjusted to neutral and spray-dried to obtain water-soluble chitosan powder. Fucoxanthin powder was dissolved in a small amount of ethanol solution, and then stearic acid was added at a mass ratio of 1:3 and stirred evenly to obtain a fucoxanthin fatty acid dispersion solution; 10.0g of dog serum albumin powder was dissolved in alkaline electrolyte water (pH 10, redox potential -700mV), and fully stirred to dissolve to obtain an alkaline protein electrolyte aqueous solution (protein concentration was 12% by mass); then the fucoxanthin-fatty acid mixture was added to the protein solution to make the mass ratio of fucoxanthin to protein 0.8:18; ultrasonic treatment was performed at a power of 400W for 1 minute to allow the fucoxanthin, fatty acid, and protein to undergo uniform self-assembly; and the reaction system was left open at room temperature to allow the pH to naturally return to 7.3 to obtain a clear fucoxanthin-protein complex aqueous solution. The prepared partially water-soluble chitosan powder is added to a fucoxanthin-protein complex aqueous solution, so that the fucoxanthin, fatty acid, protein, and water-soluble chitosan are mixed in a mass ratio of 0.8:2.4:18:0.15, wherein the concentration of chitosan is 0.1% by mass. The mixture is allowed to stand for 20 minutes to allow the chitosan to bind to the protein shell of the complex through non-covalent self-assembly to obtain a chitosan-protein-fatty acid-fucoxanthin complex aqueous solution; the mixture is spray-dried or freeze-dried after reduced pressure concentration to obtain a water-soluble fucoxanthin dry powder.

[0066] Example 7

[0067] Chitosan powder was dissolved in acetic acid solution (acetic acid concentration was 0.5% by volume), stirred evenly to fully dissolve, the concentration of chitosan was 0.5% by mass, and then mannose was added at a mass ratio of 1:6. After mixing evenly, the mixture was subjected to Maillard reaction at 60°C and pH 5.0 for 72 hours. After the reaction was completed, the pH was adjusted to neutral and spray-dried to obtain water-soluble chitosan powder. The fucoxanthin powder was dissolved in a small amount of ethanol solution, and then stearic acid was added at a mass ratio of 1:5 and stirred evenly to obtain a fucoxanthin fatty acid dispersion solution; 10.0g of porcine serum albumin powder was dissolved in alkaline electrolyte water (pH 12, redox potential -710mV), and fully stirred to dissolve to obtain an alkaline protein electrolyte aqueous solution (protein concentration was 8% by mass); the fucoxanthin-fatty acid mixture was then added to the protein solution to make the mass ratio of fucoxanthin to protein be 1.2:22; ultrasonic treatment was performed at a power of 200W for 4 minutes to allow the fucoxanthin, fatty acid, and protein to undergo uniform self-assembly; the reaction system was left open at room temperature to allow the pH to naturally return to 7.3, to obtain a clear fucoxanthin-protein complex aqueous solution. The prepared partially water-soluble chitosan powder is added to a fucoxanthin-protein complex aqueous solution, so that the fucoxanthin, fatty acid, protein, and water-soluble chitosan are mixed in a mass ratio of 1.2:6:22:11, wherein the concentration of chitosan is 4.0% by mass, and the mixture is allowed to stand for 40 minutes to allow the chitosan to bind to the protein shell of the complex through non-covalent self-assembly to obtain a chitosan-protein-fatty acid-fucoxanthin complex aqueous solution; the mixture is spray-dried or freeze-dried after reduced pressure concentration to obtain a water-soluble fucoxanthin dry powder.

[0068] Example 8

[0069] The specific method of Example 1 was used for operation, with the main difference being that the fucoxanthin-fatty acid mixture was added to the protein solution so that the mass ratio of fucoxanthin to protein was 0.5:15.

[0070] Example 9

[0071] The specific method of Example 1 was used for operation, with the main difference being that the fucoxanthin-fatty acid mixture was added to the protein solution so that the mass ratio of fucoxanthin to protein was 1.5:25.

[0072] Example 10

[0073] The specific method of Example 1 was used for operation, with the main difference being that the fucoxanthin-fatty acid mixture was added to the protein solution so that the mass ratio of fucoxanthin to protein was 0.5:15.

[0074] Example 11

[0075] The specific method of Example 1 was used for operation, with the main difference being that the fucoxanthin-fatty acid mixture was added to the protein solution so that the mass ratio of fucoxanthin to protein was 1.2:18.

[0076] Comparative Example 1: No Maillard reaction occurs

[0077] The specific method of Example 1 was used, except that chitosan powder was used directly in this comparative example without the Maillard reaction. The specific steps are as follows:

[0078] Fucoxanthin powder was dissolved in an ethanol solution, and then oleic acid was added at a mass ratio of 1:3 and stirred to obtain a fucoxanthin fatty acid dispersion solution. 10.0 g of bovine serum albumin powder was dissolved in alkaline electrolyte water (pH 11.0, redox potential -706 mV) and stirred to obtain an alkaline protein electrolyte aqueous solution (protein concentration was 10% by mass). The fucoxanthin-fatty acid mixture was then added to the protein solution to achieve a fucoxanthin-protein mass ratio of 1:20. The solution was ultrasonicated at 300 W for 3 minutes to allow uniform self-assembly of the fucoxanthin, fatty acid, and protein. The reaction system was left open at room temperature to adjust the pH to 7.8, resulting in a fucoxanthin-protein complex aqueous solution. Chitosan powder was added to the fucoxanthin-protein complex aqueous solution to a chitosan concentration of 0.5-2.5% by mass. The solution was allowed to stand for 30 minutes. Chitosan not modified by the Maillard reaction was insoluble in the aqueous solution and precipitated. The supernatant is a protein-fatty acid-fucoxanthin complex aqueous solution (the fucoxanthin embedding rate is 91.2%); spray drying is performed to obtain water-soluble fucoxanthin dry powder without chitosan.

[0079] The prepared fucoxanthin dry powder was further investigated for its physicochemical properties and absorption. After reconstitution in deionized water, the resulting solution had a particle size of 86 nm and a surface charge of -52 mV. After heating at 100°C for 15 minutes, the fucoxanthin microparticles retained 68.5%. The digestive stability, controlled release, and bioaccessibility of the water-soluble fucoxanthin were analyzed using the standardized in vitro method of simulated food digestion (Brodkorb et al.). Results showed that fucoxanthin retention was 71.6% after simulated gastric digestion and 58.4% after simulated intestinal digestion. After simulated gastrointestinal digestion, the fucoxanthin release rate was 97.6%, indicating only a 4.3-fold increase in bioaccessibility.

[0080] Comparative Example 2: Alkaline electrolyte water replaces alkaline water

[0081] The specific method of Example 1 is used, except that an alkaline aqueous solution (sodium hydroxide aqueous solution) is used to prepare the water-soluble fucoxanthin powder. The specific steps are as follows:

[0082] Chitosan powder was dissolved in acetic acid solution (1% by volume) and stirred to dissolve. The chitosan concentration was 0.5% by mass. Xylose was then added at a mass ratio of 2:1. After mixing, the mixture was subjected to a Maillard reaction at 65°C and pH 5.0 for 96 hours. After the reaction, the pH was adjusted to neutral and the mixture was spray-dried to obtain a water-soluble chitosan powder. Fucoxanthin powder was dissolved in ethanol solution, and then oleic acid was added at a mass ratio of 1:3 and stirred to obtain a fucoxanthin fatty acid dispersion solution. 10.0 g of bovine serum albumin powder was dissolved in sodium hydroxide aqueous solution (pH 11.0) and stirred to obtain an alkaline protein aqueous solution (protein concentration was 10% by mass). The fucoxanthin-fatty acid mixture was then added to the protein solution to obtain a fucoxanthin to protein mass ratio of 1:20. The mixture was then ultrasonicated at a power of 300 W for 3 minutes to obtain a fucoxanthin-protein complex aqueous solution. The prepared partially water-soluble chitosan powder was added to a fucoxanthin-protein complex aqueous solution to a chitosan concentration of 0.5% by mass. The mixture was allowed to stand for 30 minutes to allow the chitosan to bind to the protein shell of the complex through non-covalent self-assembly to obtain a chitosan-protein-fatty acid-fucoxanthin complex aqueous solution (with a fucoxanthin encapsulation efficiency of 85.6%). The mixture was then spray-dried to obtain a fucoxanthin dry powder.

[0083] The physicochemical properties and absorption efficiency of the fucoxanthin powder were further investigated. After reconstitution in deionized water, a solution was formed with a particle size of 125 nm and a surface charge of -43 mV. After heating at 100°C for 15 minutes, the water-soluble fucoxanthin microparticles retained 72.3%. The digestive stability, controlled release, and bioaccessibility of the water-soluble fucoxanthin were analyzed using the standardized in vitro method of simulated food digestion (Brodkorb et al.). Results showed that fucoxanthin retention was 74.2% after simulated gastric digestion and 63.5% after simulated intestinal digestion. After simulated gastrointestinal digestion, the fucoxanthin release rate was 97.1%, indicating only a 4.9-fold increase in bioaccessibility.

[0084] As shown in Examples 1-5 and Comparative Examples 1-2, the present invention utilizes alkaline electrolyzed water-ultrasound coupling technology, effectively avoiding the use of organic reagents, surfactants, and strong acids / bases. The provided technical solution and process offer the advantages of being both environmentally friendly and safe. Furthermore, due to the high reduction potential of alkaline electrolyzed water, it effectively prevents oxidative degradation of fucoxanthin during the preparation process. The coupling of alkaline electrolyzed water with ultrasound significantly increases the entrapment efficiency of fucoxanthin.

[0085] The present invention utilizes a simple and convenient Maillard modification technology to increase the pH solubility range of chitosan, and then adopts non-covalent self-assembly technology to cover the surface of fucoxanthin particles with chitosan, which can effectively improve the stability of fucoxanthin during gastrointestinal digestion and achieve the effect of intestinal targeted sustained release.

[0086] Although the embodiments of the present invention have been shown and described above, it will be understood that the above embodiments are illustrative and are not to be construed as limitations on the present invention. A person skilled in the art may change, modify, replace and modify the above embodiments within the scope of the present invention without departing from the principles and purpose of the present invention.

Claims

1. A method for preparing water-soluble fucoxanthin, characterized in that: The specific preparation method comprises the following steps: (1) Preparation of water-soluble chitosan powder: chitosan, acetic acid solution, and reducing sugar are mixed and subjected to Maillard reaction. After the reaction, the pH is adjusted and the mixture is dried to obtain the water-soluble chitosan powder; (2) Preparation of fucoxanthin-fatty acid mixture: fucoxanthin, ethanol and fatty acid are uniformly mixed to dissolve the fucoxanthin to obtain the fucoxanthin-fatty acid mixture; (3) Preparation of alkaline electrolyte water: using carbonate as an electrolyte, ionizing the aqueous solution of the electrolyte to obtain alkaline electrolyte water at an anode; the ionization in step (3) is performed at a voltage of 220 V for 15-30 minutes, and the pH value of the alkaline electrolyte water obtained is 10.0-12.0, and the redox potential is -750--650 mV; (4) Preparation of fucoxanthin-protein complex aqueous solution by electrolysis water coupling ultrasound technology: protein powder is mixed with the alkaline electrolyte water to obtain a protein solution; the fucoxanthin-fatty acid mixture is then added to the protein solution to obtain a mixed solution; the mixed solution is then subjected to ultrasound treatment to allow the fucoxanthin, the fatty acid and the protein in the mixed solution to self-assemble; after the reaction, the mixture is left open at room temperature to obtain the fucoxanthin-protein complex aqueous solution; (5) Preparation of chitosan-protein-fatty acid-fucoxanthin complex aqueous solution: the water-soluble chitosan powder prepared in step (1) is mixed with the fucoxanthin-protein complex aqueous solution prepared in step (4), and the mixture is reacted to bind the chitosan to the protein shell of the complex through non-covalent self-assembly to obtain the chitosan-protein-fatty acid-fucoxanthin complex aqueous solution; (6) Obtaining water-soluble fucoxanthin dry powder: drying the chitosan-protein-fatty acid-fucoxanthin complex aqueous solution to obtain the water-soluble fucoxanthin dry powder.

2. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: The reducing sugar in step (1) includes at least one of glucose, fructose, galactose, xylose or mannose.

3. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: In step (1), chitosan, acetic acid solution, and reducing sugar are mixed so that the mass ratio of chitosan to reducing sugar in the solution is (1-2):(1-3), and the pH of the reaction system after mixing is 5.0-6.

0.

4. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: In step (1), the temperature of the Maillard reaction is 60-70°C and the time is 48-96 hours.

5. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: The fatty acid in step (2) includes at least one of myristic acid, palmitic acid, stearic acid, oleic acid, linoleic acid or docosahexaenoic acid; the mass ratio of the fucoxanthin to the fatty acid is 1:(3-5).

6. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: In step (3), the pH value of the alkaline electrolyte water is 10.5 to 11.5, and the redox potential is -710 to -700 mV.

7. The method for preparing water-soluble fucoxanthin according to claim 6, characterized in that: The pH value of the alkaline electrolyte water is 11, and the redox potential is -706 mV.

8. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: The mass ratio of the fucoxanthin to the protein powder in the mixed solution in step (4) is (0.5-1.5):(15-25).

9. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: In step (4), the concentration of the protein is 8-12% by mass, and the ultrasonic treatment is performed at a power of 200-400 W for 1-5 minutes to obtain the chitosan-protein-fatty acid-fucoxanthin complex aqueous solution, wherein the fucoxanthin encapsulation rate is ≥98%.

10. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: In step (4), after the reaction, the mixture is left open at room temperature to make the pH of the system equal to 7 to 8, thereby obtaining a clear aqueous solution of the fucoxanthin-protein complex.

11. A method for preparing water-soluble fucoxanthin according to any one of claims 8 to 10, characterized in that: The mass ratio of the fucoxanthin to the protein powder is (0.8~1.2): (18~22).

12. The method for preparing water-soluble fucoxanthin according to claim 11, characterized in that: The mass ratio of the fucoxanthin to the protein powder is 1:

20.

13. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: After mixing in step (5), the concentration of chitosan in the system is 0.1-4.0% by mass. The reaction is carried out at room temperature for 20-40 minutes, and chitosan is combined with the protein shell of the complex through non-covalent self-assembly to obtain the chitosan-protein-fatty acid-fucoxanthin complex aqueous solution with nano-scale particle size.

14. The method for preparing water-soluble fucoxanthin according to claim 13, characterized in that: After mixing in step (5), the concentration of chitosan in the system is 0.5 to 2.5% by mass.

15. The method for preparing water-soluble fucoxanthin according to claim 1, characterized in that: The drying treatment in step (6) refers to spray drying, or freeze drying after vacuum concentration.

16. A water-soluble fucoxanthin dry powder prepared by the method according to any one of claims 1 to 15.

17. The water-soluble fucoxanthin dry powder according to claim 16, characterized in that: The particle size of the water-soluble fucoxanthin dry powder after redissolving in deionized water is 100 to 300 nm, and the Zeta potential of the water-soluble fucoxanthin aqueous solution is -15.0 to -45.0 mV.

18. The water-soluble fucoxanthin dry powder according to claim 16, characterized in that: After the water-soluble fucoxanthin is heated at 80-120° C. for 10-30 minutes, the retention rate of the fucoxanthin is ≥90%.

19. The water-soluble fucoxanthin dry powder according to claim 16, characterized in that: After the water-soluble fucoxanthin is subjected to simulated gastric digestion, the retention rate of the fucoxanthin is ≥89.0%.

20. The water-soluble fucoxanthin dry powder according to claim 16, characterized in that: The fucoxanthin is subjected to simulated intestinal digestion, and the retention rate of the fucoxanthin is ≥75%; the water-soluble fucoxanthin has a sustained-release effect in the gastrointestinal tract; and the fucoxanthin has digestion stability.

21. The water-soluble fucoxanthin according to any one of claims 17 to 20, characterized in that: The bioaccessibility of the water-soluble fucoxanthin dry powder is increased by 10.6 to 25.2 times compared to the raw material fucoxanthin.