A method of preparing an omega-3 fish oil emulsion composition

Abstract

The application discloses a preparation method of an Omega-3 fish oil emulsion composition. The method comprises the following steps: adding a zein solution into a hydrophilic protein aqueous phase, so that the zein is subjected to anti-solvent precipitation and is in-situ interfacially compounded with the hydrophilic protein to obtain a composite particle dispersion liquid; and then, the composite particle dispersion liquid is subjected to shearing emulsification, high-pressure homogenization and stabilization treatment together with Omega-3 fish oil containing an antioxidant to obtain the fish oil emulsion composition. The emulsion obtained by the method has small particle size, good dispersion stability, good oxidation stability and good in-vitro digestion release effect.

Description

Technical Field

[0001] This invention relates to the field of fish oil processing technology, and more particularly to a method for preparing an Omega-3 fish oil emulsion composition. Background Technology

[0002] Omega-3 fish oil is rich in polyunsaturated fatty acids such as EPA and DHA, making it highly valuable for nutritional fortification and functional food applications. However, Omega-3 fish oil is highly hydrophobic, has a pronounced fishy odor, and dispersibility in aqueous foods is poor. Furthermore, EPA and DHA are easily oxidized by factors such as oxygen, light, and metal ions, leading to flavor deterioration and nutrient loss. Therefore, constructing a stable fish oil emulsification system is an important way to improve the application performance of Omega-3 fish oil.

[0003] In existing technologies, fish oil emulsions typically use surfactants, milk proteins, hydrocolloids, etc., as emulsifiers, and are prepared using high-pressure homogenization or microfluidic homogenization. For example, CN109511858A discloses a method for preparing a high-concentration fish oil nanoemulsion, which homogenizes an aqueous phase containing emulsifiers, antioxidants, and water, and an oil phase containing fish oil and fat-soluble antioxidants, using a dual-channel microfluidic device to obtain a high-concentration fish oil nanoemulsion. This method can improve the dispersibility of fish oil and the stability of the emulsion, but it mainly relies on conventional emulsifiers to form an adsorption layer at the oil-water interface, and there is still room for further improvement in the interfacial film strength and antioxidant barrier capacity.

[0004] Pickering emulsions utilize solid particles adsorbed at the oil-water interface to form a particle interface layer, typically exhibiting good anti-agglomeration ability. CN116998699A discloses a one-step method for preparing tuna oil Pickering emulsions, which uses a pH-driven method to prepare zein / pectin composite nanoparticles, and then mixes and homogenizes them with tuna oil in one step using a dual-channel microfluidic technique to obtain the tuna oil Pickering emulsion. This method can improve the emulsion's structural stability, anti-lipid oxidation performance, and intestinal sustained-release effect. However, its composite particles are zein / pectin protein-polysaccharide composite particles, and it relies on strong acid / strong base pH driving and a dual-channel microfluidic device. It does not involve the technical solution of zein precipitating in the aqueous phase of sodium caseinate and forming protein-protein composite particles in situ at the interface.

[0005] Therefore, it remains necessary to develop a method for preparing Omega-3 fish oil emulsion compositions that differs from ordinary emulsifier nanoemulsions and zein / pectin composite particle Pickering emulsions. This invention involves dropwise addition of a zein ethanol aqueous solution to the aqueous phase of sodium caseinate, causing the zein to undergo antisolvent precipitation and in-situ complex with sodium caseinate at the interface, forming zein-sodium caseinate composite particles. These particles are then used to stabilize the Omega-3 fish oil emulsion, thereby improving the emulsion's physical stability, oxidative stability, and in vitro digestion and release efficiency. Summary of the Invention

[0006] In view of the above-mentioned deficiencies of the prior art, the technical problem to be solved by the present invention is to provide a method for preparing an Omega-3 fish oil emulsion composition, so as to improve the dispersion stability of Omega-3 fish oil in the aqueous phase, reduce the oxidative deterioration during storage, and improve its utilization effect in the in vitro simulated gastrointestinal digestion process.

[0007] To achieve the above objectives, this invention provides a method for preparing an Omega-3 fish oil emulsion composition, using OMAXorb emulsification technology; specifically, it includes the following steps: 1) Add zein to an aqueous ethanol solution and stir to dissolve, thus obtaining a zein solution; 2) Mix the hydrophilic protein, sugar protectant, and water, stir to dissolve, and obtain the hydrophilic protein aqueous phase; 3) The zein solution obtained in step 1) is added dropwise to the aqueous phase of the hydrophilic protein obtained in step 2), so that the zein undergoes antisolvent precipitation in the aqueous phase and undergoes in-situ interfacial complexation with the hydrophilic protein; then the ethanol is removed to obtain a zein-hydrophilic protein composite particle dispersion. 4) Mix Omega-3 fish oil, fat-soluble antioxidants, and plant antioxidant extracts to obtain an antioxidant fish oil phase; 5) Add the antioxidant fish oil phase obtained in step 4) to the zein-hydrophilic protein composite particle dispersion obtained in step 3), and shear emulsify to obtain Omega-3 fish oil crude emulsion; 6) The crude Omega-3 fish oil emulsion obtained in step 5) is subjected to high-pressure homogenization to obtain an Omega-3 fish oil emulsion; 7) Add thickener, stabilizer and buffer salt to the Omega-3 fish oil emulsion obtained in step 6), stir and filter to obtain Omega-3 fish oil emulsion composition.

[0008] The amounts of the above raw materials, by weight, are as follows: 0.8-2.0 parts zein, 9.6-40 parts ethanol aqueous solution, 0.4-1.2 parts hydrophilic protein, 0.5-1.5 parts saccharide protectant, 40-60 parts water, 20-40 parts Omega-3 fish oil, 0.05-0.30 parts fat-soluble antioxidant, 0.01-0.10 parts plant antioxidant extract, 0.05-0.30 parts thickener and stabilizer, and 0.05-0.50 parts buffer salt.

[0009] Preferably, in step 1), the stirring temperature is 20-30℃, the stirring speed is 400-800 r / min, and the stirring time is 30-60 min; the mass fraction of the ethanol aqueous solution is 60%-80%.

[0010] Preferably, in step 2), the hydrophilic protein is one of sodium caseinate, whey protein isolate, and β-lactoglobulin; the glycoprotectant is one or more of trehalose, sucrose, maltose, and maltodextrin. More preferably, the hydrophilic protein is sodium caseinate; and the glycoprotectant is trehalose.

[0011] Preferably, in step 3), the dripping rate of the zein solution is 0.05-0.50 L / min; the stirring speed during the dripping process is 800-1500 r / min; stirring continues for 30-90 min after the dripping is completed; and the conditions for removing ethanol are 30-40℃ and absolute pressure 0.01-0.04 MPa.

[0012] Preferably, in step 4), the total mass content of EPA and DHA in the Omega-3 fish oil is not less than 50%; the fat-soluble antioxidant is one or more of natural vitamin E, vitamin E acetate, and ascorbyl palmitate; and the plant antioxidant extract is one or more of rosemary extract, tea polyphenol oil dispersion, and astaxanthin oil dispersion.

[0013] More preferably, in step 4), the fat-soluble antioxidant is natural vitamin E; the plant antioxidant extract is rosemary extract; and the mass ratio of Omega-3 fish oil, natural vitamin E and rosemary extract is 30.00:0.15:0.05.

[0014] Preferably, in step 5), the addition rate of the antioxidant fish oil phase is 0.20-1.00 kg / min; the shear emulsification temperature is 15-25℃; the shearing speed is 8000-12000 r / min; and the shearing time is 3-10 min.

[0015] Preferably, in step 6), the high-pressure homogenization pressure is 40-80 MPa, the homogenization is performed 1-3 times, and the discharge temperature is controlled not to exceed 35℃ during the homogenization process.

[0016] Preferably, in step 7), the thickening stabilizer is one or more of xanthan gum, guar gum, sodium carboxymethyl cellulose, and sodium alginate; the buffer salt is one or more of sodium citrate, disodium hydrogen phosphate, and sodium dihydrogen phosphate; and the mass ratio of the thickening stabilizer to the buffer salt is 1:1-3.

[0017] More preferably, in step 7), the thickening stabilizer is xanthan gum and the buffer salt is sodium citrate.

[0018] The technical concept of this invention lies in utilizing the antisolvent precipitation process of zein in an aqueous phase, allowing hydrophilic proteins to simultaneously participate in interfacial complexation on the surface of newly formed particles, thereby forming composite particles with both hydrophobic anchoring and hydrophilic stabilizing effects. These composite particles can adsorb onto the surface of Omega-3 fish oil droplets, forming a stable particle interface layer, thus improving the emulsion's anti-agglomeration ability and oxidative stability. In particular, when sodium caseinate is selected as the hydrophilic protein, the resulting zein-sodium caseinate composite particle dispersion exhibits even better interfacial stabilizing effects on fish oil droplets.

[0019] The beneficial effects of this invention are as follows: Compared with fish oil emulsions stabilized by ordinary emulsifiers, the present invention stabilizes Omega-3 fish oil droplets through zein-hydrophilic protein composite particles, which can reduce droplet aggregation and oil separation, and improve the static and centrifugal stability of the emulsion.

[0020] Compared with the conventional composite method of first preparing zein particles and then adding hydrophilic proteins, the present invention enables hydrophilic proteins to participate more effectively in the composite and stabilization of the surface of newly formed particles through in-situ interfacial composite during the antisolvent precipitation process, which is conducive to the formation of a more continuous and stable fish oil droplet interface layer.

[0021] Compared to fish oil emulsions stabilized by zein-pectin composite nanoparticles, the preferred zein-sodium caseinate composite particle dispersion of this invention further improves emulsion particle size distribution, oxidative stability, and in vitro simulated gastrointestinal digestion and release performance.

[0022] The Omega-3 fish oil emulsified composition prepared by this invention has low peroxide value, anisidine value and TOTOX value during storage, and high EPA and DHA retention rates, which is beneficial to reducing the oxidative deterioration of Omega-3 fish oil.

[0023] The Omega-3 fish oil emulsion composition prepared by this invention has a low oil separation rate and EPA+DHA release rate in simulated gastric fluid, and a high EPA+DHA micellization rate in simulated intestinal fluid, which is beneficial for balancing gastric protection and intestinal release. Detailed Implementation

[0024] The parameters and sources of some raw materials in the examples are as follows: Omega-3 fish oil has a total EPA and DHA content of 60.5% and an initial peroxide value of 4.2 meq O2 / kg.

[0025] The preparation method of rosemary extract is as follows: dried rosemary leaves are pulverized and passed through a 40-mesh sieve. 1.00 kg of rosemary leaf powder is weighed and 10.00 kg of 95% ethanol aqueous solution is added. The mixture is extracted at 50°C with stirring at 300 r / min for 2 h. After filtration, the filtrate is collected. The filtrate is concentrated under reduced pressure at 45°C and 0.02 MPa until there is no obvious ethanol odor, thus obtaining rosemary concentrate. 2.00 kg of medium-chain triglyceride oil is added to the rosemary concentrate and stirred at 40°C with stirring at 300 r / min for 30 min. The mixture is then filtered through a 200-mesh filter cloth to obtain rosemary extract.

[0026] Example 1

[0027] A method for preparing an Omega-3 fish oil emulsion composition includes the following steps: 1) Preparation of zein solution Weigh 1.20 kg of zein and add it to 18.80 kg of 70% ethanol aqueous solution. Stir at 600 r / min for 40 min at 25℃ to fully dissolve the zein and obtain a zein solution. 2) Preparation of sodium caseinate aqueous phase Weigh out 0.80 kg of sodium caseinate, 1.00 kg of trehalose and 48.20 kg of deionized water, and stir at 500 r / min for 30 min at 30℃ to obtain an aqueous phase of sodium caseinate; the pH of the aqueous phase of sodium caseinate was tested to be 6.8. 3) Preparation of particulate dispersion The zein solution obtained in step 1) was added dropwise to the sodium caseinate aqueous phase obtained in step 2) at a rate of 0.20 L / min. During the dropwise addition, the stirring speed was maintained at 1200 r / min. After the dropwise addition was completed, stirring was continued for 60 min to allow the zein to undergo antisolvent precipitation in the aqueous phase and to form an interfacial complex with the sodium caseinate. Subsequently, ethanol was removed by rotary evaporation at 35 °C and an absolute pressure of 0.02 MPa to obtain a zein-sodium caseinate composite particle dispersion. 4) Preparation of antioxidant fish oil phase Weigh out 30.00 kg of Omega-3 fish oil, 0.15 kg of natural vitamin E, and 0.05 kg of rosemary extract, and stir at 25°C in the dark for 20 min to obtain the antioxidant fish oil phase; 5) Initial emulsification The antioxidant fish oil phase obtained in step 4) was added to the composite particle dispersion obtained in step 3) at a rate of 0.50 kg / min, and shearing emulsification was performed at 20°C using a high-speed shear emulsifier with a shearing speed of 10000 r / min and a shearing time of 5 min to obtain Omega-3 fish oil crude emulsion. 6) Homogenization and Refinement The Omega-3 fish oil crude emulsion obtained in step 5) was placed in a high-pressure homogenizer and homogenized twice at 60 MPa. During the homogenization process, the discharge temperature was controlled not to exceed 30℃ to obtain Omega-3 fish oil Pickering emulsion. 7) Stabilization treatment Add 0.10 kg of xanthan gum and 0.20 kg of sodium citrate to the Omega-3 fish oil Pickering emulsion obtained in step 6), stir at 300 r / min for 20 min at 25 °C, adjust the pH to 6.8 with 0.1 mol / L citric acid aqueous solution, and filter through a 100 mesh sieve to obtain the Omega-3 fish oil emulsion composition.

[0028] Example 2

[0029] A method for preparing an Omega-3 fish oil emulsion composition is basically the same as that in Example 1, except that whey protein isolate is used instead of sodium caseinate. Specifically, the method includes the following steps: 1) Preparation of zein solution Weigh 1.20 kg of zein and add it to 18.80 kg of 70% ethanol aqueous solution. Stir at 600 r / min for 40 min at 25℃ to fully dissolve the zein and obtain a zein solution. 2) Preparation of aqueous phase of whey protein isolate Weigh out 0.80 kg of whey protein isolate, 1.00 kg of trehalose, and 48.20 kg of deionized water. Stir at 500 r / min for 30 min at 30℃ to obtain the aqueous phase of whey protein isolate. The pH of the aqueous phase of whey protein isolate was tested to be 6.5. The pH was adjusted to 6.8 using 0.1 mol / L sodium hydroxide aqueous solution. 3) Preparation of particulate dispersion The zein solution obtained in step 1) was added dropwise to the whey protein isolate aqueous phase obtained in step 2) at a rate of 0.20 L / min. During the dropwise addition, the stirring speed was maintained at 1200 r / min. After the dropwise addition was completed, stirring was continued for 60 min to allow the zein to undergo antisolvent precipitation in the aqueous phase and interfacial complexation with the whey protein isolate. Subsequently, ethanol was removed by rotary evaporation at 35 °C and an absolute pressure of 0.02 MPa to obtain a zein-whey protein isolate particle dispersion. 4) Preparation of antioxidant fish oil phase Weigh out 30.00 kg of Omega-3 fish oil, 0.15 kg of natural vitamin E, and 0.05 kg of rosemary extract, and stir at 25°C in the dark for 20 min to obtain the antioxidant fish oil phase; 5) Initial emulsification The antioxidant fish oil phase obtained in step 4) was added to the particulate dispersion obtained in step 3) at a rate of 0.50 kg / min, and shearing emulsification was performed at 20°C using a high-speed shear emulsifier with a shearing speed of 10000 r / min and a shearing time of 5 min to obtain Omega-3 fish oil crude emulsion. 6) Homogenization and Refinement The Omega-3 fish oil crude emulsion obtained in step 5) was placed in a high-pressure homogenizer and homogenized twice at 60 MPa. During the homogenization process, the discharge temperature was controlled not to exceed 30℃ to obtain Omega-3 fish oil emulsion. 7) Stabilization treatment Add 0.10 kg of xanthan gum and 0.20 kg of sodium citrate to the Omega-3 fish oil emulsion obtained in step 6), stir at 300 r / min for 20 min at 25 °C, adjust the pH to 6.8 with 0.1 mol / L citric acid aqueous solution, and filter through a 100 mesh sieve to obtain the Omega-3 fish oil emulsion composition.

[0030] Example 3

[0031] A method for preparing an Omega-3 fish oil emulsion composition is basically the same as that in Example 1, except that β-lactoglobulin is used instead of sodium caseinate, specifically including the following steps: 1) Preparation of zein solution Weigh 1.20 kg of zein and add it to 18.80 kg of 70% ethanol aqueous solution. Stir at 600 r / min for 40 min at 25℃ to fully dissolve the zein and obtain a zein solution. 2) Preparation of β-lactoglobulin aqueous phase Weigh 0.80 kg of β-lactoglobulin, 1.00 kg of trehalose, and 48.20 kg of deionized water, and stir at 500 r / min for 30 min at 30℃ to obtain the aqueous phase of β-lactoglobulin. The pH of the aqueous phase of β-lactoglobulin was tested to be 6.3, and the pH was adjusted to 6.8 using 0.1 mol / L sodium hydroxide aqueous solution. 3) Preparation of particulate dispersion The zein solution obtained in step 1) was added dropwise to the β-lactoglobulin aqueous phase obtained in step 2) at a rate of 0.20 L / min. During the dropwise addition, the stirring speed was maintained at 1200 r / min. After the dropwise addition was completed, stirring was continued for 60 min to allow the zein to undergo antisolvent precipitation in the aqueous phase and interfacial complexation with β-lactoglobulin. Subsequently, ethanol was removed by rotary evaporation at 35 °C and an absolute pressure of 0.02 MPa to obtain a zein-β-lactoglobulin particle dispersion. 4) Preparation of antioxidant fish oil phase Weigh out 30.00 kg of Omega-3 fish oil, 0.15 kg of natural vitamin E, and 0.05 kg of rosemary extract, and stir at 25°C in the dark for 20 min to obtain the antioxidant fish oil phase; 5) Initial emulsification The antioxidant fish oil phase obtained in step 4) was added to the particulate dispersion obtained in step 3) at a rate of 0.50 kg / min, and shearing emulsification was performed at 20°C using a high-speed shear emulsifier with a shearing speed of 10000 r / min and a shearing time of 5 min to obtain Omega-3 fish oil crude emulsion. 6) Homogenization and Refinement The Omega-3 fish oil crude emulsion obtained in step 5) was placed in a high-pressure homogenizer and homogenized twice at 60 MPa. During the homogenization process, the discharge temperature was controlled not to exceed 30℃ to obtain Omega-3 fish oil emulsion. 7) Stabilization treatment Add 0.10 kg of xanthan gum and 0.20 kg of sodium citrate to the Omega-3 fish oil emulsion obtained in step 6), stir at 300 r / min for 20 min at 25 °C, adjust the pH to 6.8 with 0.1 mol / L citric acid aqueous solution, and filter through a 100 mesh sieve to obtain the Omega-3 fish oil emulsion composition.

[0032] Comparative Example 1 A method for preparing an Omega-3 fish oil emulsion composition is basically the same as in Example 1, except that: instead of preparing a zein-sodium caseinate composite particle dispersion, an aqueous solution of sodium caseinate is used as the emulsion stabilizing phase. The method specifically includes the following steps: 1) Preparation of sodium caseinate aqueous phase Weigh out 2.00 kg of sodium caseinate, 1.00 kg of trehalose and 67.00 kg of deionized water, and stir at 500 r / min for 30 min at 30℃ to obtain the aqueous phase of sodium caseinate; 2) Preparation of antioxidant fish oil phase Weigh out 30.00 kg of Omega-3 fish oil, 0.15 kg of natural vitamin E, and 0.05 kg of rosemary extract, and stir at 25°C in the dark for 20 min to obtain the antioxidant fish oil phase; 3) Primary emulsification The antioxidant fish oil phase obtained in step 2) was added to the sodium caseinate aqueous phase obtained in step 1) at a rate of 0.50 kg / min. The mixture was sheared and emulsified at 20°C using a high-speed shear emulsifier with a shearing speed of 10000 r / min and a shearing time of 5 min to obtain Omega-3 fish oil crude emulsion. 4) Homogenization and Refinement The Omega-3 fish oil crude emulsion obtained in step 3) was placed in a high-pressure homogenizer and homogenized twice at 60 MPa. During the homogenization process, the discharge temperature was controlled not to exceed 30℃ to obtain Omega-3 fish oil emulsion. 5) Stabilization treatment Add 0.10 kg of xanthan gum and 0.20 kg of sodium citrate to the Omega-3 fish oil emulsion obtained in step 4), stir at 300 r / min for 20 min at 25 °C, adjust the pH to 6.8 with 0.1 mol / L citric acid aqueous solution, and filter through a 100 mesh sieve to obtain the Omega-3 fish oil emulsion composition.

[0033] Comparative Example 2 A method for preparing an Omega-3 fish oil emulsion composition is basically the same as in Example 1, except that: zein and sodium caseinate only form a common composite particle dispersion, specifically including the following steps: 1) Preparation of corn glycerin particle dispersion Weigh 1.20 kg of zein and add it to 18.80 kg of a 70% ethanol aqueous solution. Stir at 600 r / min for 40 min at 25 °C to fully dissolve the zein and obtain a zein solution. Add the zein solution dropwise to 48.20 kg of deionized water at a rate of 0.20 L / min, maintaining a stirring speed of 1200 r / min during the dropwise addition. After the dropwise addition is completed, continue stirring for 60 min to allow the zein to undergo antisolvent precipitation, obtaining a zein particle dispersion. 2) Preparation of ordinary protein composite particle dispersion Add 0.80 kg of sodium caseinate and 1.00 kg of trehalose to the zein particle dispersion obtained in step 1), and stir at 500 r / min for 30 min at 30℃; then remove ethanol by rotary evaporation at 35℃ and 0.02 MPa absolute pressure to obtain a zein-sodium caseinate common composite particle dispersion. 3) Preparation of antioxidant fish oil phase Weigh out 30.00 kg of Omega-3 fish oil, 0.15 kg of natural vitamin E, and 0.05 kg of rosemary extract, and stir at 25°C in the dark for 20 min to obtain the antioxidant fish oil phase; 4) Initial emulsification The antioxidant fish oil phase obtained in step 3) was added to the ordinary composite particle dispersion obtained in step 2) at a rate of 0.50 kg / min. The dispersion was sheared and emulsified at 20°C using a high-speed shear emulsifier with a shearing speed of 10000 r / min and a shearing time of 5 min to obtain Omega-3 fish oil crude emulsion. 5) Homogenization and Refinement The Omega-3 fish oil crude emulsion obtained in step 4) was placed in a high-pressure homogenizer and homogenized twice at 60 MPa. During the homogenization process, the discharge temperature was controlled not to exceed 30℃ to obtain Omega-3 fish oil emulsion. 6) Stabilization treatment Add 0.10 kg of xanthan gum and 0.20 kg of sodium citrate to the Omega-3 fish oil emulsion obtained in step 5), stir at 300 r / min for 20 min at 25 °C, adjust the pH to 6.8 with 0.1 mol / L citric acid aqueous solution, and filter through a 100 mesh sieve to obtain the Omega-3 fish oil emulsion composition.

[0034] Comparative Example 3 A method for preparing an Omega-3 fish oil emulsion composition is basically the same as that in Example 1, except that only zein particles are used as stabilizing particles, and sodium caseinate is not added. The method specifically includes the following steps: 1) Preparation of corn glycerin particle dispersion Weigh 2.00 kg of zein and add it to 31.00 kg of a 70% ethanol aqueous solution. Stir at 600 r / min for 40 min at 25 °C to fully dissolve the zein and obtain a zein solution. Add the zein solution dropwise to 37.00 kg of deionized water at a rate of 0.20 L / min, maintaining a stirring speed of 1200 r / min during the addition. After the addition is complete, continue stirring for 60 min to allow the zein to undergo anti-solvent precipitation. Then, remove the ethanol by rotary evaporation at 35 °C and an absolute pressure of 0.02 MPa to obtain a zein particle dispersion. 2) Preparation of antioxidant fish oil phase Weigh out 30.00 kg of Omega-3 fish oil, 0.15 kg of natural vitamin E, and 0.05 kg of rosemary extract, and stir at 25°C in the dark for 20 min to obtain the antioxidant fish oil phase; 3) Primary emulsification The antioxidant fish oil phase obtained in step 2) was added to the zein particle dispersion obtained in step 1) at a rate of 0.50 kg / min. The dispersion was sheared and emulsified at 20°C using a high-speed shear emulsifier with a shearing speed of 10000 r / min and a shearing time of 5 min to obtain Omega-3 fish oil crude emulsion. 4) Homogenization and Refinement The Omega-3 fish oil crude emulsion obtained in step 3) was placed in a high-pressure homogenizer and homogenized twice at 60 MPa. During the homogenization process, the discharge temperature was controlled not to exceed 30℃ to obtain Omega-3 fish oil emulsion. 5) Stabilization treatment Add 0.10 kg xanthan gum, 0.20 kg sodium citrate, and 1.00 kg trehalose to the Omega-3 fish oil emulsion obtained in step 4), stir at 300 r / min for 20 min at 25 °C, adjust the pH to 6.8 with 0.1 mol / L citric acid aqueous solution, and filter through a 100-mesh sieve to obtain the Omega-3 fish oil emulsion composition.

[0035] Comparative Example 4 A method for preparing an Omega-3 fish oil emulsion composition is basically the same as that in Example 1, except that: zein-pectin composite nanoparticles are used as the emulsion stabilizing phase, instead of zein-sodium caseinate particle dispersion. The specific steps include: 1) Preparation of alkaline zein solution Weigh 1.20 kg of zein and add it to 18.80 kg of sodium hydroxide aqueous solution with pH 12.0. Stir at 600 r / min for 40 min at 25℃ to fully dissolve the zein and obtain an alkaline zein solution. 2) Preparation of acidic pectin solution Weigh 0.80 kg of food-grade pectin and add it to 19.20 kg of citric acid aqueous solution with pH 2.5. Stir at 500 r / min for 30 min at 30℃ to obtain an acidic pectin solution. 3) Preparation of zein-pectin composite nanoparticle dispersion The alkaline zein solution obtained in step 1) was added dropwise to the acidic pectin solution obtained in step 2) at a rate of 0.20 L / min, while maintaining a stirring speed of 1200 r / min during the dropwise addition. After the dropwise addition was completed, stirring was continued for 60 min. The pH was adjusted to 7.0 using a 5% sodium hydroxide aqueous solution. Subsequently, 1.00 kg of trehalose and 29.00 kg of deionized water were added, and the mixture was stirred at 500 r / min for 30 min at 30 °C to obtain a zein-pectin composite nanoparticle dispersion. 4) Preparation of antioxidant fish oil phase Weigh out 30.00 kg of Omega-3 fish oil, 0.15 kg of natural vitamin E, and 0.05 kg of rosemary extract, and stir at 25°C in the dark for 20 min to obtain the antioxidant fish oil phase; 5) Initial emulsification The antioxidant fish oil phase obtained in step 4) was added to the zein-pectin composite nanoparticle dispersion obtained in step 3) at a rate of 0.50 kg / min. The dispersion was then sheared and emulsified at 20°C using a high-speed shear emulsifier with a shearing speed of 10000 r / min and a shearing time of 5 min to obtain Omega-3 fish oil crude emulsion. 6) Homogenization and Refinement The Omega-3 fish oil crude emulsion obtained in step 5) was placed in a high-pressure homogenizer and homogenized twice at 60 MPa. During the homogenization process, the discharge temperature was controlled not to exceed 30℃ to obtain Omega-3 fish oil emulsion. 7) Stabilization treatment Add 0.10 kg of xanthan gum and 0.20 kg of sodium citrate to the Omega-3 fish oil emulsion obtained in step 6), stir at 300 r / min for 20 min at 25 °C, adjust the pH to 6.8 with 0.1 mol / L citric acid aqueous solution, and filter through a 100 mesh sieve to obtain the Omega-3 fish oil emulsion composition.

[0036] Test Example 1 Emulsion physical stability test 100 mL of each of the Omega-3 fish oil emulsion compositions obtained in the examples and comparative examples were placed at 25°C for 24 hours before testing. The average particle size of the emulsion was determined according to GB / T 19077-2024 "Particle Size Analysis by Laser Diffraction". PDI and Zeta potential were measured using a nanoparticle size and Zeta potential analyzer at 25°C. Each sample was measured in triplicate, and the average value was taken. Additionally, 50 mL of each sample was placed in a stoppered graduated cylinder and allowed to stand at 25°C for 30 days. The presence of stratification and oil separation was observed, and the average particle size after 30 days of standing was measured. The particle size change rate was calculated using the following formula: Particle size change rate / % = (average particle size after 30 days of standing - initial average particle size) / initial average particle size × 100%.

[0037] Take another 10.00g of each sample and place it in a centrifuge tube. Centrifuge at 4000r / min for 10min, observe the oil separation, and calculate the oil separation rate according to the following formula: Oil separation rate / % = Mass of free oil in the upper layer after centrifugation / Mass of sample × 100%.

[0038] Table 1. Physical stability test results of each sample Example 1 420 0.18 -35.2 455 8.3 No obvious oil-water separation was observed. 0.2 Example 2 475 0.21 -33.1 540 13.7 No obvious oil-water separation was observed. 0.5 Example 3 510 0.23 -30.8 610 19.6 No obvious stratification, slight flocculation 0.8 Comparative Example 1 780 0.31 -28.4 1120 43.6 Slight oil separation in the upper layer 3.8 Comparative Example 2 610 0.27 -29.6 805 32 No obvious stratification, but increased emulsion turbidity. 1.6 Comparative Example 3 950 0.39 -20.5 1680 76.8 Obvious oil-water separation was observed. 8.9 Comparative Example 4 560 0.24 -31.6 700 25.0 No obvious stratification, slight flocculation 1.0 Table 1 shows that the Omega-3 fish oil emulsion composition obtained in Example 1 had an initial average particle size of 420 nm, a PDI of 0.18, a particle size change rate of only 8.3% after standing for 30 days, and a centrifugal oil separation rate of 0.2%, exhibiting the best physical stability. Examples 2 and 3 used whey protein isolate and β-lactoglobulin, respectively, to replace sodium caseinate. Both were able to form protein-protein complex particles with zein, resulting in emulsion stability superior to the zein-pectin composite nanoparticle system in Comparative Example 4. However, their particle size, PDI, particle size change rate, and centrifugal oil separation rate were still higher than those of Example 1. This indicates that among similar protein complex particles, the composite particles formed by sodium caseinate and zein are more beneficial for improving emulsion particle size stability and anti-oil separation performance. The possible reason is that the surface of the newly formed hydrophobic particles formed by zein during the antisolvent precipitation process can adsorb sodium caseinate. The hydrophilic segments of sodium caseinate face the aqueous phase, while the hydrophobic regions of zein are more likely to face the oil phase. This may cause the resulting composite particles to exhibit an amphiphilic interface orientation similar to the Janus structure at the oil-water interface, thereby improving the stability of the fish oil droplet interface.

[0039] Test Example 2 Emulsion oxidative stability test Take 100g each of the Omega-3 fish oil emulsion compositions obtained in the examples and comparative examples, place them in 250mL brown glass bottles, retain headspace in the bottles, seal them, and store them in a 40℃ constant temperature incubator protected from light for 30 days. Samples were taken on storage day 0 and day 30, and the oil phase in the emulsion was extracted using a hexane-isopropanol mixed solvent. After recovering the oil phase, oxidation index was tested.

[0040] Peroxide value was determined according to GB 5009.227-2023, "National Food Safety Standard: Determination of Peroxide Value in Food".

[0041] The anisidine value was determined according to GB / T 24304-2024 "Determination of Anisidine Value in Animal and Vegetable Oils"; the TOTOX value was calculated according to the following formula: TOTOX value = 2 × peroxide value + anisidine value. The peroxide value is expressed as meq O2 / kg oil.

[0042] The EPA and DHA contents were determined according to GB 5009.168-2016 "National Food Safety Standard - Determination of Fatty Acids in Food", and the EPA+DHA retention rate was calculated according to the following formula: EPA+DHA retention rate / % = Total EPA and DHA content after 30 days of storage / Total EPA and DHA content at 0 days of storage × 100%.

[0043] Table 2. Results of oxidative stability tests for each sample Example 1 4.3 7.2 6.4 20.8 95.6 Example 2 4.3 8.4 7.5 24.3 94.1 Example 3 4.4 9.1 8.0 26.2 93.5 Comparative Example 1 4.4 15.8 13.6 45.2 86.7 Comparative Example 2 4.4 11.6 10.2 33.4 90.9 Comparative Example 3 4.5 19.4 17.8 56.6 82.4 Comparative Example 4 4.4 9.8 8.7 28.3 92.8 As shown in Table 2, after 30 days of storage at 40℃ in the dark, Example 1 exhibited the lowest peroxide value, anisidine value, and TOTOX value, while showing the highest EPA+DHA retention rate. The oxidative stability of Examples 2 and 3 was superior to that of Comparative Example 4, indicating that the protein-protein composite particles formed by zein and milk protein components have a better oxidative inhibition effect on Omega-3 fish oil compared to zein-pectin-polysaccharide composite particles. The superiority of Example 1 over Examples 2 and 3 may be related to the more flexible molecular chains and better interfacial spreadability of sodium caseinate, which allows for the formation of a more continuous hydrophilic stable layer on the surface of newly formed zein particles. Although Comparative Example 2 also used zein and sodium caseinate, it employed a common composite method of first preparing zein particles and then adding sodium caseinate, resulting in significantly weaker oxidative stability compared to Example 1. These results demonstrate that, under the test conditions of this application, the emulsion obtained through the in-situ interfacial composite method exhibits a lower degree of oxidative degradation. The possible reason is that during the antisolvent precipitation process, sodium caseinate simultaneously participates in the complexation of the surface of the newly formed zein particles, which may cause the complex particles to form an amphiphilic interface orientation similar to the Janus structure at the oil-water interface. This is conducive to the formation of a more stable interface barrier layer, thereby reducing the contact between fish oil and the aqueous phase, oxygen and pro-oxidation factors.

[0044] Test Example 3 In vitro simulated gastrointestinal digestion and release test Preparation of simulated gastric juice: Weigh 2.00g of sodium chloride, add it to about 800mL of deionized water and stir to dissolve. Add pepsin to make the pepsin activity 2000U / mL. Adjust the pH to 2.0 with 1.0mol / L hydrochloric acid solution. Finally, make up to 1000mL with deionized water. Prepare fresh and use immediately. Preheat to 37℃ before use.

[0045] Preparation of simulated intestinal fluid: Weigh 6.80 g of potassium dihydrogen phosphate, 5.00 g of sodium chloride and 0.20 g of calcium chloride, add to about 800 mL of deionized water and stir to dissolve. Add sodium taurocholate to make its concentration 10 mmol / L. Add pancreatic lipase to make the pancreatic lipase activity 2000 U / mL. Adjust the pH to 7.0 with 1.0 mol / L sodium hydroxide aqueous solution. Finally, make up to 1000 mL with deionized water. Prepare fresh and use immediately. Preheat to 37℃ before use.

[0046] Gastric digestion: 10.00 g of each of the Omega-3 fish oil emulsion compositions obtained in the examples and comparative examples were added to 90 mL of simulated gastric juice preheated to 37°C. The pH of the system was adjusted to 2.0 ± 0.1 using 1.0 mol / L hydrochloric acid, and the mixture was shaken and digested at 37°C and 100 r / min for 2 h. After digestion, 10.00 g of the gastric digestion fluid was taken, centrifuged at 4000 r / min for 10 min, and the upper free oil phase was collected and weighed. The gastric oil separation rate was calculated according to the following formula: Oil separation rate in the stomach stage / % = mass of free oil in the upper layer of the stomach stage / mass of fish oil in the sample × 100%.

[0047] Another 10.00 g of gastric digestive fluid was taken, and the fatty acids in the dispersible oil phase were extracted using a hexane-isopropanol mixed solvent. The EPA and DHA contents were determined according to GB 5009.168-2016 "National Food Safety Standard - Determination of Fatty Acids in Food", and the EPA+DHA release rate in the stomach was calculated using the following formula: Gastric stage EPA+DHA release rate / % = Total amount of EPA and DHA in the dispersible oil phase of gastric digestive fluid / Total amount of EPA and DHA in the sample × 100%.

[0048] Intestinal digestion: After gastric digestion, the pH of the gastric digestive fluid was adjusted to 7.0 using a 1.0 mol / L sodium hydroxide aqueous solution. Simulated intestinal fluid preheated to 37°C was added at a volume ratio of 1:1, and digestion continued with shaking at 37°C and 100 rpm for 4 hours. After digestion, the mixture was centrifuged at 10000 rpm for 20 minutes, and the supernatant micelle phase was collected. The EPA and DHA contents were determined according to GB 5009.168-2016 "National Food Safety Standard - Determination of Fatty Acids in Food," and the EPA+DHA micellarization rate was calculated using the following formula: EPA+DHA micellization rate / % = Total amount of EPA and DHA in micelles / Total amount of EPA and DHA in the sample × 100%.

[0049] Table 3 Results of in vitro simulated gastrointestinal digestion and release tests for each sample Example 1 2.8 8.6 82.4 The gastric stage is basically stable, and the intestinal stage releases sufficient nutrients. Example 2 3.5 9.7 80.1 The gastric stage is relatively stable, while the intestinal stage releases more fully. Example 3 4.1 10.2 78.8 Slight accumulation in the stomach stage, and more complete release in the intestine stage. Comparative Example 1 9.5 18.7 65.8 Oil droplets accumulate in the stomach stage. Comparative Example 2 5.6 12.4 73.6 Slight accumulation in the stomach stage, and more complete release in the intestine stage. Comparative Example 3 18.2 31.5 58.2 Significant oil separation occurs in the gastric stage, while dispersion is poor in the intestinal stage. Comparative Example 4 4.6 10.9 77.6 The gastric stage is relatively stable, while the intestinal stage releases more fully. Table 3 shows that Example 1 exhibited the lowest oil separation rate and EPA+DHA release rate in simulated gastric fluid, indicating that it better maintained the emulsion structure and reduced premature fish oil separation under gastric acid conditions. Upon entering simulated intestinal fluid, Example 1 achieved an EPA+DHA micellization rate of 82.4%, higher than other samples. Examples 2 and 3, using whey protein isolate and β-lactoglobulin instead of sodium caseinate, showed better gastric stability and intestinal micellization rate than Comparative Example 4, but still lower than Example 1. These results demonstrate that the zein-sodium caseinate composite particles not only improve the physical and oxidative stability of the emulsion but also better balance gastric protection and intestinal release, thus enhancing the in vitro bioavailability of EPA and DHA.

[0050] The preferred embodiments of the present invention have been described in detail above. It should be understood that those skilled in the art can make numerous modifications and variations based on the concept of the present invention without creative effort. Therefore, all technical solutions that can be obtained by those skilled in the art based on the concept of the present invention through logical analysis, reasoning, or limited experimentation on the basis of existing technology should be within the scope of protection defined by the claims.

Claims

1. A method for preparing an Omega-3 fish oil emulsion composition, characterized in that, Includes the following steps: 1) Add zein to an aqueous ethanol solution and stir to dissolve, thus obtaining a zein solution; 2) Mix the hydrophilic protein, sugar protectant, and water, stir to dissolve, and obtain the hydrophilic protein aqueous phase; 3) The zein solution obtained in step 1) is added dropwise to the aqueous phase of the hydrophilic protein obtained in step 2), so that the zein undergoes antisolvent precipitation in the aqueous phase and undergoes in-situ interfacial complexation with the hydrophilic protein; then the ethanol is removed to obtain a zein-hydrophilic protein composite particle dispersion. 4) Mix Omega-3 fish oil, fat-soluble antioxidants, and plant antioxidant extracts to obtain an antioxidant fish oil phase; 5) Add the antioxidant fish oil phase obtained in step 4) to the zein-hydrophilic protein composite particle dispersion obtained in step 3), and shear emulsify to obtain Omega-3 fish oil crude emulsion; 6) The crude Omega-3 fish oil emulsion obtained in step 5) is subjected to high-pressure homogenization to obtain an Omega-3 fish oil emulsion; 7) Add thickener and buffer salt to the Omega-3 fish oil emulsion obtained in step 6), stir and filter to obtain Omega-3 fish oil emulsion composition; The raw materials, by weight, are: 0.8-2.0 parts zein, 9.6-40 parts ethanol aqueous solution, 0.4-1.2 parts hydrophilic protein, 0.5-1.5 parts saccharide protectant, 40-60 parts water, 20-40 parts Omega-3 fish oil, 0.05-0.30 parts fat-soluble antioxidant, 0.01-0.10 parts plant antioxidant extract, 0.05-0.30 parts thickener and stabilizer, and 0.05-0.50 parts buffer salt.

2. The method for preparing the Omega-3 fish oil emulsified composition as described in claim 1, characterized in that: In step 1), the stirring temperature is 20-30℃, the stirring speed is 400-800 r / min, and the stirring time is 30-60 min; the mass fraction of the ethanol aqueous solution is 60%-80%.

3. The method for preparing the Omega-3 fish oil emulsified composition as described in claim 1, characterized in that: In step 2), the hydrophilic protein is one of sodium caseinate, whey protein isolate, and β-lactoglobulin; the sugar protectant is one or more of trehalose, sucrose, maltose, and maltodextrin.

4. The method for preparing the Omega-3 fish oil emulsified composition as described in claim 1, characterized in that: In step 3), the dripping rate of the corn gliadin solution is 0.05-0.50 L / min; the stirring speed during the dripping process is 800-1500 r / min; after the dripping is completed, stirring is continued for 30-90 min; the conditions for removing ethanol are 30-40℃ and absolute pressure 0.01-0.04 MPa.

5. The method for preparing the Omega-3 fish oil emulsified composition as described in claim 1, characterized in that: In step 4), the total mass content of EPA and DHA in the Omega-3 fish oil is not less than 50%; the fat-soluble antioxidant is one or more of natural vitamin E, vitamin E acetate, and ascorbyl palmitate; and the plant antioxidant extract is one or more of rosemary extract, tea polyphenol oil dispersion, and astaxanthin oil dispersion.

6. The method for preparing the Omega-3 fish oil emulsified composition as described in claim 5, characterized in that: The fat-soluble antioxidant is natural vitamin E; the plant antioxidant extract is rosemary extract.

7. The method for preparing the Omega-3 fish oil emulsified composition as described in claim 1, characterized in that: In step 5), the addition rate of the antioxidant fish oil phase is 0.20-1.00 kg / min; the shear emulsification temperature is 15-25℃; the shearing speed is 8000-12000 r / min; and the shearing time is 3-10 min.

8. The method for preparing the Omega-3 fish oil emulsified composition as described in claim 1, characterized in that: In step 6), the high-pressure homogenization pressure is 40-80 MPa, the homogenization is performed 1-3 times, and the discharge temperature is controlled not to exceed 35℃ during the homogenization process.

9. The method for preparing the Omega-3 fish oil emulsified composition as described in claim 1, characterized in that: In step 7), the thickening stabilizer is one or more of xanthan gum, guar gum, sodium carboxymethyl cellulose, and sodium alginate; the buffer salt is one or more of sodium citrate, disodium hydrogen phosphate, and sodium dihydrogen phosphate.

10. An Omega-3 fish oil emulsified composition, characterized in that, Prepared by the method according to any one of claims 1-9.