Fish oil emulsion stabilized by soybean protein-herbaceous grass polysaccharide and preparation method thereof

Abstract

The present invention discloses a fish oil emulsion stabilized by soybean protein-herb polysaccharide and a preparation method thereof. The fish oil emulsion is mainly made of fish oil, soybean protein isolate, herb polysaccharide and water, and its preparation method comprises: (1) adding fish oil to a soybean protein isolate aqueous solution under stirring and then sequentially performing dispersion treatment and homogenization treatment to obtain a primary fish oil emulsion; (2) mixing and stirring it with herb polysaccharide to obtain. When preparing the fish oil emulsion, the present invention combines soybean protein isolate and herb polysaccharide, and the two cooperate with each other to significantly increase the physical stability and oxidative stability of the fish oil emulsion. The present invention further finds that after the soybean protein isolate is ultrasonically treated and then combined with herb polysaccharide to prepare the fish oil emulsion, the binding ability of soybean protein powder and herb polysaccharide can be significantly improved compared with that without ultrasonic treatment, and the prepared ultrasonic soybean protein-herb polysaccharide stabilized fish oil emulsion has higher physical stability and oxidative stability.

Description

Technical Field

[0001] The invention relates to a fish oil emulsion preparation, in particular to a soybean protein-herbaceous grass polysaccharide-stabilized fish oil emulsion with high physical stability and oxidation stability and a preparation method thereof, belonging to the field of fish oil emulsion and its preparation. Background Art

[0002] Fish oil is rich in various unsaturated fatty acids such as linoleic acid, linolenic acid, arachidonic acid, eicosapentaenoic acid and docosahexaenoic acid, and has various health benefits such as lowering blood pressure and blood lipids. However, it has low water solubility and poor stability, and is easily affected by the external environment (light, oxygen, heat) and oxidized, resulting in decreased physiological activity and reduced bioavailability. Therefore, it is necessary to construct a suitable carrier form to reduce fish oil oxidation.

[0003] The emulsion has a small particle size and a large specific surface area. It has a good embedding, protection and transfer effect on the carried substances, and improves the biological activity and bioavailability of the carried substances. Therefore, the emulsion is a good delivery system for fish oil. The emulsification activity and emulsification stability of the emulsifier are the key to ensuring the good delivery effect of fish oil. At present, most commonly used emulsifiers are chemically synthesized small molecule surfactants, such as Tween, ounce, etc., which have certain food safety risks. In contrast, natural food-based emulsifiers are favored by consumers due to their higher food safety. Soy protein has good emulsification activity, low cost, and high biosafety, but poor emulsification stability, and is easily denatured by environmental influences, resulting in decreased emulsification activity. Therefore, the physical stability of the emulsion stabilized only by soy protein decreases during long-term storage and in the presence of certain salt ions, and it is easy to break the emulsion, resulting in a deeper degree of oxidation of the emulsion and a decrease in the activity of the carried functional ingredients.

[0004] Grass jelly polysaccharide (also known as jelly grass polysaccharide) is a water-soluble anionic heteropolysaccharide extracted from jelly grass (Mesona chinensis Benth.), rich in uronic acid, has a certain thickening ability in aqueous solution, and has good gel properties. However, the current application of grass jelly polysaccharide is mainly concentrated in gel food and health tea, such as grass jelly, tortoise age paste, herbal tea, etc., and there is no application of grass jelly polysaccharide in nutrient delivery emulsion system, especially the study of interaction with protein on the physical and oxidative stability of emulsion has not been reported. Summary of the invention

[0005] One of the objects of the present invention is to provide a fish oil emulsion with high physical stability and oxidative stability;

[0006] The second object of the present invention is to provide a method for preparing the fish oil emulsion;

[0007] The above object of the present invention is achieved through the following technical solutions:

[0008] One aspect of the present invention provides a soy protein-herb jelly polysaccharide stabilized fish oil emulsion, the main components of the fish oil emulsion include fish oil, soy protein isolate, herb jelly polysaccharide and water; preferably, in terms of mass percentage, the dosage of each component is: 2-10% of fish oil, 1.5-4% of soy protein isolate, 0.01-0.4% of herb jelly polysaccharide, and the balance is water; more preferably, the dosage of each component is: 5% of fish oil, 2.0% of soy protein isolate, and 0.05% of herb jelly polysaccharide.

[0009] As a preferred specific embodiment of the present invention, the soy protein isolate is soy protein isolate that has been treated with ultrasound; for reference, the ultrasonic treatment parameters of the ultrasonic treatment may be an ultrasonic power of 150-450W and an ultrasonic time of 1-20min; preferably, the ultrasonic power is 300W and the ultrasonic time is 15min.

[0010] Another aspect of the present invention is to provide a method for preparing the soy protein-herb jelly polysaccharide stabilized fish oil emulsion, the preparation method comprising: (1) adding fish oil to a soy protein isolate aqueous solution under stirring, and then performing dispersion treatment and homogenization treatment in sequence to obtain a primary fish oil emulsion; (2) mixing and stirring the primary fish oil emulsion with a herb jelly polysaccharide solution to obtain a soy protein-herb jelly polysaccharide stabilized fish oil emulsion.

[0011] In a preferred specific embodiment of the preparation method of the present invention, the soy protein isolate aqueous solution in step (1) is an ultrasonically treated soy protein isolate aqueous solution; for reference, the ultrasonic treatment parameters of the ultrasonic treatment may be an ultrasonic power of 150-450W and an ultrasonic time of 1-20min. Preferably, the ultrasonic power is 300W and the ultrasonic time is 15min.

[0012] In a preferred specific embodiment of the preparation method of the present invention, the stirring in step (1) is magnetic stirring, and preferably, the rotation speed of the magnetic stirring is 200-300 rpm.

[0013] In a preferred embodiment of the preparation method of the present invention, the dispersion treatment in step (1) is carried out in a high-speed shearing machine at a rotation speed of 10,000-15,000 rpm for 2-5 minutes.

[0014] In a preferred specific embodiment of the preparation method of the present invention, the homogenization treatment in step (1) is carried out in a high-pressure homogenizer at a pressure of 40-140 MPa for 2-4 times; preferably, the high-pressure homogenization pressure is 120 MPa.

[0015] In a preferred specific implementation scheme of the preparation method of the present invention, the stirring in step (2) is magnetic stirring. Preferably, the rotation speed of the magnetic stirring is 200-300 rpm, and the magnetic stirring time is 10-60 min.

[0016] The present invention finds that when preparing fish oil emulsion, adding a certain amount of grass jelly polysaccharide can significantly enhance the physical stability of the emulsion, thereby improving the oxidative stability of the emulsion.

[0017] In order to further clarify the effect of Herba Epimedii polysaccharides on improving the physical and oxidative stability of fish oil emulsion stabilized by soy protein, the present invention prepares fish oil emulsions stabilized by soy protein-carrageenan, soy protein-xanthan gum, and soy protein-Herba Epimedii polysaccharides by combining soy protein isolate with carrageenan, xanthan gum, and soy protein-Herba Epimedii polysaccharides, respectively. The results show that the fish oil emulsion prepared by soy protein-Herba Epimedii polysaccharides is significantly superior to other combinations in terms of physical and oxidative stability.

[0018] The present invention further discovered that after soy protein isolate is treated with ultrasound, an ultrasound-stabilized primary fish oil emulsion stabilized by soy protein is prepared using the soy protein, and then grass jelly polysaccharide is added to obtain an ultrasound-stabilized soy protein-grass jelly polysaccharide fish oil emulsion. Compared with the soy protein-grass jelly polysaccharide fish oil emulsion prepared without ultrasound treatment, the ultrasound-treated soy protein isolate significantly improves the binding ability of soy protein and grass jelly polysaccharide. The physical and oxidative stability of the prepared ultrasound-stabilized soy protein-grass jelly polysaccharide fish oil emulsion during storage at 37°C for 60 days and at 500mM NaCl concentration for 30 days at 25°C are significantly improved.

[0019] In the preparation of the fish oil emulsion, the present invention combines soy protein isolate and herba schoenopsis polysaccharides, and the two interact and cooperate with each other, significantly increasing the physical stability and oxidative stability of the fish oil emulsion. The present invention further finds that after the soy protein isolate is ultrasonically treated and combined with herba schoenopsis polysaccharides, the binding ability of soy protein and herba schoenopsis polysaccharides is more effectively improved. Compared with the soy protein-herba schoenopsis polysaccharides fish oil emulsion prepared without ultrasonic treatment, the soy protein-herba schoenopsis polysaccharides fish oil emulsion prepared by ultrasonic treatment has significantly improved physical and oxidative stability during storage at 37°C for 60 days and at 500mM NaCl concentration at 25°C for 30 days. The present invention provides a new idea for improving the biological activity and bioavailability of fish oil. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 The average particle size of fish oil emulsion stored at pH 7 and 37°C for 60 days; (a) the average particle size of soy protein-polysaccharide fish oil emulsion without ultrasonication, (b) the average particle size of soy protein-polysaccharide fish oil emulsion with ultrasonication.

[0021] Figure 2POV and TBARS values ​​of fish oil emulsions stored at pH 7 and 37°C for 60 days; (a) POV value of soy protein-polysaccharide fish oil emulsion without ultrasonication, (b) POV value of soy protein-polysaccharide fish oil emulsion with ultrasonication, (c) TBARS value of soy protein-polysaccharide fish oil emulsion without ultrasonication, (d) TBARS value of soy protein-polysaccharide fish oil emulsion with ultrasonication.

[0022] Figure 3 The protein fluorescence intensity values ​​of fish oil emulsions stored at pH 7 and 37°C for 60 days; (a) fluorescence intensity value of soy protein-polysaccharide fish oil emulsion without ultrasonic treatment, (b) fluorescence intensity value of soy protein-polysaccharide fish oil emulsion with ultrasonic treatment.

[0023] Figure 4 The average particle size of fish oil emulsion during storage at 500 mM NaCl concentration and 25°C; (a) the average particle size of soy protein-polysaccharide fish oil emulsion without ultrasonic treatment, (b) the average particle size of soy protein-polysaccharide fish oil emulsion with ultrasonic treatment.

[0024] Figure 5 POV and TBARS values ​​of fish oil emulsions during storage at 500 mM NaCl concentration and 25°C; (a) POV value of soy protein-polysaccharide fish oil emulsion without ultrasonication, (b) POV value of soy protein-polysaccharide fish oil emulsion with ultrasonication, (c) TBARS value of soy protein-polysaccharide fish oil emulsion without ultrasonication, (d) TBARS value of soy protein-polysaccharide fish oil emulsion with ultrasonication.

[0025] Figure 6 The protein fluorescence intensity values ​​of fish oil emulsions during storage at 500 mM NaCl concentration and 25°C; (a) fluorescence intensity value of soy protein-polysaccharide fish oil emulsion without ultrasonic treatment, (b) fluorescence intensity value of soy protein-polysaccharide fish oil emulsion with ultrasonic treatment.

[0026] Figure 7 Changes in average particle size of SPI-H fish oil emulsion treated with different ultrasonic conditions during storage at 25°C.

[0027] Figure 8 Changes in POV value of SPI-H fish oil emulsion treated with different ultrasonic conditions during storage at 25℃.

[0028] Fig. 9 Changes in TBARS values ​​of SPI-H fish oil emulsion treated with different ultrasonic conditions during storage at 25℃.

[0029] Fig.10 These are the test results for parameter screening of different homogenization pressures during the preparation of fish oil emulsion.

[0030] Fig.11These are the screening test results of different polysaccharide concentrations in the preparation process of fish oil emulsions; soy protein-herbaceous grass polysaccharide emulsion (SPI-H), soy protein-carrageenan emulsion (SPI-IC), soy protein-xanthan gum emulsion (SPI-X). DETAILED DESCRIPTION

[0031] The present invention will be further described below in conjunction with specific embodiments, and the advantages and features of the present invention will become clearer as the description proceeds. However, these embodiments are exemplary only and do not constitute any limitation to the scope of the present invention. It should be understood by those skilled in the art that the details and forms of the present invention may be modified or replaced without departing from the spirit and scope of the present invention, but these modifications and replacements all fall within the scope of protection of the present invention.

[0032] Example 1 Preparation of non-ultrasonicated soybean protein-herbaceous grass polysaccharide fish oil emulsion

[0033] Fish oil was dripped into a non-ultrasonicated soy protein isolate phosphate buffer solution (the concentration of the phosphate buffer solution was 5 mM, pH 7.0) under magnetic stirring (rotation speed was 250 rpm), and dispersed at 12,000 rpm for 3 minutes using a high-speed shearing machine to obtain a crude fish oil emulsion, which was then homogenized 3 times at 120 MPa using a high-pressure homogenizer to form a primary emulsion; the primary emulsion was then mixed with a herba schonocarpa polysaccharide solution and magnetically stirred for 30 minutes (rotation speed was 250 rpm) to obtain a non-ultrasonicated soy protein-herba schonocarpa polysaccharide fish oil emulsion, the emulsion had a pH of 7.0, was placed in a glass bottle, and stored at 37° C. in the dark; wherein the percentage content of each component in the prepared non-ultrasonicated soy protein-herba schonocarpa polysaccharide fish oil emulsion was: 5% (w / w) fish oil, 2.0% (w / w) soy protein isolate, 0.05% (w / w) herba schonocarpa polysaccharide, and the remainder was water.

[0034] Example 2 Preparation of non-ultrasonicated soybean protein-herbaceous grass polysaccharide fish oil emulsion

[0035] Fish oil is dripped into an unultrasonicated soy protein isolate aqueous solution under magnetic stirring (rotation speed of 200 rpm), and dispersed at 10,000 rpm for 5 minutes using a high-speed shearing machine, and then homogenized 4 times at 60 MPa using a high-pressure homogenizer to form a primary emulsion; the primary emulsion is then mixed with the herba schoenoprasum polysaccharide solution, and magnetic stirring is performed for 30 minutes (rotation speed of 200 rpm) to obtain an unultrasonicated soy protein-herba schoenoprasum polysaccharide fish oil emulsion, the emulsion has a pH of 7.0, is placed in a glass bottle, and is stored at 37° C. in the dark; wherein the percentage content of each component in the prepared unultrasonicated soy protein-herba schoenoprasum polysaccharide fish oil emulsion is: 2% (w / w) fish oil, 1.5% (w / w) soy protein isolate, 0.1% (w / w) herba schoenoprasum polysaccharide, and the remainder is water.

[0036] Example 3 Preparation of non-ultrasonicated soybean protein-herbaceous grass polysaccharide fish oil emulsion

[0037] Fish oil is dripped into an unultrasonicated soy protein isolate aqueous solution under magnetic stirring (rotation speed is 300 rpm), and dispersed at 15,000 rpm for 2 minutes by a high-speed shearing machine, and then homogenized twice by a high-pressure homogenizer at 100 MPa to form a primary emulsion; then the primary emulsion is mixed with a certain proportion of grass jelly polysaccharide solution, and magnetic stirring is performed for 30 minutes (rotation speed is 300 rpm) to obtain a non-ultrasonicated soy protein-grass jelly polysaccharide fish oil emulsion, the emulsion has a pH of 7.0, is placed in a glass bottle, and is stored at 37° C. in the dark; wherein the percentage content of each component in the prepared non-ultrasonicated soy protein-grass jelly polysaccharide fish oil emulsion is: 7.5% (w / w) fish oil, 3% (w / w) soy protein isolate, 0.2% (w / w) grass jelly polysaccharide, and the remainder is water.

[0038] Example 4 Preparation of Ultrasonic Soy Protein-Ceratonia Polysaccharide Fish Oil Emulsion

[0039] Under magnetic stirring (rotation speed of 250 rpm), fish oil is dropped into a soy protein isolate phosphate buffer solution (the concentration of the phosphate buffer solution is 5 mM, pH 7.0) that has been ultrasonically treated (ultrasonic treatment parameters are: ultrasonic power 300 W, ultrasonic time 15 min), and dispersed at 12,000 rpm for 3 min using a high-speed shearing machine, and then homogenized 3 times at 120 MPa using a high-pressure homogenizer to form a primary emulsion; the primary emulsion is then mixed with the herba schoenoprasum polysaccharide solution, and magnetic stirring is performed for 30 min (rotation speed of 250 rpm) to obtain an ultrasonic soy protein-herba schoenoprasum polysaccharide fish oil emulsion, the emulsion has a pH of 7.0, is placed in a glass bottle, and is stored at 37° C. in the dark; wherein the percentage content of each component in the prepared ultrasonic soy protein-herba schoenoprasum polysaccharide fish oil emulsion is: 5% (w / w) fish oil, 2.0% (w / w) soy protein isolate, 0.05% (w / w) herba schoenoprasum polysaccharide, and the remainder is water.

[0040] Example 5 Preparation of ultrasonic soybean protein-herbaceous celery polysaccharide fish oil emulsion

[0041] Fish oil was dripped into an ultrasonically treated (ultrasonic treatment parameters: ultrasonic power 150 W, ultrasonic time 20 min) soy protein isolate aqueous solution under magnetic stirring (rotation speed 200 rpm), and dispersed at 10,000 rpm for 5 min by a high-speed shearing machine, and then homogenized 4 times at 60 MPa by a high-pressure homogenizer to form a primary emulsion; the primary emulsion was then mixed with the herba schoenoprasum polysaccharide solution, and magnetically stirred for 30 min (rotation speed 200 rpm) to obtain an ultrasonic soy protein-herba schoenoprasum polysaccharide fish oil emulsion, the emulsion pH 7.0, placed in a glass bottle, and stored at 37° C. in the dark; wherein the percentage content of each component in the prepared ultrasonic soy protein-herba schoenoprasum polysaccharide fish oil emulsion is: 2% (w / w) fish oil, 1.5% (w / w) soy protein isolate, 0.1% (w / w) herba schoenoprasum polysaccharide, and the remainder is water.

[0042] Example 6 Preparation of Ultrasonic Soy Protein-Ceratonia Polysaccharide Fish Oil Emulsion

[0043] Under magnetic stirring (rotation speed of 300 rpm), fish oil is dropped into an ultrasonically treated (ultrasonic treatment parameters are: ultrasonic power 450 W, ultrasonic time 5 min) soy protein isolate aqueous solution, and dispersed at 15,000 rpm for 2 min using a high-speed shearing machine, and then homogenized twice at 100 MPa using a high-pressure homogenizer to form a primary emulsion; then the primary emulsion is mixed with a certain proportion of herba schoenoprasum polysaccharides solution, and magnetic stirring is performed for 30 min (rotation speed of 300 rpm) to obtain an ultrasonic soy protein-herba schoenoprasum polysaccharides fish oil emulsion, the emulsion has a pH of 7.0, is placed in a glass bottle, and is stored at 37° C. in the dark; wherein the percentage content of each component in the prepared ultrasonic soy protein-herba schoenoprasum polysaccharides fish oil emulsion is: 7.5% (w / w) fish oil, 3% (w / w) soy protein isolate, 0.2% (w / w) herba schoenoprasum polysaccharides, and the remainder is water.

[0044] Experimental Example 1 Comparative test on the performance of fish oil emulsions with ultrasonically treated soy protein isolate and without ultrasonically treated soy protein isolate in the preparation of fish oil emulsion

[0045] 1 Test method

[0046] 1.1 Preparation of fish oil emulsion without NaCl

[0047] Under magnetic stirring (speed 250rpm), an appropriate amount of fish oil was dropped into the soy protein isolate aqueous solution without ultrasonic treatment and ultrasonic treatment (ultrasonic treatment parameters: ultrasonic power 300W, ultrasonic time 15min) respectively, and dispersed for 3min at 12,000rpm with a high-speed shearing machine, and then homogenized three times at 120MPa with a high-pressure homogenizer to form a primary emulsion. Subsequently, the primary emulsion was mixed with a certain proportion of Herba Ceropegiae polysaccharides, carrageenan and xanthan gum solutions, respectively, and magnetically stirred for 30 min (250 rpm) to prepare non-ultrasonicated soy protein-Herba Ceropegiae polysaccharide emulsion, non-ultrasonicated soy protein-carrageenan emulsion, non-ultrasonicated soy protein-xanthan gum emulsion, ultrasonicated soy protein-Herba Ceropegiae polysaccharide emulsion, ultrasonicated soy protein-carrageenan emulsion and ultrasonicated soy protein-xanthan gum emulsion, respectively. The percentage content of each component in the emulsion was: 5% (w / w) fish oil, 2.0% (w / w) protein, 0.05% (w / w) Herba Ceropegiae polysaccharides (H) or 0.05% (w / w) carrageenan (IC) or 0.2% (w / w) xanthan gum (X), the emulsion had a pH of 7.0, was placed in a glass bottle, and stored in the dark at 37° C. for 60 days, and the average particle size, hydroperoxide (POV), thiobarbituric acid (TBARS) value and fluorescence intensity value of the emulsion were regularly monitored.

[0048] 1.2 Preparation of fish oil emulsion containing 500 mM NaCl

[0049] Place the unsonicated soy protein-polysaccharide fish oil emulsion and the sonicated soy protein (300W ultrasound for 15 min)-polysaccharide fish oil emulsion prepared in 1.1 in a beaker, add NaCl solution, mix evenly, and make the NaCl concentration in the final sample be 500 mM. Store at 25°C in the dark for 30 days, and regularly monitor the average particle size, POV, TBARS value and fluorescence intensity value of the emulsion.

[0050] 1.3 Determination method of emulsion performance index

[0051] 1.3.1 Average particle size of emulsion

[0052] The average particle size of the emulsion was measured using a Malvern laser particle size potentiometer. The emulsion was diluted 1000 times with a buffer of the same pH and ionic strength (for particle size measurement) and measured at 25°C. The refractive index of the buffer and fish oil were 1.333 and 1.451, respectively.

[0053] 1.3.2 Determination of Hydroperoxide (POV) in Emulsion

[0054] Take 1.5mL of fish oil emulsion and add 7.5mL of mixed solution (isooctane: isopropanol = 3:1), vortex shake. Collect the upper organic solvent phase (about 0.4mL) by centrifugation, add 5.6mL of mixed solution (methanol: butanol = 2:1), add 30μL of ammonium thiocyanate (3.94M) and 30μL of ferrous solution, let stand for 20min, measure the absorbance at 510nm using a UV-visible spectrophotometer, and calculate the content of hydroperoxide using the cumene hydroperoxide standard curve.

[0055] 1.3.3 Determination of thiobarbituric acid (TBARS) value of emulsion

[0056] Take 2 mL of fish oil emulsion and mix it with 4 mL of TBA solution (15% trichloroacetic acid and 0.375% thiobarbituric acid dissolved in 0.25 M hydrochloric acid), heat the solution in a boiling water bath for 15 minutes, cool it quickly after heating, and measure the absorbance at 532 nm using a UV-visible spectrophotometer. Calculate the TBARS value based on the 1,1,3,3-tetraethoxypropane standard curve.

[0057] 1.3.4 Determination of fluorescence intensity of emulsion

[0058] The degree of oxidation of soy protein in the emulsion during storage was evaluated by using fluorescence spectroscopy to assess the loss of natural tryptophan fluorescence in the emulsion. Take 200uL of the emulsion sample, dilute it in 10mL of phosphate buffer with the same pH and ionic strength, and measure it using a fluorescence spectrophotometer. The measurement parameters are: excitation wavelength 280nm, emission wavelength 300-500nm, slit width 5nm.

[0059] 2 Experimental results and analysis

[0060] In all the figures of this experiment, the concentration of xanthan gum is 0.2%, and the concentrations of carrageenan and herba schoenoprasum polysaccharide are 0.05% (respectively, the optimal sugar concentrations for the best physical stability of the emulsions stabilized by each polysaccharide and soy protein). In the figure, SPI, SPI-IC, SPI-X, and SPI-H represent fish oil emulsions stabilized by soy protein, soy protein-carrageenan, soy protein-xanthan gum, and soy protein-herba schoenoprasum polysaccharide, respectively; USPI, USPI-IC, USPI-X, and USPI-H represent fish oil emulsions stabilized by ultrasonic soy protein, ultrasonic soy protein-carrageenan, ultrasonic soy protein-xanthan gum, and ultrasonic soy protein-herba schoenoprasum polysaccharide, respectively.

[0061] 2.1 Physical and oxidative stability of the emulsion after storage at 37°C for 60 days at pH 7 2.1.1 Particle size changes during storage at 37°C

[0062] Figure 1(a) shows that when soy protein is not ultrasonicated, after 60 days of storage, the average particle size of SPI, SPI-IC, and SPI-X emulsions increases significantly, while the average particle size of SPI-H emulsion increases only slightly, indicating that compared with xanthan gum and carrageenan, Herba Cibotii polysaccharides can better enhance the physical stability of soy protein-based emulsions. Figure 1 (b) shows that under the condition of the same concentration of Herba Epimedii polysaccharides, the average particle size of the emulsion stabilized by soy protein and Herba Epimedii polysaccharides after ultrasonic treatment was lower than that of the soy protein- Herba Epimedii polysaccharides (SPI-H) emulsion without ultrasonic treatment after 60 days of storage, indicating that the binding ability of soy protein and Herba Epimedii polysaccharides molecules was enhanced after ultrasonic treatment, resulting in more Herba Epimedii polysaccharides molecules being adsorbed on the surface of the oil droplet protein film, increasing the steric hindrance between the oil droplets and further improving the physical stability of the emulsion.

[0063] 2.1.2 Oxidation of fish oil during storage of emulsion at 37°C

[0064] Figure 2 (a) with Figure 2 (c) shows that when soy protein is not treated with ultrasound, the POV and TBARS values ​​of fish oil emulsions stabilized by SPI, SPI-IC, SPI-X, and SPI-H are lower than those of the emulsion stabilized by the small molecule surfactant Tween 20 after storage for 60 days. Among them, the SPI-H emulsion has the lowest POV and TBARS values, indicating that compared with xanthan gum and carrageenan, the fish oil emulsion stabilized by Herba Epimedii polysaccharides and soy protein has the highest oxidative stability, which is closely related to the higher physical stability of the SPI-H emulsion. The Tween 20 emulsion has the largest POV and TBARS values, and the degree of fish oil oxidation is the deepest, indicating that the small molecule surfactant has the lowest ability to inhibit fish oil oxidation. Figure 2 (b) with Figure 2 (d) The POV and TBARS values ​​of the fish oil emulsion stabilized by ultrasonically treated soy protein and Herba Epimedii polysaccharides (USPI-H) were significantly lower than those of the emulsion without ultrasonic treatment (SPI-H) after 60 days of storage, indicating that even when the concentration of Herba Epimedii polysaccharides added was the same, the physical stability of the emulsion was improved due to the enhanced binding ability between soy protein and Herba Epimedii polysaccharides by ultrasonic treatment (see 2.1.1 Particle size results). At the same time, the adsorption of more Herba Epimedii polysaccharides molecules on the surface of oil droplets thickened the oil-water interface film, reducing the contact opportunity between the pro-oxidant and the fish oil in the film. Therefore, the oxidation rate of the fish oil was reduced and the oxidative stability was improved.

[0065] 2.1.3 Protein oxidation degree during storage of emulsion at 37°C

[0066] The loss of native tryptophan fluorescence intensity during emulsion storage is a direct result of tryptophan oxidative degradation.

[0067] The fluorescence intensity of the soy protein emulsion on the 0th day reflects the amount of tryptophan exposed on the protein surface. After the addition of polysaccharides, due to the interaction between polysaccharides and proteins, the polysaccharides are adsorbed on the protein surface, reducing the amount of tryptophan exposed by the soy protein, and thus the fluorescence intensity decreases. That is, the fluorescence intensity of the emulsion stabilized by soy protein and polysaccharides on the 0th day represents the binding ability of polysaccharides and proteins. The stronger the binding ability of polysaccharides and proteins, the lower the fluorescence intensity of the emulsion.

[0068] Figure 3 (a) with Figure 3 (b) shows that when the emulsion was stored at pH 7 and 37°C on day 0, SPI-H had the lowest fluorescence intensity among all the unsonicated protein emulsions, and USPI-H had the lowest fluorescence intensity among all the sonicated protein emulsions, indicating that compared with other polysaccharides, the binding ability of Herba Cibotii polysaccharides with soybean protein is the strongest. 3+ , Cu 2+ ) can act as a catalyst to convert tryptophan into free radicals, which can directly react with molecular oxygen to produce tryptophan peroxide, resulting in tryptophan loss; at the same time, during storage, the free radicals produced by lipid oxidation will promote protein oxidation, resulting in tryptophan loss and a decrease in the fluorescence intensity of the emulsion. Therefore, lipid oxidation is inevitably accompanied by protein oxidation and a decrease in tryptophan fluorescence intensity. Changes in tryptophan fluorescence intensity have been used as an indicator to reflect the degree of protein oxidation in lipoproteins and liposomes. Figure 3 (a) shows that when the protein is not ultrasonicated, the fluorescence intensity of all emulsions decreases significantly with the increase of storage time, indicating that the protein in all emulsions is oxidized, but the fluorescence intensity of SPI-H emulsion decreases the least after storage for 60 days (the difference between the fluorescence intensity of the emulsion on the 0th day and the fluorescence intensity on the 60th day), indicating that the degree of protein oxidation in SPI-H emulsion is the lightest. This is because the degree of oil oxidation in SPI-H emulsion is the lowest, the number of free radicals produced is small, which reduces the degree of protein oxidation, and the ability of Herba Epimedii polysaccharides to bind to proteins is the strongest. More Herba Epimedii polysaccharides molecules are adsorbed on the surface of the protein film at the oil-water interface. The polysaccharide has the function of chelating metal ions, which reduces the probability of metal ions and free radicals in the aqueous phase contacting proteins. Therefore, the degree of protein oxidation in SPI-H emulsion is the lightest. Similarly, after storage at 37°C for 60 days, the fluorescence intensity of USPI-H emulsion decreases the least among all ultrasonic protein emulsions ( Figure 3 (b)), that is, among all the ultrasonic protein emulsions, the degree of protein oxidation in USPI-H emulsion was the lightest.

[0069] 2.2 Physical and oxidative stability of emulsions under 500 mM NaCl storage conditions (25°C) Salt ions often exist during emulsion processing, so it is necessary to consider the physical and oxidative stability of emulsions under certain salt ion concentration conditions.

[0070] 2.2.1 Changes in emulsion particle size under 500 mM NaCl storage conditions (25 °C)

[0071] Figure 4 (a) shows that when the soy protein is not treated with ultrasound, the average particle size of the SPI emulsion is the largest after the emulsion contains 500 mM NaCl and is stored for 30 days. The average particle size of the SPI-H emulsion is significantly smaller than that of the SPI-IC and SPI-X emulsions, indicating that the physical stability of the SPI-H emulsion is better than that of other soy protein-polysaccharide emulsions. Figure 4 (b) shows that the average particle size of the emulsions stabilized by soy protein and various polysaccharides after ultrasonic treatment is significantly smaller than that of the emulsions stabilized by soy protein and various polysaccharides without ultrasonic treatment after storage at 500 mM NaCl concentration for 30 days. In particular, the average particle size of the USPI-H emulsion did not change significantly during storage, and was significantly smaller than that of the SPI-H emulsion without ultrasonic treatment, as well as the USPI-X, USPI-IC, and USPI emulsions without ultrasonic treatment. This indicates that the soy protein has an enhanced ability to bind to the herb polysaccharide molecules after ultrasonic treatment, which improves the stability of the soy protein-herb polysaccharide fish oil emulsion in high NaCl concentration. + Physical stability at concentrations.

[0072] 2.2.2 Oxidation of fish oil in emulsions under 500 mM NaCl storage conditions (25 °C)

[0073] Figure 5 (a) with Figure 5 (c) shows that when the soy protein was not ultrasonically treated, after all the fish oil emulsion samples were stored in 500 mM NaCl for 30 days, the POV and TBARS values ​​of the emulsion stabilized by Tween 20 were the largest, that is, the degree of oxidation of the Tween 20 emulsion was the deepest; compared with the SPI, SPI-X, and SPI-IC emulsions, the POV and TBARS values ​​of the SPI-H emulsion were the lowest, indicating that even at high NaCl concentrations, the degree of fish oil oxidation in the soy protein-herbaceous celery polysaccharide emulsion was still significantly lower than that of the soy protein-xanthan gum emulsion and the soy protein-carrageenan emulsion; Figure 5 (b) with Figure 5 (d) shows that under the same concentration of Herba Epimedii polysaccharides, the POV and TBARS values ​​of the sonicated soy protein-Herba Epimedii polysaccharides fish oil emulsion after storage in 500 mM NaCl for 30 days were significantly lower than those of the soy protein-Herba Epimedii polysaccharides fish oil emulsion without sonication, indicating that the sonicated soy protein improved the oxidative stability of the fish oil emulsion stabilized by soy protein and Herba Epimedii polysaccharides at high NaCl concentrations. This is mainly due to the enhanced interaction between soy protein and Herba Epimedii polysaccharides molecules after sonication, which increased the physical stability of the fish oil emulsion, and more Herba Epimedii polysaccharides molecules were adsorbed on the oil-water interface, resulting in the thickening of the interfacial film, which reduced the chance of the pro-oxidant contacting the internal oil droplets, thereby reducing the oxidation rate of the fish oil.

[0074] 2.2.3 Oxidation degree of protein in emulsion under 500 mM NaCl storage conditions (25 °C) Figure 6 The results showed that when stored at 500 mM NaCl concentration for 0 days (25°C), SPI-H had the lowest fluorescence intensity among all the unsonicated protein emulsions, and USPI-H had the lowest fluorescence intensity among all the sonicated protein emulsions, indicating that even under high NaCl concentration conditions, compared with other polysaccharides, the binding ability of Herba Cibotii polysaccharides with soybean protein was still the strongest. After storage for 30 days, the fluorescence intensity of SPI-H fish oil emulsion decreased the least among all the unsonicated protein emulsions ( Figure 6 (a)), that is, the degree of protein oxidation in SPI-H emulsion is the lightest. This is because the degree of oil oxidation in SPI-H emulsion is the lowest, the free radicals produced are less, and the soy protein has the strongest binding ability with the herb polysaccharide molecules. The shielding and chelation produced by the adsorption of the herb polysaccharide molecules on the protein surface reduces the probability of metal ions and free radicals in the aqueous phase contacting with the protein, so the degree of protein oxidation is the lightest. Similarly, after storage at 500mM NaCl concentration for 30 days (25°C), the fluorescence intensity of USPI-H fish oil emulsion decreased the least among all ultrasonic protein emulsions ( Figure 6 (b)), that is, the degree of protein oxidation in USPI-H emulsion was the lightest among all ultrasonic protein emulsions.

[0075] Experimental Example 2 Effect of different ultrasonic power and ultrasonic time on the storage physical stability and oxidative stability of fish oil emulsion during preparation

[0076] 1 Test method

[0077] Disperse the soy protein isolate in a phosphate buffer (the concentration of the phosphate buffer is 5mM, pH 7.0), stir at room temperature for 1h, and then treat at different ultrasonic powers (150W, 300W, 450W) and ultrasonic times (0min, 5min, 10min, 15min, 20min). During the ultrasonic treatment, the soy protein solution is always kept in an ice bath state. After the treated sample is further stirred at room temperature for 2h, it is placed in a 4°C refrigerator for hydration overnight. The soy protein solution hydrated overnight is taken out of the refrigerator and slowly cooled to room temperature, and then the ultrasonic soy protein-herb polysaccharide-stabilized fish oil emulsion is prepared according to the method of Example 4. The percentage of each component in the final emulsion is: 5%

[0078] (w / w) fish oil, 2% (w / w) soy protein (SPI), 0.05% (w / w) herba hyssop polysaccharides (H), adjust the pH of the emulsion to 7.0, pour into a glass bottle, and store at 25°C away from light.

[0079] 2 Test results

[0080] Figure 7The results show that the average particle size of the fish oil emulsion stabilized by soy protein and herba jelly polysaccharide after ultrasonic treatment is smaller than that of the soy protein-herba jelly polysaccharide emulsion without ultrasonic treatment after 30 days of storage at 25°C, indicating that the soy protein after ultrasonic treatment has an improved ability to bind to herba jelly polysaccharide molecules, which improves the physical stability of the fish oil emulsion. The fish oil emulsion stabilized by soy protein and herba jelly polysaccharide treated with ultrasonic power of 300W for 10min or 15min has the smallest average particle size after storage at 25°C for 30 days, indicating that the physical stability of the SPI-H emulsion treated with this ultrasonic condition is the best. This is mainly because appropriate ultrasonic treatment can stretch the protein molecular structure, expose more polar and hydrophobic groups, improve the binding ability of protein with oil bodies and herba jelly polysaccharide molecules, make more herba jelly polysaccharide molecules adsorbed on the protein membrane surface at the oil-water interface, increase the steric hindrance between oil bodies, inhibit the aggregation between oil bodies, and thus improve the physical stability of the emulsion.

[0081] Figure 8 and Fig. 9 The results show that under different ultrasonic power and ultrasonic time conditions, the fish oil emulsion stabilized by soybean protein and Herba Ceropegiae polysaccharides treated at 300W for 15min has the lowest POV and TBARS values ​​after storage at 25℃ for 30 days, indicating that the SPI-H emulsion under this ultrasonic treatment condition has the best oxidation stability. This is mainly because appropriate ultrasonic treatment can stretch the protein molecular structure, expose more polar and hydrophobic groups, improve the binding ability of protein and Herba Ceropegiae polysaccharides molecules, make more Herba Ceropegiae polysaccharides molecules adsorbed on the protein membrane surface at the oil-water interface, increase the steric hindrance between oil bodies, inhibit the aggregation between oil bodies, and improve the physical stability of the emulsion. At the same time, the adsorption of more Herba Ceropegiae polysaccharides molecules on the protein membrane surface increases the thickness of the oil-water interface film, reduces the probability of oxygen and free radicals penetrating the oil-water interface film into the oil body and contacting the oil droplets, thereby reducing the oil oxidation rate.

[0082] Experimental Example 3 Parameter screening test of homogenization pressure in the preparation of fish oil emulsion

[0083] 1 Test method

[0084] According to the method of Example 1, multiple portions of soybean protein and fish oil crude emulsions were prepared, and homogenized three times under 40MPa, 60MPa, 80MPa, 100MPa, 120MPa, and 140MPa conditions, respectively, to obtain soybean protein and fish oil primary emulsions under different homogenization conditions, and then slowly added with the aqueous solution of Herba Ceropegiae polysaccharides, and magnetically stirred for 30 minutes, so that the percentage content of each component in the final soybean protein-herba Ceropegiae polysaccharides emulsion was 5% (w / w) fish oil, 2% (w / w) SPI, and 0.05% (w / w) Herba Ceropegiae polysaccharides. The effects of different homogenization pressures on the average particle size and PDI of soybean protein-herba Ceropegiae polysaccharides emulsions were investigated.

[0085] 2 Test results

[0086] Fig.10 It shows that the average particle size of the soybean protein-herb polysaccharide emulsion prepared at a homogenization pressure of 40-140MPa gradually decreases with the increase of homogenization pressure, and the physical stability of the emulsion is enhanced. The emulsion is subjected to mechanical forces such as high-speed shear, cavitation, vortex, and impact in the homogenizer, and the larger particle size droplets in the emulsion are broken into small droplets. The higher the pressure, the stronger the mechanical force on the emulsion, so the smaller the emulsion particle size. Similarly, with the increase of homogenization pressure, the emulsion dispersion index (PDI) shows a downward trend in the range of 40-120Mpa, indicating that the increase of homogenization pressure reduces the difference in droplet size, makes the size more uniform, and enhances the stability of the emulsion. Although the average particle size of the fish oil emulsion homogenized at 140MPa is not significantly different from that of the emulsion under 120Mpa homogenization conditions (p>0.05), it still has an increasing trend, so the physical stability of the emulsion prepared under 120MPa homogenization pressure is optimal.

[0087] Test Example 4 Screening test of the concentration of celery polysaccharide in the preparation of fish oil emulsion

[0088] 1 Test method

[0089] Different soy protein-polysaccharide emulsions (soy protein-herbaceous grass polysaccharide emulsion (SPI-H), soy protein-carrageenan emulsion (SPI-IC), soy protein-xanthan gum emulsion (SPI-X)) were prepared according to the method of Example 1, and the percentage content of each component in the emulsion was 5% (w / w) fish oil, 2% (w / w) SPI, 0-0.3% (w / w) polysaccharide, and the pH value of each emulsion was adjusted to 7.0 or 3.5, respectively.

[0090] 2 Test results

[0091] Fig.11 (a) with Fig.11 (b) shows that no matter the emulsion is at pH 7 or pH 3.5, the average particle size of the emulsion is the smallest when the concentration of Herba Epimedii polysaccharides is 0.05%. Especially at pH 3.5, the average particle size of the SPI-H emulsion is much smaller than that of the fish oil emulsion stabilized by SPI alone, indicating that the addition of 0.05% Herba Epimedii polysaccharides significantly enhances the physical stability of the emulsion under acidic conditions.

Claims

1. A fish oil emulsion stabilized by soybean protein and grass jelly polysaccharide, It is characterized in that The fish oil emulsion is composed of fish oil, soy protein isolate, herba jellyfish polysaccharide and water; the amount of each component is as follows: 2-10% fish oil, 1.5-4% soy protein isolate, 0.01-0.4% herba jellyfish polysaccharide, and the balance is water; The preparation method comprises: (1) adding fish oil to a soy protein isolate aqueous solution under stirring, and then performing dispersion treatment and homogenization treatment in sequence to obtain a primary fish oil emulsion; (2) mixing and stirring the primary fish oil emulsion with a herba jellyfish polysaccharide solution to obtain a soy protein-herba jellyfish polysaccharide stabilized fish oil emulsion; The soy protein isolate aqueous solution in step (1) is a soy protein isolate aqueous solution that has been treated with ultrasound; wherein the ultrasonic treatment parameters are an ultrasonic power of 150-450 W and an ultrasonic time of 15-20 min; The stirring in step (1) is magnetic stirring; wherein the rotation speed of the magnetic stirring is 200-300 rpm; The dispersion treatment in step (1) is performed in a high-speed shearing machine at a rotation speed of 10,000-15,000 rpm for 2-5 min; The homogenization treatment in step (1) is carried out in a high pressure homogenizer at a pressure of 40-140 MPa for 2-4 times.

2. The fish oil emulsion stabilized by soybean protein-herbaceous grass polysaccharide according to claim 1, It is characterized in that The dosage of each ingredient is: fish oil 5%, soy protein isolate 2.0%, and grass jelly polysaccharide 0.05%.

3. The fish oil emulsion stabilized by soybean protein-herbaceous grass polysaccharide according to claim 1, It is characterized in that The ultrasonic power is 300 W, and the ultrasonic time is 15 min.

4. The fish oil emulsion stabilized by soybean protein-herbaceous grass polysaccharide according to claim 1, It is characterized in that The homogenization process is carried out in a high pressure homogenizer at a pressure of 120 MPa for 2-4 times.

5. The fish oil emulsion stabilized by soybean protein-herbaceous grass polysaccharide according to claim 1, It is characterized in that The stirring described in step (2) is magnetic stirring.

6. The fish oil emulsion stabilized by soybean protein-herbaceous grass polysaccharide according to claim 5, It is characterized in that The rotation speed of the magnetic stirring in step (2) is 200-300 rpm, and the stirring time is 10-60 min.

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