Nutrient enrichment type defatted DHA egg yolk powder for old people and preparation method thereof

Abstract

The invention provides nutrient enrichment type defatted DHA (docosahexaenoic acid) egg yolk powder for old people and a preparation method thereof. According to the preparation method, ultrasonic-assisted ethyl acetate degreasing is adopted, triglyceride is selectively removed, active phospholipid is reserved, degreasing is completed in a short time at low temperature, meanwhile, sulfydryl oxidation and pigment oxidation are relieved, protein denaturation and aggregation are weakened, and protein degeneration and aggregation are reduced; the problems that traditional egg yolk powder is high in neutral lipid content, low in phospholipid content and long in degreasing time, and a traditional degreasing mode has large damage to active phospholipid and protein are solved. A W / O / W type double emulsion is prepared by constructing an amphiphilic delivery system, taking hydrophilic nutrient elements as an inner water phase, taking egg yolk liquid as an outer water phase and taking DHA as an oil phase, and the W / O / W type double emulsion is rich in DHA, hydrophilic nutrient elements and other bioactive components and can better meet the requirements of human bodies for nutrient substances; the egg yolk is used as a natural carrier, so that the problem of co-encapsulation of nutrients with different polarities is solved. The preparation method is simple, short in period and suitable for industrial production.

Description

Technical Field

[0001] The present invention relates to the technical field of functional food preparation, and more specifically, to an elderly nutrition fortified defatted DHA egg yolk powder and a preparation method thereof. Background Art

[0002] In addition to containing high-quality nutrients such as proteins, vitamins, and minerals, egg yolks also contain a large amount of lipids, including 65% neutral lipids, 30% phospholipids, and 4% cholesterol. As the core functional component in egg-derived lipids, the unique biological activity of egg yolk phospholipids has shown significant value in the field of metabolic health. However, the neutral lipid content in egg yolks is relatively high, and long-term high-fat intake may induce hypertension and other cardiovascular and cerebrovascular diseases. Therefore, defatting the egg yolk powder and optimizing the lipid composition in egg yolks are beneficial to expanding the application scenarios of egg yolk powder and improving consumer acceptance. Traditionally, ethanol solvent is mainly used for defatting defatted egg yolk powder, but there are problems such as long process time, high temperature, serious denaturation of egg yolk proteins, and the removal of active phospholipids in egg yolks. Moreover, the defatted egg yolk powder almost loses its functional properties, which is not conducive to subsequent enzymatic hydrolysis. Therefore, how to selectively remove neutral lipids while minimizing the impact of the solvent on the protein structure and functional properties is a difficult point in the defatted egg yolk industry.

[0003] With consumers' emphasis on the "functional, precise, nutritious and healthy" requirements of food, DHA egg yolk powder for nutritional supplementation of special populations such as the elderly can promote the development of the egg product industry towards a high-level and high-value direction. However, there are technical challenges such as poor stability in the co-emulsification of different polar nutritional elements. DHA (docosahexaenoic acid) is a polyunsaturated fatty acid that is extremely important for human health and has various physiological functions, such as preventing cardiovascular diseases and anti-cancer. However, DHA is easily oxidized and degraded, and has a strong fishy smell, which limits its application in food. In addition to fat-soluble components such as DHA essential for the nervous system, the fortified supplementation of hydrophilic nutrients is of irreplaceable strategic significance for delaying the aging process. Specifically, ferrous gluconate, as the core matrix for heme synthesis, can effectively improve senile anemia and oxygen transport efficiency; zinc gluconate is crucial for immune homeostasis and cell repair by regulating the activities of more than 300 enzyme systems; calcium is not only the material basis for bone density, but also participates in physiological processes such as nerve conduction and muscle contraction; VC and VE, as the main antioxidants, play multiple roles in scavenging free radicals, promoting collagen synthesis and enhancing immune response; selenium builds an antioxidant defense barrier for the cardiovascular system by activating glutathione peroxidase. As people age, the gastrointestinal absorption function of the elderly declines and the basal metabolic rate decreases, but the body's requirement for the bioavailability of micronutrients increases instead. This contradiction between supply and demand makes the scientific proportioning of hydrophilic nutrients particularly important. Therefore, how to co-load DHA and hydrophilic nutritional elements and solve the co-encapsulation of different polar nutrients is a challenge in the field of nutritional delivery.

[0004] Aiming at the industrialization problems in the processing of egg yolk powder, research is carried out around the industrial chain requirements of "defatting of egg yolk powder - co-delivery of DHA - hydrophilic nutritional elements - creation of precise nutrition products". The design is carried out with the functional egg yolk powder for nutritional supplementation of the elderly population as the main line. Based on the construction of a multiphase synergistic emulsification system and the co-loading and delivery mechanism of functional factors, the egg yolk powder is endowed with functional properties such as antioxidant, realizing the synergistic enhancement of nutritional fortification and processing adaptability. This technical path can expand its application scenarios in the fields of special dietary foods, nutritional supplements, etc., promote the upgrading and transformation of the egg source functional ingredient industry towards high added value and precise nutrition, and promote the extension of the value chain of the entire industrial chain.

[0005] Chinese Patent CN 110938091 A discloses a method for separating egg yolk oil by treating egg yolk liquid with subcritical carbon dioxide fluid to complete defatting; then the defatted egg yolk liquid is vacuum freeze-dried to obtain egg yolk powder; finally, supercritical carbon dioxide fluid is used to extract and purify the egg yolk powder to obtain egg yolk lecithin. This defatting method has a complex process, large equipment investment, and high cost. Chinese Patent CN 113214895 A discloses the preparation of defatted egg yolk powder using ethanol. The defatting time in its process flow is long, the efficiency is low, the solubility of the prepared egg yolk powder is low, and the emulsifying performance is poor. Chinese Patent CN 115669940 B discloses a walnut polypeptide and DHA composite emulsion with the efficacy of assisting in improving memory. The emulsion prepared by ultrasonic emulsification after mixing EPEVLR and DHA in a molar ratio of 1:1 to 2:1 has a synergistic brain-strengthening effect, and the effect is better than that of using EPEVLR and DHA alone, achieving the improvement of the brain-strengthening effect while reducing the dosage of EPEVLR and DHA. Chinese Patent CN 107383187B discloses a method for jointly extracting IgY, phosvitin, lecithin, egg yolk oil, and defatted egg yolk powder from egg yolk. First, the egg yolk liquid is diluted with water and centrifuged, and the obtained supernatant is subjected to salting out and polyethylene glycol precipitation to extract IgY; the lower layer is egg yolk pulp. First, ethanol is used to remove the moisture of the egg yolk pulp, then a mixed solvent of ethanol and ethyl acetate is used to extract the total lipids of the egg yolk, and then two products, lecithin and egg yolk oil, can be obtained by treating with ethyl acetate; finally, phosvitin can be obtained by treating the protein after lipid removal with sodium chloride solution. However, this method has the problems of solvent residue risk and protein denaturation. Summary of the Invention

[0006] Aiming at the problems existing in the background technology, the purpose of the present invention is to provide an elderly nutrition fortified defatted DHA egg yolk powder and its preparation method. This method solves the problems of high neutral lipid content, low phospholipid content, long preparation cycle, and great damage to active phospholipids and proteins by traditional defatting methods in defatted egg yolk powder.

[0007] To achieve the above purpose, the present invention adopts the following technical solutions:

[0008] The first aspect of the present invention provides a preparation method of an elderly nutrition fortified defatted DHA egg yolk powder, comprising the following steps:

[0009] S1. Selectively defat the egg yolk powder with ultrasonic-assisted ethyl acetate to obtain defatted egg yolk powder, and then dissolve it with deionized water to obtain defatted egg yolk liquid;

[0010] S2. Dissolve hydrophilic nutrient elements with deionized water as the inner aqueous phase, use DHA algal oil containing an emulsifier as the oil phase, mix the inner aqueous phase and the oil phase, and perform homogenization treatment to obtain a water-in-oil emulsion containing DHA and hydrophilic nutrient elements;

[0011] S3. Mix the defatted egg yolk liquid as an emulsifier with a water-in-oil emulsion containing DHA and hydrophilic nutrient elements, and deionized water as the external aqueous phase, and perform homogenization treatment to obtain a water-in-oil-in-water double emulsion, that is, a W / O / W double emulsion;

[0012] S4. Add sodium alginate to the obtained water-in-oil-in-water double emulsion, dissolve and mix evenly, and perform spray drying to obtain an aged nutrition fortified defatted DHA egg yolk powder.

[0013] Preferably, in step S1, the egg yolk powder is obtained through the following steps: Wash and shell the eggs, separate the egg white and egg yolk, roll the egg yolk on filter paper to remove the residual egg white, pierce the egg yolk membrane with tweezers to collect the egg yolk liquid, dilute the obtained egg yolk liquid and then perform spray drying, collect the spray-dried egg yolk powder, and store it frozen at -20 °C for later use; wherein, the spray flow rate is 10 - 25 mL / min, the dilution ratio of the egg yolk powder is 2 - 5:1, and the inlet air temperature is 180 - 200 °C.

[0014] Preferably, in step S1, the selective defatting of the egg yolk powder with ultrasonic-assisted ethyl acetate is specifically carried out as follows: Dilute the egg yolk powder with ethyl acetate, perform ultrasonic treatment, then perform suction filtration, drying, and pulverization to obtain defatted egg yolk powder, dissolve it with deionized water to obtain defatted egg yolk liquid.

[0015] Preferably, the material-liquid ratio of the egg yolk powder to ethyl acetate is 1 g: 4 - 8 mL, and more preferably 1 g: 5 mL.

[0016] Preferably, the ultrasonic temperature is 0 - 25 °C, and more preferably 0 °C.

[0017] Preferably, the ultrasonic power is 80 - 400 w, and more preferably 320 w.

[0018] Preferably, the ultrasonic time is 5 - 25 min, and more preferably 15 min.

[0019] Preferably, in the defatted egg yolk liquid, the mass ratio of the defatted egg yolk powder to deionized water is 1: 3 - 5.

[0020] Preferably, in step S2, the homogenization rate is 9000 - 12000 r / min, and the homogenization time is 1 - 6 min. Preferably, in step S3, the homogenization rate is 8000 - 11000 r / min, and the homogenization time is 2 - 8 min.

[0021] Preferably, in step S3, the volume ratio of the water-in-oil emulsion containing DHA and hydrophilic nutrient elements to the external aqueous phase is 1: 1 - 2.

[0022] Preferably, in step S4, the addition amount of sodium alginate is 0.1-0.5% of the mass of egg yolk.

[0023] Preferably, in step S4, the temperature of spray drying is 150-200 °C, and the sample injection flow rate is 400-800 mL / h.

[0024] Preferably, the hydrophilic nutrient elements include, but are not limited to, one or more combinations of selenium, vitamin C, vitamin E, calcium, iron, zinc, etc.

[0025] Preferably, the calcium is derived from calcium citrate, the iron is derived from ferrous gluconate, and the zinc is derived from zinc gluconate.

[0026] Preferably, the DHA is selected from any one or a combination of two or more of DHA extracted from algal oil (abbreviated as algal oil DHA), DHA extracted from fish oil, DHA extracted from perilla seeds, or DHA extracted from flax seeds.

[0027] The second aspect of the present invention also provides the aged nutrition fortified defatted DHA egg yolk powder prepared by the above preparation method.

[0028] The beneficial effects of the present invention are:

[0029] (1) The present invention provides a preparation method for aged nutrition fortified defatted DHA egg yolk powder. On the one hand, by selectively defatting the egg yolk and screening and optimizing the parameter conditions in the preparation process, the lipid composition in the egg yolk is optimized. Specifically, ultrasonic-assisted ethyl acetate defatting is used to selectively remove triglycerides and retain active phospholipids, enabling the defatting process to be efficiently completed at low temperature and short time, reducing thiol oxidation and pigment oxidation, weakening protein denaturation and aggregation. Finally, the prepared defatted DHA egg yolk powder product has an increased phospholipid content, a decreased triglyceride content, a shortened defatting time, and improved solubility and emulsifying properties, solving the problems of high neutral lipid content, low phospholipid content, long defatting time, and great damage to active phospholipids and proteins in traditional egg yolk powder by traditional defatting methods, providing a new idea for improving the added value of egg products, and can be widely applied to the development of functional foods, health foods, etc. On the other hand, using the egg yolk as a natural carrier solves the problem of co-encapsulation of different polar nutrients. The whole preparation method is simple, has a short cycle, is suitable for industrial production, improves economic benefits, and is suitable for preparing high-quality aged nutrition fortified defatted DHA egg yolk powder.

[0030] (2) The present invention prepares a W / O / W double emulsion by using hydrophilic nutrient elements as the internal aqueous phase, DHA as the oil phase, and egg yolk liquid as the external aqueous phase, and finally prepares a defatted DHA egg yolk powder product rich in various bioactive components, which can better meet the human body's needs for nutrients.

[0031] (3) The present invention uses process technologies such as ultrasonic-assisted selective degreasing with ethyl acetate and two-step emulsification and homogenization. While selectively removing neutral lipids, it reduces the impact of solvents on the protein structure and functional properties. Using defatted egg yolk as the wall material and emulsifying different polar nutrients, it realizes the development of functional foods rich in various bioactive substances, and also solves bottleneck problems such as encapsulation, emulsification, and microcapsule forming of multiple nutrients. The defatted DHA egg yolk powder for elderly nutritional fortification obtained by the present invention has a suitable taste, is easy to disperse, has no peculiar smell, and is easily digested by the human body. It is of great significance for realizing the high-quality and high-value-added development of egg yolk, improving the nutritional value and economic value of egg powder, and broadening the application of egg yolk in nutritionally fortified foods. Brief Description of the Drawings

[0032] Figure 1 shows the content of total lipids in defatted egg yolk powder obtained under different ultrasonic conditions;

[0033] Figure 2 shows the content of phospholipids in the total lipids of defatted egg yolk powder obtained under different ultrasonic conditions;

[0034] Figure 3 shows the chromaticity of defatted egg yolk powder obtained by different degreasing processes (where EYP is the original egg yolk powder, U-EYP is the egg yolk powder after ultrasonic-assisted degreasing with ethyl acetate, and E-EYP is the egg yolk powder after ethanol degreasing);

[0035] Figure 4 is Figure 3 the solubility and emulsification performance diagrams of the three egg yolk powders shown in;

[0036] Figure 5 is Figure 3 the protein structure characterization diagrams of the three egg yolk powders shown in (including surface hydrophobicity, free sulfhydryl content, protein secondary structure, protein tertiary structure);

[0037] Figure 6 is the optical microscope of the W / O / W multi-layer emulsion system;

[0038] Figure 7 is the potential of the emulsions obtained by different homogenization methods (where S is the emulsion obtained after high-speed shearing, and H is the emulsion obtained by subjecting S to high-pressure homogenization again). Detailed Embodiments

[0039] In order to make the objectives, technical solutions, and advantages of the present invention clearer, the present invention will be further described in detail below with reference to the drawings and specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0040] Example 1

[0041] A preparation method of an elderly nutrition fortified defatted DHA egg yolk powder, comprising the following steps:

[0042] (1) Wash and shell the eggs, separate the egg whites and yolks, and roll the yolks on filter paper to remove the remaining egg whites. Pierce the yolk membrane with tweezers to collect the yolk liquid. The spray flow rate is 15 mL / min, the dilution ratio of the egg yolk powder is 2:1, and the inlet air temperature is 185 °C. Collect the spray-dried egg yolk powder and name it EYP, and store it frozen at -20 °C for later use;

[0043] (2) Defat the egg yolk powder with ethyl acetate under the conditions of a material-liquid ratio (egg yolk powder g: ethyl acetate mL) of 1:5, an extraction temperature of 0 °C, an ultrasonic power of 320 W, and an ultrasonic time of 15 min to obtain a defatted egg yolk liquid. After the defatted egg yolk liquid is filtered by a G3 Buchner funnel, place the filter cake in a vacuum drying oven and evaporate the solvent at 40 °C for about 24 h, and then crush it to obtain defatted egg yolk powder, named U-EYP;

[0044] (3) Weigh each hydrophilic nutrient active component (including 7500 mg calcium citrate, 4100 mg ferrous gluconate, 2470 mg zinc gluconate, 4500 mg vitamin C, 1500 mg vitamin E, and 1.5 mg selenium element), and completely dissolve it in 24 mL of deionized water to prepare a uniform internal aqueous phase solution. Take another oil phase system: Pre-dissolve polyglycerol ricinoleate (PGPR) emulsifier accounting for 6% of the volume of algal oil in 55 mL of DHA algal oil, and continuously stir at room temperature for 2 hours until completely dissolved to form a homogeneous oil phase. Mix according to the volume ratio of internal aqueous phase: oil phase = 3:7, and use a high-speed homogenizing emulsifier to continuously process at a rotation speed of 11000 r / min for 2 min to obtain a DHA-hydrophilic active substance delivery system emulsion, that is, a water-in-oil (W / O) emulsion containing DHA and hydrophilic nutrient elements; Use the pre-prepared defatted egg yolk liquid as the external aqueous phase carrier, control the volume mixing ratio of the primary emulsion to the external aqueous phase to be 1:1.5, and perform high-speed shear homogenizing emulsification treatment at 10000 r / min for 3 min to construct a stable W / O / W double emulsion, marked as S;

[0045] (4) Add 0.1% of sodium alginate based on the mass of the egg yolk to the W / O / W double emulsion, and perform high-temperature spray drying to obtain an elderly nutrition fortified defatted DHA egg yolk powder; After testing, every 5 g of the obtained elderly nutrition fortified defatted DHA egg yolk powder contains 200 mg of DHA, 40 mg of vitamin C, 13 mg of vitamin E, 40 mg of iron, 35 mg of zinc, 80 mg of calcium, and 13 μg of selenium; It can meet the daily required intake of essential nutrients for the elderly.

[0046] Example 2

[0047] The procedure is basically the same as that of Example 1, except that the material-liquid ratio of egg yolk powder to ethyl acetate is 1 g: 4 mL.

[0048] Example 3

[0049] The procedure is basically the same as that of Example 1, except that the material-liquid ratio of egg yolk powder to ethyl acetate is 1 g: 6 mL.

[0050] Example 4

[0051] The procedure is basically the same as that of Example 1, except that the material-liquid ratio of egg yolk powder to ethyl acetate is 1 g: 7 mL.

[0052] Example 5

[0053] The procedure is basically the same as that of Example 1, except that the material-liquid ratio of egg yolk powder to ethyl acetate is 1 g: 8 mL.

[0054] Example 6

[0055] The procedure is basically the same as that of Example 1, except that the ultrasonic temperature is 4 °C.

[0056] Example 7

[0057] The procedure is basically the same as that of Example 1, except that the ultrasonic temperature is 25 °C.

[0058] Example 8

[0059] The procedure is basically the same as that of Example 1, except that the ultrasonic power is 80 W.

[0060] Example 9

[0061] The procedure is basically the same as that of Example 1, except that the ultrasonic power is 160 W.

[0062] Example 10

[0063] The procedure is basically the same as that of Example 1, except that the ultrasonic power is 240 W.

[0064] Example 11

[0065] The procedure is basically the same as that of Example 1, except that the ultrasonic power is 400 W.

[0066] Example 12

[0067] The procedure is basically the same as that of Example 1, except that the ultrasonic treatment time is 5 min.

[0068] Example 13

[0069] The steps are basically the same as those in Example 1, except that the ultrasonic treatment time is 10 min.

[0070] Example 14

[0071] The steps are basically the same as those in Example 1, except that the ultrasonic treatment time is 15 min.

[0072] Example 15

[0073] The steps are basically the same as those in Example 1, except that the ultrasonic treatment time is 20 min.

[0074] Example 16

[0075] The steps are basically the same as those in Example 1, except that the ultrasonic treatment time is 25 min.

[0076] Comparative Example 1

[0077] The steps are basically the same as those in Example 1, except that the degreasing method is different. Specifically:

[0078] EYP and ethanol were added to the jacket reactor at (1 g / mL ethanol), and then the mixture was stirred at 65 °C for 10 h. After that, the filter cake and the filtrate were separated. The filter cake was mashed and the solvent was volatilized in a vacuum drying oven at 40 °C for about 24 h. Then it was pulverized to obtain E-EYP.

[0079] Comparative Example 2

[0080] The steps are basically the same as those in Example 1, except that the homogenization method is different. Specifically: The primary emulsion prepared in step (3) of Example 1 (i.e., the water-in-oil (W / O) emulsion containing DHA and hydrophilic nutrients) was mixed with the pre-prepared defatted egg yolk liquid, and was subjected to high-speed shear homogenization treatment at 10000 r / min for 3 min, and then high-pressure homogenization was carried out for 3 cycles under the condition of 50 bar to construct a W / O / W double emulsion, labeled as H.

[0081] (a) Measure the total lipid content and phospholipid content of the defatted egg yolk obtained after degreasing the egg yolk powder in Examples 1-16 above under the conditions of different solid-liquid ratios (egg yolk powder g: ethyl acetate mL is 1:4, 1:5, 1:6, 1:7, 1:8), ultrasonic temperature (0 °C, 4 °C, 25 °C), ultrasonic power (80 W, 160 W, 240 W, 320 W, 400 W), and ultrasonic treatment time (5 min, 10 min, 15 min, 20 min, 25 min). The results are shown in Figure 1 and Figure 2 .

[0082] Method for determining total lipid content: Take 2 g of egg yolk powder and place it in a 50 mL centrifuge tube. Add 20 mL of n-hexane-isopropanol (n-hexane: isopropanol = 3:2 v / v). Homogenize at 6000 rpm for 6 min. Filter the solution through a G3 sintered glass funnel and wash the filter with n-hexane-isopropanol solution. Pour the filtrate into a beaker (already weighed), first evaporate most of the solvent in a 60 °C water bath, and then place it in an oven at 105 °C for constant weight (usually 2 h).

[0083]

[0084] Where: m1: Weight of the sample and beaker after constant weight (g); m2: Weight of the beaker (g).

[0085] Method for determining phospholipid content: The determination of phospholipid content was carried out by referring to the colorimetric method for the determination of phospholipids in exported foods in SNT3851-2014.

[0086] The proportion of phospholipids in the total lipids was calculated according to the following formula

[0087]

[0088] From Figure 1 the results, it can be seen that the lipid content of all groups is lower than that of the original egg yolk powder, and as the amount of ethyl acetate solvent gradually increases, the total lipid content of the egg yolk powder continuously decreases; as the temperature increases, the lipid extraction rate gradually increases; as the ultrasonic power increases, the total lipid content of the egg yolk powder gradually decreases and tends to be stable; as the ultrasonic extraction time prolongs, the egg yolk lipids gradually dissolve in the solvent, and the total lipid content of the egg yolk powder decreases.

[0089] From Figure 2 the results, it can be seen that the proportion of phospholipids shows an inverted U shape. When the solid-liquid ratio reaches 1:5 (w / v), the phospholipid content is the highest. As the amount of solvent further increases, the selectivity gradually decreases; at 0 °C, the selectivity of ethyl acetate is the strongest and the phospholipid content is the highest; when the ultrasonic power is less than 320 W, a large amount of TG is extracted. When the ultrasonic power is 400 W, the cavitation effect is too strong, and more phospholipids dissolve in ethyl acetate, resulting in a significant decrease in the phospholipid content in the egg yolk powder; the phospholipid content reaches the peak at 15 min and then decreases.

[0090] (b) Analyze the components, protein content, chromaticity, solubility, stability coefficient, emulsifying properties, surface hydrophobicity, free sulfhydryl group content, protein secondary structure, and tertiary structure of E-EYP prepared in Comparative Example 1, U-EYP prepared in Example 1, and EYP. The results are shown in Table 1 and Figures 3 - 5 .

[0091] Method for determining protein content: Refer to GB5009.5-2016 and determine the protein content of egg yolk powder according to the Kjeldahl method.

[0092] Powder solubility measurement method: Take 1 g of egg yolk powder in a 50 mL centrifuge tube, and add 30 mL of deionized water. Vortex for 5 min. Then centrifuge at 3000 rpm for 10 min to collect the precipitate. After taking it out, remove the supernatant, add 30 mL of deionized water, shake up and down, and centrifuge again for 10 min. Place it in a blast dryer and dry for 8 h, then take it out and weigh it.

[0093]

[0094] Where: m is the sample mass (g); m1 is the mass of the centrifuge tube + precipitate after constant weight (g); m2 is the mass of the centrifuge tube (g).

[0095] Stability coefficient measurement method: Take 1 g of egg yolk powder and dissolve it in 100 mL of deionized water. Centrifuge at 2000 rpm for 10 min, then take the supernatant with a 10 mL centrifuge tube and measure the absorbance of the supernatant with a UV spectrophotometer. The stability coefficient is calculated according to the following formula.

[0096]

[0097] Where: A1 is the absorbance of the supernatant after centrifugation at the maximum absorption peak at 250 - 350 nm; A0 is the absorbance at the maximum absorption peak at 250 - 350 nm before centrifugation.

[0098] Emulsifying property measurement method: Measure the emulsifying activity (EAI) and emulsifying stability index (ESI) of the sample according to the turbidimetry method. Disperse EYP (2% w / v) in water and stir magnetically at room temperature for 2 h. Take 30 mL of the sample solution and 10 mL of sunflower seed oil. Homogenize at 10000 rpm for 1 min with a homogenizer. Pipette 100 μL of the crude emulsion from the bottom of the test tube and transfer it to 10 mL of SDS solution (0.1%, w / v) for dilution. Measure the turbidity of the diluted solution at 500 nm. EAI is the absorbance A0 measured immediately after emulsification. ESI is calculated using the following formula.

[0099]

[0100] Where: A0 is the absorbance measured immediately at 500 nm for the sample; A t is the absorbance measured after the sample has been placed for 30 min.

[0101] Method for measuring surface hydrophobicity: The surface hydrophobicity of egg yolk powder protein was measured using ANS as a hydrophobic fluorescent probe. 100 mg of egg yolk powder was dissolved in 100 mL of 10 mmol / L phosphate buffer solution with pH 7.0, and stirred for 30 min to prepare a stock solution of egg yolk protein solution with a concentration of 1 mg / mL. Then it was diluted with 10 mmol / L phosphate buffer solution with pH = 7.0 to the sample concentrations between 0.1, 0.2, 0.3, 0.4, and 0.5 mg / mL. Take 4 mL of each diluted sample, add 20 μL of ANS solution (prepared with 10 mmol / L phosphate buffer solution with pH 7.0 and a concentration of 8 mmol / L), shake well, and react in the dark for 10 min. The fluorescence intensity of the sample was measured with a fluorescence spectrophotometer at an excitation wavelength of 390 nm and an emission wavelength of 475 nm. Plot the fluorescence intensity on the vertical axis and the protein concentration on the horizontal axis, and the slope of the initial stage of the curve is the surface hydrophobicity index H0 of the protein molecule.

[0102] Method for measuring the content of free sulfhydryl groups: Weigh about 100 mg of protein sample and dissolve it in 20 mL of 0.086 mol / L Tris-Gly buffer solution. Vortex for 5 min. Take 6 mL of the above protein solution and mix it with 60 μL of Ellman solution. The mixed solution was stored in the dark at room temperature for 15 min. Pipette 1 mL of the protein solution, dilute it 10 times, and measure the absorbance at 412 nm.

[0103]

[0104] In the formula: A 412 is the absorbance at 412 nm; C is the protein concentration in the sample (mg / mL); D is the dilution factor (1.01).

[0105] Fourier transform infrared spectroscopy analysis method: Mix the egg yolk powder sample with potassium bromide, grind it evenly and press it into a tablet, and use Fourier transform infrared light to scan the sample to obtain an infrared spectrum. Test conditions: Scanning range 4000 - 400 cm -1 , resolution 4 cm -1 , cumulative scanning times 64 times, using air as the background, and the background was subtracted before scanning each sample. Take 5 mg of the sample and 500 mg of potassium bromide, and potassium bromide was placed in an oven at 60 °C overnight in advance. Mix the sample and potassium bromide evenly and then press it into a tablet for measurement.

[0106] Fluorescence spectroscopy analysis method: A solution with a sample concentration of 0.5 mg / mL was prepared using 10 mmol / L phosphate buffer (pH = 7.4). At room temperature, the intrinsic fluorescence emission spectra of each diluted sample were detected using a fluorescence spectrophotometer. The excitation wavelength and emission wavelength ranges were set to 290 nm and 400 nm, respectively. In addition, the slit width and scanning speed were 5 nm and 1200 nm / min, respectively. The test temperature was 25 °C. Phosphate buffer was used as the blank for all samples.

[0107] The obtained defatted egg yolk powder has an appearance as Figure 3 in the "U-EYP" group in Figure 4 The stability and emulsifying properties of the defatted egg yolk powder are as Figure 5 in the "U-EYP" group in

[0108] From Figures 3 - 4 it can be seen that the defatted DHA egg yolk powder prepared in Example 1 of the present invention has a lighter color, lower yellowness and brightness, and improved solubility and emulsifying properties.

[0109] Table 1 Total lipid, phospholipid and protein contents of egg yolk powder samples prepared by different defatting processes

[0110]

[0111] From the results in Table 1, it can be seen that there are significant differences in the contents of each component in the defatted egg yolk powder obtained under different defatting process conditions. The total lipid of the original egg yolk powder is 58.77%, and the protein content is 26.46%. After ultrasonic-assisted ethyl acetate extraction, the total lipid content decreased by about 36.58%. For the egg yolk powder E-EYP prepared by the traditional ethanol defatting process, the total lipid content decreased by about 29.38%. And compared with the extraction time of up to 10 h required by the traditional ethanol defatting process, the ultrasonic-assisted ethyl acetate defatting process used in the present invention takes a shorter time (15 min) and has higher efficiency. In addition, there are significant differences in the phospholipid contents of the egg yolk powder obtained by the two processes. Therefore, compared with E-EYP, the phospholipid content of U-EYP increased by about 133.09%. Compared with the original egg yolk powder, the phospholipid content of U-EYP increased by about 75.14%, the proportion of phospholipid in the total lipid increased from 18.68% to 51.60%, and the protein content increased by about 79.21%.

[0112] From Figure 3 it can be seen that compared with the control group, both ultrasonic-assisted defatting and ethanol defatting reduced the brightness of the powder. There is a large color difference between E-EYP and U-EYP and EYP, showing a redder color.

[0113] From Figure 4It can be seen that the solubility of the original egg yolk powder is about 23.97%. After degreasing, the solubility of U-EYP decreased by 23.86%, while that of E-EYP decreased by about 56.74%. After the original egg yolk powder was dissolved, the particles gradually floated up, while after the degreased egg yolk powder was dissolved, the particles gradually sank. This also reflects the difference in lipids between the two egg yolk powders. The original egg yolk powder has a higher lipid content and a lower density, tending to float up. Compared with E-EYP, U-EYP has better dissolution performance. Compared with the original egg yolk powder, there is no significant difference in the EAI of U-EYP, while the EAI of E-EYP decreases. The emulsification stability of U-EYP decreased slightly, while E-EYP almost lost its emulsification performance.

[0114] It can be seen from Figure 5 that after ethanol treatment, the surface hydrophobicity decreased significantly. On the contrary, after ultrasonic-assisted degreasing, the surface hydrophobicity of U-EYP increased significantly; after ultrasonic-assisted degreasing and ethanol degreasing, the total sulfhydryl content decreased, and sulfhydryl oxidation occurred, but the sulfhydryl oxidation of E-EYP was more serious; obvious differences can be seen from the lipid absorption peaks at 2800 cm -1 -3000 cm -1 . The peak intensity is EYP > E-EYP > U-EYP, indicating that the egg yolk powder with ultrasonic degreasing has the lowest lipid content. The amide I band peak at 1600 cm -1 -1700 cm -1 decreased, indicating that both ultrasonic and long-term degreasing will destroy hydrogen bonds; the influence of U-EYP on the protein secondary structure is less than that of E-EYP, and the traditional ethanol degreasing process increases the content of protein β-sheets; after ultrasonic-assisted ethyl acetate degreasing, the maximum absorption wavelength did not change, but the absorption intensity decreased, while the maximum absorption wavelength of E-EYP became 335 nm, showing a 2 nm blue shift compared with the original egg yolk powder, resulting in a significant decrease in solubility.

[0115] (c) Microstructure and Zeta potential analysis were carried out on the W / O / W double emulsion H prepared in Comparative Example 2 and the W / O / W double emulsion S prepared in Example 1. The results are shown in Figures 6 - 7 .

[0116] Optical microscope measurement method: Use an optical microscope to observe the microscopic morphology of the emulsion. The W / O / W double emulsion was observed with a 40× objective lens. Use a pipette to suck 3 μL of the emulsion onto a glass slide, cover it with a coverslip, and use an inverted fluorescence microscope to observe the structure of the W / O / W multi-layer emulsion system.

[0117] Measurement of Zeta potential: The potential of different samples was measured using a multi-angle particle size and high-sensitivity Zeta potential analyzer to determine the surface charge of the droplets. Before the test, the egg yolk sample was diluted 100 times with deionized water, and then the diluted sample was placed in a plastic cuvette. After equilibration for 20 s, data were collected at 25 °C. The Smoluchowsky model was used to convert the mobility measurement values into Zeta potential values.

[0118] It can be seen from Figure 6 that the defatted egg yolk solution serves as the external aqueous phase to encapsulate nutrients. The droplets in the microscopic image are water-in-oil emulsions. The internal aqueous phase contains hydrophilic nutrients such as calcium citrate, ferrous gluconate, zinc gluconate, V C , V E and selenium. The co-encapsulation of hydrophilic nutrients in the internal aqueous phase and defatted egg yolk solution in the external aqueous phase is achieved by constructing a DHA amphiphilic delivery system. Among them, the internal aqueous phase is relatively dispersed in the water-in-oil emulsion. After the emulsion is treated by high-speed shearing, the formed droplets are larger, and there are many large droplets distributed in the field of view. When high-pressure homogenization treatment is carried out, the emulsion droplets become significantly smaller and more uniform. Therefore, from a microscopic perspective, the present invention successfully prepared a W / O / W emulsion.

[0119] It can be seen from Figure 7 that the absolute values of the potentials of the two emulsions are close, but after high-pressure homogenization, the PDI of the emulsion is larger, which may be due to the destruction of the W / O / W emulsion system by high-pressure homogenization, resulting in the instability of the emulsion. Therefore, high-speed shearing treatment performs better in forming smaller droplets and improving the overall stability of the emulsion.

[0120] Compared with the prior art, the total lipid content of the elderly nutrition fortified defatted DHA egg yolk powder provided by the present invention can be reduced by about 36.58%, and the phospholipid content can be increased by about 75.14%. The defatting process is efficiently completed at low temperature and in a short time, selectively retaining phospholipids while reducing damage to the protein structure. In addition, the elderly nutrition fortified defatted DHA egg yolk powder provided by the present invention is rich in various bioactive components such as DHA and hydrophilic nutrients, which can better meet the human body's demand for nutrients.

[0121] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, and improvements made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

Claims

1. A preparation method of an elderly nutrition fortified defatted DHA egg yolk powder, characterized in that It includes the following steps: S1. Use ultrasonic-assisted ethyl acetate to selectively degrease egg yolk powder to obtain degreased egg yolk powder, and then dissolve it with deionized water to obtain degreased egg yolk liquid; S2. Dissolve hydrophilic nutrient elements with deionized water as the inner aqueous phase, use DHA algal oil containing an emulsifier as the oil phase, mix the inner aqueous phase and the oil phase, and perform homogenization treatment to obtain a water-in-oil emulsion containing DHA and hydrophilic nutrient elements; S3. Mix the degreased egg yolk liquid as an emulsifier with the water-in-oil emulsion containing DHA and hydrophilic nutrient elements, and deionized water as the outer aqueous phase, and perform homogenization treatment to obtain a water-in-oil-in-water double emulsion; S4. Add sodium alginate to the obtained water-in-oil-in-water double emulsion, dissolve and mix evenly, and perform spray drying to obtain elderly nutrition fortified degreased DHA egg yolk powder.

2. The preparation method of the elderly nutrition fortified defatted DHA egg yolk powder according to claim 1, characterized in that, In step S1, the egg yolk powder is obtained through the following steps: Wash the eggs, remove the shells, separate the egg whites and yolks, roll the yolks on filter paper to remove the remaining egg whites, pierce the egg yolk membrane with tweezers to collect the egg yolk liquid, dilute the obtained egg yolk liquid and perform spray drying, collect the spray-dried egg yolk powder, and store it frozen at -20°C for standby; among them, the spray flow rate is 10 - 25 mL / min, the dilution ratio of the egg yolk powder is 2 - 5:1, and the inlet air temperature is 180 - 200°C.

3. The preparation method of the elderly nutrition fortified defatted DHA egg yolk powder according to claim 1, characterized in that, In step S1, the specific steps for using ultrasonic-assisted ethyl acetate to selectively degrease the egg yolk powder include: Dilute the egg yolk powder with ethyl acetate, perform ultrasonic treatment, then perform suction filtration, drying, and pulverization to obtain degreased egg yolk powder, and dissolve it with deionized water to obtain degreased egg yolk liquid.

4. The preparation method of the elderly nutrition fortified defatted DHA egg yolk powder according to claim 3, wherein, The material-liquid ratio of the egg yolk powder to ethyl acetate is 1 g: 4 - 8 mL.

5. The preparation method of the elderly nutrition fortified defatted DHA egg yolk powder according to claim 1, characterized in that, The ultrasonic temperature is 0 - 25°C, the ultrasonic power is 80 - 320 w, and the ultrasonic time is 5 - 25 min.

6. The preparation method of the elderly nutrition fortified defatted DHA egg yolk powder according to claim 1, characterized in that, In step S2, the homogenization rate is 9000 - 12000 r / min, and the homogenization time is 1 - 6 min.

7. The preparation method of the geriatric nutrition fortified defatted DHA egg yolk powder according to claim 1, characterized in that, In step S3, the homogenization rate is 8000 - 11000 r / min, and the homogenization time is 2 - 8 min.

8. The preparation method of the elderly nutrition fortified defatted DHA egg yolk powder according to claim 1, characterized in that, In step S3, the volume ratio of the water-in-oil emulsion containing DHA and hydrophilic nutrient elements to the outer aqueous phase is 1: 1 - 2.

9. The preparation method of the elderly nutrition fortified defatted DHA egg yolk powder according to claim 1, characterized in that, The hydrophilic nutrient elements are one or a combination of two or more of selenium, vitamin C, vitamin E, calcium, iron, and zinc. The calcium is derived from calcium citrate, the iron is derived from ferrous gluconate, and the zinc is derived from zinc gluconate; the DHA is selected from any one or a combination of two or more of DHA extracted from algal oil, DHA extracted from fish oil, DHA extracted from perilla seeds, or DHA extracted from flax seeds.

10. An elderly nutrition fortified degreased DHA egg yolk powder prepared by the preparation method according to any one of claims 1 - 9.

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