Preparation method of water-in-oil-in-water emulsion loaded with procyanidine

Abstract

The invention discloses a preparation method of a procyanidine-loaded water-in-oil-in-water emulsion, which comprises the following steps: 1) dissolving procyanidine in deionized water to prepare an inner water phase; 2) heating and melting diglyceride, and dispersing the diglyceride and polyglycerol ricinoleate into oil to obtain an oil phase; 3) adding the internal water phase into the oil phase while stirring, uniformly stirring, and emulsifying to obtain a water-in-oil primary emulsion; 4) dissolving the separated whey protein in PBS to obtain a protein solution, stirring at room temperature, standing, mixing with lecithin, heating to 75-85 DEG C, and stirring; cooling to room temperature to obtain an outer water phase; 5) adding the water-in-oil primary emulsion into the external water phase, and emulsifying to obtain a water-in-oil-in-water emulsion loaded with procyanidine; the procyanidine-loaded water-in-oil-in-water emulsion prepared by the method has the advantages of high embedding rate, strong stability, small emulsion particle size and simple preparation process, and the emulsion system is compounded with diglyceride, so that the emulsion system has the characteristics of low saturation and health.

Description

Technical Field

[0001] The invention belongs to the technical field of emulsions, and in particular relates to a method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins. Background Art

[0002] Proanthocyanidins are highly effective natural antioxidants. Relevant studies have shown that the antioxidant capacity of oligomeric proanthocyanidins is 50 times that of vitamin E and 20 times that of vitamin C. In addition, a large number of in vivo and in vitro studies have shown that proanthocyanidins also have other biological activities and pharmacological effects such as anti-inflammatory, antibacterial, anticancer, neuroprotective, and cardiovascular protection. However, since the structure of proanthocyanidins contains a large number of active phenolic hydroxyl groups and unsaturated bonds, oxidation, polymerization, condensation and other reactions are prone to occur, resulting in poor stability of proanthocyanidins. At the same time, external factors such as pH, temperature, oxygen, and light will also affect the stability of proanthocyanidins during processing and storage. This instability greatly reduces the bioavailability of proanthocyanidins and limits their application in food, medicine and other fields.

[0003] The emulsion systems for encapsulating water-soluble polyphenols reported in current research mainly include oil-in-water and water-in-oil-in-water systems. Compared with single-layer emulsions, water-in-oil-in-water systems have a unique two-membrane three-phase structure, which can better play a role in sustained release of active substances. However, the current water-in-oil-in-water emulsion system loaded with proanthocyanidins also has some shortcomings, one of which is that the embedding rate is not high and the particle size is large. For example, the embedding rate of the water-in-oil-in-water emulsion loaded with proanthocyanidins prepared by Xu et al. is 87.74±3.12% (Molecules, 2018, 23(11): 2808-2808). The encapsulation stability of the water-in-oil-in-water emulsion loaded with proanthocyanidins prepared with whey protein isolate and konjac glucomannan complex as hydrophilic emulsifier is 87.70±1.68% (Li Xueqian, Jilin University, 2023). The encapsulation rate of the water-in-oil-in-water emulsion loaded with proanthocyanidins prepared with a cross-linked complex of betaine-soluble polysaccharides and whey protein isolate as a hydrophilic emulsifier can reach more than 90%, but the emulsion particle size is relatively large (11.30 μm) (Huang Yaocheng, South China University of Technology, 2021). Secondly, the oil phase currently used in the emulsion system is triglyceride, which is the main cause of obesity and related diseases. Therefore, constructing a low-saturated, healthy and highly stable functional delivery system based on functional lipids is an urgent problem to be solved. Summary of the invention

[0004] The purpose of the present invention is to overcome the deficiencies of the prior art and provide a method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins.

[0005] The technical solution of the present invention is summarized as follows:

[0006] A method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins comprises the following steps:

[0007] 1) dissolving proanthocyanidins in deionized water to prepare an inner water phase;

[0008] 2) heating and melting the diglyceride and polyglycerol ricinoleate, dispersing them in the oil to obtain an oil phase;

[0009] 3) Add the inner water phase to the oil phase under stirring, stir evenly, and emulsify at 10000-12000 rpm for 5-8 minutes to obtain water-in-oil colostrum;

[0010] 4) Dissolve the isolated whey protein in a 0.1 M PBS buffer solution at pH = 7.2 to obtain a protein solution, stir at room temperature for 2-4 hours, stop stirring, let stand for 10-12 hours, mix with lecithin, heat to 75-85° C., stir for 30-50 minutes, and cool to room temperature to obtain an external aqueous phase;

[0011] 5) adding the water-in-oil colostrum to the external aqueous phase, emulsifying at 6000-8000 rpm for 3-5 min to obtain a water-in-oil-in-water emulsion loaded with proanthocyanidins.

[0012] Preferably, the content of proanthocyanidins in the inner aqueous phase of step 1) is 1-3 mg / mL.

[0013] Preferably, in step 2), the mass content of diglyceride in the oil phase is 5% to 15%; the mass content of polyglycerol ricinoleate is 3% to 6%.

[0014] Preferably, the oil in step 2) is rapeseed oil, olive oil or soybean oil.

[0015] Preferably, in step 3), the mass ratio of the inner water phase to the oil phase is 1-5:9-5.

[0016] Preferably, the mass content of isolated whey protein in the protein solution of step 4) is 1% to 3%; the mass ratio of isolated whey protein to lecithin is 1:0.06 to 0.4.

[0017] Preferably, in step 5), the mass ratio of water-in-oil colostrum to the external aqueous phase is 7-4:3-6.

[0018] Compared with the prior art, the present invention has the following beneficial technical effects:

[0019] Based on the method of the present invention, the prepared water-in-oil-in-water emulsion loaded with proanthocyanidins has the advantages of high embedding rate, strong stability, small emulsion particle size and simple preparation process. Glycerol diglyceride is compounded in the emulsion system to make it low in saturation and healthy, and has good application prospects in the food field. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 These are the appearance photos of WPI+PL, WPI and PL.

[0021] Figure 2 The graph shows the viscosity measurement results of WPI+PL, WPI and PL.

[0022] Figure 3 This is a graph showing the results of determination of the encapsulation rate of proanthocyanidins in WPI+PL, WPI and PL at different temperatures.

[0023] Figure 4 The droplet size measurement results of WPI+PL, WPI and PL are shown. DETAILED DESCRIPTION

[0024] The present invention is further described in detail below in conjunction with specific embodiments, which are intended to explain the present invention rather than to limit it.

[0025] Example 1

[0026] A method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins comprises the following steps:

[0027] 1) Dissolve proanthocyanidins in deionized water to prepare an inner aqueous phase; the content of proanthocyanidins in the inner aqueous phase is 2 mg / mL

[0028] 2) heating and melting diglyceride and polyglycerol ricinoleate (PGPR) (molecular weight of polyglycerol ricinoleate: 520.69638) and dispersing them in rapeseed oil to obtain an oil phase; the mass content of diglyceride in the oil phase is 10%; the mass content of the polyglycerol ricinoleate is 5%;

[0029] 3) Add the inner water phase to the oil phase under stirring, stir evenly, and emulsify at 11000 rpm for 7 minutes to obtain water-in-oil colostrum; the mass ratio of the inner water phase to the oil phase is 3:7;

[0030] 4) Dissolving the isolated whey protein in a 0.1M PBS buffer solution with a pH of 7.2 to obtain a protein solution, stirring at room temperature for 3 hours, stopping stirring, standing for 11 hours, mixing with lecithin, heating to 80°C, stirring for 40 minutes; cooling to room temperature to obtain an external aqueous phase; the mass content of the isolated whey protein in the protein solution is 1%; the mass ratio of the isolated whey protein to the lecithin is 1:0.1;

[0031] 5) adding the water-in-oil colostrum to the external aqueous phase, emulsifying at 7000 rpm for 4 min to obtain a water-in-oil-in-water emulsion loaded with proanthocyanidins (abbreviated as WPI+PL), wherein the mass ratio of the water-in-oil colostrum to the external aqueous phase is 5:5.

[0032] Example 2

[0033] A method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins comprises the following steps:

[0034] 1) Dissolve proanthocyanidins in deionized water to prepare an inner aqueous phase; the content of proanthocyanidins in the inner aqueous phase is 1 mg / mL

[0035] 2) heating and melting the diglyceride and polyglycerol ricinoleate (PGPR) (molecular weight of polyglycerol ricinoleate: 520.69638), dispersing them into olive oil to obtain an oil phase; the mass content of the diglyceride in the oil phase is 5%; the mass content of the polyglycerol ricinoleate is 6%;

[0036] 3) Add the inner water phase to the oil phase under stirring, stir evenly, and emulsify at 10000 rpm for 8 minutes to obtain water-in-oil colostrum; the mass ratio of the inner water phase to the oil phase is 1:9;

[0037] 4) Dissolving the isolated whey protein in a 0.1M PBS buffer solution at pH=7.2 to obtain a protein solution, stirring at room temperature for 2 hours, stopping stirring, standing for 12 hours, mixing with lecithin, heating to 75°C, stirring for 50 minutes; cooling to room temperature to obtain an external aqueous phase; the mass content of the isolated whey protein in the protein solution is 1%; the mass ratio of the isolated whey protein to the lecithin is 1:0.06;

[0038] 5) adding the water-in-oil colostrum to the external aqueous phase, emulsifying at 6000 rpm for 5 min to obtain a water-in-oil-in-water emulsion loaded with proanthocyanidins, wherein the mass ratio of the water-in-oil colostrum to the external aqueous phase is 7:3.

[0039] Example 3

[0040] A method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins comprises the following steps:

[0041] 1) Dissolve proanthocyanidins in deionized water to prepare an inner water phase; the content of proanthocyanidins in the inner water phase is 3 mg / mL

[0042] 2) heating and melting the diglyceride and polyglycerol ricinoleate (PGPR) (molecular weight of polyglycerol ricinoleate, 520.69638), dispersing them in soybean oil to obtain an oil phase; the mass content of the diglyceride in the oil phase is 15%; the mass content of the polyglycerol ricinoleate is 3%;

[0043] 3) Add the inner water phase to the oil phase under stirring, stir evenly, and emulsify at 12000 rpm for 5 minutes to obtain water-in-oil colostrum; the mass ratio of the inner water phase to the oil phase is 5:5;

[0044] 4) Dissolving the isolated whey protein in a 0.1M PBS buffer solution with a pH of 7.2 to obtain a protein solution, stirring at room temperature for 4 hours, stopping stirring, standing for 10 hours, mixing with lecithin, heating to 85°C, stirring for 30 minutes; cooling to room temperature to obtain an external aqueous phase; the mass content of the isolated whey protein in the protein solution is 3%; the mass ratio of the isolated whey protein to the lecithin is 1:0.4;

[0045] 5) adding the water-in-oil colostrum to the external aqueous phase, emulsifying at 8000 rpm for 3 min to obtain a water-in-oil-in-water emulsion loaded with proanthocyanidins, wherein the mass ratio of the water-in-oil colostrum to the external aqueous phase is 4:6.

[0046] Experimental Example 1

[0047] A method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins comprises the following steps:

[0048] 1) dissolving proanthocyanidins in deionized water to prepare an inner aqueous phase; the content of proanthocyanidins in the inner aqueous phase is 2 mg / mL;

[0049] 2) heating and melting diglyceride and polyglycerol ricinoleate (PGPR) (molecular weight of polyglycerol ricinoleate: 520.69638) and dispersing them in rapeseed oil to obtain an oil phase; the mass content of diglyceride in the oil phase is 10%; the mass content of the polyglycerol ricinoleate is 5%;

[0050] 3) Add the inner water phase to the oil phase under stirring, stir evenly, and emulsify at 11000 rpm for 7 minutes to obtain water-in-oil colostrum; the mass ratio of the inner water phase to the oil phase is 3:7;

[0051] 4) dissolving the isolated whey protein in a 0.1 M PBS buffer solution at pH = 7.2 to obtain a protein solution, stirring at room temperature for 3 hours, stopping stirring, and standing for 11 hours to obtain an external aqueous phase; the mass content of the isolated whey protein in the protein solution is 1%;

[0052] 5) adding the water-in-oil colostrum to the external aqueous phase, emulsifying at 7000 rpm for 4 min to obtain a water-in-oil-in-water emulsion (WPI) loaded with proanthocyanidins, wherein the mass ratio of the water-in-oil colostrum to the external aqueous phase is 5:5.

[0053] Experimental Example 2

[0054] A method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins comprises the following steps:

[0055] 1) dissolving proanthocyanidins in deionized water to prepare an inner aqueous phase; the content of proanthocyanidins in the inner aqueous phase is 2 mg / mL;

[0056] 2) heating and melting diglyceride and polyglycerol ricinoleate (PGPR) (molecular weight of polyglycerol ricinoleate: 520.69638) and dispersing them in rapeseed oil to obtain an oil phase; the mass content of diglyceride in the oil phase is 10%; the mass content of the polyglycerol ricinoleate is 5%;

[0057] 3) Add the inner water phase to the oil phase under stirring, stir evenly, and emulsify at 11000 rpm for 7 minutes to obtain water-in-oil colostrum; the mass ratio of the inner water phase to the oil phase is 3:7;

[0058] 4) Dissolve 1 gram of lecithin in 100 mL of 0.1 M PBS buffer at pH 7.2 and stir evenly to form the external aqueous phase;

[0059] 5) adding the water-in-oil colostrum to the external aqueous phase, emulsifying at 7000 rpm for 4 min to obtain a water-in-oil-in-water emulsion loaded with proanthocyanidins (abbreviated as PL), wherein the mass ratio of the water-in-oil colostrum to the external aqueous phase is 5:5.

[0060] Experimental Example 3

[0061] The WPI prepared in Experimental Example 1, the PL prepared in Experimental Example 2, and the water-in-oil-in-water emulsion loaded with proanthocyanidins (WPI+PL for short) prepared in Example 1 were placed at room temperature for 0 days and 7 days. Figure 1 .

[0062] Depend on Figure 1 It can be seen that: no stratification occurred in the three samples at 0 days. After being placed at room temperature for seven days, PL showed obvious stratification, WPI showed slight stratification, and WPI+PL showed no stratification.

[0063] Experimental Example 4

[0064] The viscosity of the three samples was measured using a digital viscometer.

[0065] Depend on Figure 2 It can be seen that:

[0066] WPI+PL>WPI>PL,

[0067] Combination Figure 1 It can be seen that WPI+PL has stronger emulsification ability and better emulsification stability, which may be due to the fact that the hydrogen bonds or hydrophobic interactions between whey protein isolate and lecithin enhance the stability of the interfacial film.

[0068] Experimental Example 5

[0069] Take 1mL of WPI+PL, WPI and PL, add 2mL of deionized water to dilute, mix with vortex, centrifuge at 4000r / min for 10min, collect the lower clear liquid with a syringe, and filter through a 0.45μm MCE water filter to obtain a filtrate. Then the vanillin-hydrochloric acid method was used to determine the proanthocyanidin content in the filtrate. Subsequently, the effect of different temperature conditions (initial, 30℃, 40℃, 50℃, 60℃ and 70℃) on the proanthocyanidin embedding rate was determined, and the free proanthocyanidin aqueous solution was used as a control.

[0070] The calculation formula of embedding rate is:

[0071]

[0072] Depend on Figure 3 It can be seen that proanthocyanidin is a heat-sensitive substance that is easily degraded under high temperature conditions. The embedding rate of the free proanthocyanidin solution gradually decreases as the temperature increases, and the decrease is most significant when the temperature rises to 50°C. When the temperature rises to 60°C, the proanthocyanidin embedding rates of WPI+PL and WPI are both higher than those of the free aqueous solution, among which the embedding rate of WPI+PL is the highest, indicating that the two-membrane three-phase structure formed by the water-in-oil-in-water emulsion loaded with proanthocyanidins prepared by the method of the present invention has a good protective effect on proanthocyanidins.

[0073] Experimental Example 6

[0074] The particle size distribution was measured using a Malvern laser particle size analyzer. Deionized water was used as the dispersant, and the refractive index of water was 1.33. Each sample was measured three times in parallel and the average value was taken. The particle size distribution of WPI+PL, WPI and PL was obtained as shown in the figure below: Figure 4 shown.

[0075] Depend on Figure 4 It can be seen that compared with WPI and PL, the particle size distribution of WPI+PL shifts to the left, indicating that the emulsion particle size formed by WPI+P is smaller and a more structurally stable emulsion system is formed compared with WPI and PL.

Claims

1. A method for preparing a water-in-oil-in-water emulsion loaded with proanthocyanidins, characterized in that: The following steps are involved: 1) dissolving proanthocyanidins in deionized water to prepare an inner water phase; 2) heating and melting the diglyceride and polyglycerol ricinoleate, dispersing them in the oil to obtain an oil phase; 3) Add the inner water phase to the oil phase under stirring, stir evenly, and emulsify at 10000-12000 rpm for 5-8 minutes to obtain water-in-oil colostrum; 4) Dissolve the isolated whey protein in a 0.1 M PBS buffer solution at pH = 7.2 to obtain a protein solution, stir at room temperature for 2-4 hours, stop stirring, let stand for 10-12 hours, mix with lecithin, heat to 75-85° C., stir for 30-50 minutes, and cool to room temperature to obtain an external aqueous phase; 5) adding the water-in-oil colostrum to the external aqueous phase, emulsifying at 6000-8000 rpm for 3-5 min to obtain a water-in-oil-in-water emulsion loaded with proanthocyanidins.

2. The method according to claim 1, characterized in that Step 1) The content of proanthocyanidins in the inner aqueous phase is 1-3 mg / mL.

3. The method according to claim 1, characterized in that Step 2) The mass content of diglyceride in the oil phase is 5% to 15%; the mass content of polyglycerol ricinoleate is 3% to 6%.

4. The method according to claim 1, characterized in that Step 2) The oil is rapeseed oil, olive oil or soybean oil.

5. The method according to claim 1, characterized in that Step 3) The mass ratio of the inner water phase to the oil phase is 1-5:9-5.

6. The method according to claim 1, characterized in that Step 4) The mass content of isolated whey protein in the protein solution is 1% to 3%; the mass ratio of isolated whey protein to lecithin is 1:0.06 to 0.

4.

7. The method according to claim 1, characterized in that In step 5), the mass ratio of water-in-oil colostrum to external water phase is 7-4:3-6.

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