W / O / W emulsion for efficiently delivering active substances to intestinal tracts and preparation method of W / O / W emulsion

Abstract

The invention belongs to the technical field of carrier delivery, and provides a W / O / W emulsion for efficiently delivering active substances to intestinal tracts and a preparation method of the W / O / W emulsion. The W / O / W emulsion provided by the invention is prepared by constructing an outer water phase through hydrated whey protein isolate and cellulose nanocrystals, taking a glycerol-trehalose aqueous solution as an inner water phase, compounding an emulsifier oil phase and performing high-speed dispersion twice. According to the preparation method, the storage stability (TSilt, 5, Zeta potential absolute value gt, 30mV for storage in 5d) and digestion tolerance of the emulsion can be remarkably improved through the synergistic effect of the protein-cellulose nanocrystals, and internal-phase bioactive substances are prevented from being inactivated through dual protection; the external water-phase cellulose nanocrystals form a physical barrier, so that the digestion rate of whey protein is delayed (the digestion rate at the gastric digestion stage is reduced by more than 40%), the controlled release of functional components is realized, and a structural guarantee is provided for an oral delivery system.

Description

Technical Field

[0001] This invention belongs to the field of carrier delivery technology, and specifically relates to a W / O / W emulsion for efficiently delivering active substances to the intestine and its preparation method. Background Technology

[0002] Traditional W / O / W emulsion technology has the following three drawbacks: Firstly, during the digestion of a single protein in the stomach, the disordered coiled structure of the external aqueous phase is easily overstretched and broken due to the mechanical stress generated by gastric acid (pH<2) and gastric peristalsis. This leads to a sudden drop in the viscosity of the external phase of the emulsion and the rupture of the interfacial membrane, causing rapid demulsification. As a result, the physical barrier function of the double emulsion completely fails within 15-30 minutes. Secondly, after demulsification, the inner aqueous phase is directly exposed to pepsin (concentration >0.5mg / mL) and an extremely acidic environment, resulting in a decrease of more than 80% in the survival rate of the loaded probiotics, irreversible damage to the secondary structure of functional proteins (such as insulin), and a reduction in bioavailability to less than 20% of the original value. Finally, the unmodified protein aqueous phase undergoes rapid degradation during gastric digestion (half-life <30 minutes), making it impossible to achieve programmed release of functional components in the intestinal target area. This makes it difficult to meet the precision medicine needs such as oral colon-targeted delivery, thus limiting its application value in the fields of functional foods and pharmaceutical preparations. Summary of the Invention

[0003] To address the above-mentioned technical problems, this invention proposes a W / O / W emulsion for efficiently delivering active substances to the intestines and its preparation method.

[0004] The W / O / W emulsion provided by this invention is prepared by constructing an outer aqueous phase with hydrated whey protein isolate and cellulose nanocrystals, an inner aqueous phase with a glycerol-trehalose aqueous solution, and an oil phase with a compound emulsifier through two high-speed dispersions. This preparation method significantly improves the emulsion's storage stability (5-day storage TSI < 5, absolute value of Zeta potential > 30 mV) and digestibility through the synergistic effect of protein and cellulose nanocrystals, providing dual protection against the inactivation of bioactive substances in the inner phase. The cellulose nanocrystals in the outer aqueous phase form a physical barrier, slowing down the digestion rate of whey protein (reducing the digestion rate by more than 40% during gastric digestion), thus achieving controlled release of functional components.

[0005] The technical solution of this invention is: A W / O / W emulsion for highly efficient delivery of active substances to the intestine, wherein the W / O / W emulsion is obtained by the following method: A W / O / W emulsion was prepared by constructing an outer aqueous phase with hydrated whey protein isolate and cellulose nanocrystals, using a glycerol-trehalose aqueous solution as the inner aqueous phase, and then mixing an emulsifier with an oil phase and dispersing twice at high speed.

[0006] The above-mentioned method for preparing W / O / W emulsions includes the following steps: (1) Dissolve whey protein isolate in water and stir at room temperature for 1.5-2.5 h. After it is fully dissolved, place the prepared whey protein isolate solution at 4℃ and hydrate for 6-10 h. (2) Preparation of cellulose nanocrystals: Microcrystalline cellulose and sulfuric acid solution with a concentration of 9 mol / L are hydrolyzed at a ratio of 1:(15-25) at 40-60℃ for 3.5-4.5 h. (3) Add cellulose nanocrystals to the hydrated whey protein isolate solution, and use the resulting solution as the external aqueous phase of the emulsion; (4) Dissolve glycerol and trehalose in water, and use the resulting solution as the inner aqueous phase of the emulsion; (5) Dissolve sucrose fatty acid ester, polyglycerol ricinoleate and lecithin in oil, and use the resulting solution as the oil phase of the emulsion; (6) Mix the internal aqueous phase and oil phase, disperse at 10000-15000 r / min for 5-15 min to prepare a W / O emulsion; The W / O emulsion is then mixed with the external aqueous phase and dispersed at 8000-12000 r / min for 3-8 min to obtain a W / O / W emulsion.

[0007] Preferably, in (1), the mass fraction of whey protein isolate in the whey protein isolate solution is 2.5-8%.

[0008] Preferably, in (3), the amount of cellulose nanocrystals added is 0.5wt%-4wt%.

[0009] Preferably, in (4), the mass ratio of glycerol to trehalose is 2:5-15.

[0010] Preferably, in (5), the mass ratio of sucrose fatty acid ester: polyglycerol ricin glycolate: lecithin is 1:(1-5):(1-5).

[0011] Preferably, in (5), the oil is selected from any one of soybean oil, sunflower oil, and rapeseed oil; The dissolution conditions are: stirring in a water bath at 45-80℃ for 25-45 minutes.

[0012] Preferably, in (6), the mass ratio of the internal aqueous phase to the oil phase is 1:3-5; The mass ratio of W / O emulsion to external aqueous phase is 2:2-5.

[0013] Preferably, in (6), the mixing of the internal aqueous phase and the oil phase requires premixing, and the premixing operation is as follows: In a water bath at 40°C, the aqueous phase is added dropwise to the oil phase while stirring, until the two are mixed evenly.

[0014] The present invention has the following advantages and effects compared with the prior art: (1) Through the synergistic effect of protein-cellulose nanocrystals, the storage stability of the emulsion is significantly improved (5d storage TSI<5, absolute value of Zeta potential>30mV), and digestion tolerance is enhanced simultaneously. The microstructure is maintained after gastric digestion, and the dual protection mechanism effectively prevents the inactivation of bioactive substances in the internal phase. (2) Cellulose nanocrystals in the external aqueous phase form a physical barrier, which slows down the digestion rate of whey protein (the digestion rate is reduced by more than 40% during the gastric digestion stage), thereby achieving the controlled release effect of functional components and providing structural protection for oral delivery systems. (3) Using widely available whey protein and cellulose nanocrystals as core raw materials ensures material availability and reduces dependence on a single stabilizer through particle-protein composite, making costs controllable. Attached Figure Description

[0015] Figure 1 This is the Fourier transform infrared spectrum of the present invention; Figure 2 S1, S2, and S3 represent the particle size and PDI of the cellulose nanocrystals in this invention, where S1, S2, and S3 represent three repeated measurements. Figure 3 The TSI variation diagram shows the changes in W / O / W emulsions prepared by adding different amounts of sucrose fatty acid esters to the oil phase during short-term storage in this invention. Figure 4 The backscattered light reference change diagram shows the changes in W / O / W emulsions prepared with different amounts of sucrose fatty acid esters added to the oil phase during short-term storage in Experiment 2 of this invention. In the diagram, A: blank control group, B: W / O / W emulsion with 1% SE-1 added, C: W / O / W emulsion with 2% SE-1 added, D: W / O / W emulsion with 3% SE-1 added, and E: W / O / W emulsion with 4% SE-1 added. Figure 5 The particle size variation diagram of W / O / W emulsions prepared with different amounts of sucrose fatty acid esters added to the oil phase during short-term storage in Experiment 2 of this invention is shown in the figure. A: blank control group, B: W / O / W emulsion with 1% SE-1 added, C: W / O / W emulsion with 2% SE-1 added, D: W / O / W emulsion with 3% SE-1 added, E: W / O / W emulsion with 4% SE-1 added. Figure 6 The potential change graph shows the changes in the potential values ​​of W / O / W emulsions prepared by adding different amounts of sucrose fatty acid esters to the oil phase during short-term storage in Experiment 2 of this invention. Figure 7Laser confocal scanning microstructure of W / O / W emulsions prepared with different amounts of sucrose fatty acid esters added to the oil phase in Experiment 2 of this invention, wherein, a: blank control group, b: W / O / W emulsion with 1% SE-1 added, c: W / O / W emulsion with 2% SE-1 added, d: W / O / W emulsion with 3% SE-1 added, e: W / O / W emulsion with 4% SE-1 added; Figure 8 The images show the microstructures of W / O / W emulsions with different amounts of sucrose fatty acid esters added during storage in Experiment 2 of this invention. In the images, A: blank control group, B: W / O / W emulsion with 1% SE-1 added, C: W / O / W emulsion with 2% SE-1 added, D: W / O / W emulsion with 3% SE-1 added, and E: W / O / W emulsion with 4% SE-1 added. Figure 9 The optical microscope images and images of W / O / W emulsions prepared by adding different amounts of cellulose nanocrystals in the external aqueous phase during short-term storage in Experiment 3 of this invention are shown. Figure 10 The TSI variation graph shows the changes in W / O / W emulsions prepared by adding different amounts of cellulose nanocrystals in the external aqueous phase during short-term storage in Experiment 3 of this invention. Figure 11 These are laser confocal scanning microstructure images of the emulsions at each digestion stage in Experiment 4 of this invention. Among them, A: blank control group, B: W / O / W emulsion with 1% cellulose nanocrystals, C: W / O / W emulsion with 2% cellulose nanocrystals, D: W / O / W emulsion with 3% cellulose nanocrystals, and E: W / O / W emulsion with 4% cellulose nanocrystals. Detailed Implementation

[0016] To enable those skilled in the art to better understand the present invention, the present invention will now be further described in conjunction with specific embodiments.

[0017] Example 1 Table 1. Reagents and pharmaceuticals used in this invention reagents and medicines source Whey protein isolate Shanghai Yuanye Biotechnology Co., Ltd. Cellulose nanocrystals Cellulose nanocrystals were extracted from microcrystalline cellulose prepared from conifer pulp, as shown in Experimental Example 1. glycerin Sinopharm Chemical Reagent Co., Ltd. Trehalose Japan Hayashihara Co., Ltd. Sucrose fatty acid esters Liuzhou Aigefu Food Technology Co., Ltd. Polyglycerol ricinoleate Zhengzhou Dahe Food Technology Co., Ltd. Lecithin Shanghai Youchuang Industrial Co., Ltd. soybean oil Yihai Kerry Golden Dragon Fish Food Group Co., Ltd.

[0018] This embodiment provides a method for preparing a W / O / W emulsion that efficiently delivers active substances to the intestine, comprising the following steps: (1) Dissolve whey protein isolate in water and stir continuously at room temperature for 2 hours to ensure complete dissolution. Place the prepared whey protein isolate solution with a mass fraction of 5% in a refrigerator at 4°C and hydrate for 8 hours. The hydrated whey protein isolate solution is then ready for use. (2) Add 1% by mass of cellulose nanocrystals to the whey protein isolate solution after hydration for 8 h, and use the resulting solution as the external aqueous phase of the emulsion; (3) Dissolve glycerol and trehalose in water at a certain mass fraction. The mass fraction of glycerol in the resulting solution is 2% and the mass fraction of trehalose is 10%. Use this solution as the inner aqueous phase of the emulsion. (4) Dissolve sucrose fatty acid ester, polyglycerol ricinoleate and lecithin in soybean oil at a mass ratio of 1:3:3, stir in a water bath at 60°C for 30 min, and use the resulting solution as the oil phase of the emulsion; (5) Mix the inner aqueous phase and the oil phase at a mass ratio of 1:4 and disperse them for 10 min using a high-speed disperser at a speed of 13000 r / min to prepare a W / O emulsion. Premixing is required during the preparation of the W / O emulsion. That is, add the inner aqueous phase dropwise to the oil phase in a 40℃ magnetic stirring water bath and stir them evenly. The prepared W / O emulsion was mixed with the external aqueous phase at a mass ratio of 2:3 and dispersed in an ice bath at a speed of 10000 r / min for 5 min to obtain the W / O / W emulsion.

[0019] The preparation environment for W / O emulsions and W / O / W emulsions is an ice bath.

[0020] Example 2 A method for preparing a W / O / W emulsion for efficiently delivering active substances to the intestine includes the following steps: (1) Dissolve whey protein isolate in water and stir continuously at room temperature for 1.5 h to ensure complete dissolution. Place the prepared whey protein isolate solution with a mass fraction of 2.5% in a refrigerator at 4℃ and hydrate for 6 h. The hydrated whey protein isolate solution is ready for use. (2) Add cellulose nanocrystals to the hydrated whey protein isolate solution at a mass fraction of 0.5%, and use the resulting solution as the external aqueous phase of the emulsion; (3) Dissolve glycerol and trehalose in water at a mass ratio of 2:5. The resulting solution contains 2% glycerol and 5% trehalose by mass. Use this solution as the inner aqueous phase of the emulsion. (4) Dissolve sucrose fatty acid ester, polyglycerol ricinoleate and lecithin in sunflower seed oil at a mass ratio of 1:1:1, stir in a water bath at 45°C for 25 min, and use the resulting solution as the oil phase of the emulsion; (5) Mix the inner aqueous phase and the oil phase at a mass ratio of 1:3 and disperse them for 15 min using a high-speed disperser at a speed of 10000 r / min to prepare a W / O emulsion. Premixing is required during the preparation of the W / O emulsion. That is, add the inner aqueous phase dropwise to the oil phase in a 40℃ magnetic stirring water bath and stir them evenly. The prepared W / O emulsion was mixed with the external aqueous phase at a mass ratio of 2:2 and dispersed in an ice bath at 8000 r / min for 8 min to obtain the W / O / W emulsion.

[0021] The preparation environment for W / O emulsions and W / O / W emulsions is an ice bath.

[0022] Example 3 A method for preparing a W / O / W emulsion for efficiently delivering active substances to the intestine includes the following steps: (1) Dissolve whey protein isolate in water and stir continuously at room temperature for 2.5 h to ensure complete dissolution. Place the prepared whey protein isolate solution with a mass fraction of 8% in a refrigerator at 4℃ and hydrate for 10 h. The hydrated whey protein isolate solution is then ready for use. (2) Add 4% by mass of cellulose nanocrystals to the hydrated whey protein isolate solution, and use the resulting solution as the external aqueous phase of the emulsion; (3) Dissolve glycerol and trehalose in water at a certain mass fraction. The mass fraction of glycerol in the resulting solution is 2% and the mass fraction of trehalose is 15%. Use this solution as the inner aqueous phase of the emulsion. (4) Dissolve sucrose fatty acid ester, polyglycerol ricinoleate and lecithin in soybean oil at a mass ratio of 1:5:5, stir in a water bath at 80°C for 45 min, and use the resulting solution as the oil phase of the emulsion; (5) Mix the inner aqueous phase and the oil phase at a mass ratio of 1:5 and disperse them for 5 min at a speed of 15000 r / min using a high-speed disperser to prepare a W / O emulsion. Premixing is required during the preparation of the W / O emulsion, i.e., the inner aqueous phase is added drop by drop to the oil phase in a 40℃ magnetic stirring water bath to make the two evenly mixed. The prepared W / O emulsion was mixed with the external aqueous phase at a mass ratio of 2:5 and dispersed in an ice bath at 12000 r / min for 3 min to obtain the W / O / W emulsion.

[0023] The preparation environment for W / O emulsions and W / O / W emulsions is an ice bath.

[0024] Experimental Example 1: Preparation and Analysis of Cellulose Nanocrystals 1.1 Preparation of cellulose nanocrystals involved in step (2) of Examples 1-3 The extraction of cellulose nanocrystals from microcrystalline cellulose prepared from conifer pulp includes the following steps: (1) Prepare a 9 mol / L sulfuric acid solution, and hydrolyze the 9 mol / L sulfuric acid solution with microcrystalline cellulose at a ratio of 1:20 at 55°C with magnetic stirring for 4 hours; (2) After the reaction, a pale yellow liquid was obtained. 20 times the volume of cold distilled water was added to stop the reaction. The mixture was allowed to stand at room temperature for 12 hours. The upper pale yellow liquid was discarded. The lower white suspension was placed in a dialysis bag with a molecular cutoff of 3000 Da-4000 Da and dialyzed until a neutral cellulose nanocrystal suspension was obtained. (3) The neutral cellulose nanocrystal suspension was rotary evaporated at 45°C to concentrate the prepared cellulose nanocrystal solution. The solution was then ultrasonically disrupted for 30 minutes using a cell disruptor at a power of 67% (800W) and the prepared cellulose nanocrystal solution was freeze-dried.

[0025] 1.2 Analysis of cellulose nanocrystals Fourier transform infrared spectroscopy was used to compare the prepared cellulose nanocrystals with microcrystalline cellulose and commercially available cellulose nanocrystals.

[0026] like Figure 1 As shown, the prepared cellulose nanocrystals were found to possess the characteristic peaks of cellulose nanocrystals, with a peak at 812 cm⁻¹. -1 The presence of a characteristic peak of a sulfonic acid group indicates that a sulfonic acid group was introduced onto the -OH group of cellulose after sulfuric acid hydrolysis.

[0027] A 0.0005 wt% cellulose nanocrystal solution was prepared, and the particle size and dispersibility coefficient of the prepared cellulose nanocrystals were measured using a nanoparticle size analyzer.

[0028] like Figure 2 As shown, the cellulose nanocrystals prepared by ultrasonic crushing all have nanoscale particle sizes, and their polydispersity index (PDI) indicates that the prepared cellulose nanocrystals have excellent dispersibility.

[0029] Experiment Example 2: Screening Experiment on the Addition Ratio of Sucrose Fatty Acid Esters Based on Example 1, the addition ratios of substances in the internal and external aqueous phases remained unchanged. The addition amounts of sucrose fatty acid esters in the oil phase were set at 1 wt%, 2 wt%, 3 wt%, 4 wt%, and a blank control group. The W / O / W emulsions prepared with different addition amounts of sucrose fatty acid esters in the oil phase were analyzed using multiple light scattering, particle size, potential, microstructure, and macroscopic morphology measurements. A comprehensive analysis of the measured indicators was then conducted.

[0030] The effect of different amounts of sucrose fatty acid esters added to the oil phase on the total sulfide concentration (TSI) of W / O / W emulsions was determined using a Turbiscan stability analyzer. Figure 3As shown in the figure, during the 5-day storage period, the addition of sucrose fatty acid esters had a significant impact on the TSI of the W / O / W emulsion, with the W / O / W emulsion containing 1% sucrose fatty acid esters exhibiting the lowest TSI. The effect of the amount of sucrose fatty acid esters added on the backscattered light of the W / O / W emulsion is shown in the figure. Figure 4 As shown, during the 5-day storage period, all W / O / W emulsions exhibited a floating phenomenon.

[0031] The particle size distribution of the emulsion during storage was detected using a laser particle size analyzer. The effect of sucrose fatty acid ester addition on the particle size of the W / O / W emulsion during short-term storage was investigated. Figure 5 As shown, the particle size of both emulsions gradually decreases with increasing storage time. Furthermore, the addition of sucrose fatty acid esters leads to an increase in the droplet size of the W / O / W emulsion. However, during a 5-day storage period, the droplet size distribution of the W / O / W emulsion with 1% sucrose fatty acid esters showed less change compared to other W / O / W emulsions with different sucrose fatty acid ester addition levels.

[0032] The potential values ​​of the emulsion during storage were measured using a Malvern nanoparticle size analyzer. The effect of sucrose fatty acid ester addition on the potential values ​​of W / O / W emulsions during short-term storage was investigated. Figure 6 As shown, the absolute potential value of the double emulsion increases with storage time, and the absolute potential value of all emulsions is greater than 25mV. Among them, the absolute potential value of the W / O / W emulsion with 1% sucrose fatty acid ester added changes more significantly during storage, and reaches its maximum at 5 days.

[0033] The microstructure of W / O / W emulsions with different amounts of sucrose fatty acid esters was examined using laser confocal scanning microscopy. The effect of sucrose fatty acid ester content on the microstructure of W / O / W emulsions is as follows: Figure 7 As shown. The oil phase of the W / O / W emulsion was stained with 1 mg / mL Nile Red, and the proteins in the W / O / W emulsion were stained with 1 mg / mL FITC. A comprehensive analysis was conducted by comparing the microstructures of W / O / W emulsions prepared with different amounts of sucrose fatty acid esters. The results showed that the W / O / W emulsion with 1% sucrose fatty acid esters exhibited a better dual structure.

[0034] The microstructure of W / O / W emulsions with different amounts of sucrose fatty acid esters added during storage was examined using an optical microscope. The effects of sucrose fatty acid ester addition on the microstructure and macroscopic state of the W / O / W emulsions were investigated as follows: Figure 8 As shown, after 5 days of storage, the double emulsions with different concentrations of SE-1 all exhibited a dual structure. With increasing storage time, the emulsions separated into layers; the separation height gradually decreased with increasing sucrose fatty acid ester content.

[0035] W / O / W emulsions prepared with different amounts of sucrose fatty acid esters added to the oil phase during short-term storage were analyzed using multiple light scattering, particle size, potential, microstructure, and macroscopic morphology. The measured indicators were then comprehensively analyzed. Compared to W / O / W emulsions prepared with other addition amounts, the W / O / W emulsion with 1% sucrose fatty acid esters exhibited the lowest TSI and the highest absolute potential value while possessing a better microstructure, demonstrating that the W / O / W emulsion with 1% sucrose fatty acid esters exhibited better stability.

[0036] Experimental Example 3: Screening Experiment for the Addition Ratio of Cellulose Nanocrystals Based on Example 2, the addition ratios of substances in the internal aqueous and oil phases remained unchanged. The addition amounts of cellulose nanocrystals in the external aqueous phase were set at 1 wt%, 2 wt%, 3 wt%, 4 wt%, and a blank control group. The W / O / W emulsions prepared with different addition amounts of cellulose nanocrystals in the external aqueous phase were subjected to multiple light scattering, microstructure, and macroscopic morphology analysis, and the measured indicators were comprehensively analyzed.

[0037] Macroscopic and microscopic images of W / O / W emulsions prepared with different amounts of CNCs added to the external aqueous phase are shown below. Figure 9 As shown, during short-term storage, both the emulsions prepared with WPI as the external aqueous phase and the emulsions prepared with WPI-CNC exhibited a relatively obvious dual structure. No phase separation was observed in the freshly prepared emulsions, but with prolonged storage, phase separation occurred in all emulsions except for the one without CNC. This demonstrates that adding CNC to the external aqueous phase helps stabilize the emulsion.

[0038] Using a Turbiscan stability analyzer, multiple light scattering analysis was performed on W / O / W emulsions prepared with different CNC addition amounts in the external aqueous phase to determine the effect of CNC addition amount on the TSI of W / O / W emulsions. Figure 10 As shown in the figure, during storage, the TSI of the emulsion without CNC was significantly higher than that of the emulsion with CNC, indicating that the addition of CNC can enhance the stability of the emulsion.

[0039] During short-term storage, W / O / W emulsions prepared with different amounts of CNC (carbon nanoparticle) in the external aqueous phase were analyzed for multiple light scattering, microstructure, and macroscopic morphology. The measured indicators were then comprehensively analyzed. Compared to W / O / W emulsions without CNC, those with CNC exhibited improved overall stability despite possessing a distinct dual structure; however, the overall stability of W / O / W emulsions with different CNC addition amounts did not differ significantly. To achieve efficient delivery to the intestines, in vitro simulated digestion experiments were conducted to further screen the appropriate CNC addition amount.

[0040] Experimental Example 4: In vitro simulated digestion experiment Using the optimal sucrose fatty acid ester addition amount obtained in Experiment 1, W / O / W emulsions were prepared with cellulose nanocrystal addition amounts of 1wt%, 2wt%, 3wt%, and 4wt% in the external aqueous phase, as well as a blank control group. In vitro simulated digestion experiments were conducted on the prepared W / O / W emulsions, and the microstructure of the W / O / W emulsions after in vitro simulated digestion was determined. The in vitro simulated digestion characteristics of the prepared W / O / W emulsions were analyzed.

[0041] Oral digestion: Mix 5.0 g emulsion, 15 mL PBS buffer and 4 mL oral simulation solution (containing 0.1 g mucin) and stir in a constant temperature water bath at 37 °C for 30 s (100 r / min).

[0042] Gastric digestion: 20 mL of the product after oral digestion was mixed with 6.4 mL of gastric simulated solution (containing 0.064 g pepsin) and reacted in a constant temperature water bath at 37 °C for 1 h (100 r / min).

[0043] Small intestinal digestion: Take 30 mL of the product after gastric digestion, add 6.4 mL of intestinal digestion simulation solution (0.064 g pancreatic enzyme), and react in a constant temperature water bath at 37℃ for 2 h (100 r / min). The temperature is kept constant at 37℃ throughout the process.

[0044] The microstructure of W / O / W emulsions in the undigested, oral, gastric, and intestinal digestion stages was examined using laser confocal scanning microscopy. The results are as follows: Figure 11 As shown in the laser confocal scanning microscopy images after each digestion stage, the W / O / W emulsion with 1% cellulose nanocrystals still retains its dual structure during gastric digestion and is digested during intestinal digestion. This demonstrates that adding 1% sucrose fatty acid ester to the oil phase and 1% cellulose nanocrystals to the aqueous phase in a W / O / W emulsion can achieve efficient delivery to the intestines.

[0045] In summary, this invention provides a W / O / W emulsion for efficient delivery of active substances to the intestine and its preparation method. This method significantly improves the emulsion's storage stability (5-day storage TSI < 5, absolute Zeta potential > 30 mV) and digestive tolerance through the synergistic effect of protein-cellulose nanocrystals, providing dual protection against the inactivation of bioactive substances in the inner phase. The outer aqueous phase cellulose nanocrystals form a physical barrier, slowing down the digestion rate of whey protein (reducing the digestion rate by more than 40% during gastric digestion), achieving controlled release of functional components and providing structural assurance for oral delivery systems.

[0046] The above are merely preferred embodiments of the present invention and do not limit the scope of the patent. All equivalent changes and modifications made within the scope of the present invention should still fall within the scope of the present invention.

Claims

1. A W / O / W emulsion for efficiently delivering active substances to the intestines, characterized in that, The W / O / W emulsion is obtained by the following method: A W / O / W emulsion was prepared by constructing an outer aqueous phase with hydrated whey protein isolate and cellulose nanocrystals, and an inner aqueous phase with glycerol-trehalose aqueous solution, and a compound emulsifier oil phase through two high-speed dispersions. The cellulose nanocrystals were extracted from microcrystalline cellulose prepared from conifer pulp.

2. The method for preparing the W / O / W emulsion as described in claim 1, characterized in that, The steps include the following: (1) Dissolve whey protein isolate in water and stir at room temperature for 1.5-2.5 h. After it is fully dissolved, place the prepared whey protein isolate solution at 4℃ and hydrate for 6-10 h. (2) Preparation of cellulose nanocrystals: Microcrystalline cellulose and sulfuric acid solution with a concentration of 9 mol / L are hydrolyzed at a ratio of 1:(15-25) at 40-60℃ for 3.5-4.5 h. (3) Add cellulose nanocrystals to the hydrated whey protein isolate solution, and use the resulting solution as the external aqueous phase of the emulsion; (4) Dissolve glycerol and trehalose in water, and use the resulting solution as the inner aqueous phase of the emulsion; (5) Dissolve sucrose fatty acid ester, polyglycerol ricinoleate and lecithin in oil, and use the resulting solution as the oil phase of the emulsion; (6) Mix the internal aqueous phase and oil phase, disperse at 10000-15000 r / min for 5-15 min to prepare a W / O emulsion; The W / O emulsion is then mixed with the external aqueous phase and dispersed at 8000-12000 r / min for 3-8 min to obtain a W / O / W emulsion.

3. The preparation method as described in claim 2, characterized in that, In (1), the whey protein isolate solution contains 2.5-8% whey protein isolate by mass.

4. The preparation method as described in claim 2, characterized in that, In (3), the amount of cellulose nanocrystals added is 0.5wt%-4wt%.

5. The preparation method as described in claim 2, characterized in that, In (4), the mass ratio of glycerol to trehalose is 2:5-15.

6. The preparation method as described in claim 2, characterized in that, In (5), the mass ratio of sucrose fatty acid ester: polyglycerol ricin glycolate: lecithin is 1:(1-5):(1-5).

7. The preparation method as described in claim 2, characterized in that, (5) The oil is selected from any one of soybean oil, sunflower seed oil, and rapeseed oil; The dissolution conditions are: stirring in a water bath at 45-80℃ for 25-45 minutes.

8. The preparation method as described in claim 2, characterized in that, (6) In this process, the mass ratio of the internal aqueous phase to the oil phase is 1:3-5; The mass ratio of W / O emulsion to external aqueous phase is 2:2-5.

9. The preparation method as described in claim 2, characterized in that, In (6), the mixing of the internal aqueous phase and the oil phase requires premixing. The premixing operation is as follows: In a water bath at 40°C, the aqueous phase is added dropwise to the oil phase while stirring, until the two are mixed evenly.

Images