Double-gel system for regulating intestinal flora balance as well as preparation method and application of double-gel system

Abstract

The invention discloses a double-gel system for regulating intestinal flora balance as well as a preparation method and application thereof, and relates to the technical field of gel. The invention provides a double-gel system for regulating intestinal flora balance, the double-gel system comprises hydrogel and oleogel, the ratio of the weight of the oleogel to the total weight of the hydrogel and the oleogel is X, and X is greater than or equal to 30%; the hydrogel comprises the following components in percentage by weight: 5-20% of black bean protein, 0.1-5% of bifidobacterium longum and the balance of deionized water, the oil gel comprises the following components in percentage by weight: 5-10% of rice bran wax and the balance of rice bran oil. According to the invention, the intestinal flora balance is regulated through a specific double-gel system, and the intestinal health is improved.

Description

Technical Field

[0001] The present invention relates to the technical field of gels, in particular to a double gel system for regulating the balance of intestinal flora, its preparation method and application. Background Art

[0002] With the continuous improvement of people's attention to health, the importance of the balance of intestinal flora to human health has become increasingly prominent. As a beneficial intestinal bacterium, Bifidobacterium longum can regulate the balance of intestinal flora, enhance immunity, promote nutrient absorption, etc. However, Bifidobacterium longum belongs to strict anaerobes and faces many challenges in survival and colonization in the intestine, such as the destruction by digestive juices such as gastric acid and bile, and the competition in the complex intestinal environment. Therefore, how to effectively deliver Bifidobacterium longum to the intestine and make it play its role is one of the current research hotspots.

[0003] At present, a variety of delivery systems have been used for the delivery of Bifidobacterium longum, such as microcapsules, hydrogels, oleogels, etc. Among them, hydrogels and oleogels have attracted attention due to their unique properties. Hydrogels have good biocompatibility and adjustable physicochemical properties, and can protect the bacteria from the external environment; oleogels have good lubricity and sustained release performance, and can prolong the residence time of the bacteria in the intestine. Most of the existing delivery systems using single hydrogels and oleogels to deliver Bifidobacterium longum have some disadvantages. For example, the mechanical strength of hydrogels is insufficient and it is easy to break during digestion, resulting in premature release of the bacteria. And the dispersibility of oleogels in the intestine is poor, affecting the colonization effect of the bacteria.

[0004] In view of this, this application is proposed. Summary of the Invention

[0005] Based on this, the purpose of the present invention is to overcome the above-mentioned deficiencies of the prior art and provide a double gel system for regulating the balance of intestinal flora, its preparation method and application.

[0006] To achieve the above purpose, the technical solution adopted by the present invention is: a double gel system for regulating the balance of intestinal flora, including a hydrogel and an oleogel, the weight ratio of the oleogel to the total weight of the hydrogel and the oleogel is X, and X≥30%; the hydrogel includes the following components in weight percentage: 5-20% of black bean protein, 0.1-5% of Bifidobacterium longum, and the balance of deionized water; the oleogel includes the following components in weight percentage: 5-10% of rice bran wax and the balance of rice bran oil.

[0007] The present application provides a double-gel system for regulating the balance of intestinal flora, which has the following advantages: (1) Excellent protection performance: The hydrogel formed by black bean protein can effectively protect Bifidobacterium longum from the destruction of digestive juices such as gastric acid and bile, and improve the survival rate of the bacteria in the intestine; the organogel composed of rice bran wax and rice bran oil provides a relatively stable microenvironment for the bacteria, further enhancing the protection effect. Compared with a single black bean protein hydrogel system, the double-gel system has stronger stability and combines the advantages of both organogel and hydrogel. By adjusting the ratio and structure of the two phases, a gel system conducive to protecting the bacterial strain can be constructed. (2) Good delivery effect: The double-gel has good dispersibility in the intestine and provides double-network protection for the bacteria, enabling the directional release of Bifidobacterium longum and allowing it to better colonize in the intestine, thereby effectively regulating the balance of intestinal flora. (3) Simple preparation process: The preparation method of the present invention does not require complex equipment and cumbersome steps, is easy to industrialize, and reduces production costs. (4) Wide source of raw materials: Black bean protein, rice bran wax, and rice bran oil are all derived from natural plants, with rich resources, low prices, and good biocompatibility and safety, meeting people's requirements for health and environmental protection.

[0008] The inventors found in actual research that the type of the double-gel system varies with the ratio of the organogel to the hydrogel. The double-gels with different organogel contents exhibit three types: O / W, bicontinuous phase, and W / O, respectively. When the weight ratio of the organogel to the total weight of the hydrogel and the organogel increases from 30% to 50%, a phase change occurs from O / W to the bicontinuous phase. At the same time, the aqueous phase changes from a continuous state to a blocky region.

[0009] The inventors found in actual research that Bifidobacterium longum, as the key active ingredient of the present invention, has an insignificant effect on regulating the intestinal flora when its content is less than 0.1%; when the content is higher than 5%, it may cause excessive aggregation of the bacteria, affecting their colonization and diffusion in the intestine.

[0010] The inventors found in actual research that black bean protein is a high-quality plant protein with good biocompatibility and adjustable gel properties. In the present invention, as the main component of the hydrogel, black bean protein can form a stable three-dimensional network structure, effectively encapsulating Bifidobacterium longum and protecting it from the destruction of the external environment. When the content is less than 5%, the formed hydrogel has low strength and is difficult to effectively protect the bacteria; when the content is higher than 20%, the porosity of the hydrogel decreases, which is not conducive to the release and colonization of the bacteria.

[0011] In actual research, the inventors found that rice bran wax is a natural wax extracted from rice bran, which has good film-forming properties and stability. In the oleogel, rice bran wax, as the main structural component, can form a stable crystal network, jointly constituting the framework of the oleogel with rice bran oil, providing a relatively stable microenvironment for Bifidobacterium longum. When the content is less than 5%, the strength and stability of the oleogel are insufficient; when the content is higher than 10%, the flexibility and dispersibility of the oleogel become poor. Rice bran oil is a vegetable oil rich in unsaturated fatty acids, with good lubricity and biocompatibility. In the oleogel, rice bran oil, as the main component of the oil phase, can interact with rice bran wax to form a uniform oleogel system, and at the same time provide essential nutrients for Bifidobacterium longum.

[0012] Preferably, 60% ≤ X ≤ 80%; more preferably, X = 70%.

[0013] In actual research, the inventors found that when the weight ratio of the oleogel to the total weight of the hydrogel and the oleogel is 60%, the double gel begins to exhibit a W / O structure. The W / O system has a better protective effect on Bifidobacterium longum, the survival rate of Bifidobacterium longum in the double gel system is higher, and the release rate in the intestine is moderate, and it can be gradually released after reaching the intestine to maintain its activity.

[0014] Preferably, the weight percentage of Bifidobacterium longum in the hydrogel is 0.5 - 2%.

[0015] Preferably, the weight percentage of black bean protein in the hydrogel is 10 - 15%; more preferably, the weight percentage of black bean protein in the hydrogel is 12%.

[0016] Preferably, the weight percentage of rice bran wax in the oleogel is 6 - 8%.

[0017] In actual research, the inventors found that the weight percentage of Bifidobacterium longum and black bean protein in the hydrogel, and the weight percentage of rice bran wax in the oleogel all affect the performance of the prepared double gel system. When the weight percentage of Bifidobacterium longum in the hydrogel is 0.5 - 2%, and / or the weight percentage of black bean protein in the hydrogel is 10 - 15%, and / or the weight percentage of rice bran wax in the oleogel is 6 - 8%, the prepared double gel system has advantages such as better texture properties, better embedding effect, more stable, and better directional release effect.

[0018] Preferably, the black bean protein, rice bran wax, and rice bran oil can all be commercially available products, and the rice bran wax and rice bran oil can also be obtained by conventional extraction methods from rice bran.

[0019] In addition, the present application provides a method for preparing the double gel system for regulating the balance of intestinal flora, comprising the following steps:

[0020] S1. Dissolve black bean protein in deionized water to obtain a black bean protein solution, and add Bifidobacterium longum to the black bean protein solution, and mix evenly to obtain a hydrogel;

[0021] S2. Heat rice bran wax and rice bran oil until completely melted to prepare an oleogel;

[0022] S3. Mix and homogenize the hydrogel in S1 and the oleogel in S2, and refrigerate to obtain the double gel system for regulating the balance of intestinal flora.

[0023] Preferably, in S1, the weight percentage content of black bean protein is 5-20%, and the weight percentage content of Bifidobacterium longum is 0.1-5%.

[0024] Preferably, in S2, the weight percentage content of rice bran wax is 5-10%.

[0025] Preferably, in S3, the ratio of the weight of the oleogel of the double gel system for regulating the balance of intestinal flora to the total weight of the hydrogel and the oleogel is ≥30%; and / or, the homogenization time is 2-5 min, and the refrigeration temperature is 2-8 °C.

[0026] More preferably, the refrigeration temperature is 4 °C.

[0027] Furthermore, the present invention provides the application of the double gel system for regulating the balance of intestinal flora in the preparation of foods, drugs, and health products.

[0028] Preferably, it further comprises excipients; the excipients are at least one of flavoring agents, thickening agents, disintegrating agents, and fillers.

[0029] Preferably, the dosage forms of the foods, drugs, and health products are one of granules, capsules, tablets, powders, oral liquids, ointments, and beverages.

[0030] Compared with the prior art, the beneficial effects of the present invention are as follows: The present application provides a double gel system for regulating the balance of intestinal flora, which has the following advantages: (1) Excellent protection performance: The hydrogel formed by black bean protein can effectively protect Bifidobacterium longum from being damaged by digestive juices such as gastric acid and bile, and improve the survival rate of the bacteria in the intestine; The oleogel composed of rice bran wax and rice bran oil provides a relatively stable microenvironment for the bacteria, further enhancing the protection effect. Compared with a single black bean protein hydrogel system, the double gel system has stronger stability and combines the advantages of oleogel and hydrogel. By adjusting the ratio and structure of the two phases, a gel system conducive to protecting the bacterial strain is constructed. (2) Good delivery effect: The double gel has good dispersibility in the intestine and has a double-layer network protection for the bacteria, and can release Bifidobacterium longum directionally, enabling it to better colonize in the intestine, thereby effectively regulating the balance of intestinal flora. (3) Simple preparation process: The preparation method of the present invention does not require complex equipment and cumbersome steps, is easy to industrialize, and reduces the production cost. (4) Wide range of raw material sources: Black bean protein, rice bran wax and rice bran oil are all derived from natural plants, with rich resources, low prices, and good biocompatibility and safety, meeting people's requirements for health and environmental protection. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 Are actual pictures of the double gel systems prepared in Examples 1-3; among them, Figure (a) is a picture of the double gel system prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 30%; Figure (b) is a picture of the double gel system prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 50%; Figure (c) is a picture of the double gel system prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 70%;

[0032] Figure 2 Are optical microscope pictures of the double gel systems prepared in Examples 1-3; among them, Figure (a) is a picture of the double gel system prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 30%; Figure (b) is a picture of the double gel system prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 50%; Figure (c) is a picture of the double gel system prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 70%;

[0033] Figure 3Confocal laser scanning microscopy images of the double gel systems prepared in Examples 1-3; among them, Figure (a) is the double gel system diagram prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 30%; Figure (b) is the double gel system diagram prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 50%; Figure (c) is the double gel system diagram prepared when the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is 70%.

[0034] Figure 4 It is a Fourier transform infrared spectrum diagram. Detailed implementation manners

[0035] To better illustrate the purpose, technical solutions and advantages of the present invention, the present invention will be further described below in conjunction with the drawings and specific embodiments. The purpose is to understand the content of the present invention in detail, rather than a limitation to the present invention. All other embodiments obtained by those of ordinary skill in the art without creative efforts fall within the protection scope of the present invention. The experimental reagents and instruments designed in the embodiments and comparative examples of the present invention are all common ordinary reagents and instruments unless otherwise specified, and can be obtained from commercial channels. In the embodiments and comparative examples, the experimental methods used are all conventional methods unless otherwise specified; and unless otherwise specified, the raw materials used in parallel experiments are the same batch of raw materials.

[0036] Now, further description will be made on the raw materials used in the experimental process of this application, but not limited to the following raw materials:

[0037] Bifidobacterium longum - 1: Preservation number CICC 6202, purchased from China Center for Industrial Culture Collection.

[0038] Bifidobacterium longum - 2: Preservation number CICC 6196, purchased from China Center for Industrial Culture Collection.

[0039] Bifidobacterium longum - 3: Preservation number CICC 6199, purchased from China Center for Industrial Culture Collection.

[0040] Black bean protein: Protein content 80%, purchased from Xi'an Xinyiran Biotechnology Co., Ltd.

[0041] Rice bran wax: Purchased from Jitalai (Shenzhen) Industrial Co., Ltd.

[0042] Rice bran oil: Purchased from Anhui Gutianxia Food Co., Ltd.

[0043] Examples and comparative examples

[0044] Example 1

[0045] A preparation method of a double gel system for regulating the balance of intestinal flora, comprising the following steps:

[0046] S1. Dissolve black bean protein in deionized water to obtain a black bean protein solution, and add Bifidobacterium longum - 1 to the black bean protein solution, and mix evenly to obtain a hydrogel; the weight percentage of the black bean protein is 12%, and the weight percentage of the Bifidobacterium longum - 1 is 1%;

[0047] S2. Heat rice bran wax and rice bran oil until completely melted to prepare an oleogel; the weight percentage of the rice bran wax is 7%;

[0048] S3. Mix and homogenize the hydrogel in S1 and the oleogel in S2 for 3 min, and refrigerate at 4 °C to obtain the double gel system for regulating the balance of intestinal flora; the ratio of the weight of the oleogel to the total weight of the hydrogel and the oleogel is X, and the X = 30%.

[0049] Example 2

[0050] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 1, the difference is only that in S3, the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is different, and the X = 50%.

[0051] Example 3

[0052] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 1, the difference is only that in S3, the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is different, and the X = 70%.

[0053] Example 4

[0054] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S1, the weight percentage of black bean protein is 15%.

[0055] Example 5

[0056] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S1, the weight percentage of black bean protein is 5%.

[0057] Example 6

[0058] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S1, the weight percentage of black bean protein is 20%.

[0059] Example 7

[0060] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S1, the weight percentage content of Bifidobacterium longum - 1 is 4%.

[0061] Example 8

[0062] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S2, the weight percentage content of rice bran wax is 8%.

[0063] Example 9

[0064] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S2, the weight percentage content of rice bran wax is 5%.

[0065] Example 10

[0066] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S2, the weight percentage content of rice bran wax is 10%.

[0067] Example 11

[0068] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S1, the type of Bifidobacterium longum is different, which is Bifidobacterium longum - 2.

[0069] Example 12

[0070] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S1, the type of Bifidobacterium longum is different, which is Bifidobacterium longum - 3.

[0071] Comparative Example 1

[0072] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S3, the ratio X of the weight of the oleogel to the total weight of the hydrogel and the oleogel is different, and X = 20%.

[0073] Comparative Example 2

[0074] A preparation method of a double gel system for regulating the balance of intestinal flora, compared with Example 3, the difference is only that in S1, the hydrogel is a soy protein hydrogel.

[0075] Specifically, soy protein is dissolved in deionized water to obtain a soy protein solution, and Bifidobacterium longum - 1 is added to the soy protein solution and mixed evenly to obtain a hydrogel; the weight percentage of the soy protein is 12%, and the weight percentage of the Bifidobacterium longum - 1 is 1%.

[0076] Comparative Example 3

[0077] A preparation method of a double - gel system for regulating the balance of intestinal flora. Compared with Example 3, the only difference is that in S2, rice bran oil is replaced by soybean oil.

[0078] Comparative Example 4

[0079] A preparation method of a double - gel system for regulating the balance of intestinal flora. Compared with Example 3, the only difference is that in S2, rice bran oil is replaced by sesame oil.

[0080] Comparative Example 5

[0081] A preparation method of a double - gel system for regulating the balance of intestinal flora. Compared with Example 3, the only difference is that in S2, rice bran wax is replaced by candelilla wax.

[0082] Comparative Example 6

[0083] A preparation method of a double - gel system for regulating the balance of intestinal flora. Compared with Example 3, the only difference is that in S2, rice bran wax is replaced by carnauba wax.

[0084] Comparative Example 7

[0085] A preparation method of a double - gel system for regulating the balance of intestinal flora. Compared with Example 3, it does not contain an oil gel. Specifically, it includes the following steps: Black bean protein is dissolved in deionized water to obtain a black bean protein solution, and Bifidobacterium longum - 1 is added to the black bean protein solution and mixed evenly to obtain a hydrogel; the weight percentage of the black bean protein is 12%, and the weight percentage of the Bifidobacterium longum - 1 is 1%.

[0086] Comparative Example 8

[0087] A preparation method of a double - gel system for regulating the balance of intestinal flora. Compared with Example 3, it does not contain a hydrogel. Specifically, it includes the following steps:

[0088] It includes the following steps: Rice bran wax and rice bran oil are heated until completely melted. When the oil phase cools to 40°C, Bifidobacterium longum - 1 is added to the gel and stirred at a speed of 500 rpm for 10 minutes to ensure that the bacterial cells are evenly distributed throughout the oil phase to prepare an oil gel; the weight percentage of the rice bran wax is 7%, and the weight percentage of the Bifidobacterium longum - 1 is 1%.

[0089] Performance Test-1 Macroscopic and microstructural observation of the double gel loaded with Bifidobacterium longum.

[0090] The double gel systems prepared in Examples 1-3 were observed.

[0091] (1) Macroscopic observation: observation by photography.

[0092] (2) Microstructure observation: Optical microscopy The microstructure of the bigels was analyzed using an optical microscope (XPH-25C, Puda, China Ltd.) equipped with a digital camera, with a focus on the bigel type. This process involves placing a coverslip on a small drop of hot-melt bigel sample on a microscope slide. After the sample was stored at 4 °C for 24 h, it was observed under a microscope and imaged at room temperature (25 °C) at a magnification of 40×. The distribution of the aqueous and nonpolar phases was observed using a confocal laser scanning microscope (LSM 710, Carl Zeiss Jena, Germany). Prior to analysis, the oil gel was stained with a Nile red solution (0.1 mg / mL ethanol) using an argon laser with an excitation wavelength of 520 nm. The hydrogel was stained with a FITC solution (0.1 mg / mL ethanol) using an argon laser with an excitation wavelength of 470 nm. A 10 μL drop was placed on a slide and covered with a coverslip. All fluorescent images were captured at a magnification of 20 × 10.

[0093] The results are as follows Figures 1-3 shown.

[0094] Figure 1 This is a real shot of the double gel system prepared in Example 1-3. Figure 2 This is an optical microscope image of the double gel system prepared in Example 1-3. Figure 3 This is a confocal laser scanning microscopy image of the double gel system prepared in Example 1-3. The inversion test shows that ( Figure 1 ), all gels have a self-supporting structure, which can be attributed to the interwoven structure of hydrogel and oleogel. Black soybean protein is connected by hydrogen bonds and forms a three-dimensional network structure to form hydrogel, and rice bran wax crystallizes in rice bran oil and blocks the fluidity of lipophilic liquid.

[0095] The type of dual gel varies with the ratio of oil gel to hydrogel ( Figure 2 , Figure 3 ). The bigels with different oil gel contents showed three types: O / W, bicontinuous phase and W / O. When the oil gel ratio increased from 30% to 50%, the phase transition occurred from O / W to bicontinuous phase. At the same time, the water phase changed from a continuous state to a block region. When the oil gel fraction was 70%, the bigel showed a W / O structure.

[0096] Performance Test-2: Test on the initial encapsulation rate of Bifidobacterium longum by the double gel system.

[0097] Weigh 1 g of the double gel sample, add it to 9 mL of the capsule-dissolving solution with a pH of 7.4, and oscillate for 10 minutes until complete capsule dissolution. Gradient dilute the mixed solution after capsule dissolution, and then coat it on MRS agar and anaerobically culture it at 37 °C for 72 hours. Perform viable cell counting on the cultured plate to obtain the total number of viable cells (N) embedded in the double gel.

[0098] Calculate the encapsulation rate: The encapsulation rate is calculated according to the following formula:

[0099]

[0100] Among them, C 1 is the number of viable cells (CFU / g) in the dilution solution, and C 0 is the number of viable cells (CFU / g) in the capsule-dissolving solution.

[0101] The test results of the encapsulation rate are shown in Table 1.

[0102] Table 1

[0103]

[0104]

[0105] As can be seen from the above table, Examples 1 to 3 show that with the increase in the proportion of the oil gel, the encapsulation rate gradually increases. This indicates that the proportion of the oil gel has a certain positive effect on improving the encapsulation rate. The encapsulation rate of Comparative Example 1 decreased significantly, further verifying the positive effect of the oil gel proportion on the encapsulation rate.

[0106] The weight percentage content of black bean protein in Examples 3 - 6 is different. When the weight percentage content of black bean protein is 12 - 15%, the encapsulation rate is higher. The weight percentage content of Bifidobacterium longum in Examples 3 and 7 is different, and the encapsulation rates are not much different. The weight percentage content of rice bran wax in Examples 3 and 8 - 10 is different. When the weight percentage content of rice bran wax is 6 - 8%, the encapsulation rate is higher.

[0107] The hydrogel and oleogel components also have a great impact on the encapsulation rate. In Comparative Example 2, soy protein was used instead of black bean protein in the hydrogel, and the encapsulation rate decreased significantly. This indicates that black bean protein is superior to soy protein in encapsulating Bifidobacterium longum. In Comparative Examples 3 and 4, rice bran oil was replaced with soybean oil and sesame oil respectively, and the encapsulation rate decreased significantly. This indicates that rice bran oil is superior to soybean oil and sesame oil in encapsulating Bifidobacterium longum. In addition, in Comparative Examples 5 and 6, rice bran wax was replaced with candelilla wax and carnauba wax respectively, and the encapsulation rate decreased significantly. This indicates that rice bran wax is superior to candelilla wax and carnauba wax in encapsulating Bifidobacterium longum. In addition, the encapsulation rates of Comparative Example 7 without oleogel and Comparative Example 8 without hydrogel were significantly lower than those of the double-gel system. This indicates that the double-gel system is superior to the single-gel system in encapsulating Bifidobacterium longum.

[0108] Performance test - 3 Bioaccessibility of Bifidobacterium longum during in vitro digestion.

[0109] In vitro digestion simulation experiment of the biphasic gel to simulate and explore the directional release of Bifidobacterium longum.

[0110] Oral digestion (SSF) components include: phosphate buffer solution, 12.0 mg / mL KCl, 1.6 mg / mL NaCl, 28.0 mg / mL NaCO 3 、0.3 mg / mL uric acid, 0.4 mg / mL mucin, 3.0 mg / mL KSCN, 15.0 mg / mL NaH 2 PO 4 、3.0 mg / mL urea and 4.8 mg / mL α-amylase, and adjust the pH to 6.8;

[0111] Gastric digestion (SGF) components include: 3.2 mg / mL pepsin and 2.0 mg / mL NaCl, and adjust the pH to 2.0 with HCl;

[0112] The components of small intestine digestion (SIF) include: 0.4 mg / mL lipase, 0.5 mg / mL pancreatin, 0.7 mg / mL porcine bile salt and 750.0 mmol / L CaCl 2 ,and adjust the pH to 7.0.

[0113] The temperature is controlled at 37 ± 0.5 °C throughout the simulated digestion process.

[0114] The test results are shown in Table 2.

[0115] Table 2

[0116]

[0117]

[0118] As can be seen from the above table, in Examples 1-3, as the proportion of oleogel (X) gradually increases, the release rates in the oral cavity and stomach gradually decrease, while the release rate in the small intestine gradually increases. This indicates that the higher the proportion of oleogel, the better the stability of the gel system in the oral cavity and stomach, and the better the release effect in the small intestine. The stomach release rate of Comparative Example 1 is significantly higher than that of Example 3, and the small intestine release rate is significantly lower than that of Example 3, further verifying the importance of the oleogel proportion. In Examples 3-6, the weight percentage of black bean protein is different. When the weight percentage of black bean protein is 12-15%, the release rates in the oral cavity and stomach are lower, while the release rate in the small intestine is higher. This indicates that the higher the concentration of black bean protein, the better the stability of the gel system in the oral cavity and stomach, and the better the release effect in the small intestine. In Examples 3, 8-10, the weight percentage of rice bran wax is different. When the weight percentage of rice bran wax is 6-8%, the release rates in the oral cavity and stomach are lower, while the release rate in the small intestine is higher.

[0119] In Comparative Example 2, when soy protein hydrogel is used instead of black bean protein hydrogel, the stomach release rate is significantly higher than that of Example 3, and the small intestine release rate is significantly lower than that of Example 3. This indicates that the protective effect and release effect of black bean protein hydrogel are better than those of soy protein hydrogel. In Comparative Examples 3-4, when soybean oil or sesame oil is used instead of rice bran oil, the stomach release rate increases significantly, and the small intestine release rate decreases significantly. This indicates that the protective effect and release effect of rice bran oil are better than those of soybean oil and sesame oil. In Comparative Examples 5-6, when candelilla wax or carnauba wax is used instead of rice bran wax, the stomach release rate increases significantly, and the small intestine release rate decreases significantly. This indicates that the protective effect and release effect of rice bran wax are better than those of candelilla wax and carnauba wax. In Comparative Examples 7-8, without oleogel and hydrogel, the stomach release rate increases significantly, and the small intestine release rate decreases significantly. This indicates that the protective effect and release effect of the double gel system are better than those of the single gel system.

[0120] Performance test - 4 Texture analysis of the double gel loaded with Bifidobacterium longum.

[0121] The measurement was carried out using a texture analyzer equipped with a 50 kg load cell, and a p-0.5 probe was used for measurement. The sample was placed in a 100 mL cylindrical glass beaker for measurement, and the height of the sample in the beaker was 2.0 cm. At a pre-test speed of 2.0 mm / s, the sample was then compressed at a test speed and a post-test speed of 2.0 mm / s, and the trigger force was 5.0 g. The probe was immersed 1.2 cm (60%) into the sample and then returned to the starting position. Each sample was measured three times repetitively.

[0122] The test results are shown in Table 3.

[0123] Table 3

[0124]

[0125]

[0126] As can be seen from the above table, in Examples 1-3, as the proportion of oleogel increased, both the hardness and adhesiveness increased significantly, indicating that increasing the oleogel proportion can enhance the mechanical strength and cohesiveness of the double gel. The hardness and adhesiveness of Comparative Example 1 were much lower than those of Example 3, indicating that when the oleogel proportion was less than 30%, the structure was loose and could not effectively support the double gel network. The weight percentage content of black bean protein in Examples 3-6 was different. When the weight percentage content of black bean protein was 12-15%, the hardness and adhesiveness were higher and the structural stability was better.

[0127] The weight percentage content of Bifidobacterium longum in Example 3 and Example 7 was different, and the difference in hardness and adhesiveness was not significant. The weight percentage content of rice bran wax in Example 3 and Examples 8-10 was different. When the weight percentage content of rice bran wax was 6-8%, the hardness and adhesiveness were higher, and too high or too low values both weakened the structural integrity.

[0128] In addition, in Comparative Example 2, soy protein was used to replace black bean protein, and the hardness was only 41% of that in Example 3, proving that the unique amino acid composition of black bean protein was more conducive to forming a stable gel network. In Comparative Examples 3-4, rice bran oil was replaced with soybean oil / sesame oil, and the hardness decreased by more than 50%, indicating that the highly unsaturated fatty acids in rice bran oil and rice bran wax synergistically enhanced cross-linking. In Comparative Examples 5-6, rice bran wax was replaced with candelilla wax / carnauba wax, and the hardness decreased significantly, indicating that the crystal form of rice bran wax was more suitable for the oleogel structure. The hardness of only the hydrogel or only the oleogel was less than 25% of that in Example 3, confirming that the synergistic effect of the double gel was the key to improving the mechanical properties.

[0129] Performance test - 5 Analysis of the intermolecular forces of the double gel loaded with Bifidobacterium longum.

[0130] The Nicolet 67 Fourier transform infrared spectrometer equipped with a Smart iTX ATR sampling accessory was used to analyze the possible structural interactions in the gel systems prepared in Examples 1-3 and Comparative Examples 7-8. The instrument was operated in the attenuated total reflection mode (ATR), and the spectral range was collected within the wavenumber range of 4000-650 cm -1 and scanned 32 times.

[0131] The test results are as Figure 4 shown.

[0132] As Figure 4 can be seen, the O / W double gel (Example 1) showed a lower transmittance in the range of 1000 cm -1 to 1500 cm -1 which may be related to the C-H bending vibration, while at 3000 cm -1The absorption peaks nearby may be related to the stretching vibrations of O-H or N-H. The overall transmittance of the bicontinuous double gel is relatively high, especially in the range of 2800 - 3000 cm -1 The medium-intensity absorption peaks within this range may be related to the stretching vibration of C-H. The W / O double gel (Example 3) shows multiple absorption peaks between 1500 cm -1 and 2000 cm -1 , which may be related to the stretching vibrations of C=C or C=O. The high transmittance above 3000 cm -1 indicates less absorption at these wavelengths.

[0133] The spectrum of reference H (Comparative Example 7) shows that its overall transmittance is relatively low, especially between 1500 cm -1 and 3000 cm -1 , showing multiple absorption peaks, which may indicate that H contains various functional groups such as C=C, C=O, O-H, etc. The spectrum of reference O (Comparative Example 8) shows that the transmittance changes relatively greatly, especially in the high wavenumber region (about 3000 cm -1 to 3600 cm -1 ), where there may be absorption peaks of C-H stretching vibration. The transmittance may decrease significantly in these regions, indicating that the vibrations corresponding to these regions are absorbed by the sample molecules. Near 1700 cm -1 , the emerging absorption peaks are related to the stretching vibration of C=O. The appearance of such peaks indicates that the sample may contain carbonyl-containing functional groups such as ketones, aldehydes, or acids. In addition, there are relatively strong absorption peaks near 1000 cm -1 related to the bending vibration of C-H or the bending vibration of C-O. There are relatively large absorption peaks between 600 cm -1 and 800 cm -1 , which may be related to the vibration of the molecular skeleton (such as C-C stretching vibration).

[0134] Through comparative analysis, we found that the differences in absorption peaks of different double gel types at specific wavelengths reflect their differences in chemical structures. However, no new absorption peaks appeared and no absorption peaks disappeared in all double gels. In addition, combining the common properties of oil gels and water gels and enhancing the bond energy may also be the reason why double gels are more stable than single-phase gels.

[0135] Performance test - 6: Oil-binding capacity (OBC) and water-holding capacity (WHC) of the double gel loaded with Bifidobacterium longum.

[0136] The stability of the double gel was determined by measuring the oil-binding capacity (OBC) and water-holding capacity (WHC) under centrifugal force, which helped to evaluate the retention capacity of oil and water in the double gel sample. The centrifugation operation was carried out at 25 °C at a speed of 9200 revolutions per minute for 15 minutes. Then, the oil released at the top of the sample was collected using a syringe, and the weight of the resulting test tube (Wc) was then measured. Finally, the water released at the bottom of the test tube was removed, and the final weight of the test tube (Wd) was measured. The oil-binding capacity (OBC) and water-holding capacity (WHC) were calculated using formulas (1) and (2), respectively.

[0137]

[0138] Where Wa is the weight of the centrifuge tube (g), Wb is the weight of the test tube containing the sample before centrifugation (g), OP represents the density of vegetable oil, and WP represents the density of distilled water.

[0139] The test results are shown in Table 4.

[0140] Table 4

[0141]

[0142]

[0143]

[0144]

[0145] As can be seen from the above table, in Examples 1-3, as the proportion of oleogel increased, the long-term stability of OBC and WHC was significantly improved. For example, in Example 3, OBC and WHC remained the highest after 30 days, indicating that a high oleogel proportion effectively locked the oil phase and water phase through a hydrophobic network, reducing phase separation. In contrast, the OBC and WHC of Comparative Example 1 after 30 days were much lower than those of Example 3, indicating that when the oleogel proportion was less than 30%, the double gel structure was loose and could not resist the oil-water loss caused by centrifugal force.

[0146] The weight percentage of black bean protein in Examples 3-6 was different. When the weight percentage of black bean protein was 12-15%, the long-term stability of OBC and WHC was higher. The weight percentage of Bifidobacterium longum in Examples 3 and 7 was different, and the long-term stability of OBC and WHC differed little. The weight percentage of rice bran wax in Examples 3 and 8-10 was different. When the weight percentage of rice bran wax was 6-8%, the long-term stability of OBC and WHC was higher.

[0147] In Comparative Example 2, soy protein was used to replace black bean protein, and the OBC and WHC after 30 days were only 53% and 35% of those in Example 3, respectively, proving that the hydrophilic-hydrophobic balance of black bean protein is more conducive to the stability of the double gel network. In Comparative Examples 3-4, rice bran oil was replaced with soybean oil and sesame oil, and the OBC and WHC after 30 days were close to those in Comparative Example 1, indicating that the high γ-oryzanol content in rice bran oil is crucial for the stability of the oleogel. In Comparative Examples 5-6, rice bran wax was replaced with candelilla wax and carnauba wax, and the performance was much worse than that in Example 3, indicating that the specific fatty acid composition of rice bran wax (such as high octacosanol) is irreplaceable for the formation of the gel network. The WHC and OBC of only the hydrogel or only the oleogel after 30 days were less than 35% of those in Example 3, confirming that the synergistic effect of the double gel is the core of stability. In addition, the high oil / water holding rate indicates that the double gel can effectively protect Bifidobacterium longum from the digestive environment and delay intestinal release, thereby improving the colonization efficiency of probiotics.

[0148] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit the protection scope of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those of ordinary skill in the art should understand that the technical solutions of the present invention can be modified or equivalently replaced without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A double gel system for regulating the balance of intestinal flora, characterized in that: Comprising a hydrogel and an oleogel, wherein the weight ratio of the oleogel to the total weight of the hydrogel and the oleogel is X, and X is ≥ 30%; The hydrogel comprises the following components in percentage by weight: 5-20% black bean protein, 0.1-5% Bifidobacterium longum, and the balance deionized water; The oil gel comprises the following components in percentage by weight: 5-10% of rice bran wax and the remainder of rice bran oil.

2. The double gel system according to claim 1, characterized in that The 60%≤X≤80%.

3. The double gel system according to claim 1, characterized in that: The weight percentage of the black bean protein in the hydrogel is 10-15%.

4. The double gel system according to claim 1, characterized in that The weight percentage of rice bran wax in the oil gel is 6-8%.

5. A method for preparing a double gel system for regulating the balance of intestinal flora according to any one of claims 1 to 4, characterized in that: The steps include: S1, dissolving black bean protein in deionized water to obtain a black bean protein solution, adding Bifidobacterium longum to the black bean protein solution, and mixing well to obtain a hydrogel; S2, heating rice bran wax and rice bran oil until they are completely melted to prepare an oil gel; S3, the hydrogel in S1 and the oil gel in S2 are mixed and homogenized, and refrigerated to obtain the dual gel system for regulating the balance of intestinal flora.

6. The method for preparing the double gel system for regulating the balance of intestinal flora according to claim 5, characterized in that: In the S1, the weight percentage of black bean protein is 5-20%, and the weight percentage of Bifidobacterium longum is 0.1-5%.

7. The method for preparing the double gel system for regulating the balance of intestinal flora according to claim 5, characterized in that: In the S2, the weight percentage of rice bran wax is 5-10%.

8. The method for preparing the double gel system for regulating the balance of intestinal flora according to claim 5, characterized in that: In said S3, in said dual gel system for regulating the balance of intestinal flora, the weight ratio of the oil gel to the total weight of the hydrogel and the oil gel is ≥30%; And / or, the homogenization time is 2-5 min, and the refrigeration temperature is 2-8°C.

9. Use of the double gel system for regulating the balance of intestinal flora as claimed in any one of claims 1 to 4 in the preparation of food, medicine and health care products.

10. The use according to claim 9, characterized in that The invention also includes auxiliary materials, which are at least one of flavoring agents, thickeners, disintegrants and fillers.

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