Microecological preparation for inhibiting alpha-glucosidase as well as preparation method and application of microecological preparation

Abstract

The invention belongs to the technical field of natural product extraction and preparation and biological medicine, and particularly relates to a microecological preparation for inhibiting alpha-glucosidase as well as a preparation method and application of the microecological preparation. Specifically, the microecological preparation capable of effectively inhibiting alpha-glucosidase is successfully prepared from three raw materials including Chinese yam, cape jasmine and coix seeds through the processes of water extraction, alcohol precipitation, microbial fermentation and the like, the microecological preparation gathers active ingredients of the three traditional Chinese medicines, and the alpha-glucosidase can be effectively inhibited under the fermentation of lactobacillus rhamnosus. The Chinese yam, cape jasmine and coix seed water extract has a remarkable inhibition effect on alpha-glucosidase in vitro, has a remarkable effect of degrading macromolecular polysaccharide in the Chinese yam, cape jasmine and coix seed water extract into micromolecular sugar after fermentation, is more beneficial to absorption and utilization of a human body, and can be applied to prevention and treatment of related diseases such as diabetes, so that the Chinese yam, cape jasmine and coix seed water extract has a very good practical application value.

Description

Technical Field

[0001] The invention belongs to the field of natural product extraction and preparation and biomedicine technology, and relates to a microecological preparation for inhibiting alpha-glucosidase, and a preparation method and application thereof. Background Art

[0002] The information disclosed in this background technology section is only intended to enhance the understanding of the overall background of the invention, and should not necessarily be regarded as an admission or any form of suggestion that the information constitutes the prior art already known to a person skilled in the art.

[0003] Diabetes has become a common and frequently-occurring disease in many countries. The incidence of diabetes and its complications is on the rise worldwide, especially in developing countries. Its mortality rate is second only to cardiovascular and cerebrovascular diseases and cancer, and is considered the third largest killer of humans. Therefore, active prevention and treatment of diabetes is imminent.

[0004] α-Glucosidase (AG) is a hydrolase located on the surface of intestinal epithelial cells, which can hydrolyze the α-1,4-glycosidic bond of α-glucose to produce glucose. When the human body eats, the enzyme converts carbohydrates into glucose, which is absorbed into the blood through the intestinal wall, resulting in increased blood sugar after meals. However, enzyme inhibitors competitively inhibit the activity of α-glucosidase, inhibit / delay the hydrolysis of disaccharides and oligosaccharides into monosaccharides, reduce or inhibit the body's absorption of glucose, and thus effectively lower postprandial blood sugar. Studies have found that the pharmacological activity of α-glucosidase inhibitors is mainly reflected in regulating sugar metabolism. It is particularly noteworthy that it plays an important role in lowering blood sugar and preventing and treating diabetic complications. However, there are still few studies and reports on the preparation of α-glucosidase inhibitors using natural products. Summary of the invention

[0005] In view of the deficiencies of the above-mentioned prior art, the present invention provides a microecological preparation for inhibiting α-glucosidase and its preparation method and application. Specifically, the present invention uses three raw materials, namely, yam, gardenia, and coix seed, and successfully prepares a microecological preparation that effectively inhibits α-glucosidase through processes such as water extraction, alcohol precipitation, and microbial fermentation, so that it can be used in the prevention and treatment of related diseases such as diabetes. Based on the above research results, the present invention is completed.

[0006] In order to achieve the above technical objectives, the technical solution adopted by the present invention is as follows:

[0007] The first aspect of the present invention provides a probiotic preparation for inhibiting α-glucosidase, wherein the probiotic preparation for inhibiting α-glucosidase is prepared from yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide by fermentation with Lactobacillus rhamnosus; wherein the yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide are prepared from yam, gardenia and coix seed by water extraction and alcohol precipitation method respectively.

[0008] The second aspect of the present invention provides a method for preparing the above-mentioned microecological preparation, the preparation method comprising:

[0009] S1. Preparing yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide respectively by water extraction and alcohol precipitation method;

[0010] S2, mixing the yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide obtained in step S1, and fermenting them with Lactobacillus rhamnosus.

[0011] The present invention verifies through experiments that the microecological preparation prepared by the above preparation method has significant efficacy in inhibiting α-glucosidase.

[0012] Therefore, the third aspect of the present invention provides the use of the above-mentioned microecological preparation in any one or more of the following:

[0013] (a1) inhibiting α-glucosidase or preparing products that inhibit α-glucosidase;

[0014] (a2) Preparation of products for the prevention and / or treatment of diabetes and related diseases.

[0015] Beneficial technical effects of one or more of the above technical solutions:

[0016] (1) The Chinese medicine composition provided by the above technical solution has a good inhibitory effect on α-glucosidase in in vitro experiments and has a synergistic effect. At the same time, the above technical solution screens the proportions of different compositions to screen out compositions that produce better synergistic effects, which expands new ideas and methods for the research on innovation and development of Chinese medicines;

[0017] (2) The microecological preparation provided by the above technical solution effectively aggregates the active ingredients of the three traditional Chinese medicines. After fermentation by Lactobacillus rhamnosus, it has an extremely significant ability to degrade the macromolecular polysaccharides in the three traditional Chinese medicines into small molecular polysaccharides, which is beneficial to the absorption of polysaccharides in the body. Among them, Chinese yam, gardenia and coix seed are foods that are both medicinal and edible, which effectively improves the safety of the composition and has good practical application value;

[0018] (3) Among the above-mentioned microecological preparations, Chinese yam contains chemical components such as fatty acids, polysaccharides, proteins, amino acids, and trace elements, and has multiple biological activities. Gardenia has the effects of purging fire and relieving restlessness, clearing heat and promoting diuresis, cooling blood and detoxifying. The Compendium of Materia Medica once recorded that Gardenia has the effects of relieving five kinds of jaundice, relieving five kinds of stranguria, promoting urination, relieving thirst, and improving eyesight. Gardenia jasminoides, a terpene compound in Gardenia, is its main active ingredient, which has the effect of reducing hyperglycemia caused by adrenaline. Coix seeds are rich in polysaccharides, terpenes, flavonoids, sterols, alkaloids, and lactams, and have pharmacological effects such as anti-oxidation, liver protection, and immunity enhancement.

[0019] (4) The microecological preparation provided by the above technical solution uses Chinese yam water extract, gardenia water extract, and coix seed water extract, which gathers the active ingredients of three traditional Chinese medicines. Under the fermentation of rhamnosus Lactobacillus, experiments have shown that it not only has a significant inhibitory effect on α-glucosidase in vitro, but also has a significant effect on degrading the large molecular polysaccharides in the Chinese yam, gardenia and coix seed water extracts into small molecular sugars after fermentation, and has good practical application value. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The accompanying drawings in the specification, which constitute a part of the present invention, are used to provide a further understanding of the present invention. The exemplary embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute improper limitations on the present invention.

[0021] Figure 1 The figure shows the changes in the content of oligosaccharides in different samples before and after fermentation. DETAILED DESCRIPTION

[0022] It should be noted that the following detailed descriptions are all illustrative and intended to provide further explanation of the present invention. Unless otherwise specified, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art to which the present invention belongs.

[0023] It should be noted that the terms used herein are only for describing specific embodiments and are not intended to limit exemplary embodiments according to the present invention. As used herein, unless the context clearly indicates otherwise, the singular form is also intended to include the plural form. In addition, it should be understood that when the terms "comprising" and / or "including" are used in this specification, they indicate the presence of features, steps, operations, devices, components and / or combinations thereof. It should be understood that the scope of protection of the present invention is not limited to the specific embodiments described below; it should also be understood that the terms used in the embodiments of the present invention are intended to describe specific embodiments, rather than to limit the scope of protection of the present invention.

[0024] As mentioned above, there are still few studies and reports on the preparation of α-glucosidase inhibitors using natural products.

[0025] In view of this, in a typical specific embodiment of the present invention, a probiotic preparation for inhibiting α-glucosidase is provided, wherein the probiotic preparation for inhibiting α-glucosidase is prepared from yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide by fermentation with Lactobacillus rhamnosus; wherein the yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide are prepared from yam, gardenia and coix seed by water extraction and alcohol precipitation method, respectively.

[0026] Wherein, the mass ratio of yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide is 5-10:1-5:1-5, preferably 2:1:1.

[0027] The Lactobacillus rhamnosus may specifically be Lactobacillus rhamnosus CGMCC No.1.12734, which can be purchased from the China General Microbiological Culture Collection Center in a commercial manner.

[0028] In another specific embodiment of the present invention, a method for preparing the above-mentioned microecological preparation is provided, and the preparation method comprises:

[0029] S1. Preparing yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide respectively by water extraction and alcohol precipitation method;

[0030] S2, mixing the yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide obtained in step S1, and fermenting them with Lactobacillus rhamnosus.

[0031] The specific method of step S1 includes:

[0032] S1-1, respectively grind and sieve yam, gardenia, and coix seed, add 3-4 times of their weight of water, after ultrasonic treatment, centrifuge to remove precipitate, then add 3-4 times of water and ultrasonicate, and repeat this process 2-4 times; after removing the precipitate, concentrate the aqueous solution by rotary evaporation to obtain yam, gardenia, and coix seed water extracts, respectively;

[0033] S1-2, respectively add ethanol solution to the water extract of yam, gardenia and coix seed in step S1-1, heat and stir, centrifuge to remove the supernatant, and repeat this process 2-4 times; dry the obtained precipitate to obtain yam, gardenia and coix seed polysaccharides.

[0034] Wherein, in the step S1-1, the sieving mesh number is 150-250 mesh, preferably 200 mesh; the ultrasonic treatment conditions are: ultrasonic frequency is 20-80 Hz, and ultrasonic treatment time is 10-60 min; preferably, ultrasonic frequency is 50 Hz, and ultrasonic treatment time is 30 min; centrifugal conditions are: centrifugation at 7000-9000 rpm for 3-6 min.

[0035] In step S1-2, the ethanol solution is a high-concentration ethanol solution, specifically a 96% ethanol solution, and the stirring time is 5-30 minutes, preferably 15 minutes.

[0036] In the step S2, as mentioned above, the Lactobacillus rhamnosus may be specifically Lactobacillus rhamnosus CGMCC No. 1.12734, which can be purchased from the China General Microbiological Culture Collection Administration Center in a commercial manner.

[0037] The specific conditions during the fermentation process are: the fermentation pH is controlled to be neutral, and the initial concentration of Lactobacillus rhamnosus is controlled to be 0.1-10×10 7 cfu / ml (preferably 1×10 7 cfu / ml), the fermentation temperature is controlled at 25-35°C (preferably 30°C), and the fermentation time is 40-150h (preferably 96h).

[0038] Furthermore, in order to ensure more complete fermentation, nitrogen sources such as beef extract, yeast extract, sodium acetate, diammonium hydrogen citrate, dipotassium hydrogen phosphate, and inorganic nutrient factors may be added during the fermentation process. The specific amounts of the above components added may be the same as those used in conventional bacterial fermentation culture, and are not specifically limited here.

[0039] The present invention verifies through experiments that the microecological preparation prepared by the above preparation method has significant efficacy in inhibiting α-glucosidase.

[0040] Therefore, in another specific embodiment of the present invention, the use of the above-mentioned microecological preparation in any one or more of the following is provided:

[0041] (a1) inhibiting α-glucosidase or preparing products that inhibit α-glucosidase;

[0042] (a2) Preparation of products for the prevention and / or treatment of diabetes and related diseases.

[0043] Among them, in (a1), the product can be food, medicine or experimental reagent for non-medical use, and the experimental reagent can be used for basic research.

[0044] In (a2), the product may be a medicine; the diabetes-related diseases include but are not limited to diabetic-mediated retinopathy, diabetic nephropathy, polyneuritis, autonomic dysfunction, diabetic foot, etc.

[0045] It should be noted that, in the present invention, the term "food" used should be understood in a broad sense, and it can be understood as any form that can be eaten. For example, the food in the present invention includes ordinary food and special food. The special food in the present invention includes health food and special medical formula food; and ordinary food is relative to special food, which is food suitable for everyone.

[0046] The food includes but is not limited to solid food and liquid food; the solid food includes but is not limited to baked food (such as biscuits, bread, cakes), candy, solid beverages, etc.; the liquid food includes but is not limited to liquid beverages (such as carbonated beverages, fruit juice beverages, milk beverages), etc.

[0047] The drug can be administered in a unit dosage form, and the dosage form can be a liquid dosage form or a solid dosage form. The liquid dosage form can be a true solution, a colloid, a microparticle dosage form, an emulsion dosage form, or a suspension dosage form. Other dosage forms include tablets, capsules, dripping pills, aerosols, pills, powders, solutions, emulsions, granules, suppositories, freeze-dried powder injections, inclusion compounds, landfills, patches, and liniments.

[0048] The present invention is further explained by the following examples, but it does not constitute a limitation of the present invention. It should be understood that these examples are only used to illustrate the present invention and are not used to limit the scope of the present invention. Example 1 Effect of microecological preparations on inhibiting α-glucosidase at in vitro enzyme levels

[0049] 1. Materials and Instruments

[0050] 1.1 Materials

[0051] Chinese yam, gardenia, and coix seed were from the High-tech Industry (Pilot) Base of Shandong Academy of Sciences; Lactobacillus rhamnosus CGMCC No.1.12734 (commercially available from the China General Microbiological Culture Collection Center), α-glucosidase (Beijing Coolbo Technology Co., Ltd.), PNPG (S818110, Shanghai MacLean Biochemical Technology Co., Ltd.), acarbose (A10966, Tianjin Puxitang Biotechnology Co., Ltd.), DNS reagent (GB 20287, Shanghai Yuanye Biotechnology Co., Ltd.), and phosphate buffer (SL6100SX, Beijing Coolbo Technology Co., Ltd.).

[0052] 1.2 Instruments

[0053] Microplate reader (model: 1SPA-0093), Dynex, USA.

[0054] 1.3 Solution preparation

[0055] (1) 1 U / mL α-glucosidase: Prepare 1 U / mL stock solution in 0.1 mol / L phosphate buffer, pH 6.8, and store at -20°C.

[0056] (2) 5 mmol / L substrate (PNPG): Weigh 0.3765 g of PNPG, completely dissolve it in phosphate buffer and make up to a 50 mL volumetric flask.

[0057] (3) 0.1 mol / L Na2CO3 reaction termination solution: Weigh 0.53 g of Na2CO3 powder, dissolve it in distilled water and make up to 50 ml. Store at 4°C for later use.

[0058] 1.4 Sample preparation

[0059] After the three medicinal materials of Chinese yam, gardenia and coix seed were crushed, they were passed through a 200-mesh sieve, and three times of their weight of pure water were added in batches, ultrasonicated at 50Hz for 30min, centrifuged at 8000rpm for 5min to remove the precipitate, and then three times of pure water was added and ultrasonicated again. This process was repeated three times. The aqueous solution after the precipitate was removed was rotary evaporated until it became a viscous semi-fluid, and the water extracts of Chinese yam, gardenia and coix seed were obtained respectively.

[0060] The water extracts of Chinese yam, gardenia and coix seed were added to 5 times of 96% ethanol solution, stirred at 45℃ for 15min, centrifuged at 5000rpm for 5min to remove the supernatant, and the process was repeated 3 times. The precipitate was dried at 25℃ for 24 hours to obtain Chinese yam, gardenia and coix seed polysaccharides.

[0061] Prepare a culture medium, specifically: 2g beef extract, 2g yeast extract, 2g sodium acetate, 1g diammonium hydrogen citrate, 2g dipotassium hydrogen phosphate, and add the above-prepared yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide, add water to 1L; adjust the pH of the culture medium to 7, then add Lactobacillus rhamnosus for fermentation culture, the fermentation conditions are 28°C, 72h; the microecological preparation of α-glucosidase is obtained;

[0062] The sample types are as follows:

[0063] (1) Chinese Yam Water Extract: Weigh 4 g of Chinese Yam Water Extract and dissolve it completely in distilled water for later use. (2) Gardenia Water Extract: Weigh 4 g of Gardenia Water Extract and dissolve it completely in distilled water for later use. (3) Coix Seed Water Extract: Weigh 4 g of Coix Seed Water Extract and dissolve it completely in distilled water for later use.

[0064] (4) Chinese yam, gardenia, coix seed water extract-1: 8 g of Chinese yam water extract, 4 g of gardenia water extract, and 4 g of coix seed water extract were completely dissolved in distilled water for later use.

[0065] (5) Chinese yam, gardenia, coix seed water extract-2: 4 g Chinese yam water extract, 4 g gardenia water extract, 4 g coix seed water extract, use distilled water to completely dissolve for later use.

[0066] (6) Probiotic preparation-1: A probiotic preparation was prepared according to the above method, wherein 8 g of yam polysaccharide, 4 g of gardenia polysaccharide, 4 g of coix seed polysaccharide, and the initial cell concentration of Lactobacillus rhamnosus was 1×10 7 cfu / ml.

[0067] (7) Probiotic preparation-2: A probiotic preparation was prepared according to the above method, wherein 4 g of yam extract, 4 g of gardenia extract, 4 g of coix seed extract, and the initial cell concentration of Lactobacillus rhamnosus was 1×10 7 cfu / ml obtained by fermentation.

[0068] (8) Probiotic preparation-3: Probiotic preparation was prepared according to the above method, wherein 8 g of yam polysaccharide, 4 g of gardenia polysaccharide, 4 g of coix seed polysaccharide, and the initial cell concentration of Lactobacillus rhamnosus was 1×10 6 cfu / ml.

[0069] 2.1 Probiotics inhibit α-glucosidase

[0070] Add the dosage in the test tube according to Table 1; first add 300 μL α-amylase solution and 100 μL sample solution (polysaccharide or acarbose solution), and incubate in a water bath at 37°C for 5 minutes; then add 400 μL starch solution, mix well and incubate in a water bath at 37°C for 5 minutes; finally add 200 μL DNS reagent, take out after boiling in a water bath for 5 minutes, cool to room temperature, add distilled water to dilute the solution 25 times; mix well and add the sample to a 96-well plate, 200 μL per well, set up 3 replicate wells, and measure the absorbance at 540 nm.

[0071] Add the dosage according to Table 1 to the ELISA plate, first add the buffer solution, microecological preparation and other sample solutions and the enzyme solution in a 37℃ water bath, react at 37℃ for 15min and then take it out; add the PNPG solution and react at 37℃ for 20min; finally add the Na2CO3 solution to terminate the reaction. Measure the absorbance at 405nm. The experiment is divided into drug reaction group, drug control group, blank reaction group and blank control group, with 3 replicates in each group.

[0072] Table 1 α-glucosidase inhibition test reagent addition table

[0073] Blank reaction group Blank control group Drug response group Drug control group Sample (μL) —— —— 40 40 Distilled water (μL) 40 40 —— —— Enzyme solution (μL) 40 —— 40 —— Buffer (μL) —— 40 —— 40 PANG(μL) 60 60 60 60 DNS solution (μL) 60 60 60 60

[0074] Inhibition rate = (A blank - A sample) / A blank × 100%

[0075] A empty = A blank reaction group - A blank control group

[0076] Sample A = A drug response group - A drug control group

[0077] result:

[0078] The results are shown in Table 2, which show that each composition has a certain inhibitory activity of α-glucosidase.

[0079] Table 2 Effect of each composition on inhibiting α-glucosidase activity

[0080]

[0081]

[0082] 2.2 The probiotic preparations prepared from the three traditional Chinese medicine compositions have a synergistic inhibitory effect on α-glucosidase

[0083] The stepwise screening method starts with the fermentation of one Chinese medicine, determines the optimal dose, adds the second medicine, determines the dose of the two medicines, and then adds the third medicine. The indicator is the inhibition rate of the drug on a-amylase as the observation point, and calculates the combination index (CI).

[0084] Three plant extracts (Chinese yam, gardenia and coix seed polysaccharide) with anti-α-glucosidase activity were selected for the detection of enzyme inhibition effect. The factional inhibitory concentration index (FIC) was used to evaluate the effect of combined drug use with the help of standard chessboard titration method. FIC was calculated as follows: ∑FIC = FICA + FICB, where FICA = MICA (in the presence of B) / MICA (alone), FICB = MICB (in the presence of A) / MICB (alone). The FIC index interpretation criteria are as follows: ≤0.5, synergistic effect; 0.5-1, additive effect; 1-4.0, irrelevant effect; >4.0, antagonistic effect. The experiment was repeated 3 times.

[0085] 3. The results showed that yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide could synergistically inhibit the activity of α-glucosidase (Table 3).

[0086] Table 3 Effect of combined use of yam, gardenia and coix seed polysaccharides

[0087]

[0088]

[0089] Example 2 Probiotics promote degradation of macromolecular polysaccharides

[0090] 1. Materials and Instruments

[0091] 1.1 Materials

[0092] Chinese yam, gardenia, and coix seed were from the High-tech Industry (Pilot) Base of Shandong Academy of Sciences, Lactobacillus rhamnosus CGMCC No.1.12734, industrial alcohol, phenol (AR), concentrated sulfuric acid (AR), and glucose (AR).

[0093] 1.2 Instruments

[0094] AR-1104 electronic balance: AUX International Trading Shanghai Co., Ltd., DKB-501A super constant temperature bath: Shanghai Jinghong Experimental Equipment Co., Ltd., SHB-II circulating water multi-purpose vacuum pump: Zhengzhou Great Wall Science and Technology Industry and Trade Co., Ltd., TU-1810 UV-visible spectrophotometer: Beijing Pusi General Instrument Co., Ltd., QE-200 high-speed crusher: Wuyi County Yili Tools Co., Ltd., Hebei Province.

[0095] 2.1 Drawing of polysaccharide standard curve

[0096] Accurately weigh 5 mg of glucose dried to constant weight at 105°C and dilute to 100 ml to a concentration of 50 mg·L -1 (0.05mg / ml) glucose solution. Take 0.2, 0.4, 0.6, 0.8, 1.0, 1.2ml of the above solution into 6 test tubes respectively, add distilled water to 2ml, add 1.0ml of 5% phenol solution, shake well and quickly add 5.0ml of concentrated sulfuric acid, shake well and let stand for 10min, heat in a boiling water bath for 15min, take out and cool to room temperature. Perform the same operation with 2.0ml of distilled water as a blank control, measure the absorbance at 480nm, make a standard curve with absorbance versus concentration, and get the regression equation y=1.354x-0.1204 (r=0.998), where y represents absorbance and x is glucose concentration. Measure the y value and substitute it into the regression equation to calculate the polysaccharide content.

[0097] 2.2 Sample preparation

[0098] Carbon sources, inorganic salts and other nutritional factors were added to yam polysaccharides, gardenia polysaccharides and coix seed polysaccharides, the pH was adjusted to 7, sterile water was added in proportion, and fermented with Lactobacillus rhamnosus at 30°C for 96h. The polysaccharides were detected by standard curve analysis to evaluate whether the addition of Lactobacillus rhamnosus affected the degradation of polysaccharides.

[0099] After grinding the three medicinal materials of Chinese yam, gardenia and coix seed, the mixture was sieved through a 200-mesh sieve, and three times of their weight of purified water was added in batches. The mixture was ultrasonicated at 50 Hz for 30 minutes, centrifuged at 8000 rpm for 5 minutes to remove the precipitate, and then three times of purified water was added and ultrasonicated. This process was repeated three times. The aqueous solution after the precipitate was removed was rotary evaporated until it became a viscous semi-fluid to obtain the aqueous extracts of Chinese yam, gardenia and coix seed.

[0100] The water extracts of Chinese yam, gardenia and coix seed were added to 5 times 96% ethanol solution, heated and stirred for 15 minutes, centrifuged at 5000rpm for 5 minutes to remove the supernatant, and this process was repeated 3 times. The precipitate was dried at 25℃ for 24 hours to obtain Chinese yam, gardenia and coix seed polysaccharides.

[0101] Preparation of probiotic preparations: One or more of yam polysaccharides, gardenia polysaccharides, and coix seed polysaccharides were used as carbon sources. At the same time, 2g of beef extract, 2g of yeast extract, 2g of sodium acetate, 1g of diammonium hydrogen citrate, and 2g of dipotassium hydrogen phosphate were added to the culture medium. Water was added to 1L, and the pH of the culture medium was adjusted to 7. Then, an initial bacterial concentration of 1×10 7 cfu / ml of Lactobacillus rhamnosus was fermented and cultured at 28°C for 72h; the microecological preparation of α-glucosidase was obtained, and polysaccharides were detected by standard curve analysis to evaluate whether the addition of Lactobacillus rhamnosus affected the degradation of polysaccharides.

[0102] Sample Type:

[0103] (1) Chinese yam fermentation sample: The Chinese yam fermentation liquid was obtained by the above-mentioned preparation method of the microecological preparation, wherein the carbon source was 8 g of Chinese yam polysaccharide.

[0104] (2) Gardenia fermentation sample: The gardenia fermentation liquid was obtained by the above-mentioned preparation method of the probiotic preparation, wherein the carbon source was 4 g of gardenia polysaccharide.

[0105] (3) Coix seed fermentation sample: The coix seed fermentation liquid was obtained by using the above-mentioned preparation method of the microecological preparation, wherein the carbon source was 4 g of coix seed polysaccharide.

[0106] (4) Probiotic preparation-1: Probiotic preparation-1 was obtained by the above-mentioned preparation method of probiotic preparation, wherein the carbon source was 8 g of yam polysaccharide, 4 g of gardenia polysaccharide, and 4 g of coix seed polysaccharide.

[0107] (5) Probiotic preparation-2: Probiotic preparation-1 was obtained by the above-mentioned preparation method of probiotic preparation, wherein the carbon source was 4 g of yam polysaccharide, 4 g of gardenia polysaccharide, and 4 g of coix seed polysaccharide.

[0108] Statistical analysis:

[0109] The oligosaccharide content before and after fermentation was detected and analyzed by one-way analysis of variance using SPSS27 software (*P<0.05, **0.01<P<0.05).

[0110] 3. The results showed that the oligosaccharide content of yam fermented samples, coix seed fermented samples, probiotics-1 and probiotics-2 increased significantly after fermentation by Lactobacillus rhamnosus. Therefore, yam fermented samples, coix seed fermented samples, probiotics-1 and probiotics-2 can degrade macromolecular polysaccharides into small molecular sugars ( Figure 1 ).

[0111] It should be noted that the above examples are only used to illustrate the technical solution of the present invention rather than to limit it. Although the present invention is described in detail with reference to the given examples, those skilled in the art may modify or replace the technical solution of the present invention as needed without departing from the spirit and scope of the technical solution of the present invention.

Claims

1. A microecological preparation for inhibiting α-glucosidase, characterized in that: The microecological preparation for inhibiting α-glucosidase is obtained by fermenting yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide with Lactobacillus rhamnosus; wherein the yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide are respectively obtained by water extraction and alcohol precipitation of yam, gardenia and coix seed.

2. The microecological preparation according to claim 1, characterized in that: The mass ratio of the yam polysaccharide, the gardenia polysaccharide and the coix seed polysaccharide is 5-10:1-5:1-5.

3. The microecological preparation according to claim 1, characterized in that: The Lactobacillus rhamnosus may specifically be Lactobacillus rhamnosus CGMCC No.1.12734.

4. The method for preparing the probiotic preparation according to any one of claims 1 to 3, characterized in that: The preparation method comprises: S1. Preparing yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide respectively by water extraction and alcohol precipitation method; S2, mixing the yam polysaccharide, gardenia polysaccharide and coix seed polysaccharide obtained in step S1, and fermenting them with Lactobacillus rhamnosus.

5. The preparation method according to claim 4, characterized in that: The specific method of step S1 includes: S1-1, respectively grind and sieve yam, gardenia, and coix seed, add 3-4 times of their weight of water, after ultrasonic treatment, centrifuge to remove precipitate, then add 3-4 times of water and ultrasonicate, and repeat this process 2-4 times; after removing the precipitate, concentrate the aqueous solution by rotary evaporation to obtain yam, gardenia, and coix seed water extracts, respectively; S1-2, respectively add ethanol solution to the water extract of yam, gardenia and coix seed in step S1-1, stir, centrifuge to remove supernatant, and repeat this process 2-4 times; dry the obtained precipitate to obtain yam, gardenia and coix seed polysaccharides.

6. The preparation method according to claim 5, characterized in that: In the step S1-1, the sieve mesh is 150-250 mesh, preferably 200 mesh; the ultrasonic treatment conditions are: ultrasonic frequency is 20-80 Hz, and ultrasonic treatment time is 10-60 min; preferably ultrasonic frequency is 50 Hz, and ultrasonic treatment time is 30 min; centrifugal conditions are: centrifugation at 7000-9000 rpm for 3-6 min; In the step S1-2, the ethanol solution is a high-concentration ethanol solution, specifically a 96% ethanol solution, and the stirring time is 5-30 minutes.

7. The preparation method according to claim 5, characterized in that: In step S2, the specific conditions during the fermentation process are: the fermentation pH is controlled to be neutral, and the fermentation concentration of Lactobacillus rhamnosus is controlled to be 0.1-10×10 7 cfu / ml, the fermentation temperature was controlled at 25-35°C, and the fermentation time was 40-150h.

8. Use of the probiotic preparation according to any one of claims 1 to 3 or the probiotic preparation obtained by the preparation method according to any one of claims 4 to 7 in any one or more of the following: (a1) inhibiting α-glucosidase or preparing products that inhibit α-glucosidase; (a2) Preparation of products for the prevention and / or treatment of diabetes and related diseases.

9. The use according to claim 8, characterized in that In (a1), the product is food, medicine or experimental reagent for non-medical use, and the experimental reagent is used for basic research.

10. The use according to claim 8, characterized in that In (a2), the product is a medicine.

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