Mulberry leaf fermentation composition helpful for reversing insulin resistance and preparation method thereof

Abstract

The invention discloses a mulberry leaf fermentation composition helpful for reversing insulin resistance and a preparation method thereof, relates to the technical field of mulberry leaf fermentation treatment, and solves the problem that in the prior art, a technology for preparing auxiliary materials for assisting diabetes treatment from mulberry leaves and related products do not exist. The mulberry leaf fermentation composition is prepared from a mulberry leaf extract, a highland barley extract, a bitter gourd extract, hawthorn powder, yam flour, lemon powder, fructo-oligosaccharide, water-soluble dietary fibers, resistant dextrin and a fermentation microbial preparation, wherein the highland barley extract is prepared by mixing a highland barley flavone extract and a highland barley polysaccharide extract according to a mass ratio of (4-6): 1. Through multi-channel combination of'inhibiting sugar absorption, enhancing insulin signals, protecting islet cells and improving metabolism ', insulin signal channels can be effectively improved, inflammatory oxidative stress can be relieved, glucose and lipid metabolism disorder can be regulated, and insulin resistance can be reversed.

Description

Technical Field

[0001] This invention relates to the field of mulberry leaf fermentation technology, and specifically to a mulberry leaf fermentation composition that helps reverse insulin resistance and its preparation method. Background Technology

[0002] Insulin resistance is a core pathophysiological link in the development and progression of type 2 diabetes, and also an important common basis for many metabolic diseases such as obesity and metabolic syndrome. It is characterized by decreased sensitivity to insulin, leading to reduced glucose uptake and utilization efficiency, compensatory hyperinsulinemia, and potentially ultimately β-cell dysfunction and overt diabetes. Currently, the main clinical approaches to improving insulin resistance include lifestyle interventions and drug therapy (such as metformin and thiazolidinediones). However, these drugs may have varying degrees of side effects, including gastrointestinal reactions, weight gain, edema, or cardiovascular risks, making the exploration of safe and effective adjunctive intervention strategies crucial.

[0003] Mulberry leaves (Morus alba L.), a traditional medicinal and edible plant, are often used in Traditional Chinese Medicine as an adjunct treatment for diabetes (equivalent to diabetes in modern medicine). Modern research shows that mulberry leaves are rich in various active ingredients, including alkaloids (such as 1-deoxynojirimycin, DNJ), flavonoids, polysaccharides, and γ-aminobutyric acid (GABA). These components are believed to have multiple effects, including inhibiting α-glucosidase, delaying carbohydrate absorption, promoting insulin secretion, protecting pancreatic β-cells, regulating lipid metabolism, and anti-inflammatory and antioxidant properties, thus theoretically possessing the potential to improve insulin sensitivity.

[0004] In existing technologies, some researchers have begun to develop methods for directly using mulberry leaves or traditional mulberry leaf compound formulas to improve insulin resistance. For example, patent application CN115119938B discloses a method for reversing insulin resistance, which involves mixing mulberry leaf extract powder with raw materials such as olive oil, wakame seaweed dietary fiber powder, and soy protein powder, and then using a spraying mechanism to produce a spray. Another example is patent application CN111728214A, which discloses a medicinal and edible composition to assist in reversing diabetes. This composition uses polydextrose, sea cucumber, milk powder, whey protein, fructooligosaccharides, buckwheat extract, ginseng, polygonatum extract, yam, mulberry, wolfberry extract, poria cocos, and mulberry leaf extract to create a medicinal and edible composition to assist in reversing diabetes. For example, patent application CN121177388A discloses a tea composition for lowering blood sugar, its preparation method and application. It uses 1-5 parts of white tea, 1-9 parts of rose petals, 1-9 parts of guava leaf tea, 1-5 parts of mulberry leaves, 1-5 parts of vine tea, and 1-5 parts of Eucommia ulmoides. The raw materials are then crushed separately and mixed to obtain a mixture. The mixture is then bagged and brewed with hot water to obtain tea soup.

[0005] However, the direct application of natural mulberry leaf raw materials has the following limitations: First, the content of active ingredients in mulberry leaves is greatly affected by variety, origin, and harvesting period, making it difficult to guarantee stability and consistency; Second, active substances in mulberry leaves (such as DNJ, flavonoids, and polysaccharides) may be encapsulated by plant cell walls or bound to the matrix, making it difficult to fully release and effectively absorb them in the intestines during conventional water extraction or oral administration, resulting in insufficient actual usable concentrations in the body and affecting the stable performance of their efficacy; Third, mulberry leaves themselves may contain substances that affect taste and a small amount of anti-nutritional factors, limiting the acceptance of direct consumption or simple extract application.

[0006] In recent years, the application of microbial fermentation technology in the development of traditional Chinese medicine has provided new insights. The fermentation process utilizes microorganisms and their enzyme systems to disrupt plant cell walls, promoting the release and transformation of active ingredients, potentially generating new metabolites or enhancing the bioactivity and bioavailability of existing components. Currently, research has applied fermentation technology to various medicinal materials such as kudzu root and astragalus to enhance their hypoglycemic and immunomodulatory effects. However, research on mulberry leaf fermentation has largely focused on preparing tea, feed additives, or obtaining general active ingredients; there is still no publicly available technology that systematically applies specific microbial fermentation processes to mulberry leaves and clearly demonstrates that the resulting substances help "reverse insulin resistance."

[0007] Therefore, developing a safe and effective fermented mulberry leaf composition with significant insulin resistance reversal function is of great practical significance and application value as a new option for adjunctive treatment of diabetes. Summary of the Invention

[0008] To address the above shortcomings, this invention provides a mulberry leaf fermentation composition and its preparation method that help reverse insulin resistance. This solves the problem that existing technologies and products do not yet utilize mulberry leaves to prepare adjuvants for diabetes treatment. This invention, through a multi-pathway combination of "inhibiting glucose absorption + enhancing insulin signaling + protecting pancreatic islet cells + improving metabolism," can effectively improve the insulin signaling pathway, reduce inflammatory and oxidative stress, and regulate glucose and lipid metabolism disorders, thereby reversing insulin resistance. The specific technical solution is as follows: A mulberry leaf fermentation composition that helps reverse insulin resistance, by weight, is made from the following raw materials comprising: 55-60 parts mulberry leaf extract, 40-50 parts highland barley extract, 22-28 parts bitter melon extract, 20-25 parts hawthorn powder, 20-25 parts yam powder, 15-20 parts lemon powder, 10-15 parts fructooligosaccharides, 10-15 parts water-soluble dietary fiber, 1-5 parts resistant dextrin, and 2-8 parts fermentation microbial preparation; wherein the highland barley extract is composed of highland barley flavonoid extract and highland barley polysaccharide extract mixed in a mass ratio of (4-6):1.

[0009] Preferably, the mulberry leaf extract is prepared by moistening mulberry leaves, steam sterilizing, and inoculating with *Bacillus subtilis* (a type of bacteria). Eurotium cristatum The bacterial strain is prepared by solid-state fermentation for 4 to 6 days. After fermentation, citric acid is added, the mixture is crushed, ethanol is added for ultrasonic extraction, filtered, and the extract is concentrated and dried to obtain the final product.

[0010] More preferably, the specific preparation method of the mulberry leaf extract is as follows: the mulberry leaf extract is prepared by moistening mulberry leaves, steam sterilizing, and inoculating with *Bacillus subtilis* (a type of bacteria). Eurotium cristatum The microbial strain is prepared by solid-state fermentation for 4-6 days. After fermentation, 3-12% citric acid by weight of mulberry leaves is added, and the mixture is ball-milled at 100-300 r / min for 5-15 min. Then, 70-80% ethanol aqueous solution is added at a material-to-liquid ratio of 1:(30-50). The mixture is ultrasonically extracted for 10-20 min in a water bath at a frequency of 180-220 W and a temperature of 30-40℃. The residue is removed by filtration, and the extract is concentrated and dried by rotary evaporation to obtain a mulberry leaf extract with a 1-deoxynojirimycin content of not less than 0.1%.

[0011] Preferably, the viable count of the sugar-inhibiting *Gynostemma pentaphyllum* is (7-9) × 10⁻⁶. 8 CFU / g; the glucose-inhibiting bacteria ( Eurotium cristatum It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on July 3, 2025, with accession number GDMCC: No. 66629.

[0012] Preferably, the preparation method of the highland barley flavonoid extract is as follows: soaking healthy and intact highland barley grains at 22-28℃ for 24 hours, steam sterilizing, and inoculating with sugar-resistant bacteria (…). Streptomyces sp . The microbial strain was fermented in solid state for 70-75 hours. After fermentation, the strain was crushed and 80-88% ethanol aqueous solution was added at a material-to-liquid ratio of 1:(10-15). The mixture was then refluxed at 70-80℃ for 110-130 minutes. The reflux extraction was repeated 2-3 times. The extracts were combined, centrifuged, filtered to remove residue, and the filtrate was concentrated under reduced pressure, finely filtered, ultrafiltered, and enriched using a macroporous resin column. After washing with water, the filtrate was eluted with 70-75% ethanol. The eluent was concentrated and freeze-dried to obtain the barley flavonoid extract.

[0013] Preferably, the viable count of the anti-saccharide bacteria is (5-8) × 10⁻⁶. 8 CFU / g; the anti-saccharide bacteria ( Streptomyces sp . It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on March 26, 2025, with accession number GDMCC: No. 66062.

[0014] Preferably, the preparation method of the barley polysaccharide extract is as follows: after crushing barley, ethanol is added for reflux extraction, the residue is collected by filtration, the residue is repeatedly extracted with distilled water 1 to 2 times, all supernatants are combined, the extract is concentrated under reduced pressure, precipitated with alcohol, dialyzed, and freeze-dried to obtain the extract.

[0015] More preferably, the specific preparation method of the barley polysaccharide extract is as follows: barley is dried at 63-68℃, pulverized to pass through a sieve with a mesh size of 1-1.3 mm, added to 80% ethanol at a material-to-liquid ratio of 1:39-41 (m / m), and extracted by reflux at 84-86℃ for 1.8-2.1 h. The residue is obtained by hot filtration, washed, dried, and then added to distilled water at a material-to-liquid ratio of 1:9.5-10.5 (g / mL). The extract is then extracted at a constant temperature of 54-57℃ for 110-130 m. Centrifuge at 330–360 rpm for 13–16 min, collect the filtrate, repeat the extraction twice with the residue, combine all supernatants, concentrate under reduced pressure to one-third of the original volume, add 3.8–4.2 times the volume of the concentrate of 95% ethanol, let stand at 4°C for 14–16 h for alcohol precipitation, filter, repeat alcohol precipitation twice with the precipitate, dissolve the precipitate in water, dialyze with flowing distilled water for 58–62 h, freeze dry the liquid in the dialysate bag to obtain barley polysaccharide extract.

[0016] Preferably, the extraction method of the bitter melon extract is as follows: bitter melon is crushed and water is added, then a compound enzyme is added for enzymatic hydrolysis, centrifuged, the supernatant is taken for alcohol precipitation, the precipitate and filtrate are collected by centrifugation, the filtrate is adsorbed and eluted by LSA-20 resin, concentrated under reduced pressure, and freeze-dried to obtain powder, and the powder and precipitate are mixed to obtain the final product.

[0017] More preferably, the specific extraction method of the bitter melon extract is as follows: Fresh bitter melon is cut into 15-20mm pieces, dried at 50-60℃ until the moisture content is less than 50%, and pulverized to pass through a sieve with a mesh size of 1.0-1.5mm. The bitter melon powder is added to water with a pH of 5.0 at a material-to-water ratio of 1:(3.95-4.1). A compound enzyme of 1.3-1.5% by weight of bitter melon is added, wherein the mass ratio of cellulase to pectinase in the compound enzyme is (1-1.5):1. The mixture is kept at 60℃ and stirred for 3-4 hours for enzymatic hydrolysis. The hydrolysate is centrifuged at 200-300r / min to collect the supernatant. Under stirring, 3 times the volume of 95% ethanol is slowly added to the supernatant. The mixture is allowed to stand at 4℃ for 10-14 hours. The precipitate and filtrate are collected by centrifugation. The filtrate is then diluted with water at a concentration of 1.4-1.42×10⁻⁶. -4 The solution was adsorbed through LSA-20 resin at a flow rate of m / s, and then eluted with 4 column volumes of 80% ethanol at the same rate. The eluent was concentrated under reduced pressure and freeze-dried to obtain a powder. The powder was then mixed with the precipitate to obtain bitter melon extract.

[0018] Preferably, the fermentation microbial preparation is composed of Lactobacillus plantarum, Lactobacillus acidophilus, and Saccharomyces cerevisiae in a mass ratio of (0.5-1):(0.2-0.6):1.

[0019] Preferably, the viable count of the *Lactobacillus plantarum* is (4–7) × 10⁻⁶. 8 CFU / g, viable count of Lactobacillus acidophilus (1-4) × 10⁻⁶ 8 The CFU / g and viable count of brewer's yeast were (2-5) × 10⁻⁶. 8 CFU / g.

[0020] Preferably, the water-soluble dietary fiber is selected from one of apple pectin, grapefruit peel pectin, and blueberry pectin; the dosage form of the mulberry leaf fermentation composition is powder, granules, tablets, or capsules.

[0021] Preferably, a method for preparing a mulberry leaf fermentation composition as described above that helps reverse insulin resistance includes the following steps: (1) Mix mulberry leaf extract, highland barley extract, bitter melon extract, hawthorn powder, yam powder, lemon powder, fructooligosaccharide, water-soluble dietary fiber and resistant dextrin, sterilize, and obtain the material to be fermented; (2) Inoculate the fermentation microbial preparation into the material to be fermented in step (1), and ferment at 30-37°C for 3-7 days to obtain the fermentation product; (3) The fermentation product of step (2) is inactivated and dried to obtain the mulberry leaf fermentation composition.

[0022] Preferably, in step (2), during the fermentation process, a magnetic field with a frequency of 10kHz to 30kHz and a magnetic induction intensity of 0.5mT to 3mT is used for resonant magnetic field treatment for 10 to 20 minutes every day, and the magnetic field generating device is set on the outer wall of the fermentation tank.

[0023] Preferably, the use of a mulberry leaf fermentation composition as described above that helps reverse insulin resistance in the preparation of a food, health food, or medicine for improving insulin resistance, regulating blood sugar, or preventing / adjunctive treatment of type 2 diabetes.

[0024] The present invention achieves at least the following beneficial effects: 1. In this invention, citric acid is added during the extraction process of mulberry leaf extract. Firstly, it acts as a grinding aid. During ball milling, the solid citric acid crystals collide and rub violently with the mulberry leaf powder particles. The relatively high hardness of citric acid helps to effectively break down the tough cell walls and fiber structure of mulberry leaves, exposing encapsulated components such as 1-deoxynojirimycin (DNJ), making them easier to dissolve during subsequent extraction. Secondly, citric acid is a weak acid. Under the mechanical energy of ball milling, its acidic environment can promote the hydrolysis or chemical bond breakage of DNJ bound to cellulose and hemicellulose in mulberry leaves, transforming it from a bound state to a more soluble free state. This maximizes the dissolution of DNJ and other effective components. Furthermore, DNJ exhibits better stability and higher solubility in a weakly acidic environment. The mulberry leaf extract of this invention is rich in 1-deoxynojirimycin (DNJ) and flavonoids. DNJ can effectively inhibit α-glucosidase, delay the breakdown of carbohydrates into glucose, and directly reduce postprandial blood glucose from the source, thus alleviating the pressure on insulin secretion. Flavonoids can activate the PI3K / Akt pathway, a key channel for insulin to function in cells, enhance the sensitivity of cells to insulin, and reduce the damage of the high-glucose environment to the pancreas (pancreatic β cells) through antioxidant and anti-inflammatory effects. The bitter melon extract (rich in saponins and polysaccharides) directly phosphorylates and activates the IRS-1 / PI3K / Akt signaling cascade, promotes the transfer of glucose transporters to the cell membrane, and strongly promotes the entry of glucose into the cell for utilization.

[0025] 2. PPARγ is a classic target for improving insulin sensitivity (the target of thiazolidinediones), and AMPK is a cellular energy sensor; their combined action can promote glucose and lipid metabolism balance. The barley flavonoid extracts of this invention (such as haloflavones and saponins) can activate the PPARγ and AMPK pathways, assisting 1-deoxynojirimycin (DNJ) in mulberry leaf extract in inhibiting α-glucosidase, thereby improving lipid metabolism and insulin sensitivity. The barley polysaccharide extracts of this invention (such as β-glucan) are high-quality prebiotics that can promote the growth of beneficial bacteria and improve the intestinal microecology. Healthy flora can promote the secretion of hormones such as glucagon-like peptide-1 (GLP-1) in the intestine. GLP-1 can promote insulin secretion in a glucose-dependent manner, inhibit glucagon, and protect β cells. Simultaneously, a healthy intestinal microecology can improve the body's immunity and reduce the occurrence of chronic low-grade inflammation, thereby achieving the effect of reversing insulin resistance.

[0026] 3. Among the raw materials used in this invention, 1-deoxynojirimycin (DNJ) in mulberry leaf extract inhibits intestinal α-glucosidase, controlling sugar absorption at its source; bitter melon extract and mulberry leaf polyphenols directly activate intracellular insulin signaling pathways (such as PI3K-AKT), regulating signaling and improving sensitivity; mulberry leaf polysaccharides and barley polysaccharides protect pancreatic β-cells, while barley flavonoids and bitter melon extract reduce oxidative damage, protecting and repairing the pancreas; barley flavonoids and bitter melon extract can also regulate lipid metabolism. Therefore, the mulberry leaf fermentation composition prepared in this invention, through a multi-pathway combination of "inhibiting sugar absorption + enhancing insulin signaling + protecting pancreatic cells + improving metabolism," can effectively improve insulin signaling pathways, reduce inflammatory oxidative stress, regulate glucose and lipid metabolism disorders, and reverse insulin resistance.

[0027] 4. This invention utilizes strains such as *Gastrodinium spp.*, *Gastrodinium spp.*, and *Lactobacillus plantarum* to ferment raw materials. This process degrades macromolecular polysaccharides and proteins in the raw materials into smaller peptides and oligosaccharides that are more easily absorbed by the intestines. It also degrades substances that affect digestion and absorption or have mild irritant properties. Under the influence of these strains, the content of beneficial components such as 1-deoxynojirimycin (DNJ), flavonoids, and prebiotics is increased. Fermentation effectively degrades bitter and astringent substances in the raw materials, producing pleasant sour or aromatic substances, resulting in improved flavor and texture. The fermentation process is assisted by a resonant magnetic field, which effectively enhances the permeability of microbial cell membranes, stimulates metabolic enzyme activity and substance transport, promotes nutrient absorption and metabolic product excretion, thereby comprehensively accelerating the growth, reproduction, and metabolic rate of microorganisms, ultimately increasing the content of beneficial components in the product. Detailed Implementation

[0028] The specific embodiments of the present invention are described in detail below, but it should be understood that the scope of protection of the present invention is not limited to the specific embodiments. Unless otherwise defined, all technical terms used below have the same meaning as commonly understood by those skilled in the art. The technical terms used herein are for the purpose of describing specific embodiments only and are not intended to limit the scope of protection of the present invention. Unless otherwise specifically stated, all raw materials, reagents, instruments, and equipment used in the present invention are commercially available or can be prepared by existing methods.

[0029] The endpoints and any values ​​of the ranges disclosed herein are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.

[0030] The present invention relates to the sugar-inhibiting fungus ( Eurotium cristatum It was deposited on July 3, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC: No. 66629.

[0031] The anti-sugar bacteria involved in this invention ( Streptomyces sp . It was deposited on March 26, 2025 at the Guangdong Provincial Center for the Preservation of Microbial Cultures, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, with accession number GDMCC: No. 66062.

[0032] Example 1 A mulberry leaf fermented composition powder that helps reverse insulin resistance is made from the following raw materials in parts by weight: 55 parts mulberry leaf extract, 40 parts highland barley extract, 22 parts bitter melon extract, 20 parts hawthorn powder, 20 parts yam powder, 15 parts lemon powder, 10 parts fructooligosaccharides, 10 parts apple pectin, 1 part resistant dextrin, and 2 parts fermentation microbial preparation; the highland barley extract is a mixture of highland barley flavonoid extract and highland barley polysaccharide extract in a mass ratio of 4:1.

[0033] Specifically, the preparation method of mulberry leaf extract is as follows: mulberry leaves are moistened, sterilized by steam, and inoculated with *Bacillus subtilis* (a type of bacteria). Eurotium cristatum The microbial strain was prepared by solid-state fermentation for 4 days. After fermentation, 3% citric acid (by weight of mulberry leaves) was added, and the mixture was ball-milled at 100 r / min for 15 min. A 70% ethanol aqueous solution was added at a material-to-liquid ratio of 1:30. The mixture was then ultrasonically extracted in a water bath at 180 W and 30 °C for 20 min. The residue was removed by filtration, and the extract was concentrated and dried by rotary evaporation to obtain a mulberry leaf extract with a 1-deoxynojirimycin content of not less than 0.1%.

[0034] More specifically, the preparation method of barley flavonoid extract is as follows: healthy and intact barley grains are soaked at 22℃ for 24 hours, sterilized by steam, and inoculated with anti-glycemic bacteria (…). Streptomyces sp . The bacterial strain was fermented in solid state for 70 hours. After fermentation, the strain was crushed and 80% ethanol aqueous solution was added at a material-to-liquid ratio of 1:10. The mixture was then extracted by hot reflux at 70℃ for 130 minutes. The reflux extraction was repeated 3 times. The extracts were combined, centrifuged and filtered to remove residue. The filtrate was concentrated under reduced pressure, finely filtered, ultrafiltered, and enriched by macroporous resin column. After washing with water, it was eluted with 70% ethanol. The eluent was concentrated and freeze-dried to obtain barley flavonoid extract.

[0035] More specifically, the preparation method of barley polysaccharide extract is as follows: Barley is dried at 63℃, pulverized to pass through a 1mm sieve, added to 80% ethanol at a material-to-liquid ratio of 1:39 (m / m), and extracted by reflux at 84℃ for 2.1h. The residue is obtained by hot filtration. After washing and drying, the residue is added to distilled water at a material-to-liquid ratio of 1:9.5 (g / mL), and extracted at 54℃ for 110min. Then, it is centrifuged at 330r / min for 13min, and the filtrate is collected. The residue is extracted twice more. All supernatants are combined and concentrated under reduced pressure to one-third of the original volume. 95% ethanol, 3.8 times the volume of the concentrated liquid, is added and allowed to stand at 4℃ for 14h for alcohol precipitation. After filtration, the precipitate is precipitated twice more. The precipitate is dissolved in water and dialyzed with flowing distilled water for 58h. The liquid in the dialysate bag is freeze-dried to obtain barley polysaccharide extract.

[0036] More specifically, the extraction method for bitter melon extract is as follows: Fresh bitter melon is cut into 15mm pieces, dried at 50℃ until the moisture content is below 50%, and pulverized through a 1.0mm mesh sieve. The bitter melon powder is added to water at pH 5.0 at a material-to-water ratio of 1:3.95. 1.3% (by weight) of a compound enzyme (cellulase and pectinase in a 1:1 mass ratio) is added. The mixture is hydrolyzed at 60℃ with constant stirring for 3 hours. The hydrolysate is centrifuged at 200 rpm to collect the supernatant. Under stirring, three times the volume of 95% ethanol is slowly added to the supernatant. The mixture is allowed to stand at 4℃ for 10 hours, and the precipitate and filtrate are collected by centrifugation. The filtrate is then... -4 The solution was adsorbed through LSA-20 resin at a flow rate of m / s, and then eluted with 4 column volumes of 80% ethanol at the same rate. The eluent was concentrated under reduced pressure and freeze-dried to obtain a powder. The powder was then mixed with the precipitate to obtain bitter melon extract.

[0037] More specifically, the fermentation microbial preparation is composed of Lactobacillus plantarum, Lactobacillus acidophilus, and Saccharomyces cerevisiae in a mass ratio of 0.5:0.2:1.

[0038] More specifically, the viable count of *Bacillus thuringiensis* was 7 × 10⁻⁶. 8 CFU / g, viable count of anti-glycolytic bacteria: 5 × 10⁻⁶ 8 CFU / g, viable count of Lactobacillus plantarum 7×10 8 CFU / g, viable count of Lactobacillus acidophilus 1×10 8 The CFU / g and viable count of brewer's yeast were 2×10⁻⁶. 8 CFU / g.

[0039] A method for preparing a mulberry leaf fermented composition powder that helps reverse insulin resistance, as described above, includes the following steps: (1) Mix mulberry leaf extract, highland barley extract, bitter melon extract, hawthorn powder, yam powder, lemon powder, fructooligosaccharide, apple pectin and resistant dextrin, sterilize, and obtain the material to be fermented; (2) Inoculate the fermentation microbial preparation into the material to be fermented in step (1), ferment at 30°C for 7 days. During the fermentation process, use a magnetic field with a frequency of 10kHz and a magnetic induction intensity of 3mT for 10 minutes every day to resonate magnetic field treatment. The magnetic field generating device is set on the outer wall of the fermentation tank to obtain the fermentation product. (3) The fermentation product of step (2) is inactivated and dried to obtain the mulberry leaf fermentation composition.

[0040] Example 2 A fermented mulberry leaf composition granule that helps reverse insulin resistance is made from the following raw materials in parts by weight: 60 parts mulberry leaf extract, 50 parts highland barley extract, 28 parts bitter melon extract, 25 parts hawthorn powder, 25 parts yam powder, 20 parts lemon powder, 15 parts fructooligosaccharides, 15 parts grapefruit peel pectin, 5 parts resistant dextrin, and 2-8 parts fermentation microbial preparation; the highland barley extract is a mixture of highland barley flavonoid extract and highland barley polysaccharide extract in a mass ratio of 6:1.

[0041] Specifically, the preparation method of mulberry leaf extract is as follows: mulberry leaves are moistened, sterilized by steam, and inoculated with *Bacillus subtilis* (a type of bacteria). Eurotium cristatum The microbial strain was prepared by solid-state fermentation for 6 days. After fermentation, 12% citric acid (by weight of mulberry leaves) was added, and the mixture was ball-milled at 300 r / min for 5 min. An 80% ethanol aqueous solution was added at a material-to-liquid ratio of 1:50. The mixture was then ultrasonically extracted for 10 min in a water bath at 220 W and 40 °C. The residue was removed by filtration, and the extract was concentrated and dried by rotary evaporation to obtain a mulberry leaf extract with a 1-deoxynojirimycin content of not less than 0.1%.

[0042] More specifically, the preparation method of barley flavonoid extract is as follows: healthy and intact barley grains are soaked at 28°C for 24 hours, sterilized by steam, and inoculated with anti-glycemic bacteria (…). Streptomyces sp . The bacterial strain was fermented in solid state for 75 hours. After fermentation, the strain was crushed and 88% ethanol aqueous solution was added at a material-to-liquid ratio of 1:15. The mixture was then extracted by hot reflux at 80℃ for 110 minutes. The reflux extraction was repeated twice. The extracts were combined, centrifuged, filtered to remove residue, and the filtrate was concentrated under reduced pressure, finely filtered, ultrafiltered, and enriched by macroporous resin column. After washing with water, the filtrate was eluted with 75% ethanol. The eluent was concentrated and freeze-dried to obtain barley flavonoid extract.

[0043] More specifically, the preparation method of highland barley polysaccharide extract is as follows: Highland barley is dried at 68℃, pulverized to pass through a 1.3mm sieve, added to 80% ethanol at a material-to-liquid ratio of 1:41 (m / m), and extracted by reflux at 86℃ for 1.8h. The residue is obtained by hot filtration. After washing and drying, the residue is added to distilled water at a material-to-liquid ratio of 1:10.5 (g / mL), and extracted at 57℃ for 110min. Then, it is centrifuged at 360r / min for 13min, and the filtrate is collected. The residue is extracted twice more. All supernatants are combined and concentrated under reduced pressure to one-third of the original volume. 95% ethanol, 4.2 times the volume of the concentrated liquid, is added and allowed to stand at 4℃ for ethanol precipitation for 16h. After filtration, the precipitate is precipitated twice more. The precipitate is dissolved in water and dialyzed with flowing distilled water for 62h. The liquid in the dialysate bag is freeze-dried to obtain highland barley polysaccharide extract.

[0044] More specifically, the extraction method for bitter melon extract is as follows: Fresh bitter melon is cut into 20mm pieces, dried at 60℃ until the moisture content is below 50%, and pulverized through a 1.5mm mesh sieve. The bitter melon powder is added to water at pH 5.0 at a material-to-water ratio of 1:4.1. 1.5% of a compound enzyme (by weight of bitter melon) is added, with a mass ratio of cellulase to pectinase of (1-1.5):1. The mixture is stirred at 60℃ for 4 hours for enzymatic hydrolysis. The hydrolysate is centrifuged at 300 rpm to collect the supernatant. Under stirring, three times the volume of 95% ethanol is slowly added to the supernatant. The mixture is allowed to stand at 4℃ for 14 hours, and the precipitate and filtrate are collected by centrifugation. The filtrate is then processed at 1.42 × 10⁻⁶ ppm. -4 The solution was adsorbed through LSA-20 resin at a flow rate of m / s, and then eluted with 4 column volumes of 80% ethanol at the same rate. The eluent was concentrated under reduced pressure and freeze-dried to obtain a powder. The powder was then mixed with the precipitate to obtain bitter melon extract.

[0045] More specifically, the fermentation microbial preparation is made by mixing Lactobacillus plantarum, Lactobacillus acidophilus, and Saccharomyces cerevisiae in a mass ratio of 1:0.6:1.

[0046] More specifically, the viable count of *Bacillus thuringiensis* was 9 × 10⁻⁶. 8 CFU / g, viable count of anti-glycolytic bacteria: 8 × 10⁻⁶ 8 CFU / g, viable count of Lactobacillus plantarum 4×10 8 CFU / g, viable count of Lactobacillus acidophilus 4×10 8 The CFU / g and viable count of brewer's yeast were 5×10⁻⁶. 8 CFU / g.

[0047] A method for preparing mulberry leaf fermentation composition granules, as described above, which helps reverse insulin resistance, includes the following steps: (1) Mix mulberry leaf extract, highland barley extract, bitter melon extract, hawthorn powder, yam powder, lemon powder, fructooligosaccharide, grapefruit peel pectin and resistant dextrin, sterilize, and obtain the material to be fermented; (2) Inoculate the fermentation microbial preparation into the material to be fermented in step (1), ferment at 37°C for 3 days. During the fermentation process, use a magnetic field with a frequency of 30kHz and a magnetic induction intensity of 0.5mT for 20 minutes every day to resonate magnetic field treatment. The magnetic field generating device is set on the outer wall of the fermentation tank to obtain the fermentation product. (3) The fermentation product of step (2) is inactivated, dried, and granulated to obtain the mulberry leaf fermentation composition.

[0048] Example 3 A mulberry leaf fermentation composition tablet that helps reverse insulin resistance is made from the following raw materials in parts by weight: 56 parts mulberry leaf extract, 42 parts highland barley extract, 23 parts bitter melon extract, 21 parts hawthorn powder, 21 parts yam powder, 16 parts lemon powder, 11 parts fructooligosaccharides, 11 parts blueberry pectin, 2 parts resistant dextrin, and 2-8 parts fermentation microbial preparation; the highland barley extract is a mixture of highland barley flavonoid extract and highland barley polysaccharide extract in a mass ratio of 4:1.

[0049] Specifically, the preparation method of mulberry leaf extract is as follows: mulberry leaves are moistened, sterilized by steam, and inoculated with *Bacillus subtilis* (a type of bacteria). Eurotium cristatum The microbial strain was prepared by solid-state fermentation for 4 days. After fermentation, 5% citric acid (by weight of mulberry leaves) was added, and the mixture was ball-milled at 150 r / min for 12 min. A 73% ethanol aqueous solution was added at a material-to-liquid ratio of 1:35. The mixture was then ultrasonically extracted in a water bath at 190 W and 33 °C for 18 min. The residue was removed by filtration, and the extract was concentrated and dried by rotary evaporation to obtain a mulberry leaf extract with a 1-deoxynojirimycin content of not less than 0.1%.

[0050] More specifically, the preparation method of barley flavonoid extract is as follows: healthy and intact barley grains are soaked at 23°C for 24 hours, sterilized by steam, and inoculated with anti-glycemic bacteria (…). Streptomyces sp . The bacterial strain was fermented in solid state for 71 hours. After fermentation, the strain was crushed and 83% ethanol aqueous solution was added at a material-to-liquid ratio of 1:12. The mixture was then extracted by hot reflux at 72℃ for 125 minutes. The reflux extraction was repeated 3 times. The extracts were combined, centrifuged and filtered to remove residue. The filtrate was concentrated under reduced pressure, finely filtered, ultrafiltered, and enriched by macroporous resin column. After washing with water, it was eluted with 72% ethanol. The eluent was concentrated and freeze-dried to obtain barley flavonoid extract.

[0051] More specifically, the preparation method of highland barley polysaccharide extract is as follows: Highland barley is dried at 64℃, pulverized to pass through a 1.1mm sieve, added to 80% ethanol at a material-to-liquid ratio of 1:39 (m / m), and extracted by reflux at 84℃ for 1.9h. The residue is obtained by hot filtration. After washing and drying, the residue is added to distilled water at a material-to-liquid ratio of 1:9.8 (g / mL), and extracted at 55℃ for 125min. Then, it is centrifuged at 340r / min for 14min, and the filtrate is collected. The residue is extracted twice more. All supernatants are combined and concentrated under reduced pressure to one-third of the original volume. 95% ethanol (3.9 times the volume of the concentrated liquid) is added, and the mixture is allowed to stand at 4℃ for 14.5h for alcohol precipitation. After filtration, the precipitate is precipitated twice more. The precipitate is dissolved in water and dialyzed with flowing distilled water for 59h. The liquid in the dialysate bag is freeze-dried to obtain highland barley polysaccharide extract.

[0052] More specifically, the extraction method for bitter melon extract is as follows: Fresh bitter melon is cut into 16mm pieces, dried at 52℃ until the moisture content is below 50%, and pulverized through a 1.1mm sieve. The bitter melon powder is added to water at pH 5.0 at a material-to-water ratio of 1:3.98. A compound enzyme (1.3% by weight of bitter melon) is added, with a mass ratio of cellulase to pectinase of 1.1:1. The mixture is hydrolyzed at 60℃ with constant stirring for 3.2 hours. The hydrolysate is centrifuged at 220 rpm to collect the supernatant. Under stirring, three times the volume of 95% ethanol is slowly added to the supernatant. The mixture is allowed to stand at 4℃ for 11 hours, and the precipitate and filtrate are collected by centrifugation. The filtrate is then... -4 The solution was adsorbed through LSA-20 resin at a flow rate of m / s, and then eluted with 4 column volumes of 80% ethanol at the same rate. The eluent was concentrated under reduced pressure and freeze-dried to obtain a powder. The powder was then mixed with the precipitate to obtain bitter melon extract.

[0053] More specifically, the fermentation microbial preparation is composed of Lactobacillus plantarum, Lactobacillus acidophilus, and Saccharomyces cerevisiae in a mass ratio of 0.6:0.3:1.

[0054] More specifically, the viable count of *Bacillus saccharidogenus* was 8 × 10⁻⁶. 8 CFU / g, viable count of anti-glycolytic bacteria: 6 × 10⁻⁶ 8 CFU / g, viable count of Lactobacillus plantarum 5×10 8 CFU / g, viable count of Lactobacillus acidophilus 2×10 8 The CFU / g and viable count of brewer's yeast were 3×10⁻⁶. 8 CFU / g.

[0055] A method for preparing a mulberry leaf fermentation composition tablet, as described above, which helps reverse insulin resistance, includes the following steps: (1) Mix mulberry leaf extract, highland barley extract, bitter melon extract, hawthorn powder, yam powder, lemon powder, fructooligosaccharide, blueberry pectin and resistant dextrin, sterilize, and obtain the material to be fermented; (2) Inoculate the fermentation microbial preparation into the material to be fermented in step (1), ferment at 32°C for 6 days. During the fermentation process, use a magnetic field with a frequency of 15kHz and a magnetic induction intensity of 1mT for 12 minutes every day to resonate magnetic field treatment. The magnetic field generating device is set on the outer wall of the fermentation tank to obtain the fermentation product. (3) The fermentation product of step (2) is inactivated, dried, and compressed into tablets to obtain the mulberry leaf fermentation composition.

[0056] Example 4 A mulberry leaf fermented composition capsule that helps reverse insulin resistance is made from the following raw materials in parts by weight: 59 parts mulberry leaf extract, 49 parts highland barley extract, 27 parts bitter melon extract, 24 parts hawthorn powder, 24 parts yam powder, 19 parts lemon powder, 14 parts fructooligosaccharides, 14 parts grapefruit peel pectin, 4 parts resistant dextrin, and 7 parts fermentation microbial preparation; the highland barley extract is a mixture of highland barley flavonoid extract and highland barley polysaccharide extract in a mass ratio of 6:1.

[0057] Specifically, the preparation method of mulberry leaf extract is as follows: mulberry leaves are moistened, sterilized by steam, and inoculated with *Bacillus subtilis* (a type of bacteria). Eurotium cristatum The microbial strain was prepared by solid-state fermentation for 6 days. After fermentation, 10% citric acid (by weight of mulberry leaves) was added, and the mixture was ball-milled at 250 r / min for 8 min. A 78% ethanol aqueous solution was added at a material-to-liquid ratio of 1:45, and the mixture was ultrasonically extracted in a water bath at 210 W and 38 °C for 12 min. The residue was removed by filtration, and the extract was concentrated and dried by rotary evaporation to obtain a mulberry leaf extract with a 1-deoxynojirimycin content of not less than 0.1%.

[0058] More specifically, the preparation method of barley flavonoid extract is as follows: healthy and intact barley grains are soaked at 26℃ for 24 hours, sterilized by steam, and inoculated with anti-glycemic bacteria (…). Streptomyces sp . The bacterial strain was fermented in solid state for 74 hours. After fermentation, the strain was crushed and 86% ethanol aqueous solution was added at a material-to-liquid ratio of 1:14. The mixture was then extracted by hot reflux at 78℃ for 115 minutes. The reflux extraction was repeated 3 times. The extracts were combined, centrifuged and filtered to remove residue. The filtrate was concentrated under reduced pressure, finely filtered, ultrafiltered, and enriched by macroporous resin column. After washing with water, it was eluted with 74% ethanol. The eluent was concentrated and freeze-dried to obtain barley flavonoid extract.

[0059] More specifically, the preparation method of highland barley polysaccharide extract is as follows: Highland barley is dried at 67℃, pulverized to pass through a 1.2mm sieve, added to 80% ethanol at a material-to-liquid ratio of 1:41 (m / m), and extracted by reflux at 86℃ for 1.9h. The residue is obtained by hot filtration. After washing and drying, the residue is added to distilled water at a material-to-liquid ratio of 1:10.3 (g / mL), and extracted at 56℃ for 115min. Then, it is centrifuged at 350r / min for 15min, and the filtrate is collected. The residue is extracted twice more. All supernatants are combined and concentrated under reduced pressure to one-third of the original volume. 95% ethanol, 4.1 times the volume of the concentrated liquid, is added and allowed to stand at 4℃ for ethanol precipitation for 15.5h. After filtration, the precipitate is precipitated twice more. The precipitate is dissolved in water and dialyzed with flowing distilled water for 61h. The liquid in the dialysate bag is freeze-dried to obtain highland barley polysaccharide extract.

[0060] More specifically, the extraction method for bitter melon extract is as follows: Fresh bitter melon is cut into 19mm pieces, dried at 58℃ until the moisture content is below 50%, and pulverized through a 1.4mm mesh sieve. The bitter melon powder is added to water at pH 5.0 at a material-to-water ratio of 1:4.05. A compound enzyme of 1.5% by weight of bitter melon is added, with a mass ratio of cellulase to pectinase of 1.4:1. The mixture is hydrolyzed at 60℃ with constant stirring for 3.8 hours. The hydrolysate is centrifuged at 280 r / min to collect the supernatant. Under stirring, three times the volume of 95% ethanol is slowly added to the supernatant. The mixture is allowed to stand at 4℃ for 13 hours, and the precipitate and filtrate are collected by centrifugation. The filtrate is then processed at 1.42 × 10⁻⁶. -4 The solution was adsorbed through LSA-20 resin at a flow rate of m / s, and then eluted with 4 column volumes of 80% ethanol at the same rate. The eluent was concentrated under reduced pressure and freeze-dried to obtain a powder. The powder was then mixed with the precipitate to obtain bitter melon extract.

[0061] More specifically, the fermentation microbial preparation is made by mixing Lactobacillus plantarum, Lactobacillus acidophilus, and Saccharomyces cerevisiae in a mass ratio of 0.9:0.5:1.

[0062] More specifically, the viable count of *Bacillus saccharidogenus* was 8 × 10⁻⁶. 8 CFU / g, viable count of anti-glycolytic bacteria: 7 × 10⁻⁶ 8 CFU / g, viable count of Lactobacillus plantarum 6×10 8 CFU / g, viable count of Lactobacillus acidophilus 3×10 8 The CFU / g and viable count of brewer's yeast were 4×10⁻⁶. 8 CFU / g.

[0063] A method for preparing a capsule of mulberry leaf fermentation composition as described above, which helps reverse insulin resistance, includes the following steps: (1) Mix mulberry leaf extract, highland barley extract, bitter melon extract, hawthorn powder, yam powder, lemon powder, fructooligosaccharide, grapefruit peel pectin and resistant dextrin, sterilize, and obtain the material to be fermented; (2) The fermentation microbial preparation is inoculated into the material to be fermented in step (1), and fermented at 36°C for 4 days. During the fermentation process, a magnetic field with a frequency of 25kHz and a magnetic induction intensity of 2.5mT is used for resonance magnetic field treatment for 18 minutes every day. The magnetic field generating device is set on the outer wall of the fermentation tank to obtain the fermentation product. (3) The fermentation product of step (2) is inactivated and dried, and then packaged into capsules to obtain the mulberry leaf fermentation composition.

[0064] Example 5 A fermented mulberry leaf composition granule that helps reverse insulin resistance is made from the following raw materials in parts by weight: 58 parts mulberry leaf extract, 45 parts highland barley extract, 25 parts bitter melon extract, 23 parts hawthorn powder, 22 parts yam powder, 17 parts lemon powder, 13 parts fructooligosaccharides, 12 parts blueberry pectin, 3 parts resistant dextrin, and 5 parts fermentation microbial preparation; the highland barley extract is a mixture of highland barley flavonoid extract and highland barley polysaccharide extract in a mass ratio of 5:1.

[0065] Specifically, the preparation method of mulberry leaf extract is as follows: mulberry leaves are moistened, sterilized by steam, and inoculated with *Bacillus subtilis* (a type of bacteria). Eurotium cristatum The microbial strain was prepared by solid-state fermentation for 5 days. After fermentation, 7% citric acid (by weight of mulberry leaves) was added, and the mixture was ball-milled at 200 r / min for 10 min. A 75% ethanol aqueous solution was added at a material-to-liquid ratio of 1:40. The mixture was then ultrasonically extracted for 15 min in a water bath at 200 W and 35 °C. The residue was removed by filtration, and the extract was concentrated and dried by rotary evaporation to obtain a mulberry leaf extract with a 1-deoxynojirimycin content of not less than 0.1%.

[0066] More specifically, the preparation method of barley flavonoid extract is as follows: healthy and intact barley grains are soaked at 25°C for 24 hours, sterilized by steam, and inoculated with anti-glycemic bacteria (…). Streptomyces sp . The bacterial strain was fermented in solid state for 73 hours. After fermentation, the strain was crushed and 84% ethanol aqueous solution was added at a material-to-liquid ratio of 1:13. The mixture was then extracted by hot reflux at 75°C for 120 minutes. The reflux extraction was repeated 3 times. The extracts were combined, centrifuged, filtered to remove residue, and the filtrate was concentrated under reduced pressure, finely filtered, ultrafiltered, and enriched by macroporous resin column. After washing with water, the filtrate was eluted with 73% ethanol. The eluent was concentrated and freeze-dried to obtain barley flavonoid extract.

[0067] More specifically, the preparation method of barley polysaccharide extract is as follows: Barley is dried at 65℃, pulverized to pass through a 1.2mm sieve, added to 80% ethanol at a material-to-liquid ratio of 1:40 (m / m), and extracted by reflux at 85℃ for 2 hours. The residue is obtained by hot filtration. After washing and drying, the residue is added to distilled water at a material-to-liquid ratio of 1:10 (g / mL), and extracted at 55℃ for 2 hours. Then, it is centrifuged at 350r / min for 15 minutes, and the filtrate is collected. The residue is extracted twice more. All supernatants are combined and concentrated under reduced pressure to one-third of the original volume. Four times the volume of 95% ethanol is added, and the mixture is allowed to stand at 4℃ for 15 hours for alcohol precipitation. After filtration, the precipitate is precipitated twice more. The precipitate is dissolved in water and dialyzed with flowing distilled water for 60 hours. The liquid in the dialysate bag is freeze-dried to obtain barley polysaccharide extract.

[0068] More specifically, the extraction method for bitter melon extract is as follows: Fresh bitter melon is cut into 18mm pieces, dried at 505℃ until the moisture content is below 50%, and pulverized through a 1.3mm mesh sieve. The bitter melon powder is added to water at pH 5.0 at a material-to-water ratio of 1:4.0. A compound enzyme of 1.4% of the bitter melon weight is added, with a mass ratio of cellulase to pectinase of 1.3:1. The mixture is stirred at 60℃ for 3.5 hours for enzymatic hydrolysis. The hydrolysate is centrifuged at 250 rpm to collect the supernatant. Under stirring, three times the volume of 95% ethanol is slowly added to the supernatant. The mixture is allowed to stand at 4℃ for 12 hours, and the precipitate and filtrate are collected by centrifugation. The filtrate is then processed at 1.41 × 10⁻⁶ ppm. -4 The solution was adsorbed through LSA-20 resin at a flow rate of m / s, and then eluted with 4 column volumes of 80% ethanol at the same rate. The eluent was concentrated under reduced pressure and freeze-dried to obtain a powder. The powder was then mixed with the precipitate to obtain bitter melon extract.

[0069] More specifically, the fermentation microbial preparation is composed of Lactobacillus plantarum, Lactobacillus acidophilus, and Saccharomyces cerevisiae in a mass ratio of 0.7:0.4:1.

[0070] More specifically, the viable count of *Bacillus saccharidogenus* was 8 × 10⁻⁶. 8 The CFU / g and the viable count of anti-saccharide bacteria were 6.5 × 10⁻⁶. 8 The CFU / g and viable count of *Lactobacillus plantarum* were 5.5 × 10⁻⁶. 8 The CFU / g and viable count of Lactobacillus acidophilus were 2.5 × 10⁻⁶. 8 The CFU / g and viable count of brewer's yeast were 3.5 × 10⁻⁶. 8 CFU / g.

[0071] A method for preparing mulberry leaf fermentation composition granules, as described above, which helps reverse insulin resistance, includes the following steps: (1) Mix mulberry leaf extract, highland barley extract, bitter melon extract, hawthorn powder, yam powder, lemon powder, fructooligosaccharide, blueberry pectin and resistant dextrin, sterilize, and obtain the material to be fermented; (2) Inoculate the fermentation microbial preparation into the material to be fermented in step (1), ferment at 35°C for 5 days. During the fermentation process, use a magnetic field with a frequency of 20kHz and a magnetic induction intensity of 2mT for 15 minutes every day to resonate magnetic field treatment. The magnetic field generating device is set on the outer wall of the fermentation tank to obtain the fermentation product. (3) The fermentation product of step (2) is inactivated, dried, and granulated to obtain the mulberry leaf fermentation composition.

[0072] Comparative Example 1 The difference from Example 5 is that the raw materials include 30 parts of highland barley extract, while other conditions remain unchanged.

[0073] Comparative Example 2 The difference from Example 5 is that the raw materials contain 65 parts of barley extract, while other conditions remain unchanged.

[0074] Comparative Example 3 The difference from Example 5 is that the barley extract is made by mixing barley flavonoid extract and barley polysaccharide extract in a mass ratio of 8:1, while other conditions remain unchanged.

[0075] Comparative Example 4 The difference from Example 5 is that the raw materials contain 45 parts of mulberry leaf extract, while other conditions remain unchanged.

[0076] Comparative Example 5 The difference from Example 5 is that the raw materials contain 75 parts of mulberry leaf extract, while other conditions remain unchanged.

[0077] Comparative Example 6 The difference from Example 5 is that no saccharin-inhibiting bacteria were added during the preparation of mulberry leaf extract for fermentation, while other conditions remained unchanged.

[0078] Comparative Example 7 The difference from Example 5 is that no anti-glucosamine bacteria were added during the preparation of the barley flavonoid extract, while other conditions remained unchanged.

[0079] Comparative Example 8 The difference from Example 5 is that the fermentation microbial preparation is made by mixing *Gynostemma pentaphyllum*, *Gynostemma pentaphyllum*, *Lactobacillus plantarum*, *Lactobacillus acidophilus*, and *Saccharomyces cerevisiae* in a mass ratio of 9:7:1:0.6:1, while other conditions remain unchanged.

[0080] Comparative Example 9 The difference from Example 5 is that the frequency of the resonant magnetic field treatment during fermentation is 45kHz and the magnetic induction intensity is 5mT, while other conditions remain unchanged.

[0081] Comparative Example 10 The difference from Example 5 is that the resonant magnetic field treatment time during fermentation is 60 minutes, while other conditions remain unchanged.

[0082] I. Animal Experiments 1. Animal models Animals: SPF-grade male C57BL / 6J mice, weighing 18–22g.

[0083] Modeling and grouping: Several mice were fed a high-sugar, high-fat diet for 10 weeks, then fasted for 8 hours. Fasting blood glucose (FBG) was measured. Mice with FBG ≥ 11.1 mmol / L were considered to have successfully modeled the mice. 150 mice that successfully modeled the mice were fed for another 4 weeks and then randomly divided into 15 groups of 10 mice each. The example groups (Examples 1-5), comparative groups (Comparative Examples 1-9), and positive control groups were set up. In addition, 10 normal mice were selected as blank control groups.

[0084] 2. Administration Examples 1-5 and Comparative Examples 1-9: The fermented mulberry leaf compositions prepared by the methods of Examples 1-5 and Comparative Examples 1-9 were administered by gavage at a dose of 200 mg / (kg·d) (based on dry matter). Positive control group: metformin was administered by gavage at a dose of 200 mg / (kg·d); Blank control group: administered normal saline by gavage at a dose of 200 mg / (kg·d); Cycle: Continuous administration for 4 weeks.

[0085] 3. Indicator Testing At the end of the fourth week of drug administration, after a 12-hour fast, the following tests were performed: Fasting blood glucose (FBG, mmol / L): Blood was collected from the tail vein and measured with a blood glucose meter.

[0086] Fasting insulin (FINS, mIU / L): Serum was collected and measured using an ELISA kit.

[0087] Calculation of indices: The mouse insulin resistance index (HOMA-IR) and insulin sensitivity index (ISI) were calculated according to the formula.

[0088] Calculation formula: HOMA-IR=FBG×FINS / 22.5; ISI=ln[1 / (FBG×FINS)].

[0089] The experimental results are shown in Table 1 below.

[0090] Table 1 In summary, among the raw materials used in this invention, 1-deoxynojirimycin (DNJ) in mulberry leaf extract inhibits intestinal α-glucosidase, controlling sugar absorption at its source; bitter melon extract and mulberry leaf polyphenols directly activate intracellular insulin signaling pathways (such as PI3K-AKT), regulating signaling and improving sensitivity; mulberry leaf polysaccharides and barley polysaccharides protect pancreatic β-cells, while barley flavonoids and bitter melon extract reduce oxidative damage, protecting and repairing the pancreas; barley flavonoids and bitter melon extract can also regulate lipid metabolism. Therefore, the mulberry leaf fermentation composition prepared in this invention, through a multi-pathway combination of "inhibiting sugar absorption + enhancing insulin signaling + protecting pancreatic cells + improving metabolism," can effectively improve insulin signaling pathways, reduce inflammatory oxidative stress, regulate glucose and lipid metabolism disorders, and reverse insulin resistance.

[0091] The foregoing description of specific exemplary embodiments of the invention is for illustrative and explanatory purposes. These descriptions are not intended to limit the invention to the precise forms disclosed, and it will be apparent that many changes and variations can be made in accordance with the foregoing teachings. The exemplary embodiments were chosen and described in order to explain the specific principles of the invention and its practical application, thereby enabling those skilled in the art to implement and utilize various different exemplary embodiments of the invention, as well as various different choices and variations. The scope of the invention is intended to be defined by the claims and their equivalents.

Claims

1. A mulberry leaf fermentation composition that helps reverse insulin resistance, characterized in that, The mulberry leaf fermentation composition, by weight, is made from the following raw materials in parts by weight: 55-60 parts mulberry leaf extract, 40-50 parts highland barley extract, 22-28 parts bitter melon extract, 20-25 parts hawthorn powder, 20-25 parts yam powder, 15-20 parts lemon powder, 10-15 parts fructooligosaccharides, 10-15 parts water-soluble dietary fiber, 1-5 parts resistant dextrin, and 2-8 parts fermentation microbial preparation; the highland barley extract is a mixture of highland barley flavonoid extract and highland barley polysaccharide extract in a mass ratio of (4-6):

1.

2. The mulberry leaf fermentation composition according to claim 1, which helps reverse insulin resistance, is characterized in that, The mulberry leaf extract is prepared by moistening mulberry leaves, steam sterilizing them, and inoculating them with *Bacillus subtilis* (a type of bacteria). Eurotium cristatum The microbial strain is prepared by solid-state fermentation for 4 to 6 days. After fermentation, citric acid is added, followed by ultrasonic extraction with ethanol. The extract is then filtered, concentrated, and dried to obtain the final product.

3. The mulberry leaf fermentation composition according to claim 2, which helps reverse insulin resistance, is characterized in that, The viable count of the *Gynostemma pentaphyllum* was (7–9) × 10⁻⁶. 8 CFU / g; the glucose-inhibiting bacteria ( Eurotium cristatum It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on July 3, 2025, with accession number GDMCC: No. 66629.

4. The mulberry leaf fermentation composition according to claim 1, which helps reverse insulin resistance, is characterized in that, The preparation method of the barley flavonoid extract is as follows: healthy and intact barley grains are soaked at 22-28℃ for 24 hours, sterilized by steam, and inoculated with anti-glycemic bacteria (…). Streptomyces sp . The bacterial strain was fermented in solid state for 70-75 hours. After fermentation, the strain was pulverized and 80-88% ethanol aqueous solution was added at a material-to-liquid ratio of 1:(10-15). The mixture was then refluxed at 70-80℃ for 110-130 minutes. The reflux extraction was repeated 2-3 times. The extracts were combined, centrifuged, filtered to remove residue, and the filtrate was concentrated under reduced pressure, finely filtered, ultrafiltered, and enriched using a macroporous resin column. After washing with water, the filtrate was eluted with 70-75% ethanol. The eluent was concentrated and freeze-dried to obtain barley flavonoid extract. The preparation method of the barley polysaccharide extract was as follows: barley was pulverized and refluxed with ethanol. The residue was collected by filtration and extracted with distilled water 1-2 times. All supernatants were combined and the extracts were concentrated under reduced pressure, precipitated with alcohol, dialyzed, and freeze-dried to obtain the final product.

5. The mulberry leaf fermentation composition according to claim 4 that helps reverse insulin resistance, characterized in that, The viable count of the anti-saccharide bacteria is (5-8) × 10⁻⁶. 8 CFU / g; the anti-saccharide bacteria ( Streptomyces sp . It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on March 26, 2025, with accession number GDMCC: No. 66062.

6. The mulberry leaf fermentation composition according to claim 1, characterized in that, The extraction method of the bitter melon extract is as follows: after crushing the bitter melon, add water, then add a compound enzyme for enzymatic hydrolysis, centrifuge, take the supernatant for alcohol precipitation, centrifuge to collect the precipitate and filtrate, the filtrate is adsorbed and eluted by LSA-20 resin, concentrated under reduced pressure, and freeze-dried to obtain powder, and the powder and precipitate are mixed to obtain the final product.

7. The mulberry leaf fermentation composition according to claim 1, which helps reverse insulin resistance, is characterized in that, The fermentation microbial preparation is composed of *Lactobacillus plantarum*, *Lactobacillus acidophilus*, and *Saccharomyces cerevisiae* in a mass ratio of (0.5–1):(0.2–0.6):1; the viable count of *Lactobacillus plantarum* is (4–7) × 10⁻⁶. 8 CFU / g, viable count of Lactobacillus acidophilus (1-4) × 10⁻⁶ 8 The CFU / g and viable count of brewer's yeast were (2-5) × 10⁻⁶. 8 CFU / g.

8. The mulberry leaf fermentation composition according to claim 1, characterized in that, The water-soluble dietary fiber is selected from one of apple pectin, grapefruit peel pectin, and blueberry pectin; the dosage form of the mulberry leaf fermentation composition is powder, granules, tablets, or capsules.

9. A method for preparing a mulberry leaf fermentation composition as described in any one of claims 1 to 8 that helps reverse insulin resistance, characterized in that, Includes the following steps: (1) Mix mulberry leaf extract, highland barley extract, bitter melon extract, hawthorn powder, yam powder, lemon powder, fructooligosaccharide, water-soluble dietary fiber and resistant dextrin, sterilize, and obtain the material to be fermented; (2) Inoculate the fermentation microbial preparation into the material to be fermented in step (1), and ferment at 30-37°C for 3-7 days to obtain the fermentation product; (3) The fermentation product of step (2) is inactivated and dried to obtain the mulberry leaf fermentation composition.

10. The preparation method according to claim 9, characterized in that, In step (2), during the fermentation process, a resonant magnetic field with a frequency of 10kHz to 30kHz and a magnetic induction intensity of 0.5mT to 3mT is used for 10 to 20 minutes every day.

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