A compound spleen-strengthening and dampness-removing white hyacinth bean paste, its preparation method and application

Abstract

This invention relates to the field of deep processing technology of white hyacinth bean, specifically to a method for preparing a compound spleen-strengthening and dampness-removing white hyacinth bean paste. This invention utilizes low-temperature roasting to pretreat a specific ratio of dampness-removing medicinal herbs, including licorice, poria cocos, coix seed, and bitter almond, and prepares a compound decoction rich in dampness-removing components. This decoction is then used as a biological stressor to promote the germination of white hyacinth beans, with its concentration controlled to encourage moderate germination and the production of substances that resist adverse conditions. Simultaneously, leveraging the plant's water absorption capacity during germination, the invention incorporates the dampness-removing components of the compound decoction. Stable solid-state fermentation with pure Rhizopus further enhances the dampness-removing components of the compound white hyacinth bean and fermented soybean. This not only increases the content of major dampness-removing components such as polyphenols, proteins, and polysaccharides in the white hyacinth bean and fermented soybean, but also increases the content of trace dampness-removing substances such as rutin, quercetin, kaempferol, luteolin, and picolinic acid. It also promotes the dissolution of dampness-removing amino acids—alanine and glycine. The dampness-removing compound white hyacinth bean and fermented soybean components are clearly defined, and the content of spleen-strengthening and dampness-removing substances is richer.

Description

Technical fields:

[0001] This invention relates to the field of deep processing technology of white hyacinth bean, specifically to a method for preparing a compound spleen-strengthening and dampness-removing white hyacinth bean paste. Background technology:

[0002] Spleen deficiency and dampness accumulation is a complex metabolic syndrome that severely hinders normal metabolism and immune regulation. It generally requires multi-target treatment with compound traditional Chinese medicine formulas targeting water metabolism, immunity, and the gastrointestinal tract. Currently, spleen-strengthening and dampness-removing formulas mainly consist of single-herb foods and compound formulas containing different proportions of herbs, such as decoctions, tea beverages, granules, thick ointments, and lactic acid bacteria fermented beverages. White hyacinth bean, a commonly used herb for removing dampness, has diuretic, dampness-removing, spleen-strengthening, and stomach-nourishing effects. It is often used as an adjuvant in dampness-removing formulas to strengthen the spleen and remove dampness, and is also a main ingredient in dampness-removing bread, meal replacement powders, tea beverages, enzymes, wine, yogurt, or other dampness-removing compound formulas.

[0003] Currently developed products have drawbacks such as simple mixing and compounding, crude processing, and low added value. Some of the deep processing technologies used mainly involve liquid lactic acid bacteria or yeast fermentation, and only one or two major categories of dampness-removing components, such as polysaccharides, are tested for dissolution. The dampness-removing components are not clearly defined, and the extraction rate of various components in liquid is still relatively low, resulting in extremely low utilization rate of dampness-removing medicinal materials and no significant synergistic effect of strengthening the spleen and removing dampness.

[0004] Modern pharmacology shows that, in addition to major functional components such as proteins, polysaccharides, and polyphenols, some trace components are often overlooked in spleen-strengthening and dampness-removing formulas. These include pachymic acid, glycyrrhizic acid, coixol, luteolin, kaempferol, rutin, oleanolic acid, glycine, and alanine. These are the material basis for the synergistic dampness-removing effects of compound products. Currently, dampness-removing technology still has shortcomings in the research and utilization of components, but this will provide a reference for the effective substances in dampness-removing compound products for further processing.

[0005] Germination, fermentation, and compounding are common processing methods for enhancing the efficacy of traditional Chinese medicine. Bean sprouts and fermented black soybeans generally have higher overall material utilization and added value than unprocessed raw materials and their decoctions. However, research on germination technology mainly focuses on enhancing GABA in legumes and grains such as black beans, mung beans, highland barley, and oats; fermentation technology mainly focuses on enhancing efficacy through liquid fermentation with acetic acid bacteria, yeast, and lactic acid bacteria; while research on Rhizopus solid-state fermentation technology mainly focuses on improving the flavor of green tea, mulberry leaf tea, yeast, and thrombolytic fermented black soybeans. Compound-enhanced fermented black soybeans such as ginseng and mulberry leaf are already available on the market.

[0006] Sprouting and fermentation are common methods in bean processing. Bean sprouts and fermented black beans have relatively higher overall material utilization and added value than white lentils and broth. However, in industrial production, bean sprouting has a long cycle and low germination rate. Within the same batch, sprouts can be too short or too long, resulting in inconsistent nutritional quality. Ultimately, over-germination and microbial spoilage can cause significant losses to bean components. Compound processing is one method to enhance the efficiency of bean processing. This involves simple soaking and absorption of liquid additives (CN114424815 A) or mixing powdered additives onto the bean skin for fermentation (CN 107927583A). This method has a long production cycle, is prone to spoilage, and results in insufficient absorption or coating of additives, leading to resource waste and uneven fermentation. Furthermore, traditional bean fermentation processes involve mixed natural fermentation of multiple microorganisms, making it difficult to precisely control the fermentation metabolism process, resulting in inconsistent quality and efficacy between different batches of final products.

[0007] Therefore, it is necessary to improve the utilization rate of auxiliary materials, compound herbal formulas for removing dampness, control the degree of germination of beans, optimize the pure-culture fermentation conditions to enrich substances with dampness-removing effects, innovate the human body utilization form of dampness-removing formulas, and process white hyacinth beans to enhance the spleen-strengthening and dampness-removing effects. Summary of the Invention:

[0008] To solve the above-mentioned technical problems, the present invention will soak white hyacinth beans in a decoction of a decoction for removing dampness to promote germination, and then ferment them to prepare white hyacinth bean paste, thereby obtaining a health food with the function of strengthening the spleen and removing dampness.

[0009] One of the technical solutions provided by this invention is a method for preparing white hyacinth bean paste, as detailed below:

[0010] S1 Preparation of the decoction for removing dampness: Take 50-100 parts of Poria cocos, 50-100 parts of Coix lacryma-jobi, 15-60 parts of Glycyrrhiza uralensis, and 20-65 parts of Prunus armeniaca. Bake the herbs in an oven at 120-160℃ for 15-30 minutes. Then add 2000-4000 parts of water and soak for 15-60 minutes. Boil over high heat for 30-60 minutes. Filter the residue. The filtrate is the decoction for removing dampness.

[0011] Preferably, the filtrate is concentrated into a decoction of the dehumidifying compound at a concentration of 0.1–0.5 g / mL;

[0012] S2 Soaking Treatment: Remove impurities, select plump, moderately sized, and disease-free white hyacinth beans, add 1 to 2 times the weight of the white hyacinth beans in a decoction of a dehumidifying compound, and soak the white hyacinth beans at 25 to 30°C for 6 to 12 hours.

[0013] Preferably, the white hyacinth bean is the Yunnan black-browed white hyacinth bean;

[0014] Preferably, the concentration of the decoction of the dampness-removing compound is 0.1-0.2 g / mL;

[0015] S3 staged germination: Place the soaked white hyacinth beans in a glass dish, lay a damp cloth on the glass dish containing the white hyacinth beans and place it in an incubator. Control the germination temperature in the incubator to 20-30℃ and the humidity to 70-90%. After germination for 6-24 hours, replace the damp cloth and add 0.5-2 times the weight of the white hyacinth bean raw material in decoction of dehumidifying compound to soak the white hyacinth beans. Continue germination for 18-30 hours.

[0016] Preferably, the concentration of the decoction of the dampness-removing compound is 0.3-0.5 g / mL;

[0017] S4 High Temperature Sterilization: Sterilize the sprouted white hyacinth beans and the remaining dehumidifying soup at 121℃ for 15 minutes, then let them cool.

[0018] S5 Inoculation Fermentation: Add 5% to 20% (v / w) of Rhizopus seed liquid to the bean sprouts, shake well, spread the beans evenly in a bamboo sieve with a hole diameter of 1 to 3 mm, and ferment in an open incubator at 26 to 36℃ for 4 to 7 days, maintaining a humidity of 50 to 80%. Stir the fermented soybeans every other day to ensure good ventilation and uniform fermentation.

[0019] Further, the method for preparing the Rhizopus seed liquid is as follows: Rhizopus is inoculated into potato dextrose medium for expansion culture, and then cultured at a constant temperature of 30°C for 5 days; preferably, the Rhizopus is Rhizopus microsporus DU-106, with the preservation number GDMCC 60888.

[0020] Preferably, in S1, the ratio of the dampness-removing herbs is: 100 parts Poria cocos, 100 parts Coix lacryma-jobi, 35 parts Glycyrrhiza uralensis, and 40 parts Prunus armeniaca. They are baked at 140℃ for 20 minutes, soaked in 3000 parts water for 20 minutes, and decocted over high heat for 60 minutes.

[0021] Preferably, in S2, the volume ratio of the soaking dehumidifying decoction is 1, the concentration is 0.2 g / mL, and the soaking time is 8 hours;

[0022] Preferably, in S3, the germination temperature is controlled at 26℃, the moisture content at 80%, the first germination time is 12h, and the volume ratio of the dehumidifying decoction added for the second germination is 1, the concentration is 0.5g / mL, and the germination time is 24h.

[0023] Preferably, in S5, the fermentation inoculum amount is 15% (v / w), the sieve aperture is 2mm, the fermentation temperature is 30℃, the fermentation humidity is 75%, and the fermentation time is 6d.

[0024] The second technical solution provided by the present invention is white hyacinth bean paste prepared by the method described in the first technical solution.

[0025] The third technical solution provided by this invention is the application of the white hyacinth bean curd described in the second technical solution, particularly its application in the preparation of food, health products or medicines with spleen-strengthening and dampness-removing effects.

[0026] Beneficial effects:

[0027] This invention utilizes low-temperature roasting to pretreat a specific ratio of dampness-removing medicinal herbs, including licorice, poria cocos, coix seed, and bitter almond, and prepares them into a compound decoction with a special aroma, color, and rich dampness-removing components. This decoction is then used as a biological stress agent to promote the germination of white hyacinth beans. By controlling its concentration, the decoction promotes moderate germination of white hyacinth beans. It not only controls the degree of germination but also promotes the production of adversity-resistant substances such as polyphenols, GABA, and free amino acids in white hyacinth beans under stress. Meanwhile, by utilizing the plant's ability to absorb water during germination, the dehumidifying components of the compound decoction for removing dampness are combined and then further enhanced through stable pure-culture Rhizopus solid-state fermentation. This not only increases the content of major dehumidifying components such as polyphenols, proteins, and polysaccharides in white hyacinth bean and fermented soybean, but also increases the content of trace dehumidifying substances such as rutin, quercetin, kaempferol, luteolin, and picolinic acid. At the same time, it promotes the dissolution of alanine and glycine, amino acids with dehumidifying effects. The dehumidifying compound white hyacinth bean and fermented soybean components are clearly defined, and the content of substances with spleen-strengthening and dehumidifying effects is richer.

[0028] This invention evaluates the efficacy of the compound white hyacinth bean and fermented soybean product described herein using a reserpine-induced spleen deficiency model. Results show that the compound white hyacinth bean and fermented soybean product of this invention can significantly reduce fecal water content in spleen-deficient rats, promote gastric emptying and urinary D-xylose excretion, inhibit the release of the inflammatory factor TNF-α in the spleen, and increase serum MTL, thus exhibiting spleen-strengthening, stomach-nourishing, and dampness-removing effects. This application can improve reserpine-induced spleen deficiency rats with loose stools, gastrointestinal dysfunction, spleen inflammation, and intestinal hormone imbalances, exhibiting spleen-strengthening, stomach-nourishing, and dampness-removing effects, and can be developed as a health food for strengthening the spleen and removing dampness. Attached image description:

[0029] Figure 1 A bar graph showing the changes in fecal water content in each group of rats;

[0030] Figure 2 A bar chart showing the changes in gastric emptying rate in each group of rats;

[0031] Figure 3 A bar chart showing the changes in urinary D-xylose content in each group of rats;

[0032] Figure 4 The TNF-α content in the spleen tissue homogenate of rats in each group;

[0033] Figure 5 The serum MTL content of rats in each group;

[0034] Note: In the above figures, the model group showed a significant difference compared to the blank group (p < 0.001); the drug group showed significant differences compared to the model group (p < 0.05), (p < 0.01), (p < 0.01), (p < 0.001). Detailed implementation method:

[0035] The present invention is described below through specific embodiments. These embodiments should be understood as illustrative, not limiting, of the scope of the invention, which is defined solely by the claims. For those skilled in the art, various changes or modifications to the material composition and dosage in these embodiments, without departing from the spirit and scope of the invention, also fall within the scope of protection of the present invention.

[0036] The Rhizopus described in this invention and its embodiments is Rhizopus microsporus DU-106, with accession number GDMCC 60888.

[0037] The present invention will be further explained and described below through specific embodiments.

[0038] Example 1

[0039] S1 Preparation of the decoction for removing dampness: Take 100 parts of Poria cocos, 100 parts of Coix lacryma-jobi, 35 parts of Glycyrrhiza uralensis, and 40 parts of Prunus armeniaca. Bake the herbs in an oven at 140℃ for 20 minutes. Add 3000 parts of water and soak for 20 minutes. Boil over high heat for 60 minutes, then filter to remove the residue. Concentrate the decoction to 0.2 g / mL and 0.5 g / mL.

[0040] Example 2

[0041] S1 Preparation of the decoction for removing dampness: Take 100 parts of Poria cocos, 100 parts of Coix lacryma-jobi, 35 parts of Glycyrrhiza uralensis, and 40 parts of Prunus armeniaca. Bake the herbs in an oven at 140℃ for 20 minutes. Add 3000 parts of water and soak for 20 minutes. Boil over high heat for 60 minutes, then filter to remove the residue. Concentrate the decoction to 0.2 g / mL and 0.5 g / mL.

[0042] S2 Soaking: Remove impurities and select plump, moderately sized, and disease-free Yunnan black-browed white hyacinth beans. Add a 0.2g / mL concentration of decoction of a dampness-removing compound to the white hyacinth beans at 1 times their weight in water, and soak the white hyacinth beans at 26℃ for 8 hours to allow them to fully absorb the nutrients.

[0043] S3 staged biological stress germination: Lay a damp cloth on a glass dish containing white hyacinth beans, control the germination temperature in the incubator at 26℃ and the humidity at 80%. After germination for 12 hours, replace the damp cloth and add a decoction of a dampness-removing compound with a concentration of 0.5g / mL, which is 1 times the weight of the white hyacinth bean raw material, to soak the white hyacinth beans. Continue germination for 24 hours.

[0044] Example 3

[0045] S1 Preparation of the decoction for removing dampness: Take 100 parts of Poria cocos, 100 parts of Coix lacryma-jobi, 35 parts of Glycyrrhiza uralensis, and 40 parts of Prunus armeniaca. Bake the herbs in an oven at 140℃ for 20 minutes. Add 3000 parts of water and soak for 20 minutes. Boil over high heat for 60 minutes, then filter to remove the residue. Concentrate the decoction to 0.2 g / mL and 0.5 g / mL.

[0046] S2 Soaking: Remove impurities and select plump, moderately sized, and disease-free Yunnan black-browed white hyacinth beans. Add a decoction of a dampness-removing compound with a concentration of 0.2 g / mL, equal in volume to the weight of the white hyacinth beans, and soak the white hyacinth beans at 26℃ for 8 hours to allow them to fully absorb the nutrients.

[0047] S3 staged biological stress germination: Lay a damp cloth on a glass dish containing white hyacinth beans, control the germination temperature in the incubator at 26℃ and the humidity at 80%. After germination for 12 hours, replace the damp cloth and add a decoction of a dampness-removing compound with a concentration of 0.5g / mL, which is 1 times the weight of the white hyacinth bean raw material, to soak the white hyacinth beans. Continue germination for 24 hours.

[0048] S4 High-Temperature Sterilization: White hyacinth bean sprouts and residual dampness-removing soup are sterilized at 121℃ for 15 minutes and then cooled.

[0049] S5 Inoculation and Fermentation: Rhizopus microsporus DU-106 was inoculated into potato dextrose medium for expansion culture and cultured at 30℃ for 5 days. In a clean bench, 15% (v / v) Rhizopus seed culture was added to the sterilized bean sprouts, shaken well, and the beans were spread evenly on a bamboo sieve with 2mm aperture. The mixture was then fermented in an open incubator at 30℃ for 6 days, maintaining 75% humidity. The fermented soybeans were stirred every other day to ensure good ventilation and uniform fermentation.

[0050] Example 4

[0051] S1 Preparation of the decoction of the dampness-removing compound: Take 80 parts of Poria cocos, 80 parts of Coix lacryma-jobi, 60 parts of Glycyrrhiza uralensis, and 65 parts of Prunus armeniaca. Bake the dampness-removing herbs in an oven at 150℃ for 15 minutes. Then add 3000 parts of water and soak for 20 minutes. Boil over high heat for 60 minutes. Filter the residue and concentrate the decoction of the dampness-removing compound to 0.2 g / mL and 0.5 g / mL.

[0052] S2 Soaking: Remove impurities and select plump, moderately sized, and disease-free Yunnan black-browed white hyacinth beans. Add a 0.2g / mL concentration of decoction of a dampness-removing compound to the white hyacinth beans at 1 times their weight and soak the white hyacinth beans at 26℃ for 12 hours.

[0053] S3 Germination: Lay a damp cloth flat on a glass dish containing white hyacinth beans, control the germination temperature in the incubator at 28℃ and the humidity at 90%. After germination for 6 hours, replace the damp cloth and add 0.5g / mL of dehumidifying compound decoction, which is 1 times the weight of the white hyacinth bean raw material. Continue germination for 30 hours.

[0054] S4 High-Temperature Sterilization: White hyacinth bean sprouts and residual dampness-removing soup are sterilized at 121℃ for 15 minutes and then cooled.

[0055] S5 Inoculation and Fermentation: Rhizopus microsporus DU-106 was inoculated into potato dextrose medium for expansion culture and cultured at 30℃ for 5 days. 10% (v / w) Rhizopus seed culture was added to the bean sprouts in a clean bench, shaken well, and the beans were spread evenly in a bamboo sieve with 1mm aperture. The mixture was then fermented in an open incubator at 34℃ for 4 days, maintaining 70% humidity. The fermented soybeans were stirred every other day to ensure good ventilation and uniform fermentation.

[0056] Comparative Example 1

[0057] S1 Soaking: Remove impurities and select plump, medium-sized, and disease-free Yunnan black-brown white hyacinth beans. Add water equal to the weight of the white hyacinth beans and soak them at 26℃ for 8 hours to allow them to fully absorb the nutrients.

[0058] S2 germination: Lay a damp cloth on a glass dish containing white hyacinth beans, control the germination temperature in the incubator at 26℃ and the humidity at 80%. After 12 hours of germination, replace the damp cloth, add water equal to the weight of the white hyacinth beans to soak them, and continue germination for 24 hours.

[0059] S3 High-Temperature Sterilization: White bean sprouts and residual water are sterilized at 121℃ for 15 minutes and then cooled.

[0060] S4 Inoculation and Fermentation: Rhizopus microsporus DU-106 was inoculated into potato dextrose medium for expansion culture and cultured at 30℃ for 5 days. In a clean bench, 15% (v / w) Rhizopus seed culture was added to the bean sprouts, shaken well, and the beans were spread evenly in a bamboo sieve with 2mm aperture. The mixture was then fermented in an open incubator at 30℃ for 6 days, maintaining 75% humidity. The fermented soybeans were stirred every other day to ensure good ventilation and uniform fermentation.

[0061] Comparative Example 2

[0062] S1 Preparation of a decoction of a traditional Chinese medicine formula for dispelling dampness: Take 100 parts Poria cocos, 100 parts Coix lacryma-jobi, 35 parts Glycyrrhiza uralensis, and 40 parts Prunus armeniaca. Bake the herbs in an oven at 140℃ for 20 minutes. Add 3000 parts water and soak for 20 minutes. Boil over high heat for 60 minutes, then filter to remove the residue. Concentrate the decoction to 0.2 g / mL.

[0063] S2 Soaking: Remove impurities and select plump, moderately sized, and disease-free Yunnan black-browed white hyacinth beans. Add a decoction of a dampness-removing compound with a concentration of 0.2 g / mL, equal in volume to the weight of the white hyacinth beans, and soak the white hyacinth beans at 26℃ for 8 hours to allow them to fully absorb the nutrients.

[0064] S3 High-Temperature Sterilization: White hyacinth bean sprouts and residual dampness-removing soup are sterilized at 121℃ for 15 minutes and then cooled.

[0065] S4 Inoculation and Fermentation: Rhizopus microsporus DU-106 was inoculated into potato dextrose medium for expansion culture and cultured at 30℃ for 5 days. In a clean bench, 15% (v / w) Rhizopus seed culture was added to the bean sprouts, shaken well, and the beans were spread evenly in a bamboo sieve with 2mm aperture. The mixture was then fermented in an open incubator at 30℃ for 6 days, maintaining 75% humidity. The fermented soybeans were stirred every other day to ensure good ventilation and uniform fermentation.

[0066] Comparative Example 3

[0067] S1 Preparation of the decoction for removing dampness: Take 100 parts Poria cocos, 100 parts Coix lacryma-jobi, 35 parts Glycyrrhiza uralensis, and 40 parts Prunus armeniaca. Bake the herbs in an oven at 140℃ for 20 minutes. Add 3000 parts water and soak for 20 minutes. Boil over high heat for 60 minutes, then filter to remove the residue. Concentrate the decoction to 0.2 g / mL.

[0068] S2 Soaking: Remove impurities and select plump, moderately sized, and disease-free Yunnan black-browed white hyacinth beans. Add a 0.2g / mL concentration of decoction of a dampness-removing compound, equal in volume to the weight of the white hyacinth beans, and soak the white hyacinth beans at 26℃ for 8 hours to allow them to fully absorb the nutrients.

[0069] S3 Stress Germination: Lay a damp cloth on a glass dish containing white hyacinth beans, control the germination temperature in the incubator at 26℃ and the humidity at 80%. After germination for 12 hours, replace the damp cloth, add water equal to the weight of the white hyacinth beans to soak them, and continue germination for 24 hours.

[0070] S4 High-Temperature Sterilization: White hyacinth bean sprouts and residual dampness-removing soup are sterilized at 121℃ for 15 minutes and then cooled.

[0071] S5 Inoculation and Fermentation: Rhizopus microsporus DU-106 was inoculated into potato dextrose medium for expansion culture and cultured at 30℃ for 5 days. In a clean bench, 15% (v / w) Rhizopus seed culture was added to the bean sprouts, shaken well, and the beans were spread evenly in a bamboo sieve with 2mm aperture. The mixture was then fermented in an open incubator at 30℃ for 6 days, maintaining 75% humidity. The fermented soybeans were stirred every other day to ensure good ventilation and uniform fermentation.

[0072] Comparative Example 4

[0073] S1 Preparation of the decoction for removing dampness: Take 100 parts Poria cocos, 100 parts Coix lacryma-jobi, 35 parts Glycyrrhiza uralensis, and 40 parts Prunus armeniaca. Bake the herbs in an oven at 140℃ for 20 minutes. Add 3000 parts water and soak for 20 minutes. Boil over high heat for 60 minutes, then filter to remove the residue. Concentrate the decoction to 0.5 g / mL.

[0074] S2 Soaking: Remove impurities and select plump, moderately sized, and disease-free Yunnan black-brown white hyacinth beans. Add water equal to the weight of the white hyacinth beans and soak at 26℃ for 8 hours to allow them to fully absorb the nutrients.

[0075] S3 Stress Germination: Lay a damp cloth flat on a glass dish containing white hyacinth beans, control the germination temperature in the incubator at 26℃ and the humidity at 80%. After germination for 12 hours, replace the damp cloth and add a decoction of a dampness-removing compound with a concentration of 0.5g / mL equal to the weight of the white hyacinth beans to soak the beans. Continue germination for 24 hours.

[0076] S4 High-Temperature Sterilization: White hyacinth bean sprouts and residual dampness-removing soup are sterilized at 121℃ for 15 minutes and then cooled.

[0077] S5 Inoculation and Fermentation: Rhizopus microsporus DU-106 was inoculated into potato dextrose medium for expansion culture and cultured at 30℃ for 5 days. In a clean bench, 15% (v / w) Rhizopus seed culture was added to the bean sprouts, shaken well, and the beans were spread evenly in a bamboo sieve with 2mm aperture. The mixture was then fermented in an open incubator at 30℃ for 6 days, maintaining 75% humidity. The fermented soybeans were stirred every other day to ensure good ventilation and uniform fermentation.

[0078] Experiment Example 1 Germination Rate Determination

[0079] 1.1 Experimental Methods

[0080] After germination was completed in each embodiment and comparative example, germinated white hyacinth beans were selected based on the bud tip breaking through the seed coat, and the germination rate was calculated. The formula for calculating the germination rate of white hyacinth beans is: (Number of germinated seeds / Total number of seeds tested) × 100%.

[0081] 1.2 Experimental Results

[0082] In nature, there is a biological stress effect between organisms. Plants can promote or inhibit the germination and growth of competing plants by releasing their own secondary metabolites. As the concentration of secondary metabolites increases, there is a trend of first promoting and then inhibiting. This not only controls the degree of germination and growth of competing plants, but also enables plants under stress to produce substances that resist adverse conditions, such as polyphenols, GABA, and free amino acids.

[0083] The compound decoction made from licorice, poria cocos, coix seed, and bitter almond is rich in nutrients and allelopathic stress substances, including glycosides, organic acids, straight-chain alcohols, fatty acids, naphthoquinones, anthraquinones, polyphenols, flavonoids, steroids, terpenes, amino acids, and polypeptides. These substances can promote plant germination and growth at low concentrations, but inhibit cell division and water and ion absorption and transport at high concentrations. Therefore, controlling the concentration of the compound decoction to induce staged stress germination in white hyacinth beans not only promotes moderate germination and the release of active substances, but also increases the absorption of the compound decoction.

[0084] The results are shown in Table 1. The germination rate of Example 3 was significantly higher than that of Comparative Examples 1, 3, and 4. In Example 3, the white hyacinth beans were first treated with 0.2 g / mL of a dehumidifying decoction to promote germination, and then further stressed with 0.5 g / mL of the same dehumidifying decoction, resulting in a higher germination rate than Comparative Example 1, which germinated in plain water. In Comparative Example 3, the germination rate was higher than that of Comparative Example 1, which germinated in plain water, due to the diffusion of nutrients from the dehumidifying decoction and the white hyacinth beans from the high-osmotic to low-osmotic water. However, in the later stages of germination with plain water, the germination rate was lower than that of Example 3 because the white hyacinth beans contained nutrients from the dehumidifying decoction and their own nutrients diffused from the high-osmotic to the low-osmotic water. In Comparative Example 4, after germination in plain water, the beans were subjected to a 0.5 g / mL high-osmotic stress environment, which significantly inhibited germination and growth, resulting in a germination rate significantly lower than that of Example 3, Comparative Examples 1, and 3.

[0085] Table 1 Germination rate of white hyacinth bean in different embodiments and comparative examples

[0086]

[0087] Furthermore, network pharmacology indicates that phenolic substances are important components for strengthening the spleen and removing dampness. However, most phenolic substances in plants are bound to fatty acids, sugars, proteins, or other macromolecules through insoluble bonds, making them difficult for the human body to absorb and utilize. The pretreatment method for the white hyacinth bean curd in this application is stress germination, and different germination rates will have a certain promoting effect on the release of free phenolic substances and other secondary metabolites.

[0088] Experiment Example 2: Determination of Main Nutrients in the Product

[0089] 2.1 Experimental Methods

[0090] After the preparation of each embodiment and comparative example, the total phenol content of the samples was determined using the Folin-Ciocalteu method described by Wang Jianfei. The polysaccharide content was determined according to the phenol-sulfuric acid method of SN / T 4260-2015. The starch content was determined according to the acid hydrolysis method for food starch in GB5009.9-2016. The protein content was determined using the Dumas combustion method described by Han Bo. The cellulose content was determined according to the acid-base hydrolysis method described in GB / T 5515-2008. The ash content was determined according to the method shown in national standard GB 5009.4-2016.

[0091] 2.2 Experimental Results

[0092] This invention enhances the efficacy of legumes through compounding, germination, and fermentation. It utilizes the complex components of the raw materials to provide rich nutrition for germination and fermentation, and generates more active ingredients through the release, dissolution, and biotransformation of active enzyme systems. After processing, the protein, polysaccharides, polyphenols, and other dehumidifying components of white hyacinth beans change, resulting in higher absorption and utilization rates and dehumidifying effects in bean sprouts and fermented soybeans compared to unprocessed raw materials and their decoctions. Li Haiyang used silica gel column chromatography and mass spectrometry to separate and identify the chemical structures of white hyacinth bean components, and verified and compared the efficacy using a senna-induced diarrhea model in mice. The study showed that sweet amino acids such as glycine and alanine are effective components for strengthening the spleen and stopping diarrhea, consistent with the description in traditional Chinese medicine pharmacology that white hyacinth beans are sweet and warm in nature, and can tonify the spleen and stomach. Furthermore, the white hyacinth beans of this invention, after stress germination and fermentation, show a significant increase in the content of sweet amino acids. GABA is an important functional amino acid with sedative, hypoglycemic, hypotensive, antitumor, and brain function-improving effects. Although GABA is widely distributed in plants, its content is generally low. Plants only produce specific chemical substances to resist adversity when they are subjected to environmental stress and germination, and they generate large amounts of GABA by breaking down proteins.

[0093] After white hyacinth beans were germinated with different concentrations of compound dehumidifying decoction, different culture media were provided for the growth and fermentation of Rhizopus microsporus DU-106, resulting in different dehumidifying substances in the compound white hyacinth bean fermented soybean after fermentation.

[0094] Example 1 is a decoction of dampness-removing Chinese medicinal herbs, which mainly contains a large amount of water. Its active ingredients are polysaccharides, polyphenols and free amino acids, but the dissolution of active ingredients is low and the utilization rate of medicinal ingredients is low.

[0095] Example 2 uses 0.2 g / mL and 0.5 g / mL of a dehumidifying decoction to force white hyacinth beans to sprout in stages, which significantly enhances the polysaccharides, polyphenols, and functional amino acids such as alanine, glycine, and GABA in white hyacinth beans.

[0096] Examples 3 and 4, based on stress germination, further fermented the sprouts with Rhizopus microsporus DU-106. The staged stress germination using low and high concentrations of dehumidifying decoction resulted in the production of oligosaccharides and small peptides readily available for Rhizopus growth in the white hyacinth bean sprouts. The active lipases, amylases, cellulases, and glycosidases in Rhizopus microsporus DU-106 further promoted the dissolution of free phenols, polysaccharides, and free amino acids. Simultaneously, these carbohydrates were used to synthesize Rhizopus mycelial proteins. Therefore, the total phenol, polysaccharide, protein, glycine, alanine, and GABA content in Examples 3 and 4 was higher than in Examples 1, 2, and Comparative Examples 1-4.

[0097] Table 2. Differences in the content of polysaccharides, polyphenols, proteins, and free amino acids of white hyacinth bean in different embodiments and comparative examples.

[0098]

[0099] Experimental Example 3: Identification of Functional Substances

[0100] 3.1 Experimental Methods

[0101] After the processing of each embodiment and comparative example was completed, the determination of rutin, luteolin, kaempferol, quercetin, oleanolic acid, and coixol was performed according to the determination method described by Shang Shu. The samples were extracted with methanol and then determined by high performance liquid chromatography. The content of picolinic acid was determined according to the determination method of picolinic acid in white hyacinth bean described by Chen Dan. The samples were derivatized before column analysis and then determined by high performance liquid chromatography.

[0102] 3.2 Experimental Results

[0103] In nature, saponins, flavonoids, and polyphenols are mainly bound to fats, proteins, glycosides, and other substances through insoluble bonds, which hinders the dissolution of active substances in traditional Chinese medicine and their absorption by the human body. Flavonoid aglycones are part of the total flavonoid structure and belong to a subgroup of phenolic substances. Compared with conjugated flavonoids, they have higher bioavailability, which is conducive to the full and effective exertion of functional activities. Esters such as coixol, rutin, luteolin, kaempferol, and oleanolic acid, as well as free phenols and secondary saponins, are common substances in Poria cocos, licorice, bitter almond, coix seed, and white hyacinth bean. However, under natural conditions, flavonoid aglycones such as quercetin, kaempferol, and luteolin often combine with glycosides to form conjugated flavonoids such as quercetin, rutin, and luteolin, while oleanolic acid can combine with glycosides to form conjugated saponins. Picalinic acid is a unique spleen-strengthening and dampness-removing component of white hyacinth bean. It has the effects of strengthening the spleen and stopping diarrhea, moistening the intestines and promoting bowel movements, and regulating nerves. It is an important metabolite of lysine. Germination and fermentation help to release lysine into free amino acids and bioconvert them into picalinic acid.

[0104] During germination and Rhizopus fermentation, active metabolic enzymes are produced. On the one hand, these enzymes hydrolyze and destroy the cell walls of white hyacinth bean sprouts, promoting the dissolution of bound flavonoids, with a significant increase in the amount of rutin and other bound flavonoids dissolved. On the other hand, they also promote the biotransformation of some bound flavonoids. The germination and Rhizopus fermentation process activates various phenolic enzymes in the plant, such as rutin-degrading enzymes and phenylalanine ammonia-lyase. Rutin-degrading enzymes break down rutin into quercetin or quercetin, while phenolic enzymes promote the production of new phenolic substances such as kaempferol and luteolin. Therefore, after compound processing, the content of rutin and the new phenols produced by its degradation are significantly increased. This invention can release total phenols, total flavonoids, total saponins, esters, and polypeptides through germination and Rhizopus fermentation, and then biotransform them into free phenols, secondary flavonoids, secondary saponins, and esters. The absorption capacity and spleen-strengthening and dampness-removing effects of fermented black soybeans can be evaluated by measuring the content of secondary metabolites of total phenols, total flavonoids, amino acids, and esters, such as rutin, luteolin, kaempferol, quercetin, oleanolic acid, picolinic acid, and coixol.

[0105] The traditional processing method for Chinese medicinal herbs is decoction. Example 1 shows a decoction of dampness-removing herbs. The contents of rutin, luteolin, quercetin, kaempferol, oleanolic acid, and coixol in the decoction were all low, significantly lower than in other examples and comparative examples. Example 2 shows that after sprouting white hyacinth beans under stress with both low and high concentrations of the dampness-removing decoction from Example 1, the amount of dampness-removing trace substances in the white hyacinth beans was significantly increased. Example 3 shows a compound dampness-removing white hyacinth bean fermented with Rhizopus mold using white hyacinth bean sprouts from Example 2, further increasing the amount of dampness-removing trace substances.

[0106] Comparative Examples 3 and 4, respectively, used 0.2 g / mL and 0.5 g / mL of a dampness-removing decoction to force the white hyacinth beans to germinate. The resulting compound dampness-removing white hyacinth bean fermented soybeans had a much higher content of dampness-removing substances than the white hyacinth bean fermented soybeans sprouted in water in Comparative Example 1 and the white hyacinth bean fermented soybeans that absorbed the dampness-removing decoction in Comparative Example 2. Because Example 3 utilized 0.2 g / mL and 0.5 g / mL of the dampness-removing decoction to force germination in stages and Rhizopus fermentation, it greatly enriched the components of the dampness-removing decoction and the dampness-removing components of the white hyacinth bean itself. The contents of rutin, luteolin, quercetin, kaempferol, oleanolic acid, and coixol were much higher than those in Comparative Examples 3 and 4.

[0107] Table 3. Content of dampness-removing trace substances in white hyacinth bean from different embodiments and comparative examples. (μg / mL、μg / g)

[0108]

[0109]

[0110] Experiment Example 4: In vivo experiment

[0111] In summary, the efficacy was verified by comparing Examples 1-4 and Comparative Examples 1-4.

[0112] 4.1 Experimental animals

[0113] 120 SPF-grade male SD rats, 4 weeks old, weighing 120 - 150 g, were provided by the Guangdong Provincial Center for Laboratory Animals. License number: SCXK(Guangdong)2018 - 0002.

[0114] 4.2 Experimental methods

[0115] An experimental model of spleen-deficiency SD rats induced by intraperitoneal injection of reserpine was used. Two weeks of adaptive feeding was carried out before the formal experiment began. Two weeks later, the 120 rats were evenly divided into 10 groups according to body weight: blank group (K), spleen-deficiency group (L), group of fermented root of germinated white hyacinth bean with clear water by Rhizopus (Comparative Example 1), group of white hyacinth bean fermented with soaking compound Qushi Decoction (Comparative Example 2), group of germinated white hyacinth bean fermented with 0.2 g / mL compound Qushi Decoction (Comparative Example 3), group of germinated white hyacinth bean fermented with 0.5 g / mL compound Qushi Decoction (Comparative Example 4), Qushi Decoction group (Example 1), compound Qushi white hyacinth bean sprout group (Example 2), compound Qushi white hyacinth bean fermented group (Example 3), compound Qushi white hyacinth bean fermented group (Example 4).

[0116] In the blank group, normal saline was intraperitoneally injected and intragastrically administered. In the other groups, while the rats were intraperitoneally injected with 0.5 mg / kg reserpine to induce spleen deficiency, the corresponding drugs were intragastrically administered at a volume of 10 mL / kg (the decoction was concentrated into a concentrated solution containing 0.036 g / mL of dampness-eliminating medicinal materials, and the fermented soybean powder was prepared into a suspension of 0.036 g / mL). In the spleen-deficiency group, the drug was replaced with an equal volume of normal saline, and this continued for 16 days. After fasting for 12 h on the 14th day, 3% D-xylose (10 mL / kg) was intragastrically administered for the urinary D-xylose excretion experiment. Urine was collected 5 h later, and the content of D-xylose was determined by the phloroglucinol method to compare the water metabolism and small intestine absorption function of rats in each group. On the 15th day, feces were taken and the water content was determined by the direct drying method. On the 17th day, before dissection, 0.5 mL of phenol red was intragastrically administered to each rat, and the gastric emptying rate was measured after washing the stomach 30 min later.

[0117] The content of TNF-α in the spleen tissue homogenate was determined by enzyme-linked immunosorbent assay. 100 mg of spleen tissue was weighed and added to a centrifuge tube together with grinding beads. 1000 μL of normal saline was added, and it was ground frozen at -4 °C to make a homogenate. After that, it was centrifuged frozen at 4000 rpm for 15 min, and the supernatant was taken for determination. The content of motilin in the serum was determined by enzyme-linked immunosorbent assay.

[0118] 4.3 Experimental results

[0119] The spleen is an important site for immune responses, storing, producing, and purifying blood, and is closely related to the metabolism of nutrients and waste products in the body. Traditional Chinese medicine states that "the spleen governs transformation and transportation, and is the source of life and transformation." Symptoms of spleen deficiency include poor mental state, diarrhea, indigestion, edema, and so on. The mechanisms by which traditional Chinese medicine strengthens the spleen and eliminates dampness mainly include improving the spleen's immune function, regulating gastrointestinal hormones, regulating ion levels to adjust osmotic pressure, upregulating aquaporin expression, and regulating intestinal flora. These mechanisms help the spleen's function of transformation and transportation, gastrointestinal water metabolism, and thus promote urination, thereby exerting the effect of strengthening the spleen and eliminating dampness.

[0120] TNF-α is the earliest pro-inflammatory cytokine to appear in the inflammatory response, increasing vascular endothelial cell permeability and promoting the release of other cytokines. The TNF-α level in the spleen can reflect the spleen's immune function. Motilin is distributed in the small intestine and promotes the transport of water and electrolytes in the gastrointestinal tract, as well as strong gastric contractions and small intestinal catabolic movements. D-xylose is a pentose sugar; after oral administration, its absorption site is mainly in the upper jejunum. It is not metabolized by the liver and is mainly excreted through urine, reflecting the absorptive function of the small intestine. Motilin, urinary D-xylose content, gastric emptying rate, and fecal water content can reflect the gastrointestinal motility and digestive and absorptive functions of rats with spleen deficiency from both microscopic and macroscopic perspectives.

[0121] Experimental results are as follows Figure 1-5 As shown. Figure 1 A bar graph showing the changes in fecal water content in each group of rats; Figure 2 A bar chart showing the changes in gastric emptying rate in each group of rats; Figure 3 A bar chart showing the changes in urinary D-xylose content in each group of rats; Figure 4 The TNF-α content in the spleen tissue homogenate of rats in each group; Figure 5 The serum MTL content of rats in each group is shown.

[0122] Compared with the control group, the reserpine-induced spleen deficiency model group showed significantly increased levels of TNF-α, an inflammatory factor in the spleen, and significantly increased fecal water content, while significantly decreased levels of motilin (MTL), urinary D-xylose, and gastric emptying rate, indicating that the model group exhibited severe spleen deficiency diarrhea and gastrointestinal dysfunction symptoms.

[0123] Compared with the model group, each example and comparative example improved the symptoms of spleen deficiency and weak stomach to varying degrees. Among them, the compound spleen-strengthening and dampness-removing fermented soybean prepared in Example 3 had the strongest effect in improving spleen deficiency diarrhea and intestinal dysfunction. In Example 3, the spleen inflammatory factor (TNF-α) and fecal moisture of the spleen-deficient rats were significantly reduced, while the excretion of motilin, urinary D-xylose and gastric emptying were significantly increased. The function and state of the spleen and gastrointestinal tract were closest to the normal group, and it had a strong spleen-strengthening and dampness-removing effect. From the component determination of Example 3 in Experiment 2, it can be seen that the compound spleen-strengthening and dampness-removing fermented soybean was the richest in main components and trace components. This indicates that the compound spleen-strengthening and dampness-removing fermented soybean after sprouting under the stress of dampness-removing decoction and fermentation with Rhizopus microsporus DU-106 overcomes the shortcomings of traditional dampness-removing decoctions with low dissolution and traditional fermented soybean processing technology with simple and poor efficacy.

[0124] Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make various changes, modifications, substitutions and variations in form and detail to these embodiments without departing from the spirit and principles of the present invention. The scope of the present invention is defined by the claims and their equivalents.

Claims

1. A method for preparing white hyacinth bean paste, characterized in that, Specifically as follows: S1 Preparation of the decoction for removing dampness: Take 50-100 parts of Poria cocos, 50-100 parts of Coix lacryma-jobi, 15-60 parts of Glycyrrhiza uralensis, and 20-65 parts of Prunus armeniaca. Bake the herbs in an oven at 120-160℃ for 15-30 minutes. Then add 2000-4000 parts of water and soak for 15-60 minutes. Boil over high heat for 30-60 minutes. Filter the residue. The filtrate is the decoction for removing dampness. S2 Soaking Treatment: Remove impurities, select plump white hyacinth beans with moderate particle size and free from pests and diseases, add a decoction of a dampness-removing compound with a volume equal to the weight of the white hyacinth beans, and soak the white hyacinth beans at 25~30℃ for 8 hours; the concentration of the decoction of the dampness-removing compound is 0.2 g / mL. S3 Staged Germination: Soaked white hyacinth beans are placed in a glass dish, a damp cloth is laid on top of the dish, and the dish is placed in an incubator. The germination temperature and humidity are controlled at 26 ℃ and 80%, respectively. After germination for 12 hours, the damp cloth is replaced, and the white hyacinth beans are soaked in a decoction of a dehumidifying compound with a concentration of 0.5 g / mL. Germination continues for another 24 hours. S4 High-Temperature Sterilization: Sterilize the sprouted white hyacinth beans and the remaining dehumidifying soup at 121 ℃ for 15 min, then let cool. S5 Inoculation Fermentation: Add 15% of Rhizopus seed liquid (based on white hyacinth bean raw material) to the bean sprouts, shake well, spread the beans evenly in a bamboo sieve with a pore size of 2mm, and ferment in an open incubator at 30℃ for 6 days, maintaining a humidity of 75%. Stir the fermented soybeans every other day to ensure good ventilation and uniform fermentation.

2. The method for preparing white hyacinth bean paste as described in claim 1, characterized in that, In S1, the filtrate is concentrated to a decoction of the dehumidifying compound at a concentration of 0.1~0.5 g / mL.

3. The method for preparing white hyacinth bean paste as described in claim 1, characterized in that, The Rhizopus in S5 has the preservation number GDMCC 60888.

4. The method for preparing white hyacinth bean paste as described in claim 1, characterized in that, In S1, the ratio of herbs for removing dampness is: 100 parts Poria cocos, 100 parts Coix lacryma-jobi, 35 parts Glycyrrhiza uralensis, and 40 parts Prunus armeniaca. They are baked at 140℃ for 20 minutes, soaked in 3000 parts water for 20 minutes, and then decocted over high heat for 60 minutes.

5. White hyacinth bean paste prepared by the method according to any one of claims 1-4.

6. The application of the white hyacinth bean paste as described in claim 5, characterized in that, It is used in the preparation of food or health products with spleen-strengthening and dampness-removing effects.

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