Yixinshu granules and preparation method thereof

Abstract

The invention provides Yixinshu granules and a preparation method thereof, and belongs to the technical field of medicines. Ginseng, astragalus membranaceus, radix ophiopogonis, hawthorn, salvia miltiorrhiza, schisandra chinensis and ligusticum wallichii are subjected to immobilized enzyme enzymolysis and enzyme-assisted fermentation, a product is filtered, an ionic liquid solid-phase extraction agent is added, carbon dioxide is introduced, stirring adsorption, filtering, solid desorption and drying are conducted, extract is prepared, the extract, maltodextrin and stevioside are evenly mixed, granulation and drying are conducted, and the Yixinshu granules are prepared. The Yixinshu granules prepared by the invention are simple in preparation method, wide in raw material source, easy to realize industrial application, relatively low in cost and high in patient compliance, have the characteristics of quick dissolution, quick absorption, high bioavailability and the like, have the effects of tonifying qi and restoring pulse, promoting blood circulation to remove blood stasis and nourishing yin and promoting salivation, and have a wide application prospect.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to Yixinshu granules and a preparation method thereof. Background Art

[0002] Cardiovascular and cerebrovascular diseases are one of the most important factors affecting human health and causing death. According to literature reports, more than 20 million people die from cardiovascular and cerebrovascular diseases each year worldwide, and nearly 400,000 in my country. Traditional Chinese medicine has accumulated rich clinical experience in the diagnosis and treatment of this disease, and Chinese herbal prescriptions have unique advantages in the treatment of cardiovascular and cerebrovascular diseases. Chest pain and angina pectoris due to coronary heart disease belong to the category of "chest pain" in traditional Chinese medicine. The cause of chest pain is related to the invasion of cold evil, improper diet, emotional disorder, and physical weakness due to old age. According to the symptoms, it can be divided into heart blood stasis, phlegm turbidity obstruction, yin cold stagnation, heart and kidney yin deficiency, qi and yin deficiency, and yang qi deficiency. "Heart meridian obstruction" is the main pathological mechanism of this disease. Most of the clinical manifestations are mixed with deficiency and excess, showing symptoms of fundamental deficiency and superficial excess. The treatment of this disease advocates the principle of tonification as the main method and unblocking as the main method. Tonification mainly refers to tonifying Qi and Yin, aiming to regulate the excessive or insufficient Qi and blood in the internal organs to treat the symptoms of fundamental deficiency. Unblocking mainly refers to activating blood circulation and unblocking Yang, aiming to unblock the obstruction in the meridians to treat the symptoms of superficial excess.

[0003] Yixinshu is made by adding blood-activating drugs to the ancient prescription "Shengmai Yin", which treats both the symptoms and the root cause. "Shengmai Yin" is a commonly used basic prescription for treating chest pain. There are many reports on the clinical research of Shengmai Yin in coronary heart disease. It has significant efficacy, good effect, and is easy to take, and is deeply favored by patients.

[0004] Chinese invention patent application CN101804152A discloses a preparation method of Yixinshu tablets, which discloses 150-250g of ginseng, 150-250g of ophiopogon japonicus, 95-150g of schisandra chinensis, 150-250g of astragalus, 240-285g of salvia miltiorrhiza, 95-150g of chuanxiong, 150-250g of hawthorn, 20-60g of microcrystalline cellulose, 1-3g of magnesium stearate, and an appropriate amount of starch. Granulation is performed using 80% ethanol. During the operation, the higher the ethanol concentration, the lower the viscosity after wetting, resulting in a wide and uneven distribution range of the mesh number of the obtained wet granules, and more fine powder, resulting in the occurrence of flower pieces, cracks, etc. during tableting. The obtained Yixinshu tablets have a long disintegration time and will have unqualified disintegration time, poor stability, unqualified microorganisms, etc. after being stored for a period of time. Summary of the invention

[0005] The purpose of the present invention is to propose a Yixinshu granule and a preparation method thereof. The preparation method is simple, the raw material source is wide, and it is easy to realize industrial application, the cost is low, and the patient compliance is high. The drug has the characteristics of rapid dissolution, fast absorption, high bioavailability, etc., has the effects of invigorating qi and restoring pulse, promoting blood circulation and removing blood stasis, nourishing yin and promoting body fluid, and has broad application prospects.

[0006] The technical solution of the present invention is achieved in this way:

[0007] The invention provides a preparation method of Yixinshu granules. The method comprises the following steps: subjecting ginseng, astragalus, ophiopogon, hawthorn, salvia miltiorrhiza, schisandra chinensis and ligusticum chuanxiong to enzymolysis and enzyme-assisted fermentation with immobilized enzymes, filtering the products, adding an ionic liquid solid phase extractant, introducing carbon dioxide, stirring and adsorbing, filtering, solid desorption, drying to obtain an extract, uniformly mixing the extract with maltodextrin and stevioside, granulating, and drying to obtain Yixinshu granules.

[0008] As a further improvement of the present invention, the following steps are included:

[0009] S1. Enzymatic hydrolysis: Wash, dry and crush ginseng, astragalus, ophiopogon, hawthorn, salvia miltiorrhiza, schisandra chinensis and ligusticum chuanxiong to obtain Chinese medicine powder; add the immobilized enzyme to water, add the Chinese medicine powder, heat and stir for enzymatic hydrolysis to obtain a mixture;

[0010] S2. Enzyme-assisted fermentation: inoculating the mixture with the fermentation bacteria seed solution, enzyme-assisted fermentation, separating the immobilized enzyme with a magnet, filtering, and collecting the filtrate;

[0011] S3. Ionic liquid solid phase extractant adsorption: add an ionic liquid solid phase extractant to the filtrate, introduce carbon dioxide, stir for adsorption, filter, and collect solids;

[0012] S4. Desorption: adding the solid to an ethanol aqueous solution, heating and stirring for desorption, filtering, recovering ethanol from the filtrate, and drying to obtain an extract;

[0013] S5. Preparation of Yixinshu Granules: Evenly mix the extract, maltodextrin and stevioside, granulate and dry to obtain Yixinshu Granules.

[0014] As a further improvement of the present invention, the mass ratio of ginseng, astragalus, ophiopogon, hawthorn, salvia miltiorrhiza, schisandra chinensis, ligusticum chuanxiong and immobilized enzyme in step S1 is 10-20: 10-20: 10-20: 10-20: 15-25: 5-15: 5-15: 3-5, the temperature of the heated stirring enzymolysis is 25-35°C, the time is 2-4h, and the preparation method of the immobilized enzyme is as follows:

[0015] T1. Under the protection of inert gas, ferric chloride and ferrous chloride are dissolved in water, the pH value of the solution is adjusted, the reaction is heated and stirred, centrifuged, washed, dried, and calcined to obtain magnetic particles;

[0016] T2. 1-(3-aminopropyl) imidazole, 3-bromopropyltrimethoxysilane and a silane coupling agent were mixed and added to water, magnetic particles were added, stirred and mixed evenly, the pH value was adjusted, and heated for hydrolysis to obtain ionic liquid / magnetic composite particles;

[0017] T3. Add ionic liquid / magnetic composite particles into water, add NHS and EDC, stir to activate, add composite enzyme, stir to react, separate with a magnet, freeze-dry, and obtain immobilized enzyme.

[0018] As a further improvement of the present invention, the mass ratio of ferric chloride to ferrous chloride in step T1 is 3.24:1.26, the pH value of the adjusted solution is 9.5-10.5, the temperature of the heated and stirred reaction is 80-90°C, the time is 4-6h, the calcination temperature is 400-600°C, and the time is 2-4h; the mass ratio of 1-(3-aminopropyl)imidazole, 3-bromopropyltrimethoxysilane, silane coupling agent, and magnetic particles in step T2 is 5-7:3-6:2-4:15-20, the adjusted pH value is 9-10, and the The temperature of heating hydrolysis is 50-60°C, the time is 5-7h, the silane coupling agent is selected from at least one of KH550, KH602, and KH792; the mass ratio of the ionic liquid / magnetic composite particles, NHS, EDC and composite enzyme in step T3 is 12-15:2-4:2-4:4-7, the composite enzyme is selected from at least one of cellulase, pectinase, neutral protease, ficin, and bromelain, preferably, cellulase and neutral protease, the mass ratio is 10:3-5, and the stirring reaction time is 8-10h.

[0019] As a further improvement of the present invention, the fermentation bacteria in step S2 are Bacillus subtilis and Lactobacillus plantarum, and the bacterial content of the bacterial seed liquid is 10 8 -10 9 cfu / mL, the inoculation amounts were 1-2v / v% and 0.5-1.5v / v%, respectively, and the enzyme-assisted fermentation conditions were 25-30°C, 150-250r / min, and the enzyme-assisted fermentation was 36-48h.

[0020] As a further improvement of the present invention, the mass ratio of the filtrate and the ionic liquid solid phase extractant in step S3 is 100:10-13, the ventilation rate of the carbon dioxide is 15-25 mL / min, the stirring adsorption time is 1-2 h, and the preparation method of the ionic liquid solid phase extractant is as follows:

[0021] U1. adding graphene oxide to dichloromethane, adding thionyl chloride, stirring the reaction, and removing the solvent and excess thionyl chloride under reduced pressure to obtain chlorinated graphene oxide;

[0022] U2. adding chlorinated graphene oxide to acetonitrile, adding acryl alcohol and a base, heating and stirring to react, filtering, washing, and drying to obtain olefinic graphene oxide;

[0023] U3. Adding nano-silica to water, adding olefinic graphene oxide, stirring and dispersing evenly, spray drying, and obtaining wrinkled olefinic graphene oxide-coated nano-silica;

[0024] U4. 1-vinylimidazole and 1,12-dibromododecane were mixed and added to acetonitrile, heated to reflux under inert gas protection, the solvent was removed under reduced pressure, filtered, washed, and dried to obtain 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide;

[0025] U5. Add wrinkled olefinic graphene oxide-coated nano-silica and 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide to acetone, add initiator, heat and stir to react under inert gas protection, add salt, stir to react, filter, wash and dry to obtain an ionic liquid solid phase extractant.

[0026] As a further improvement of the present invention, the mass ratio of graphene oxide to thionyl chloride in step U1 is 10:2-3, and the stirring reaction time is 1-2 hours; the mass ratio of chlorinated graphene oxide, acryl alcohol and base in step U2 is 12-15:4-6:7-10, and the base is selected from at least one of triethylamine, diethylamine, tert-butanol, NaOH and KOH, and the temperature of the heating stirring reaction is 45-55°C and the time is 3-5 hours; the mass ratio of nano-silica to alkenyl graphene oxide in step U3 is 10:4- 7, the average particle size of the nano-silica is 200-500nm; the molar ratio of 1-vinylimidazole and 1,12-dibromododecane in step U4 is 2:0.9-1; the mass ratio of the wrinkled olefinic graphene oxide-coated nano-silica, 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide, initiator and salt in step U5 is 10-12:13-15:0.1-0.2:18-20, the temperature of the heating and stirring reaction is 65-75°C, the time is 3-5h, and the salt is NaBF 4 and KPF 6 The stirring reaction time is 12-15 hours, and the initiator is azobisisobutyronitrile or azobisisoheptanenitrile.

[0027] As a further improvement of the present invention, the concentration of the ethanol aqueous solution in step S4 is 60-70wt%, the temperature of the heating and stirring desorption is 45-55°C, the time is 0.5-1.5h, and the relative water density of the extract is 1.2-1.22.

[0028] As a further improvement of the present invention, the mass ratio of the extract, maltodextrin and steviol glycoside in step S5 is 100:70-120:0.2-0.4.

[0029] The present invention further protects Yixinshu granules prepared by the above-mentioned preparation method.

[0030] The present invention has the following beneficial effects:

[0031] The invention uses the Yixinshu classic prescription as a raw material and adopts a new extraction technology. The Yixinshu granules obtained by the invention have a high content of effective ingredients, which greatly improves the drug efficacy. The granules have the effects of invigorating qi and restoring pulse, promoting blood circulation and removing blood stasis, nourishing yin and promoting body fluid production. The granules have significant drug effects on cardiovascular diseases such as angina pectoris caused by coronary heart disease, anxiety and depression caused by coronary heart disease, viral myocarditis, arrhythmia, heart failure, etc. by protecting vascular endothelial function, protecting myocardial cell function, inhibiting inflammatory response, regulating calcium homeostasis, and reducing fibrosis level. The invention has good patient compliance, small side effects, and a small dosage. The drug has the characteristics of rapid dissolution, fast absorption, high bioavailability, etc., which reduces the drug cost for patients.

[0032] The invention prepares a 1-(3-aminopropyl) imidazole and 3-bromopropyl trimethoxysilane for reaction, and then mixes with a silane coupling agent for reaction and hydrolysis to form a silicon dioxide layer coated on the surface of magnetic ferroferric oxide particles, thereby greatly improving the mechanical properties of the immobilized enzyme and preventing the enzyme from being loose and structurally broken during long-term enzymolysis. Then, the surface amino groups are condensed with the carboxyl groups of the composite enzyme protein, so that the composite enzyme is fixed on the magnetic material, so that the prepared immobilized enzyme is easy to be separated by a magnet and is convenient for repeated use, thereby greatly improving the utilization rate of the enzyme. In addition, the carrier modified by the ionic liquid also greatly increases the loading amount of the composite enzyme, thereby improving the enzyme activity. At the same time, the composite enzyme includes cellulase and neutral protease, so that the cell wall and macromolecular protein can be enzymatically decomposed, so that a large amount of active components are dissolved, and the utilization rate of the active components of traditional Chinese medicine is greatly improved. At the same time, the ionic liquid on the immobilized enzyme can also be complexed with cellulose in the cell wall, destroying the hydrogen bonds between cellulose molecules, so that the cellulose can be dissolved in the ionic liquid, so that the target substance can be better dissolved out of the cell wall, thereby improving the extraction rate.

[0033] After enzymatic hydrolysis, the Yixinshu raw materials of the present invention are further fermented with bacteria, including Bacillus subtilis and Lactobacillus plantarum, which can promote the breaking of the cell walls of traditional Chinese medicine plants and the generation of highly active rare components, such as promoting the conversion of ordinary ginsenosides into rare saponins, thereby greatly improving the efficacy of the prepared Yixinshu and having a better use effect.

[0034] The invention prepares an ionic liquid solid phase extractant, chlorinates the carboxyl groups on the surface of graphene oxide, and reacts with allyl alcohol to prepare graphene oxide with alkenyl groups, sprays and dries the liquid droplets through an atomizer of a spray dryer to form fine droplets, and the droplets have a large specific surface area. When the droplets contact with hot air, the solvent evaporates rapidly, resulting in rapid drying of alkenyl graphene oxide and nano-silicon dioxide suspended droplets in the droplets, thereby forming wrinkled alkenyl graphene oxide-coated nano-silicon dioxide, and the wrinkled structure greatly increases the specific surface area of ​​the carrier, thereby increasing the loading amount of the ionic liquid. It further undergoes copolymerization reaction with 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide to form a polymer structure with two imidazole rings, which can provide more action sites and more types of action forces. It can be used as a solid phase adsorbent and has good separation and adsorption capabilities for active components such as saponins, phenolic acids, glycosides, flavonoids, polyphenols, etc. in the fermentation broth. Under the action of carbon dioxide, carbon dioxide combines with the imidazole structure to increase the polarity of the ionic liquid solid phase extractant and greatly improve its adsorption capacity for polar active components. After desorption by heating ethanol, carbon dioxide dissipates and the active components enter ethanol with a small impurity content. The desorbed ionic liquid solid phase extractant can also be reused after activation, greatly reducing the extraction cost.

[0035] The preparation method of the Yixinshu granules prepared by the invention is simple, the raw material source is wide, and it is easy to realize industrial application, the cost is low, and the patient compliance is high. The drug has the characteristics of rapid dissolution, rapid absorption, high bioavailability, etc., has the effects of invigorating qi and restoring pulse, promoting blood circulation and removing blood stasis, nourishing yin and promoting body fluid, and has broad application prospects. DETAILED DESCRIPTION

[0036] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0037] NHS, N-hydroxysuccinimide; EDC, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide.

[0038] The average particle size of nano-silicon dioxide is 500nm.

[0039] Bacillus subtilis and Lactobacillus plantarum, 20 billion cfu / g, preparation method of bacterial seed liquid: inoculate the bacterial strain into Gao's medium, culture at the optimum temperature, 200r / min, 18-24h, and obtain a bacterial content of 10 8 -10 9 cfu / mL of bacterial seed solution.

[0040] Preparation Example 1 Preparation of immobilized enzyme

[0041] Here’s how:

[0042] T1. Under nitrogen protection, 3.24 g of ferric chloride and 1.26 g of ferrous chloride were dissolved in 200 mL of water, the pH value of the solution was adjusted to 9.5, heated to 80 ° C, stirred for 4 h, centrifuged, washed, dried, and calcined at 400 ° C for 2 h to obtain magnetic particles;

[0043] T2. 5 g of 1-(3-aminopropyl)imidazole, 3 g of 3-bromopropyltrimethoxysilane, and 2 g of silane coupling agent KH602 were mixed into 200 mL of water, 15 g of magnetic particles were added, the mixture was stirred and mixed evenly, the pH value was adjusted to 9, heated to 50 ° C, and hydrolyzed for 5 h to obtain ionic liquid / magnetic composite particles;

[0044] T3. 12 g of ionic liquid / magnetic composite particles were added to 250 mL of water, 2 g of NHS and 2 g of EDC were added, and the mixture was stirred for activation for 30 min, 4 g of the composite enzyme was added, and the mixture was stirred for reaction for 8 h, separated by a magnet, and freeze-dried to obtain an immobilized enzyme;

[0045] The complex enzyme is cellulase and neutral protease, and the mass ratio is 10:3.

[0046] Preparation Example 2 Preparation of immobilized enzyme

[0047] Here’s how:

[0048] T1. Under nitrogen protection, 3.24 g of ferric chloride and 1.26 g of ferrous chloride were dissolved in 200 mL of water, the pH value of the solution was adjusted to 10.5, heated to 90 ° C, stirred for 6 h, centrifuged, washed, dried, and calcined at 600 ° C for 4 h to obtain magnetic particles;

[0049] T2. 7 g of 1-(3-aminopropyl)imidazole, 6 g of 3-bromopropyltrimethoxysilane, and 4 g of silane coupling agent KH792 were added to 200 mL of water, 20 g of magnetic particles were added, stirred and mixed, the pH value was adjusted to 10, heated to 60 ° C, and hydrolyzed for 7 h to obtain ionic liquid / magnetic composite particles;

[0050] T3. 15 g of ionic liquid / magnetic composite particles were added to 250 mL of water, 4 g of NHS and 4 g of EDC were added, and the mixture was stirred for activation for 30 min. 7 g of the composite enzyme was added, and the mixture was stirred for reaction for 10 h. The mixture was separated by a magnet and freeze-dried to obtain an immobilized enzyme.

[0051] The complex enzyme is cellulase and neutral protease, and the mass ratio is 10:5.

[0052] Preparation Example 3 Preparation of immobilized enzyme

[0053] Here’s how:

[0054] T1. Under nitrogen protection, 3.24 g of ferric chloride and 1.26 g of ferrous chloride were dissolved in 200 mL of water, the pH value of the solution was adjusted to 10, heated to 85 ° C, stirred for 5 h, centrifuged, washed, dried, and calcined at 500 ° C for 3 h to obtain magnetic particles;

[0055] T2. 6 g of 1-(3-aminopropyl)imidazole, 5 g of 3-bromopropyltrimethoxysilane, and 3 g of silane coupling agent KH550 were added to 200 mL of water, 17 g of magnetic particles were added, stirred and mixed, the pH value was adjusted to 9.5, heated to 55 ° C, and hydrolyzed for 6 h to obtain ionic liquid / magnetic composite particles;

[0056] T3. 13 g of ionic liquid / magnetic composite particles were added to 250 mL of water, 3 g of NHS and 3 g of EDC were added, and the mixture was stirred for activation for 30 min. 5.5 g of the composite enzyme was added, and the reaction was stirred for 9 h. The mixture was separated by a magnet and freeze-dried to obtain an immobilized enzyme.

[0057] The complex enzyme is cellulase and neutral protease, and the mass ratio is 10:4.

[0058] Comparative Preparation Example 1

[0059] Compared with Preparation Example 3, the difference is that 1-(3-aminopropyl)imidazole and 3-bromopropyltrimethoxysilane are not added in Step T2.

[0060] The details are as follows:

[0061] T1. Under nitrogen protection, 3.24 g of ferric chloride and 1.26 g of ferrous chloride were dissolved in 200 mL of water, the pH value of the solution was adjusted to 10, heated to 85 ° C, stirred for 5 h, centrifuged, washed, dried, and calcined at 500 ° C for 3 h to obtain magnetic particles;

[0062] T2. 14 g of silane coupling agent KH550 was mixed with 200 mL of water, 17 g of magnetic particles were added, the mixture was stirred and mixed, the pH value was adjusted to 9.5, heated to 55 ° C, and hydrolyzed for 6 h to obtain modified magnetic composite particles;

[0063] T3. 13 g of modified magnetic composite particles were added to 250 mL of water, 3 g of NHS and 3 g of EDC were added, and the mixture was stirred for activation for 30 min, 5.5 g of composite enzyme was added, and the reaction was stirred for 9 h, separated by a magnet, and freeze-dried to obtain an immobilized enzyme;

[0064] The complex enzyme is cellulase and neutral protease, and the mass ratio is 10:4.

[0065] Preparation Example 4 Preparation of ionic liquid solid phase extractant

[0066] Here’s how:

[0067] U1. Add 10 g of graphene oxide to 200 mL of dichloromethane, add 2 g of thionyl chloride, stir and react for 1 h, remove the solvent and excess thionyl chloride under reduced pressure to obtain chlorinated graphene oxide;

[0068] U2. Add 12 g of chlorinated graphene oxide to 300 mL of acetonitrile, add 4 g of propylene alcohol and 7 g of NaOH, heat to 45 ° C, stir and react for 3 h, filter, wash, and dry to obtain olefinic graphene oxide;

[0069] U3. 10 g of nano-silica was added to 150 mL of water, 4 g of olefinic graphene oxide was added, ultrasonic stirring and dispersion was performed at 1000 W for 15 min, and then spray dried to obtain wrinkled olefinic graphene oxide-coated nano-silica;

[0070] U4. 20 mmol 1-vinylimidazole and 9 mmol 1,12-dibromododecane were mixed and added into 100 mL acetonitrile. The mixture was heated under reflux for 7 h under nitrogen protection. The solvent was removed under reduced pressure, filtered, washed and dried to obtain 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide.

[0071] U5. 10g of wrinkled olefin-based graphene oxide-coated nanosilica and 13g of 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide were mixed and added to 300mL of acetone, and 0.1g of azobisisobutyronitrile was added. Under nitrogen protection, the mixture was heated to 65°C, stirred for 3h, and 18g of NaBF was added. 4 , stir the reaction for 12 hours, filter, wash and dry to obtain an ionic liquid solid phase extractant.

[0072] Preparation Example 5 Preparation of ionic liquid solid phase extractant

[0073] Here’s how:

[0074] U1. Add 10 g of graphene oxide to 200 mL of dichloromethane, add 3 g of thionyl chloride, stir and react for 2 h, remove the solvent and excess thionyl chloride under reduced pressure to obtain chlorinated graphene oxide;

[0075] U2. Add 15 g of chlorinated graphene oxide to 300 mL of acetonitrile, add 6 g of propylene alcohol and 10 g of KOH, heat to 55 ° C, stir and react for 5 h, filter, wash, and dry to obtain olefinic graphene oxide;

[0076] U3. 10 g of nano-silica was added to 150 mL of water, 7 g of olefinic graphene oxide was added, 1000 W ultrasonic stirring was used for 15 min, and then spray dried to obtain wrinkled olefinic graphene oxide-coated nano-silica;

[0077] U4. 20 mmol 1-vinylimidazole and 10 mmol 1,12-dibromododecane were mixed and added into 100 mL acetonitrile. The mixture was heated under reflux for 7 h under nitrogen protection. The solvent was removed under reduced pressure, filtered, washed and dried to obtain 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide.

[0078] U5. 12g of wrinkled olefin-based graphene oxide-coated nanosilica and 15g of 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide were mixed and added to 300mL of acetone, and 0.2g of azobisisoheptanenitrile was added. Under nitrogen protection, the mixture was heated to 75°C, stirred for 5h, and 20g of KPF was added. 6 , stir the reaction for 15 hours, filter, wash and dry to obtain an ionic liquid solid phase extractant.

[0079] Preparation Example 6 Preparation of ionic liquid solid phase extractant

[0080] Here’s how:

[0081] U1. Add 10 g of graphene oxide to 200 mL of dichloromethane, add 2.5 g of thionyl chloride, stir and react for 1.5 h, remove the solvent and excess thionyl chloride under reduced pressure to obtain chlorinated graphene oxide;

[0082] U2. Add 13 g of chlorinated graphene oxide to 300 mL of acetonitrile, add 5 g of propylene alcohol and 8.5 g of triethylamine, heat to 50°C, stir and react for 4 h, filter, wash and dry to obtain olefinic graphene oxide;

[0083] U3. 10 g of nano-silica was added to 150 mL of water, 5.5 g of olefinic graphene oxide was added, 1000 W ultrasonic stirring was used for 15 min, and the mixture was spray dried to obtain wrinkled olefinic graphene oxide-coated nano-silica;

[0084] U4. 20 mmol 1-vinylimidazole and 9.5 mmol 1,12-dibromododecane were mixed and added into 100 mL acetonitrile. The mixture was heated under reflux for 7 h under nitrogen protection. The solvent was removed under reduced pressure, filtered, washed and dried to obtain 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide.

[0085] U5. 11g of wrinkled olefin-based graphene oxide-coated nanosilica and 14g of 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide were mixed and added to 300mL of acetone, and 0.15g of azobisisobutyronitrile was added. Under nitrogen protection, the mixture was heated to 70°C, stirred for 4h, and 19g of NaBF was added. 4 , stir the reaction for 13 hours, filter, wash and dry to obtain an ionic liquid solid phase extractant.

[0086] Comparative Preparation Example 2

[0087] Compared with Preparation Example 6, the difference is that the olefinic graphene oxide in step U3 is replaced by an equal mass of KH570.

[0088] The details are as follows:

[0089] U3. Add 10g of nano-silica into 150mL of ethanol, add 5.5g of silane coupling agent KH570, heat and stir at 50°C for 2h, centrifuge, wash, and dry to obtain modified nano-silica.

[0090] Comparative Preparation Example 3

[0091] Compared with Preparation Example 6, the difference is that the 1,12-dibromododecane in U4 is replaced by an equal molar amount of 1-bromododecane.

[0092] The details are as follows:

[0093] U4. Mix 20 mmol 1-vinylimidazole and 9.5 mmol 1-bromododecane and add into 100 mL acetonitrile. Heat and reflux for 7 h under nitrogen protection. Remove the solvent under reduced pressure, filter, wash and dry to obtain monoimidazole bromide.

[0094] Example 1

[0095] This embodiment provides a method for preparing Yixinshu granules, comprising the following steps:

[0096] S1. Enzymatic hydrolysis: 10 g of ginseng, 10 g of astragalus, 10 g of ophiopogon, 10 g of hawthorn, 15 g of salvia miltiorrhiza, 5 g of schisandra chinensis, and 5 g of chuanxiong were washed, dried, and crushed to obtain Chinese medicine powder; 3 g of the immobilized enzyme prepared in Preparation Example 1 was added to 500 mL of water, and the above Chinese medicine powder was added, heated to 25 ° C, and stirred for enzymatic hydrolysis for 2 h to obtain a mixture;

[0097] S2. Enzyme-assisted fermentation: The mixture was inoculated with Bacillus subtilis and Lactobacillus plantarum seed liquid at inoculation amounts of 1 v / v% and 0.5 v / v%, respectively, at 25°C, 150 r / min, and the enzyme-assisted fermentation was carried out for 36 h. The immobilized enzyme was separated by a magnet, filtered, and the filtrate was collected;

[0098] S3. Ionic liquid solid phase extractant adsorption: 10 g of the ionic liquid solid phase extractant prepared in Preparation Example 4 was added to 100 g of the filtrate, carbon dioxide was introduced, the carbon dioxide ventilation rate was 15 mL / min, stirred for 1 h, filtered, and the solid was collected;

[0099] S4. Desorption: The solid was added to a 60 wt% ethanol aqueous solution, heated to 45 ° C, stirred for desorption for 0.5 h, filtered, the filtered solid was washed, regenerated, recycled, the filtrate was recovered ethanol, dried, and an extract was obtained with a relative water density of 1.2;

[0100] S5. Preparation of Yixinshu Granules: 100 g of extract, 70 g of maltodextrin and 0.2 g of stevioside were uniformly mixed, granulated and dried to obtain Yixinshu Granules.

[0101] Example 2

[0102] This embodiment provides a method for preparing Yixinshu granules, comprising the following steps:

[0103] S1. Enzymatic hydrolysis: 20 g of ginseng, 20 g of astragalus, 20 g of ophiopogon, 20 g of hawthorn, 25 g of salvia miltiorrhiza, 15 g of schisandra chinensis, and 15 g of chuanxiong were washed, dried, and crushed to obtain Chinese medicine powder; 5 g of the immobilized enzyme prepared in Preparation Example 2 was added to 500 mL of water, the Chinese medicine powder was added, heated to 35 ° C, and stirred for enzymatic hydrolysis for 4 h to obtain a mixture;

[0104] S2. Enzyme-assisted fermentation: The mixture was inoculated with Bacillus subtilis and Lactobacillus plantarum seed liquid at inoculation amounts of 2 v / v% and 1.5 v / v%, respectively, at 30°C, 250 r / min, and the enzyme-assisted fermentation was performed for 48 h. The immobilized enzyme was separated by a magnet, filtered, and the filtrate was collected;

[0105] S3. Ionic liquid solid phase extractant adsorption: 13 g of the ionic liquid solid phase extractant prepared in Preparation Example 5 was added to 100 g of the filtrate, carbon dioxide was introduced, the carbon dioxide ventilation rate was 25 mL / min, stirred and adsorbed for 2 h, filtered, and the solid was collected;

[0106] S4. Desorption: The solid was added to a 70 wt% ethanol aqueous solution, heated to 55 ° C, stirred for desorption for 1.5 h, filtered, the filtered solid was washed, regenerated, recycled, the filtrate was recovered ethanol, dried, and an extract was obtained with a relative water density of 1.22;

[0107] S5. Preparation of Yixinshu Granules: 100 g of extract, 120 g of maltodextrin and 0.4 g of stevioside were mixed evenly, granulated and dried to obtain Yixinshu Granules.

[0108] Example 3

[0109] This embodiment provides a method for preparing Yixinshu granules, comprising the following steps:

[0110] S1. Enzymolysis: Wash 15g of ginseng, 15g of astragalus, 15g of ophiopogon, 15g of hawthorn, 20g of salvia miltiorrhiza, 10g of schisandra chinensis, and 10g of chuanxiong, dry them, and grind them to obtain Chinese medicine powder; add 4g of the immobilized enzyme prepared in Preparation Example 3 to 500mL of water, add the above Chinese medicine powder, heat to 30°C, stir and enzymolyze for 3h to obtain a mixture;

[0111] S2. Enzyme-assisted fermentation: The mixture was inoculated with Bacillus subtilis and Lactobacillus plantarum seed liquid at inoculation amounts of 1.5 v / v% and 1 v / v%, respectively, at 27°C, 200 r / min, and the enzyme-assisted fermentation was performed for 42 h. The immobilized enzyme was separated by a magnet, filtered, and the filtrate was collected;

[0112] S3. Ionic liquid solid phase extractant adsorption: 12 g of the ionic liquid solid phase extractant prepared in Preparation Example 6 was added to 100 g of the filtrate, carbon dioxide was introduced, the carbon dioxide ventilation rate was 20 mL / min, adsorption was stirred for 1.5 h, filtered, and the solid was collected;

[0113] S4. Desorption: The solid was added to a 65wt% ethanol aqueous solution, heated to 50°C, stirred for desorption for 1h, filtered, the filtered solid was washed, regenerated, recycled, the filtrate was recovered for ethanol, dried, and an extract was obtained, the relative density of which was 1.21 with respect to water;

[0114] S5. Preparation of Yixinshu Granules: 100 g of extract, 100 g of maltodextrin and 0.3 g of stevioside were uniformly mixed, granulated and dried to obtain Yixinshu Granules.

[0115] Comparative Example 1

[0116] Compared with Example 3, the difference is that the immobilized enzyme in step S1 is prepared by Comparative Preparation Example 1.

[0117] Comparative Example 2

[0118] Compared with Example 3, the difference is that the immobilized enzyme is replaced by a composite enzyme of equal mass, and the composite enzyme is cellulase and neutral protease, and the mass ratio is 10:4.

[0119] The details are as follows:

[0120] S1. Enzymatic hydrolysis: Wash, dry and crush 15g ginseng, 15g astragalus, 15g ophiopogon, 15g hawthorn, 20g salvia miltiorrhiza, 10g schisandra chinensis and 10g chuanxiong to obtain Chinese medicine powder; add 4g complex enzyme to 500mL water, add the above Chinese medicine powder, heat to 30°C, stir and enzymolyze for 3h to obtain a mixture;

[0121] The complex enzyme is cellulase and neutral protease, and the mass ratio is 10:4.

[0122] Comparative Example 3

[0123] Compared with Example 3, the difference is that no immobilized enzyme is added in step S1.

[0124] The details are as follows:

[0125] S1. Wash, dry and crush 15g of ginseng, 15g of astragalus, 15g of ophiopogon, 15g of hawthorn, 20g of salvia miltiorrhiza, 10g of schisandra chinensis and 10g of chuanxiong respectively to obtain Chinese medicine powder, add it into 500mL of water, heat it to 30°C, stir and enzymatically hydrolyze for 3h to obtain a mixture.

[0126] Comparative Example 4

[0127] Compared with Example 3, the difference is that the Bacillus subtilis seed liquid is not inoculated in step S2.

[0128] The details are as follows:

[0129] S2. Enzyme-assisted fermentation: inoculate the mixture with Lactobacillus plantarum seed solution at an inoculation rate of 2.5 v / v%, at 27°C, 200 r / min, and ferment for 42 h. Separate the immobilized enzyme with a magnet, filter, and collect the filtrate.

[0130] Comparative Example 5

[0131] Compared with Example 3, the difference is that the Lactobacillus plantarum seed liquid is not inoculated in step S2.

[0132] The details are as follows:

[0133] S2. Enzyme-assisted fermentation: inoculate the mixture with Bacillus subtilis seed solution at an inoculation rate of 2.5 v / v%, ferment at 27°C, 200 r / min, and perform enzyme-assisted fermentation for 42 h. Separate the immobilized enzyme with a magnet, filter, and collect the filtrate.

[0134] Comparative Example 6

[0135] Compared with Example 3, the difference is that step S2 is not performed.

[0136] The details are as follows:

[0137] S1. Enzymolysis: 15 g of ginseng, 15 g of astragalus, 15 g of ophiopogon, 15 g of hawthorn, 20 g of salvia miltiorrhiza, 10 g of schisandra chinensis, and 10 g of chuanxiong were washed, dried, and crushed to obtain Chinese medicine powder; 4 g of the immobilized enzyme prepared in Preparation Example 3 was added to 500 mL of water, and the above Chinese medicine powder was added, heated to 30 ° C, stirred for enzymolysis for 3 h, a mixture was obtained, and filtered to obtain a filtrate;

[0138] S2. Ionic liquid solid phase extractant adsorption: 12 g of the ionic liquid solid phase extractant prepared in Preparation Example 6 was added to 100 g of the filtrate, carbon dioxide was introduced, the carbon dioxide ventilation rate was 20 mL / min, stirred and adsorbed for 1.5 h, filtered, and the solid was collected;

[0139] S3. Desorption: The solid was added to a 65wt% ethanol aqueous solution, heated to 50°C, stirred for desorption for 1h, filtered, the filtered solid was washed, regenerated, recycled, the filtrate was recovered for ethanol, dried, and an extract was obtained, the relative density of which was 1.21 with respect to water;

[0140] S4. Preparation of Yixinshu Granules: 100 g of extract, 100 g of maltodextrin and 0.3 g of stevioside were uniformly mixed, granulated and dried to obtain Yixinshu Granules.

[0141] Comparative Example 7

[0142] Compared with Example 3, the difference is that steps S3 and S4 are not performed.

[0143] The details are as follows:

[0144] S1. Enzymolysis: Wash 15g of ginseng, 15g of astragalus, 15g of ophiopogon, 15g of hawthorn, 20g of salvia miltiorrhiza, 10g of schisandra chinensis, and 10g of chuanxiong, dry them, and grind them to obtain Chinese medicine powder; add 4g of the immobilized enzyme prepared in Preparation Example 3 to 500mL of water, add the above Chinese medicine powder, heat to 30°C, stir and enzymolyze for 3h to obtain a mixture;

[0145] S2. Enzyme-assisted fermentation: The mixture was inoculated with Bacillus subtilis and Lactobacillus plantarum seed liquids at inoculation amounts of 1.5 v / v% and 1 v / v%, respectively, at 27°C, 200 r / min, and the enzyme-assisted fermentation was carried out for 42 h. The immobilized enzyme was separated by a magnet, filtered, and the filtrate was collected and concentrated to obtain an extract having a relative water density of 1.21;

[0146] S3. Preparation of Yixinshu Granules: 100 g of extract, 100 g of maltodextrin and 0.3 g of stevioside were uniformly mixed, granulated and dried to obtain Yixinshu Granules.

[0147] Comparative Example 8

[0148] Compared with Example 3, the difference is that no carbon dioxide is introduced in step S3.

[0149] The details are as follows:

[0150] S3. Ionic liquid solid phase extractant adsorption: Add 12 g of the ionic liquid solid phase extractant prepared in Preparation Example 6 to 100 g of the filtrate, stir and adsorb for 1.5 h, filter, and collect the solid.

[0151] Comparative Example 9

[0152] Compared with Example 3, the difference is that the ionic liquid solid phase extractant is prepared by Comparative Preparation Example 2.

[0153] Comparative Example 10

[0154] Compared with Example 3, the difference is that the ionic liquid solid phase extractant is prepared by Comparative Preparation Example 3.

[0155] Test Example 1

[0156] Wistar rats were selected as experimental subjects, 10 were randomly selected as blank group, and all the other were carried out left renal artery stenosis (anesthetized by 2% sodium pentobarbital 40mg / kg intraperitoneal injection, fixed in supine position, skin and base layer were cut along the abdominal midline 1.5cm below the xiphoid process after skin disinfection, fascia of renal pedicle was separated, renal artery was separated, left renal artery was clamped near the aorta end, the left renal artery was partially narrowed, abdominal cavity was closed in layers, and the contralateral kidney was not touched during the operation. Penicillin was injected intramuscularly within 3 days after operation to prevent infection), blood pressure of rats was stabilized in 30 days after operation, and those who were defined as hypertension at 160mmHg or more were removed if systolic pressure did not meet the experimental requirements. The rats with successful modeling were randomly divided into model group, embodiment 1-3 group, comparative example 1-10 group, positive drug group, 10 in every group.

[0157] The positive drug group was gavaged with 0.0099 / kg of Captopril daily, the Example 1-3 group and the Comparative Example 1-10 group were gavaged with 15g / kg of the corresponding Yixinshu granules, and the model group and the blank group were gavaged with the corresponding amount of normal water. In order to observe the reversal effect of the drug on left ventricular hypertrophy, each group began to be gavaged at the same time in the 6th week after surgery, with a gavage volume of 1mL / 100g body weight, and gavage was administered once a day at a fixed time. During the experiment, each group was fed with ordinary feed and had free access to water. The drug was administered continuously for 6 weeks.

[0158] The systolic blood pressure of the rat tail artery was measured in the awake state after the last afternoon drug infusion.

[0159] The rats' eyeballs were removed to collect blood, which was centrifuged and the serum was separated. The contents of nitric oxide NO (kit), endothelin ET (kit), and angiotensin AngII (homogeneous competitive radioimmunoassay) were measured.

[0160] Rats were dissected, the hearts were taken out, the right atrium and right ventricle tissues were removed along the muscular end of the ventricular septum, and the left ventricle and ventricular septum were retained. The left ventricle was accurately weighed, and the LVW (left ventricular weight) / BW (body weight) value was used as a quantitative indicator to evaluate the degree of left ventricular hypertrophy (LVH).

[0161] The results are shown in Table 1.

[0162] Table 1

[0163]

[0164] Note: * compared with the blank group, P < 0.05; # compared with the model group, P < 0.05.

[0165] It can be seen from the above table that the Yixinshu granules prepared in Examples 1-3 of the present invention have the effects of lowering blood pressure, reversing left ventricular hypertrophy, reducing the ET and AngII contents in rats, and increasing the NO content.

[0166] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for preparing Yixinshu granules, characterized in that: Ginseng, astragalus, ophiopogon, hawthorn, salvia miltiorrhiza, schisandra chinensis and ligusticum chuanxiong are subjected to immobilized enzyme hydrolysis and enzyme-assisted fermentation, the product is filtered, an ionic liquid solid phase extractant is added, carbon dioxide is introduced, the product is stirred for adsorption, filtered, the solid is desorbed and dried to obtain an extract, which is evenly mixed with maltodextrin and stevioside, granulated and dried to obtain Yixinshu granules.

2. The preparation method according to claim 1, characterized in that: The following steps are involved: S1. Enzymatic hydrolysis: Wash, dry and crush ginseng, astragalus, ophiopogon, hawthorn, salvia miltiorrhiza, schisandra chinensis and ligusticum chuanxiong to obtain Chinese medicine powder; add the immobilized enzyme to water, add the Chinese medicine powder, heat and stir for enzymatic hydrolysis to obtain a mixture; S2. Enzyme-assisted fermentation: inoculating the mixture with the fermentation bacteria seed solution, enzyme-assisted fermentation, separating the immobilized enzyme with a magnet, filtering, and collecting the filtrate; S3. Ionic liquid solid phase extractant adsorption: add an ionic liquid solid phase extractant to the filtrate, introduce carbon dioxide, stir for adsorption, filter, and collect solids; S4. Desorption: adding the solid to an ethanol aqueous solution, heating and stirring for desorption, filtering, recovering ethanol from the filtrate, and drying to obtain an extract; S5. Preparation of Yixinshu Granules: Evenly mix the extract, maltodextrin and stevioside, granulate and dry to obtain Yixinshu Granules.

3. The preparation method according to claim 1, characterized in that: In step S1, the mass ratio of ginseng, astragalus, ophiopogon, hawthorn, salvia miltiorrhiza, schisandra chinensis, ligusticum chuanxiong and immobilized enzyme is 10-20:10-20:10-20:10-20:15-25:5-15:5-15:3-5, the temperature of heating and stirring enzymolysis is 25-35°C, the time is 2-4h, and the preparation method of the immobilized enzyme is as follows: T1. Under the protection of inert gas, ferric chloride and ferrous chloride are dissolved in water, the pH value of the solution is adjusted, the reaction is heated and stirred, centrifuged, washed, dried, and calcined to obtain magnetic particles; T2. 1-(3-aminopropyl) imidazole, 3-bromopropyltrimethoxysilane and a silane coupling agent were mixed and added to water, magnetic particles were added, stirred and mixed evenly, the pH value was adjusted, and heated for hydrolysis to obtain ionic liquid / magnetic composite particles; T3. Add ionic liquid / magnetic composite particles into water, add NHS and EDC, stir to activate, add composite enzyme, stir to react, separate with a magnet, freeze-dry, and obtain immobilized enzyme.

4. The preparation method according to claim 3, characterized in that: The mass ratio of ferric chloride to ferrous chloride in step T1 is 3.24:1.26, the pH value of the solution is adjusted to 9.5-10.5, the temperature of the heated and stirred reaction is 80-90°C, the time is 4-6h, the calcination temperature is 400-600°C, and the time is 2-4h; the mass ratio of 1-(3-aminopropyl)imidazole, 3-bromopropyltrimethoxysilane, silane coupling agent, and magnetic particles in step T2 is 5-7:3-6:2-4:15-20, the pH value is adjusted to 9-10, and the temperature of the heated hydrolysis is The temperature is 50-60°C, the time is 5-7h, the silane coupling agent is selected from at least one of KH550, KH602, and KH792; the mass ratio of the ionic liquid / magnetic composite particles, NHS, EDC and composite enzyme in step T3 is 12-15:2-4:2-4:4-7, the composite enzyme is selected from at least one of cellulase, pectinase, neutral protease, ficin, and bromelain, preferably, cellulase and neutral protease, the mass ratio is 10:3-5, and the stirring reaction time is 8-10h.

5. The preparation method according to claim 1, characterized in that: The fermentation bacteria in step S2 are Bacillus subtilis and Lactobacillus plantarum, and the bacterial content of the bacterial seed liquid is 10 8 -10 9 cfu / mL, the inoculation amounts were 1-2v / v% and 0.5-1.5v / v%, respectively, and the enzyme-assisted fermentation conditions were 25-30°C, 150-250r / min, and the enzyme-assisted fermentation was 36-48h.

6. The preparation method according to claim 1, characterized in that: The mass ratio of the filtrate to the ionic liquid solid phase extractant in step S3 is 100:10-13, the ventilation rate of the carbon dioxide is 15-25 mL / min, the stirring adsorption time is 1-2 h, and the preparation method of the ionic liquid solid phase extractant is as follows: U1. adding graphene oxide to dichloromethane, adding thionyl chloride, stirring the reaction, and removing the solvent and excess thionyl chloride under reduced pressure to obtain chlorinated graphene oxide; U2. adding chlorinated graphene oxide to acetonitrile, adding acryl alcohol and a base, heating and stirring to react, filtering, washing, and drying to obtain olefinic graphene oxide; U3. Adding nano-silica to water, adding olefinic graphene oxide, stirring and dispersing evenly, spray drying, and obtaining wrinkled olefinic graphene oxide-coated nano-silica; U4. 1-vinylimidazole and 1,12-dibromododecane were mixed and added to acetonitrile, heated to reflux under inert gas protection, the solvent was removed under reduced pressure, filtered, washed, and dried to obtain 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide; U5. Add wrinkled olefinic graphene oxide-coated nano-silica and 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide to acetone, add initiator, heat and stir to react under inert gas protection, add salt, stir to react, filter, wash and dry to obtain an ionic liquid solid phase extractant.

7. The preparation method according to claim 6, characterized in that: In step U1, the mass ratio of graphene oxide to thionyl chloride is 10:2-3, and the stirring reaction time is 1-2 hours; in step U2, the mass ratio of chlorinated graphene oxide, acryl alcohol and base is 12-15:4-6:7-10, and the base is selected from at least one of triethylamine, diethylamine, tert-butanol, NaOH and KOH, and the temperature of the heating stirring reaction is 45-55° C. and the time is 3-5 hours; in step U3, the mass ratio of nano-silica to alkenyl graphene oxide is 10:4-7, and the average particle size of the nano-silica is 200-500nm; The molar ratio of 1-vinylimidazole and 1,12-dibromododecane in U4 is 2:0.9-1; the mass ratio of the wrinkled olefinic graphene oxide coated nano-silica, 3,3'-dodecane-1,1'-di-1-vinylbisimidazole bromide, initiator and salt in step U5 is 10-12:13-15:0.1-0.2:18-20, the temperature of the heating and stirring reaction is 65-75°C, the time is 3-5h, the salt is NaBF4 and KPF6, the stirring reaction time is 12-15h, and the initiator is azobisisobutyronitrile or azobisisoheptanenitrile.

8. The preparation method according to claim 1, characterized in that: The concentration of the ethanol aqueous solution in step S4 is 60-70wt%, the temperature of the heating and stirring desorption is 45-55°C, the time is 0.5-1.5h, and the relative water density of the extract is 1.2-1.

22.

9. The preparation method according to claim 1, characterized in that: The mass ratio of the extract, maltodextrin and steviol glycoside in step S5 is 100:70-120:0.2-0.

4.

10. Yixinshu granules prepared by the preparation method according to any one of claims 1 to 9.

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