Compound isatis root granules and preparation method thereof

Abstract

The invention belongs to the technical field of medicines, and provides compound radix isatidis granules and a preparation method thereof. The preparation method of the compound isatis root granules comprises the following steps: (1) washing isatis root and folium isatidis, drying, crushing, mixing, adding ethanol with the concentration of 30-85%, carrying out microwave extraction at the temperature of 50-70 DEG C, and filtering to obtain first filtrate and filter residues; (2) adding water at 65-70 DEG C into the filter residue, soaking for 10 minutes, then carrying out microwave extraction, naturally cooling to room temperature, and filtering to obtain a second filtrate; and (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate into thick paste, and granulating. The preparation method is ingenious in design and linked with one another, dissolution of effective components, especially antiviral effective components, of the raw material medicine can be greatly improved, the preparation time is shortened, and the medicine effect and the patent medicine quality are improved. The prepared compound isatis root granules have excellent antiviral effect, good quality and stable drug effect.

Description

Technical Field

[0001] The invention belongs to the technical field of medicine, and in particular relates to a compound Radix Isatidis granules and a preparation method thereof. Background Art

[0002] Compound Isatis Root Granules are a Chinese patent medicine, which mainly has the effects of cooling blood, detoxifying and clearing away heat. It is suitable for wind-heat colds, sore throats, and is effective in treating upper respiratory tract infections and tonsillitis.

[0003] The prescription of Compound Isatis Root Granules contains only two medicinal materials, of which Isatis Folium accounts for 60% and Isatis Root accounts for 40%, and the auxiliary materials are starch and sucrose. Isatis Folium is the leaf of Isatis indigotica, and Isatis Root is the root of Isatis indigotica, which are mostly used clinically to fight against viral infections.

[0004] Modern research shows that the chemical composition of compound isatis root granules is diverse, among which the isatis leaf mainly contains indoles, quinazolinones, mustard glycosides, organic acids and amino acid compounds, etc.; the isatis root mainly contains alkaloids, organic acids, amino acids, nucleosides, phenylpropanoids, etc. The compound isatis root granules can have antiviral, antibacterial, anti-inflammatory, anti-endotoxin, immune system regulation, anti-tumor, anti-platelet aggregation and other pharmacological activities, which are inseparable from the joint action of its various chemical components.

[0005] Years of research have shown that among the numerous chemical components of compound isatis root granules, the components that can play a major antiviral role are indigo and indigotin in isatis leaf and isatis root, as well as isatis root polysaccharide, (R,S)-goitrin and nucleosides in isatis root. In particular, (R,S)-goitrin, as the component with the strongest antiviral activity and the largest content among the total alkaloid components of isatis root, has been used as an indicator component for evaluating the quality of isatis root medicinal materials since the 2010 edition of the Pharmacopoeia.

[0006] At present, the preparation process of compound isatis root granules adopts the traditional water extraction and alcohol precipitation method. The water extraction and alcohol precipitation method has the following main disadvantages: (1) The water extraction and alcohol precipitation method is used to extract isatis root and isatis indigotica simultaneously, and the dissolution rate of the effective ingredients is low, especially the effective ingredients mainly used for antiviral dissolution are less; (2) The water decoction method takes a long time to process the medicinal materials, which can easily cause the cork layer and cortex of isatis root to be boiled, and the resin, plant protein and starch granules in the xylem parenchyma cells will be dissolved in large quantities, affecting the dissolution of the effective ingredients; (3) The water extraction and alcohol precipitation method usually requires a long production cycle, which will increase time and labor costs; (4) After alcohol precipitation treatment, the extract is more viscous and may encounter difficulties in granulation.

[0007] Therefore, finding a preparation process that can improve the dissolution rate of the effective ingredients of compound Isatis granules, especially the antiviral effective ingredients, has become an urgent problem to be solved. Summary of the invention

[0008] In view of the above-mentioned deficiencies in the prior art, the first purpose of the present invention is to provide a method for preparing compound isatis root granules. The preparation method is ingeniously designed and interconnected, which can greatly improve the dissolution of the active ingredients of the raw materials, especially the antiviral active ingredients, shorten the preparation time, and improve the efficacy and quality of the finished drug.

[0009] In view of the above-mentioned deficiencies in the prior art, the second object of the present invention is to provide a compound isatis root granule, which has excellent antiviral effect, good quality and stable efficacy.

[0010] In order to achieve the above object, the solution adopted by the present invention is:

[0011] A preparation method of compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, adding ethanol with a concentration of 30-85%, performing microwave extraction at a temperature of 50-70°C, and filtering to obtain a first filtrate and filter residue; (2) adding 65-70°C water to the filter residue, soaking it for 10 minutes, then performing microwave extraction, naturally cooling it to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0012] A compound isatis root granule is prepared by the preparation method of the compound isatis root granule.

[0013] The compound Radix Isatidis granules and the preparation method thereof provided by the present invention have the following beneficial effects:

[0014] The preparation method of the compound Radix Isatidis granules provided by the present invention adopts microwave + organic solvent extraction with a wide concentration range, and then the filter residue is extracted with water at a specific temperature by microwave and then cooled:

[0015] From the microscopic perspective of cell disruption, microwave heating will cause polar substances inside the cells and cell walls, especially water molecules, to absorb microwave energy and generate a large amount of heat, causing the temperature inside the cells to rise rapidly. The pressure generated by the vaporization of liquid water will break through the cell membrane and cell wall, forming tiny pores; the solvent outside the cell can quickly enter the cell through these pores to extract the effective ingredients.

[0016] In the present application, a relatively wide range of ethanol concentrations is used for extraction, which can greatly improve the dissolution of the active ingredients in Folium Isatidis and Radix Isatidis, especially the dissolution of the antiviral ingredients indigo and indigotin. In addition, the dissolution of other anti-inflammatory and antioxidant ingredients such as isovitexin is also significantly improved.

[0017] Then the filter residue is soaked in 65-70℃ water for 10min and then subjected to microwave extraction. At this time, the drug cell wall is in a state of multiple micropores being opened. At this time, 65-70℃ water is directly added to facilitate the rapid entry of water into the cell, and the main antiviral components such as (R, S)-gaoitrin and nucleoside substances are quickly extracted. Then, during the natural cooling process, total alkaloids, organic acids and other antibacterial and water-soluble substances such as toxins and peroxide free radicals can be extracted. Under the specific technical conditions of this application, not only more effective ingredients can be extracted, but also the multiple decoctions in the water extraction and alcohol precipitation method can easily lead to the dissolution of starch in the cells, affecting the extraction of effective ingredients, and decoction or boiling water can inactivate the enzymes in the root of isatis, thereby affecting the synthesis of (R, S)-gaoitrin. DETAILED DESCRIPTION

[0018] In order to make the purpose, technical scheme and advantages of the embodiments of the present invention clearer, the technical scheme in the embodiments of the present invention will be described clearly and completely below. If the specific conditions are not specified in the embodiments, they are carried out according to conventional conditions or conditions recommended by the manufacturer. If the manufacturer of the reagents or instruments used is not specified, they are all conventional products that can be purchased commercially.

[0019] The following is a detailed description of a compound Radix Isatidis granule and a preparation method thereof provided in an embodiment of the present invention.

[0020] A preparation method of compound isatis root granules comprises: (1) washing, drying, crushing and mixing isatis root and isatis leaf, firstly adding 30-45% ethanol which is 6-10 times the mass of isatis root and isatis leaf, extracting at 600-700W microwave for 6-8min at 50-60℃, then filtering, then adding 50-65% ethanol which is 6-10 times the mass of isatis root and isatis leaf, extracting at 700W-800W microwave for 5-7min at 60-65℃, then filtering, then adding 70-85% ethanol which is 6-10 times the mass of isatis root and isatis leaf, extracting at 900W-1000W microwave for 3-5min at 65-70℃, then filtering, collecting the filtrates obtained three times and combining them to obtain the first filtrate, and collecting the filter residue.

[0021] In this embodiment, a step-by-step heating method is used, and different concentrations of ethanol and different microwave powers are used in different stages to extract Radix Isatidis and Folium Isatidis. On the basis of the holes produced on the cell surface by microwaves, the step-by-step heating can cause the cells to shrink further, thereby causing cracks on the cell surface, thereby accelerating the entry and exit of the solvent, dissolving and releasing the intracellular products, and quickly and effectively promoting the dissolution of the effective ingredients, so as to extract the effective ingredients such as indigo, indirubin, and isovitexin to the greatest extent in the least time.

[0022] (2) Add 20-25 times the weight of 65-70℃ water to the filter residue and soak for 10 minutes. Extract at 500-600W microwave for 10-12 minutes. After cooling naturally to room temperature, filter to obtain the second filtrate. (3) Collect the first filtrate and the second filtrate, recover ethanol, concentrate the filtrate to a thick paste, and granulate. The granulation can be carried out by the existing technology of compound isatis root granulation technology. The obtained drug granules should comply with the relevant provisions under the granule (Appendix ⅠC of the first volume of the Chinese Pharmacopoeia 2000 edition).

[0023] A compound isatis root granule is prepared by the preparation method of the compound isatis root granule.

[0024] The features and performance of the present invention are further described in detail below in conjunction with the embodiments.

[0025] Example 1

[0026] The present embodiment provides a preparation method of compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, first adding 85% ethanol with a concentration of 24 times the mass of isatis root and isatis leaf, extracting at 50°C, 1000W microwave for 14 minutes and filtering, collecting the obtained filtrates and combining them to obtain a first filtrate, and collecting filter residues; (2) adding 68°C water with a concentration of 22 times the mass of the filter residues and soaking for 10 minutes, extracting at 550W microwave for 11 minutes, naturally cooling to room temperature, filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0027] Example 2

[0028] The present embodiment provides a preparation method of compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, first adding 30% ethanol with a concentration of 30% which is 30 times the mass of isatis root and isatis leaf, extracting at 70°C, 600W microwave for 20 minutes and filtering, collecting the obtained filtrates and combining them to obtain a first filtrate, and collecting filter residues; (2) adding 68°C water with a mass of 22 times the mass of the filter residues to soak for 10 minutes, extracting at 550W microwave for 11 minutes, cooling naturally to room temperature, filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0029] Example 3

[0030] The present embodiment provides a method for preparing compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, first adding 50% ethanol in an amount 26 times the mass of isatis root and isatis leaf, extracting at 60°C, 800W microwave for 18 minutes and filtering, collecting the obtained filtrates and combining them to obtain a first filtrate, and collecting filter residues; (2) adding 68°C water in an amount 22 times the mass of the filter residue and soaking it for 10 minutes, extracting at 550W microwave for 11 minutes, naturally cooling to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0031] Example 4

[0032] The present embodiment provides a preparation method of compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, firstly adding 45% ethanol at a concentration of 6 times the mass of isatis root and isatis leaf, extracting at 50°C, 700W microwave for 6 minutes and filtering, then adding 65% ethanol at a concentration of 6 times the mass of isatis root and isatis leaf, extracting at 60°C, 800W microwave for 5 minutes and filtering, then adding 85% ethanol at a concentration of 6 times the mass of isatis root and isatis leaf, extracting at 65°C, 1000W microwave for 3 minutes and filtering, collecting the filtrates obtained three times and combining them to obtain a first filtrate, and collecting filter residue; (2) adding 68°C water at a concentration of 22 times the mass of the filter residue to soak for 10 minutes, extracting at 550W microwave for 11 minutes, naturally cooling to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0033] Example 5

[0034] The present embodiment provides a preparation method of compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, firstly adding 30% ethanol with a concentration of 10 times the mass of isatis root and isatis leaf at 60°C, extracting with microwave at 600W for 8min and filtering, then adding 65% ethanol with a concentration of 6 times the mass of isatis root and isatis leaf at 60°C, extracting with microwave at 800W for 5min and filtering, then adding 70% ethanol with a concentration of 10 times the mass of isatis root and isatis leaf at 70°C, extracting with microwave at 900W for 5min and filtering, collecting the filtrates obtained three times and combining them to obtain a first filtrate, and collecting filter residue; (2) adding 68°C water with a concentration of 22 times the mass of the filter residue and soaking it for 10min, extracting with microwave at 550W for 11min, naturally cooling to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0035] Example 6

[0036] The present embodiment provides a preparation method of compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, firstly adding 40% ethanol in an amount 8 times the mass of isatis root and isatis leaf, extracting at 55°C, 650W microwave for 7 minutes and filtering, then adding 60% ethanol in an amount 8 times the mass of isatis root and isatis leaf, extracting at 63°C, 750W microwave for 6 minutes and filtering, then adding 80% ethanol in an amount 8 times the mass of isatis root and isatis leaf, extracting at 68°C, 950W microwave for 4 minutes and filtering, collecting the filtrates obtained three times and combining them to obtain a first filtrate, and collecting filter residue; (2) adding 68°C water in an amount 22 times the mass of the filter residue and soaking it for 10 minutes, extracting at 550W microwave for 11 minutes, naturally cooling to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0037] Example 7

[0038] The present embodiment provides a preparation method of compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, firstly adding 40% ethanol in an amount 8 times the mass of isatis root and isatis leaf at 55°C, extracting with microwave at 650W for 7min and filtering, then adding 60% ethanol in an amount 8 times the mass of isatis root and isatis leaf at 63°C, extracting with microwave at 750W for 6min and filtering, then adding 80% ethanol in an amount 8 times the mass of isatis root and isatis leaf at 68°C, extracting with microwave at 950W for 4min and filtering, collecting the filtrates obtained three times and combining them to obtain a first filtrate, and collecting filter residue; (2) adding 70°C water in an amount 20 times the mass of the filter residue to soak for 10min, extracting with microwave at 500W for 12min, naturally cooling to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0039] Example 8

[0040] The present embodiment provides a preparation method of compound isatis root granules, comprising: (1) washing, drying, crushing and mixing isatis root and isatis leaf, firstly adding 40% ethanol in an amount 8 times the mass of isatis root and isatis leaf at 55°C, extracting with microwave at 650W for 7min and filtering, then adding 60% ethanol in an amount 8 times the mass of isatis root and isatis leaf at 63°C, extracting with microwave at 750W for 6min and filtering, then adding 80% ethanol in an amount 8 times the mass of isatis root and isatis leaf at 68°C, extracting with microwave at 950W for 4min and filtering, collecting the filtrates obtained three times and combining them to obtain a first filtrate, and collecting filter residue; (2) adding 65°C water in an amount 25 times the mass of the filter residue to soak for 10min, extracting with microwave at 600W for 10min, naturally cooling to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0041] Comparative Example 1

[0042] The present comparative example provides a preparation method of compound Radix Isatidis granules, comprising: (1) washing Radix Isatidis and Folium Isatidis clean, drying, crushing and mixing, firstly adding 90% ethanol in a concentration of 20 times the mass of Radix Isatidis and Folium Isatidis, extracting at 40°C, 1200W for 10 minutes and filtering, collecting the obtained filtrates and combining them to obtain a first filtrate, and collecting filter residues; (2) adding 68°C water in a concentration of 22 times the mass of the filter residues and soaking them for 10 minutes, extracting at 550W for 11 minutes, naturally cooling to room temperature, filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0043] Comparative Example 2

[0044] The present comparative example provides a preparation method of compound Radix Isatidis granules, comprising: (1) washing, drying, crushing and mixing Radix Isatidis and Folium Isatidis, firstly adding 50% ethanol in an amount 5 times the mass of Radix Isatidis and Folium Isatidis by weight, extracting at 45°C, 800W for 5 minutes and filtering, then adding 70% ethanol in an amount 5 times the mass of Radix Isatidis and Folium Isatidis by weight, extracting at 55°C, 900W for 4 minutes and filtering, then adding 90% ethanol in an amount 5 times the mass of Radix Isatidis by weight, extracting at 60°C, 1100W for 2 minutes and filtering, collecting the filtrates obtained three times and combining them to obtain a first filtrate, and collecting filter residue; (2) adding 68°C water in an amount 22 times the mass of the filter residue and soaking it for 10 minutes, extracting at 550W for 11 minutes, naturally cooling to room temperature and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0045] Comparative Example 3

[0046] The present comparative example provides a preparation method of compound Radix Isatidis granules, comprising: (1) washing, drying, crushing and mixing Radix Isatidis and Folium Isatidis, firstly adding 40% ethanol in an amount 8 times the mass of Radix Isatidis and Folium Isatidis, extracting at 55°C, 650W microwave for 7 minutes and filtering, then adding 60% ethanol in an amount 8 times the mass of Radix Isatidis and Folium Isatidis, extracting at 63°C, 750W microwave for 6 minutes and filtering, then adding 80% ethanol in an amount 8 times the mass of Radix Isatidis, extracting at 68°C, 950W microwave for 4 minutes and filtering, collecting the filtrates obtained three times and combining them to obtain a first filtrate, and collecting filter residue; (2) adding 22 times the mass of boiling water to the filter residue, soaking it for 10 minutes, extracting at 550W microwave for 11 minutes, naturally cooling it to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0047] Comparative Example 4

[0048] The present comparative example provides a preparation method of compound Radix Isatidis granules, comprising: (1) washing, drying, crushing and mixing Radix Isatidis and Folium Isatidis, firstly adding 40% ethanol in an amount 8 times the mass of Radix Isatidis and Folium Isatidis at 55°C, extracting with microwave at 650W for 7min and filtering, then adding 60% ethanol in an amount 8 times the mass of Radix Isatidis and Folium Isatidis at 63°C, extracting with microwave at 750W for 6min and filtering, then adding 80% ethanol in an amount 8 times the mass of Radix Isatidis at 68°C, extracting with microwave at 950W for 4min and filtering, collecting the filtrates obtained three times and combining them to obtain a first filtrate, and collecting filter residue; (2) adding 18 times the mass of 55°C water to the filter residue, soaking it for 10min, extracting with microwave at 480W for 15min, naturally cooling it to room temperature, and filtering to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering ethanol, concentrating the filtrate to a thick paste, and granulating.

[0049] Experimental Example 1

[0050] (1) Determination of Indigo Red:

[0051] 1. Test conditions

[0052] 1.1. Instruments and reagents: Agilent C 18 Column (250 mm×4.6 mm, 5 μm); detection wavelength: 293 nm.

[0053] 1.2. Chromatographic conditions: octadecyl silica bonded silica as filler; mobile phase: methanol-water solution (69:31, v / v); flow rate: 1.0 ml / min; detection wavelength: 293 nm; room temperature; injection volume: 10 μl; the number of theoretical plates based on the indigo red peak should not be less than 2000, and the separation degree should be greater than 1.5.

[0054] 1.3. Preparation of reference solution: Accurately weigh about 10 mg of indigo red reference substance, place it in a 100 ml volumetric flask, add methanol to dilute to the scale, and shake well; accurately pipette 1 ml, place it in a 10 ml volumetric flask, and add methanol to make an indigo red reference solution containing 10 μg per 1 ml.

[0055] 1.4. Preparation of test solution: Take appropriate amount of compound isatis root granules collected in Examples 1-8 and Comparative Examples 1-4 respectively, grind them into powder, take about 10 g, accurately weigh, put into a 50 ml volumetric flask, add 30 ml of methanol, ultrasonically treat for 20 minutes, add methanol to the scale, shake well, centrifuge, and take the supernatant.

[0056] 1.5 Test: Inject the reference solution and the test solution into the liquid chromatograph, record the chromatogram, calculate the content, and the test results are shown in Table 1.

[0057] (2) Determination of (R,S)-coitrin:

[0058] 2. Test conditions

[0059] 2.1. Instruments and reagents: Agilent C 18 Column (250 mm x 4.6 mm, 5 μm).

[0060] 2.2. Chromatographic conditions: Octadecyl silica bonded silica as filler; column temperature: 35°C; detection wavelength: 245nm; mobile phase: methanol-water (15:85); flow rate: 1.0ml / min; the number of theoretical plates based on (R,S)-godiffusion peak should not be less than 6000, and the separation degree should be greater than 1.5.

[0061] 2.3. Preparation of reference solution: Accurately weigh about 12.5 mg of (R, S)-gazodone reference substance and place it in a conical flask. Accurately add 100 ml of methanol to prepare a (R, S)-gazodone standard solution with a mass concentration of 0.1250 g / L. Shake well. Accurately pipette 1 ml and place it in a 10 ml volumetric flask. Add water to make a (R, S)-gazodone reference solution containing 50 μg per 1 ml.

[0062] 2.4. Preparation of test solution: Take appropriate amount of compound isatis root granules collected in Examples 1-8, Comparative Examples 1-4, and Pharmacopoeia (2015 edition of "Chinese Pharmacopoeia (Part I)"), grind into fine powder and pass through No. 4 sieve, take about 1 g, accurately weigh, put in a stoppered conical flask, accurately add 50 ml of water, weigh the mass, extract by ultrasound (power 250 W, frequency 30 kHz) for 1 h, let it cool to room temperature, weigh the mass, make up the lost mass with water, shake well, filter, and take the filtrate to obtain.

[0063] 2.5 Test: Inject the reference solution and the test solution into the liquid chromatograph, record the chromatogram, calculate the content, and the test results are shown in Table 1.

[0064] (3) Determination of nucleoside substances (adenosine, uridine and guanosine): Determination was carried out according to the content detection method of Isatis Root Granules in the Chinese Pharmacopoeia 2020 Edition Part 1. The determination results are shown in Table 1.

[0065] Table 1 Contents of indirubin, (R,S)-goitrin and nucleosides in each group

[0066]

[0067] It can be seen from the data in Table 1 that the compound isatis root granules prepared by the compound isatis root granules preparation method provided in Examples 1-8 of the present application have significantly improved dissolution of effective ingredients, especially antiviral effective ingredients, compared with the compound isatis root granules prepared by the pharmacopoeia preparation method.

[0068] Experimental Example 2

[0069] 1. Experimental animals: 160 KM mice, half male and half female, weighing 18-22 g, provided by Chengdu Dashuo Experimental Animal Co., Ltd.; virus, influenza A virus (A1 / Beijing Defense / 97-53H1N1), preserved by the Department of Microbiology and Immunology, Shanxi Medical University.

[0070] 2. Experimental methods:

[0071] 2.1 Animal grouping and dosing: The mice were randomly divided into 16 groups (10 mice in each group): normal control group, virus group, positive control group (acyclovir injection, Chengdu Haipu Pingyuan Pharmaceutical Co., Ltd., 100 mg / kg), pharmacopoeia group (Isatis root granules, Sichuan Good Doctor Panxi Pharmaceutical Co., Ltd., 8 g / kg); experimental groups 1-12 (Isatis root granules (8 g / kg) provided in Examples 1-8 and Comparative Examples 1-4).

[0072] 2.2 Measurement indicators and methods: Except for the normal control group, the mice in the other groups were lightly anesthetized with ether and inoculated with influenza A virus (0.03 ml / mouse) intranasally.

[0073] The normal control group and the virus group were given distilled water by enema once 2 hours before virus infection, and then twice a day, 0.2 ml / time, for 5 days; the positive control group and experimental groups 1-12 were given distilled water by enema once 2 hours before virus infection, and then twice a day, 0.2 ml / time, for 5 days.

[0074] The mice were observed for 15 consecutive days from the time of virus infection. The number of mice that became ill and died was observed and recorded. The mortality rate and average survival time were calculated (the statistics were calculated from the first day to the day of death). The results are shown in Table 2.

[0075] 2.3 Statistical methods: The experimental data were analyzed using SPSS 21.0 statistical software. The data results were expressed as mean ± standard deviation (x ± s). One-way ANOVA was used for analysis, and the differences between the groups were compared using t-test. P < 0.05 was considered statistically significant.

[0076] 3. Experimental results:

[0077] Table 2 Mortality and average survival days of mice in each group

[0078]

[0079]

[0080] Note: Compared with the normal control group, ## P<0.01; compared with the virus group, * P<0.05, ** P<0.01.

[0081] The results in Table 2 show that the mortality rate and average survival days of mice in different groups were analyzed. It can be seen that the normal control group grew normally and no mice died within 15 days; all mice in the virus group died within 15 days; compared with the virus group, the mortality rate of mice in the positive control group decreased significantly (P < 0.01), and the average survival days increased significantly (P < 0.01); compared with the virus group, the mortality rate of mice in experimental groups 1-8 decreased significantly, and the average survival days increased significantly. It is explained that the compound isatis granules prepared by the preparation method of the compound isatis granules provided in Examples 1-8 of the present application are beneficial to improving the antiviral therapeutic effect of the isatis granules.

[0082] In summary, the preparation method of the compound isatis root granules provided by the present invention is cleverly designed and closely linked, which can greatly improve the dissolution of the active ingredients of the raw materials, especially the antiviral active ingredients, shorten the preparation time, and improve the efficacy and quality of the finished drug.

[0083] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for preparing compound Radix Isatidis granules, characterized in that: include: (1) Washing, drying, crushing and mixing Radix Isatidis and Folium Isatidis, adding 30-85% ethanol, performing microwave extraction at a temperature of 50-70° C., and filtering to obtain a first filtrate and a filter residue; (2) adding 65-70° C. water to the filter residue and soaking it for 10 min, then subjecting it to microwave extraction, cooling it naturally to room temperature, and filtering it to obtain a second filtrate; (3) collecting the first filtrate and the second filtrate, recovering the ethanol, concentrating the filtrate into a thick paste, and granulating.

2. The method for preparing compound Radix Isatidis granules according to claim 1, characterized in that: In step (1), the Radix Isatidis and the Folium Isatidis are washed, dried, crushed, and then mixed, and 24-30 times the mass of the ethanol is added.

3. The method for preparing compound Radix Isatidis granules according to claim 2, characterized in that: In step (1), the conditions for microwave extraction include: microwave extraction at 600-1000W for 14-20min.

4. The method for preparing compound Radix Isatidis granules according to claim 3, characterized in that: In step (1), first, 6-10 times the mass of ethanol with a concentration of 30-45% is added at 50-60°C, and microwave extraction is performed at 600-700W for 6-8min, and then filtration is performed. Then, 6-10 times the mass of ethanol with a concentration of 50-65% is added at 60-65°C, and microwave extraction is performed at 700W-800W for 5-7min, and then filtration is performed. Then, 6-10 times the mass of ethanol with a concentration of 70-85% is added at 65-70°C, and microwave extraction is performed at 900W-1000W for 3-5min, and then filtration is performed. The filtrates obtained three times are collected and combined to obtain the first filtrate.

5. The method for preparing the compound Radix Isatidis granules according to claim 1, characterized in that: In step (2), 20-25 times the mass of water is added to the filter residue.

6. The method for preparing the compound Radix Isatidis granules according to claim 1, characterized in that: In step (2), the microwave extraction conditions include: 500-600W microwave extraction for 10-12 min.

7. A compound Radix Isatidis granules, characterized in that: The compound isatis root granules are prepared by the preparation method of any one of claims 1 to 6.

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