CN113171437A

CN113171437A - Application of bupleurum tenue composition in preparing antiviral preparation for resisting coronavirus

Info

Publication number: CN113171437A
Application number: CN202110505803.2A
Authority: CN
Inventors: 林德良
Original assignee: Beijing Handian Pharmaceutical Co ltd
Current assignee: Beijing Handian Pharmaceutical Co ltd
Priority date: 2021-05-10
Filing date: 2021-05-10
Publication date: 2021-07-27

Abstract

The invention relates to the technical field of new application of a traditional Chinese medicine composition, in particular to application of a bupleurum composition in preparing an antiviral preparation for resisting coronavirus. The invention provides an application of a bupleurum composition in preparing an antiviral preparation for resisting coronavirus, wherein the bupleurum composition is an extract obtained by extracting traditional Chinese medicine raw materials comprising 6-10 parts of bupleurum, 2-4 parts of ginger processed pinellia, 2-4 parts of scutellaria, 2-4 parts of codonopsis pilosula, 2-4 parts of liquorice, 2-4 parts of ginger and 2-4 parts of Chinese date in parts by weight. The bupleurum tenue composition has obvious effect of inhibiting coronavirus, especially has excellent effect of inhibiting novel coronavirus, and can be further used for treating diseases such as pneumonia or acute respiratory syndrome caused by coronavirus.

Description

Application of bupleurum tenue composition in preparing antiviral preparation for resisting coronavirus

Technical Field

The invention relates to the technical field of new application of a traditional Chinese medicine composition, in particular to application of a bupleurum composition in preparing an antiviral preparation for resisting coronavirus.

Background

Xiaochaihu granules are used as a harmonizing agent and have the effects of relieving exterior syndrome, dissipating heat, soothing liver and harmonizing stomach. It can be used for treating exogenous diseases, with symptoms of alternating cold and heat, fullness in chest and hypochondrium, anorexia, vexation, vomiting, bitter taste of mouth, and dry throat.

Coronaviruses are a large family of viruses known to cause the common cold and more serious diseases such as Middle East Respiratory Syndrome (MERS) and Severe Acute Respiratory Syndrome (SARS). The novel coronavirus is a new strain of coronavirus that has not been previously discovered in humans. The prior art has no specific treatment method for diseases caused by the novel coronavirus.

In view of this, the invention is particularly proposed.

Disclosure of Invention

The invention aims to provide application of a bupleurum composition in preparing an antiviral preparation for resisting coronavirus. The bupleurum composition provided by the embodiment of the invention has obvious effect of inhibiting coronavirus, especially has excellent effect of inhibiting novel coronavirus, and can be further used for treating diseases such as pneumonia or acute respiratory syndrome caused by coronavirus.

The invention is realized by the following steps:

in a first aspect, the invention provides an application of a bupleurum composition in preparing an antiviral preparation against coronavirus, wherein the bupleurum composition is an extract obtained by extracting traditional Chinese medicine raw materials comprising 6-10 parts by weight of bupleurum, 2-4 parts by weight of pinellia ternate, 2-4 parts by weight of scutellaria baicalensis, 2-4 parts by weight of codonopsis pilosula, 2-4 parts by weight of liquorice, 2-4 parts by weight of ginger and 2-4 parts by weight of Chinese date. For example, 8 parts of radix bupleuri, 3 parts of ginger processed pinellia, 3 parts of radix Scutellariae, 3 parts of radix Codonopsis, 3 parts of radix Glycyrrhizae, 3 parts of ginger and 3 parts of fructus Jujubae; or 6 parts of radix bupleuri, 2 parts of rhizoma pinelliae preparata, 2 parts of radix scutellariae, 2 parts of radix codonopsis, 2 parts of liquorice, 2 parts of ginger and 2 parts of Chinese date; or 10 parts of radix bupleuri, 4 parts of ginger processed pinellia, 4 parts of radix scutellariae, 4 parts of radix codonopsis, 4 parts of liquorice, 4 parts of ginger and 4 parts of Chinese date.

The raw materials only provide active ingredients for the bupleurum tenue composition, and the bupleurum tenue composition can also comprise pharmaceutically acceptable auxiliary materials, including but not limited to wetting agents, lubricants, disintegrants, coatings, flavoring agents, fillers and the like.

In the bupleurum tenue composition, the bupleurum tenue has the effect of dispelling pathogenic factors from the exterior, so that the pathogenic factors in the exterior half can be diffused from the exterior, and the scutellaria baicalensis has the effect of clearing heat, so that the pathogenic factors in the interior half can be diffused from the interior; pinellia ternata clears away phlegm and lowers turbid qi to arrest vomiting; radix Codonopsis is effective in invigorating qi and strengthening body resistance, and radix Glycyrrhizae is effective in regulating stomach; furthermore, the ginger and the Chinese date can help shaoyang grow qi and prevent the transmission of pathogenic factors.

In an alternative embodiment, the bupleurum tenue composition is an extract obtained by extracting 8 parts of bupleurum, 3 parts of pinellia ternate, 3 parts of scutellaria, 3 parts of codonopsis pilosula, 3 parts of liquorice, 3 parts of ginger and 3 parts of jujube in parts by weight.

In an alternative embodiment, the preparation of said minor radix bupleuri composition comprises: preparing aqueous extracts of the radix bupleuri, the radix scutellariae, the radix codonopsitis, the liquorice and the Chinese date; preparing alcohol extracts of the ginger pinellia and the ginger; and then mixing the water extract and the alcohol extract.

In an alternative embodiment, the preparation of the aqueous extract comprises: mixing the radix bupleuri, the scutellaria baicalensis, the codonopsis pilosula, the liquorice, the Chinese dates and water, decocting, and then carrying out solid-liquid separation.

The decoction provided by the embodiment of the invention is the existing decoction mode, and the decoction can be obtained by respectively mixing the raw materials with water for decoction and then mixing the decoction, or can be obtained by mixing the raw materials and then mixing the raw materials with water for decoction.

In an alternative embodiment, the preparation of the alcohol extract comprises: soaking the ginger pinellia tuber and the ginger with an alcohol solvent, percolating, and collecting percolate.

The percolate can be prepared by respectively mixing ginger processed pinellia tuber and ginger with alcohol solvents to prepare a percolate corresponding to a single raw material, and then mixing to obtain a required mixed percolate, or mixing ginger processed pinellia tuber and ginger with alcohol solvents to perform percolation.

In an alternative embodiment, the alcoholic solvent is a monohydric alcohol solvent, preferably ethanol.

It should be noted that ethanol is only an example of the embodiment of the present invention, and other alcohol solvents capable of extraction are also within the scope of the embodiment of the present invention, and for example, methanol may also be used.

In an alternative embodiment, the coronavirus is a COVID-19 virus.

In an alternative embodiment, the effective concentration of the bupleurum tenue composition in the cell model is between 2.5 μ g/mL and 50 μ g/mL.

In a second aspect, the embodiment of the present invention provides an application of the bupleurum tenue composition in preparing a medicament for treating diseases caused by coronavirus;

preferably, the disease comprises pneumonia and respiratory syndrome.

In a third aspect, the embodiment of the present invention provides an application of the bupleurum tenue composition in preparing a medicine for treating pneumonia caused by a novel coronavirus.

The invention has the following beneficial effects: the bupleurum composition provided by the embodiment of the invention has obvious effect of inhibiting coronavirus, especially has excellent effect of inhibiting novel coronavirus, and can be further used for treating diseases such as pneumonia or acute respiratory syndrome caused by coronavirus.

Drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings needed to be used in the embodiments will be briefly described below, it should be understood that the following drawings only illustrate some embodiments of the present invention and therefore should not be considered as limiting the scope, and for those skilled in the art, other related drawings can be obtained according to the drawings without inventive efforts.

FIG. 1 shows the results of the test according to example 1 of the present invention.

Detailed Description

In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.

The features and properties of the present invention are described in further detail below with reference to examples.

Experimental example 1

This embodiment provides a method for preparing a composition of bupleurum tenue, which comprises:

weighing 80 parts of radix bupleuri, 30 parts of ginger processed pinellia, 30 parts of radix scutellariae, 30 parts of radix codonopsis, 30 parts of liquorice, 30 parts of ginger and 30 parts of Chinese date.

Mixing bupleuri radix, Scutellariae radix, radix Codonopsis, Glycyrrhrizae radix and fructus Jujubae, adding 12 times of water, decocting twice for 1.5 hr, mixing decoctions obtained by solid-liquid separation, and concentrating to relative density of 1.05-1.15(50 deg.C).

Mixing rhizoma Zingiberis recens and rhizoma Pinelliae Preparatum, mixing with 70% ethanol solution, soaking for 24 hr, percolating at a speed of 1-3ml/min, collecting percolate, and concentrating to relative density of 1.05-1.15(50 deg.C).

Mixing the above water extract and ethanol extract, concentrating to relative density of 1.15-1.20(50 deg.C), and drying to obtain bupleuri radix composition.

Experimental example 1 in vitro pharmacodynamic test of Small Bupleurum composition against novel coronavirus

Testing the reagent: xiaochaihu composition (supplied by Beijing Han dynasty pharmaceutical Co., Ltd., lot number 190401); cell line: the cell strain HEK293 for stably expressing ACE2 receptor is provided by the pharmacological efficacy center of the pharmaceutical technology Limited of Hebei pharmaceutical technology, China science; COVID-19 pseudovirus (batch number: LV-nCov1-2), supplied by Guangzhou Parian Biotechnology, Inc.

The method comprises the following steps: d1, taking HEK293-ACE2 cells in logarithmic growth phase, and adjusting cell concentration to 6.25 × 10⁴Per mL, cells were added at 5000/well/80. mu.L in rows E-G of columns 2-7 and 9 of a 96-well plate, and medium was added at 80. mu.L in rows E-G of column 8 at 37 ℃ with 5% CO₂The culture was carried out overnight. D2, adding 10 μ L of test sample (1-50 μ G/mL of drug, 2-20 μ G/mL of drug, 3-10 μ G/mL of drug, 4-5 μ G/mL of drug, 5-2.5 μ G/mL of drug) with corresponding concentration into the test sample group in column 2-6, E-G, and adding 10 μ L of pseudovirus solution; adding 10 mu L of chloroquine phosphate into the positive control group of the E-G row in the 7 columns, and simultaneously adding 10 mu L of pseudovirus solution; adding 20 μ L of culture medium into blank control group; negative control group was added with 10. mu.L of pseudovirus solution and 10. mu.L of solvent control containing 0.25% DMSO. 37 ℃ and 5% CO₂Incubating for 48h, adding 40 μ L of 1 × Reporter lysine Buffer, standing at-80 deg.C for 2h, thawing at 4000r/min, centrifuging for 5min, sucking 20 μ L of supernatant to 96-well white enzyme standard plate, adding 100 μ L of Luciferase Assay Reagent, and detecting LuciferaseActivity (RLU, high RLU values indicate more pseudovirus infecting the incoming cells). The plate layout is as follows:

and (3) test results: after 48h of drug action it showed: the RLU/RLUc value of the positive control group (the ratio of the fluorescence value of the experimental group to the average fluorescence intensity of the negative control group) is 0.05 +/-0.01, and the RLU/RLUc ratio of the test sample is 0.14 +/-0.05, 0.08 +/-0.00, 0.13 +/-0.05, 0.25 +/-0.15 and 0.16 +/-0.02 in sequence from high concentration to low concentration (50 mu g/mL, 20 mu g/mL, 10 mu g/mL, 5 mu g/mL and 2.5 mu g/mL); compared with the negative control group, the RLU/RLUc ratio of each dose group of the test article is significantly different from that of the positive control group (P <0.001 or P <0.01), see figure 1.

From the results, under the test conditions, the inhibitory rate of the bupleurum tenue composition on the COVID-19 pseudovirus can reach more than 50% when the concentration is 2.5 mu g/mL-50 mu g/mL, which shows that the bupleurum tenue composition has good inhibitory effect on the infection of the COVID-19 pseudovirus with the cell strain HEK293 which stably expresses the ACE2 receptor.

Experimental example 2 clinical treatment of novel Corona Microcoris composition for pneumonia

Medicine preparation: bupleurum tenue particles; production enterprises: beijing Han Dynasty pharmaceutical Co., Ltd; specification: each bag is 5 g.

The design scheme follows the principle and basis: based on the diagnosis and treatment scheme (trial 6 th edition) of the novel coronavirus pneumonia, an inclusion standard and an exclusion standard are established.

The subject population: 1. the compound is suitable for the light and common western medicine diagnosticians of the new coronary pneumonia;

2. the traditional Chinese medicine is in accordance with the syndrome differentiation of typhoid fever and shaoyang disease;

3. age between 17-65 years, inclusive;

the new coronary pneumonia is mild, and the general western medicine diagnosticians:

and (3) light: the clinical symptoms are slight, and no pneumonia is shown in the imaging; the general type is as follows: has fever and respiratory tract symptoms, and pneumonia can be seen in imaging.

Cold with shaoyang syndrome:

and (3) master certificate: alternating cold and heat; secondary verification: fullness in the chest and hypochondrium, silentness, no appetite, vexation, vomiting, bitter taste in the mouth, dry throat and dizziness; tongue pulse: a thin and white coating and a wiry pulse.

Exclusion criteria: 1. primary diseases such as liver and kidney function are combined, the abnormality of the liver and kidney function is more than 2 times and more, and the psychopath is suffered;

2. pregnant or preparatory to pregnancy, and lactating women;

3. people allergic to the medicinal components and people with severe allergic constitution.

Administration: the preparation is taken with warm boiled water, 1 bag (5g) each time, 2 times a day; combining the medicines: 500 million U of interferon is atomized, and 0.5g of ribavirin is statically dropped.

The course of treatment is as follows: 14 days (note: if the clinical symptoms and the traditional Chinese medicine symptoms are relieved after the bupleurum composition is taken for 3 days, the bupleurum composition can be continuously taken and observed for 14 days at most, otherwise, the recipe needs to be stopped immediately, and other treatment methods are changed).

Grouping: in consideration of ethics, the observation is not provided with a positive medicine group and a placebo group, and only clinical observation before and after administration is carried out on patients conforming to mild and common types (typhoid fever shaoyang syndrome).

The main observation indexes are as follows: 1. the syndrome integral change of the traditional Chinese medicine (0 day, 3 days, 7 days and 14 days after the bupleurum composition is taken);

2. imaging (pulmonary CT) changes in new coronavirus pneumonias;

3. and detecting the positive negative conversion rate of the novel coronavirus nucleic acid by real-time fluorescence RT-PCR.

Secondary observation indicators: blood routine: WBC, lymphocyte count.

The safety index is as follows: blood convention, urine convention, stool convention, electrocardiogram, liver and kidney functions, and adverse events.

Observation start-stop time: the observation of 3 patients meeting the grouping condition was completed at 3/month 3, 3/month 6, and 2/month 28 days of 2020, and at 3/month 5, 3/month 11, and 3/month 7 days.

Historical medication situations: 3 patients had no historical medication.

The novel bupleurum composition is administered to patients with mild coronavirus pneumonia 2 times a day by 1 bag at a time, and after 2-8 days for common patients (typhoid shaoyang syndrome), the symptoms of the typhoid shaoyang syndrome of the patients are obviously improved, the symptoms of fever, bitter taste in mouth, dry throat, cough, red tongue with white fur and floating and rapid pulse and the like are obviously improved, and the inflammation of both lungs is improved. Adverse events did not occur during the observation period.

In view of the fact that the false negative rate of the nucleic acid detection kit is high during the experiment (3 months 2020), the detection result of the chest CT film is considered to be more meaningful. In addition, aerosol therapy is indicated for upper respiratory infections, whereas CODVID-19 is primarily for lower respiratory infections; ribavirin is not directed against the new coronavirus, so that the treatment effects of ribavirin and ribavirin are limited, and the medicine with the main treatment effect is the bupleurum tenue composition.

Specific cases are as follows:

it should be noted that: the date of CT detection and the date of discharge do not represent the time window for administration of the treatment, after CT detection and before the date of discharge.

Tian Yi (hospitalization number: xxxx), Man, age 32. The patient is admitted to the hospital for 2 days due to fever and cough. Fever, bitter taste in the mouth, poor appetite, red tongue with white, greasy coating and slippery and rapid pulse. The breast CT is examined at 2020-2-25 to show that the infection of the lower lobes of the lung is double, and the viral pneumonia is not excluded. Blood routine: LYM%: 19.0 percent. ESR: 62mm/h, CRP: 29.56 mg/L. The novel coronavirus nucleic acid is detected positively.

Hospitalization diagnosis

Traditional Chinese medicine diagnosis: spring warm (pathogen stagnate shaoyang)

And (3) Western diagnosis: 1. viral pneumonia

2. Pulmonary infection

After admission, hyperpyrexia and cough persist. The interferon needle was given nebulized and the ribavirin needle was titrated intravenously. The Chinese medicinal composition 5g is added in 3 days in 3 months, and after 2 times a day, the body temperature is gradually normal, and the cough is relieved. A 3-month 5-day breast CT review showed earlier absorption of both lung lower lobe infectious lesions. Blood routine review: NEU: 6.47*10⁹/L、 NEU％：80.3％、LYM：0.98*10⁹/L、LYM％：12.1％、RBC：3.56*10¹²/L、HGB： 117g/L, blood biochemistry: ESR: 104mm/h, CRP: 40.82 mg/L. The patient is discharged after 3 months and 12 days.

Wan (hospitalization number: xxxx), female, 35 years old. The patient is admitted to the hospital for 10 days due to cough and 1 day due to fever. Fever, dry throat, poor appetite, red tongue with thin and yellow coating and rapid pulse. Breast CT on 2020-3-6 days: infectious lesions of the left lung lower lobe and viral pneumonia are not excluded. Blood routine: RBC: 1.68*10¹²The ratio of/L HGB: 80g/L HCT: 14.1% LYM%: 16.2% MCH: 47.9 pg. Blood sedimentation: 78mm/H, CRP 51.65 mg/L. The novel coronavirus nucleic acid detection is negative.

Admission diagnosis:

And (3) Western diagnosis: 1. viral pneumonia

2. Pulmonary infection

3. Anemia (anemia)

The interferon needle is atomized after the hospital is admitted for 2020-3-6 days, and the ribavirin needle is dripped. The traditional Chinese medicine composition is 5g, and is orally taken 2 times a day, the body temperature is gradually normal after 2 days, and the cough is relieved. Breast CT was reviewed on day 3, 11: the left lung inferior lobe is infected with lesion, and the lesion is better absorbed than before (3 months and 6 days). Blood routine review: RBC: 1.3*10¹²The ratio of/L HGB: 84g/L HCT: 11.2 percent. Blood biochemistry: ESR: 84mm/h, CRP: 10.09 mg/L. The patient is discharged after 3 months and 12 days.

One (hospitalization number: xxx), female, 57 years old. The patient is admitted to the hospital for 2 days due to fever and cough. Fever, bitter taste in the mouth, dry throat, poor appetite, red tongue with thin and yellow coating and floating and rapid pulse. 2020-2-2 chest CT show: chest CT: there is little inflammation in the right lung. Blood-checking routine: white blood cell count of 2.84X 10⁹L; lymphocyte count of 0.85X 10⁹L; neutrophil count 1.33X 10⁹And L. Blood sedimentation: 48mm/H, CRP 20.15 mg/L. The novel coronavirus nucleic acid is detected positively.

Hospitalization diagnosis

And (3) Western diagnosis: 1. viral pneumonia

2. Pulmonary infection

After admission, the interferon needle is atomized and the ribavirin needle is dripped. Administering the Chinese medicinal composition for 2 months and 28 days5g of the drug is orally taken 2 times a day, the body temperature is gradually normal, and the cough disappears. Neutrophil count at 1.74X 10 after 3 months and 2 days⁹and/L, normal blood sedimentation and CRP. Breast CT was reviewed on 3 months and 7 days: the inferior lobe of the right lung has a few fibrous cord lesions. The patient is discharged after 3 months and 12 days.

In conclusion, the bupleurum tenue composition has good treatment effect on pneumonia and the like caused by the new coronavirus.

The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, or improvement made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims

1. An application of a bupleurum composition in preparing an antiviral preparation for resisting coronavirus is characterized in that the bupleurum composition is an extract obtained by extracting traditional Chinese medicine raw materials comprising 6-10 parts of bupleurum, 2-4 parts of pinellia ternate, 2-4 parts of scutellaria baicalensis, 2-4 parts of codonopsis pilosula, 2-4 parts of liquorice, 2-4 parts of ginger and 2-4 parts of Chinese date in parts by weight.

2. The use as claimed in claim 1, wherein the bupleurum tenue composition is an extract obtained by extracting 8 parts of bupleurum, 3 parts of pinellia ternate, 3 parts of scutellaria, 3 parts of codonopsis pilosula, 3 parts of glycyrrhiza, 3 parts of ginger and 3 parts of jujube in parts by weight.

3. The use as claimed in claim 1 or claim 2, wherein the minor bupleurum root composition is prepared by: preparing aqueous extracts of the radix bupleuri, the radix scutellariae, the radix codonopsitis, the liquorice and the Chinese date; preparing alcohol extracts of the ginger pinellia and the ginger; and then mixing the water extract and the alcohol extract.

4. Use according to claim 3, wherein the preparation of the aqueous extract comprises: mixing the radix bupleuri, the scutellaria baicalensis, the codonopsis pilosula, the liquorice, the Chinese dates and water, decocting, and then carrying out solid-liquid separation.

5. The use of claim 3, wherein the preparation of the alcohol extract comprises: soaking the ginger pinellia tuber and the ginger with an alcohol solvent, percolating, and collecting percolate.

6. Use according to claim 5, characterized in that the alcoholic solvent is a monohydric alcoholic solvent, preferably ethanol.

7. The use of claim 1, wherein the coronavirus is a COVID-19 virus.

8. The use of claim 1, wherein the effective concentration of said composition of bupleurum tenue in a cell model is between 2.5 μ g/mL and 50 μ g/mL.

9. Use of a minor bupleurum composition according to claim 1 in the manufacture of a medicament for the treatment of a disease caused by a coronavirus;

preferably, the disease comprises pneumonia and respiratory syndrome.

10. Use of a minor radix bupleuri composition according to claim 1 for the manufacture of a medicament for the treatment of pneumonia caused by a novel coronavirus;

preferably, the pneumonia caused by the novel coronavirus comprises a mild, common type of pneumonia of the novel coronavirus.