CN1548036A - Medicine containing alkannin compound as active ingredient - Google Patents

Abstract

The object of the present invention is to provide a drug containing an acannin compound or a salt thereof, which is used to treat or prevent human microbial infection diseases, inflammatory diseases, malignant tumors, bleeding and blood diseases, and immunoregulatory diseases.

Description

Drugs containing arcanine compounds as active ingredients

Technical Field The present invention relates to medicines containing Alkanin compounds or their salts, in particular to medicines containing Alkanin compounds or their salts as active ingredients for the prevention and treatment of human microbial infections (including SARA), inflammation, Drugs for tumors, bleeding, blood disorders, or immunomodulatory disorders.

Background technology Arkanine compounds are compounds that have been disclosed in literature reports (Xiao Peigen, Jia Xiaoguang, etc. editor-in-chief "Zicao", China Traditional Chinese Medicine Press, 2001 edition P24-25), wherein the arcanine compounds have the following general structure :

Arkanin compounds are all insoluble in water, but easily soluble in oil, alcohol or ethers. .), Inner Mongolia comfrey (Arnebiaguttata Bunge.) and so on. It is known that the mixed extracts of Comfrey have anti-inflammatory effects, but they are all used in the form of mixed extracts when they are used in medicine. Artificial and biosynthetic arcanine compounds are used in pharmaceuticals as a single compound or a combination of a few compounds, especially for the prevention and treatment of microbial infections, inflammation, tumors, hemorrhage, blood diseases or immunomodulatory diseases in the human body There are no reports in the preparation of other drugs.

SUMMARY OF THE INVENTION Therefore, the object of the present invention is to provide a single compound or a few compounds isolated from comfrey extract for the prevention and treatment of microbial infection, inflammation, tumor, hemorrhage, blood Drugs for diseases or immunomodulatory diseases.

The present invention provides medicines for preventing and treating microbial infections, inflammations, tumors, hemorrhages, hematological diseases or immunoregulatory diseases in human body, the medicines contain arcanine compounds represented by the following formula (I) or their salts,

When R is OH in the formula, it is:

Alkannin;

When R is (CH ₃ ) ₂ C=CHC(O)O-:

β, β-dimethylacrylalkannin (β, β-dimethylacrylalkannin);

When R is CH ₃ C(O)O-:

Acetylalkannin (acetylalkannin);

When R is (CH ₃ ) ₂ C=C(CH ₃ )CH ₂ C(O)O-:

2,3-Dimethylpentenoyl alkannin (teracrylalkannin);

When R is (CH ₃ ) ₂ COHCH ₂ C(O)O-:

β-hydroxyisovalerylalkannin (β-hydroxyisovalerylalkannin):

When R is (CH ₃ ) ₂ C[OC(O)CH ₃ ]CH ₂ C(O)O-:

β-Acetoxyisovalerylalkannin (β-acetoxyisovalerylalkannin):

Any combination of 1-5 of the above compounds; among them: preferably containing 1-3 of arcanin, β, β-dimethylacryloyl arcanin and acetyl arcanin; more preferably containing β, β- Dimethacryloylarkanin and/or acetylarkanin; most preferably containing acetylarkanin. The salts of arcanine compounds in the present invention include alkali metal, alkaline earth metal and ammonium salts and the like.

The medicine of the present invention can contain any single compound in the arcanine compounds as a bulk drug, and can also contain a combination of a small number of compounds thereof, wherein the purity of each individual compound is 80% or more, preferably the purity is 90% or more. When it contains a combination of a few kinds of compounds, the content of the active ingredient in the composition of the few kinds of compounds is 70% or more.

The medicament of the present invention may also contain other medicaments as active ingredients as necessary. The other drugs are not particularly limited, and can be appropriately selected by those skilled in the art based on prior art knowledge.

The content of the arcanine compounds contained in the medicine of the present invention ranges from 0.0001% to 75% (percentage by weight), which can be properly selected according to different preparations and diseases. When used in human body, the daily dosage of the arcanine compound can be controlled between 10 μg-20 g, preferably 100 μg-10 g, more preferably 1 mg-8 g, most preferably 5 g. It can be appropriately selected according to the different conditions of the patient, such as age, weight, and condition. Single or divided administration can be used. The medicament of the present invention can be administered internally, externally, by injection, inhalation or transdermally.

The medicament containing arcanine compounds of the present invention can be used for the prevention and treatment of microbial infections in the human body, the microorganisms include pathogenic Gram-positive cocci such as Staphylococcus aureus, Streptococcus pneumoniae, Staphylococcus epidermidis, Enterococcus faecalis etc.; pathogenic gram-negative cocci such as Klebsiella pneumoniae, Serratia marcescens, maltophilia, etc.; anaerobic or microaerophilic pathogenic bacteria such as: Helicobacter pylori, etc.; deep and superficial fungi Classes such as Candida, Aspergillus fumigatus, Cryptococcus, dermatophyte, Candida krusei, Seduospora apex, etc.; various mycoplasma infections such as Ureaplasma urealyticum, Mycoplasma pneumoniae, etc.; viruses such as hepatitis B virus, influenza virus , herpes virus, HIV virus, and pathogenic microorganisms that cause SARS, such as: coronavirus and its mutant viruses.

The medicine containing arcanine compounds of the present invention is sensitive to pathogenic microorganisms, but insensitive to beneficial microorganisms such as lactobacilli and bifidobacteria.

The medicine containing the arcanine compound of the present invention can be used for preventing and treating inflammation in human body. The inflammation includes, for example, phlebitis, vascular purpura, vaginitis, edema, and the like.

The medicine containing arcanine compounds of the present invention can also be used to prevent and treat hemorrhage and blood diseases in human body, such as burns and scalds, various dermatitis, sepsis, hemophilia, essential thrombocythemia, leukemia and the like.

The medicine containing arcanin compounds of the present invention can also be used to prevent and treat human tumors, especially malignant tumors, such as ascites tumors: liver cancer, L1210; solid tumors: W256, S180, gastric cancer 823, squamous cell carcinoma 109, Lewis Lung cancer, etc.

The medicine containing arcanin compounds of the present invention can be used to prevent and treat human immunomodulatory diseases, that is, to promote the non-specific immunity and specific cellular immunity of the body by promoting the immune response of T lymphocytes and the like.

Therefore, the medicament of the present invention can be used in the respiratory system, digestive system, urinary system, reproductive system, blood system, circulatory system, skin and mucous membrane parts of the human body.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS The preparation and pharmacodynamic tests of the pharmaceutical preparation containing arcanine compounds of the present invention will be described in detail below, but the content of the present invention is not limited thereto.

Preparation Example 1

After pulverizing 2 kg of Xinjiang Comfrey, extract it with petroleum ether until the residue of Comfrey is colorless, and recover the solvent to obtain 80 g of dark red slurry. The resulting slurry was separated and prepared by silica gel H column liquid chromatography, namely: gradient elution with 1%-20% ethyl acetate-petroleum ether, to obtain monomers of the aforementioned six arcanin compounds. That is: 0.712 grams of arcanin (yield 0.89%), 29.024 grams of β, β-dimethylacryl arcanin (yield 36.28%), 13.27 grams of acetyl arcanin (yield 16.59%), 6.032 grams 2,3-Dimethylpentenoyl arcanin (yield 7.54%), 0.776 g β-hydroxyisovaleryl arcanin (yield 0.97%), 0.792 g β-acetoxy isovaleryl arcanin Ning (yield 0.99%). After high-pressure liquid chromatography analysis, the purity is above 90%.

Preparation example 2.

After pulverizing 2 kilograms of Xinjiang Comfrey, pass through a 20-40 mesh sieve, and use CO ₂ -supercritical extraction to obtain 70 grams of red paste, which is separated and prepared by high-pressure liquid chromatography (Knauer K1001 type in Germany), and the preparation column: Silica gel H 10 μm 50×300 mm; that is, by gradient elution with 1%-20% ethyl acetate-petroleum ether, the red monomers of the six arcanin compounds mentioned above were respectively obtained. That is: 0.707 grams of arcanin (yield 1.01%), 30.877 grams of β, β-dimethylacryl arcanin (yield 44.11%), 15.869 grams of acetyl arcanin (yield 22.67%), 6.034 grams 2,3-Dimethylpentenoyl arcanin (yield 8.62%), 0.91 g of β-hydroxyisovaleryl arcanin (yield 1.30%) and 0.77 g of β-acetoxyisovaleryl arcanin Ning (yield: 1.10%) was analyzed by high-pressure liquid chromatography, and the purity was above 90%.

Example 1

According to methods known to those skilled in the art, tablets of one or a combination of a few of the six compounds are prepared, wherein the tablets can be made to contain 10%-70% of arcanine compounds according to actual needs .

Under sterile operating conditions, take 100 grams of acetylalkannin (acetylalkannin) obtained by the above Preparation Example 1 or Preparation Example 2, 100 grams of microcrystalline fiber, 30 grams of magnesium stearate, 4 grams of hydroxypropylmethylcellulose white. Tablets of 0.5 g were prepared according to known tableting techniques and equipment.

Example 2

Get 100 grams of the composition (arkanin, β, the mixing ratio of β-dimethylacryloyl akanin and acetyl akanin) obtained by the above-mentioned preparation example 1 or preparation example 2 is 1: 1: 2), other is identical with embodiment 1, prepares the tablet of 0.5 gram.

Example 3

The ointments of the six arcanine compounds were prepared according to methods known to those skilled in the art. Wherein the ointment can be prepared to contain 0.0001%-10% of arcanine compounds according to actual needs. Under aseptic conditions, take 0.5 gram of the arcanin compounds (arkanin, β, β-dimethylacryloyl arcanin, acetylarkanin and β-arkanin) obtained in the above preparation example 1 or preparation example 2 The mixing ratio of hydroxyisovaleryl arcanin is 0.7: 1: 1: 2: 0.5), 80 grams of vaseline, 10 grams of liquid paraffin and 10 grams of anhydrous lanolin, grind it evenly in a mortar, and pack it into products. For external use. The ointment can also be used to make patches for transdermal absorption according to methods known to those skilled in the art.

Example 4

The injections of the six arcanine compounds were prepared according to methods known to those skilled in the art. Under sterile conditions, take 0.5 g of the β,β-dimethylacryloylarkanin obtained in Preparation Example 1 or Preparation Example 2, 400 ml of propylene glycol, 100 ml of ethanol, 20 ml of Tween-80, and benzyl alcohol 15 milliliters, the mixture is fully dissolved, add water to 1000 milliliters, mix well, and bottle to make injection products.

The results of drug efficacy tests of drugs containing arcanine compounds are described below.

(1) Drug preparation

Weigh 5.0 mg of arckanin, β,β-dimethylacryloylarkanin and acetylarkanin obtained in Preparation Example 1 or Preparation Example 2, respectively. Drugs were dissolved in 1 mL of DMSO. After diluting it 50 times with RPMI-1640 medium, it was aliquoted and further diluted to concentrations of 100, 50, 25, 12.5, 6.25, 3.125, 1.5625, 0.78125, and 0.390625 (μg/ml).

(2) Drug sensitivity test

Dispensing the above-mentioned medicines of various concentrations into well plates, and inoculating them with prepared bacterial strains with a concentration of about 10 ³ -10 ⁶ .

The test results showed that arcanin, β, β-dimethylacryloyl arcanin and acetyl arcanin had considerable sensitivity to Gram-positive aureus, with a sensitivity range (MIC) of 0.391- 12.5μg/ml; for common Gram-negative pathogens, Klebsiella pneumoniae, its sensitivity MIC range is 0.391-6.25μg/ml, and a few strains are 12.5-50μg/ml; Serratia marcescens (Most clinical isolates) and most strains of Bacillus maltophilia have a sensitivity range of 0.391-3.125 μg/ml. Among them, it is especially effective for Bacillus maltophilus, its sensitivity range is 0.391-0.781μg/ml, while berberine is 8-32μg/ml, that is, it is significantly better than berberine; the MIC for Bacteroides, especially Bacteroides fragilis is 0.391 -6.25μg/ml; quite sensitive to Helicobacter pylori, its MIC is 0.391-0.781μg/ml.

In addition, the results of the in vitro antifungal test of β, β-dimethylacryloyl arcanin show that the MIC range for Candida and Cryptococcus is between 2.08-33.3 μg/ml, and the MIC ₉₀ is 33.3 μg/ml, The MIC range of fluconazole is between 0.125-64μg/ml, and the MIC ₉₀ is 69μg/ml; the MIC range for dermatophytes is between 4.16-8.32μg/ml, and the MIC ₉₀ is 4.16μg/ml. The MIC range of conazole against most strains of dermatophyte is between 32-64μg/ml, and the MIC ₉₀ is 64μg/ml, and the difference between the two is obvious. In addition, surprisingly, β, β-dimethylacryloyl arcanin has a good inhibitory effect on fluconazole-resistant Candida krusei, with MIC of 8.32-16.6 μg/ml, and The MIC of Scedospora apime, which is not sensitive to most antifungal drugs such as conazole, is between 4.16-8.32 μg/ml. In addition, the MIC of acetylarkanin against Cryptococcus neoformans is 3.90625 μg/ml, and the MIC against Trichophyton rubrum is 0.90625-62.5 μg/ml; The MICs of cocci and trichophyton were 3.0625-250 μg/ml. Therefore, arcanine compounds are broad-spectrum and potent antifungal agents.

In addition, the MICs of the arcanine compounds of the present invention to microorganisms beneficial to the human body, such as lactobacilli and bifidobacteria, are all above 200 μg/ml. Therefore, it can be seen from the above data that the drug containing the arcanine compounds of the present invention Sensitive to pathogenic microorganisms, but insensitive to beneficial microorganisms.

From the comparative test of the mixed extract of Comfrey and 1-3 kinds of said Akanins compounds, it can be seen that the medicine containing 1-3 kinds of said Akanins compounds of the present invention is obviously better than the mixture of Comfrey The results of the MIC (μg/ml) of the extract are shown in Table 1.

Table 1 strain name A B C Staphylococcus epidermidis 12.5 0.391 0.7812 Serratia marcescens 25 0.781 3.125 Bacteroides >200 0.391 0.391 Candida albicans 500 3.9062 250

Note: A is the mixed extract of Comfrey;

B is β, β-dimethylacryloyl arcanin;

C is a mixture of arcanine compounds (the mixing ratio of arcanin, β, β-dimethylacryloyl arcanin and acetyl arcanin is 1:1:2)

The results of the antibacterial effect test of arcanin, β, β-dimethylacryl arcanin and acetyl arcanin on mycoplasma pneumoniae showed that the minimum concentration MIC for inhibiting mycoplasma pneumoniae was 3.751 μg/ml and 2 μg respectively /ml and 7.819μg/ml, equivalent to the inhibitory effect of pure erythromycin for injection 0.1925μg/ml.

Table 2 below shows the test results of the ointment of arcanine compounds prepared in the above-mentioned Example 3 for skin topical application on some diseases.

Table 2 disease Number of subjects Efficient cure rate Treatment days Route of administration Remark Burns and scalds 300 100% 100% 6-20 Affected area direct 92 people were scalded, 186 were superficial 2 degrees, 114 were deep 2 and 3 degrees hemorrhoid 117 100% 97.4% 15 Affected area direct Three cases recurred after half a year Shingles 98 100% 100% 3-7 Affected area direct Among them, 12 cases added polymyocyte cervical erosion 80 100% 100% 10-20 Vaginal administration Pediatric nosebleed 257 99.6% 72.8% 15 nasal administration flat wart 100 96% 81% 10-30 Affected area direct chronic prostatitis 40 82.5% 57.5% 10-20 anal drug acne 50 92% 60% 15 Affected area direct bedsore 30 100% 100% 7-21 Affected area direct chapped eczema 98 94.9% 66.3% 10-30 Affected area direct Genital warts 55 100% 100% 5-35 Affected area direct baby diaper dermatitis 208 100% 100% 2-6 Affected area direct

It can be seen from the above table that the skin external use of arcanine compounds is suitable for treating most abscesses, scabies and herpes, especially for burns and scalds, and does not leave scars after healing.

The animal test results of arcanin, β,β-dimethylacryloyl arcanin and acetyl arcanin on transplanted tumors are shown in Table 3 below.

table 3 tumor type β,β-Dimethacryloylarkanin Acetyl arcanine Akanin Tumor inhibition rate life extension rate Tumor inhibition rate life extension rate Tumor inhibition rate life extension rate ascites liver cancer 113.4% 112.6% 130.8% S180 9.63% 35.7% Lewis lung cancer 42.8% 52.6% L1210 128% W256 77%

It can be seen from the above table that β, β-dimethylacryloyl arcanin has different degrees of curative effect on liver cancer, S180, Lewis lung cancer; curative effect; Akanin is only effective for liver cancer.

The test results of arcanine compounds on viruses showed that: on the 7th day after duck hepatitis B virus infected ducks, oral administration of β, β-dimethylacryl arcanine, 100 mg/kg twice a day, 10 days had a positive effect on the serum of infected ducks. The inhibitory effect of DHBV-DNA level is significant (P<0.05-0.01), and there is no toxic reaction; the 50mg/kg group has a significant inhibitory effect (P<0.05).

The in vitro test of β, β-dimethylacryloylarkanin on hepatitis B virus showed that the average inhibition rate on HBsAg was 96.2601%, and the average inhibition rate on HBeAg was 91.6056% at a concentration of 30 μg/ml.

The test results of arcanin and β,β-dimethylacryloyl arcanin against HIV-1 reverse transcriptase and integrase in vitro are shown in Table 4 below.

Table 4 _IC50 (μg/ml) anti-HIV reverse transcriptase positive control PFA 0.097 Arkanines >20 anti-HIV integrase Positive control ABPS-Y 0.922 Arkanines 12.467

The role of arcanine compounds was studied by using the mouse model of hypoimmunity induced by mitomycin C. The results show that the β of 6 mg/kg is injected intraperitoneally every day, and the β-dimethylacryloyl arcanine is injected continuously for 5 days, and the NK cell cytotoxicity of mouse splenocytes is improved by about 20% (P＜0.001), which shows that it can make mice The damage of macrophages in the peritoneal cavity has recovered, which can increase the migration ability of macrophages in the peritoneal cavity, increase the activity of T lymphocytes, and promote the immune response of T lymphocytes. It can promote the body's non-specific immunity and specific cells. Immunity.

Claims (13)

1. medicine that is used to prevent and treat human microorganism's infection, inflammation, tumor, hemorrhage, hematopathy or immune regulative disease, this medicine contains the alkannin compound or its esters of formula (I) expression,

R is for being selected from by OH, (CH in the formula ₃) ₂C=CHC (O) O-, CH ₃C (O) O-, (CH ₃) ₂C=C (CH ₃) CH ₂C (O) O-, (CH ₃) ₂COHCH ₂C (O) O-, (CH ₃) ₂C[OC (O) CH ₃] CH ₂Any 1-5 kind in C (O) group that O-formed.

2. medicine as claimed in claim 1, wherein R is for being selected from OH, (CH ₃) ₂C=CHC (O) O-and CH ₃The 1-3 kind of C (O) O-.

3. medicine as claimed in claim 2, wherein R is (CH ₃) ₂C=CHC (O) O-and/or CH ₃C (O) O-.

4. medicine as claimed in claim 3, wherein R is (CH ₃) ₂C=CHC (O) O-.

5. medicine as claimed in claim 1, the purity of each chemical compound that wherein said medicine contains are more than 80% or 80%.

6. medicine as claimed in claim 1, the purity of each chemical compound that wherein said medicine contains are more than 90% or 90%.

7. medicine as claimed in claim 1, wherein when described medicine contained several combination of compounds, the content of effective in the said composition was more than 70% or 70%

8. medicine as claimed in claim 1, this medicine also can contain the other medicines active component.

9. medicine as claimed in claim 1, when being used for human body, the day use amount of described alkannin compound can be controlled between the 10 μ g-10g.

10. medicine as claimed in claim 1 can be with the mode administration of for oral administration, external, injection, suction or transdermal.

11. medicine as claimed in claim 1, this medicine is used for the treatment of or prevents the infected by microbes of each system of human body, comprises streptococcus pneumoniae, Klebsiella Pneumoniae, helicobacter pylori, candidiasis, cryptococcus, dermatophytosis, most advanced and sophisticated many spores of match mycete; Various mycoplasma infections comprise Ureaplasma urealyticum, mycoplasma pneumoniae; Or viral infection comprises hepatitis B virus, influenza virus, herpesvirus, HIV virus.

12. medicine as claimed in claim 1, this medicine are used for the treatment of or prevent the pathogenic microorganism that may cause the SARS disease of human body, as: coronavirus and variant viral thereof, negative myxovirus, chlamydia etc.

13. medicine as claimed in claim 1, this medicine are used for the treatment of or prevent ascites tumor, hepatocarcinoma, L1210, solid tumor, W256, sarcoma 180, gastric cancer 823, scale cancer 109 or the Lewis lung cancer of human body.