CN112546202A - Complexing agent with HPV virus inhibiting function and preparation method thereof - Google Patents
Complexing agent with HPV virus inhibiting function and preparation method thereof Download PDFInfo
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/32—Burseraceae (Frankincense family)
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/1703—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
- A61K38/1709—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P15/00—Drugs for genital or sexual disorders; Contraceptives
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P15/00—Drugs for genital or sexual disorders; Contraceptives
- A61P15/02—Drugs for genital or sexual disorders; Contraceptives for disorders of the vagina
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/20—Antivirals for DNA viruses
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0652—Cells of skeletal and connective tissues; Mesenchyme
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Abstract
The invention relates to a complexing agent with the function of inhibiting HPV virus and a preparation method thereof, and the complexing agent comprises the following components in percentage by mass: 20-35% of olive extract, 0.5-2% of cell factor freeze-dried powder and the balance of sterile water. The olive extract is obtained by performing molecular modification and ion catalytic separation on high-purity olive oil. The cytokine freeze-dried powder is prepared by a stem cell culture solution through a freeze-drying technology. The compound preparation of the invention has no stimulation to vagina, safety and no side effect; the carried antivirus and antibacterial components can quickly permeate into the skin, protect mucosal cells from being invaded by viruses, quickly repair skin tissues and improve the healing and regenerating capability of wounds; the immunity of the organism can be improved, and a good antiviral effect can be achieved; in the course of viral infection, cytokines stimulate the human immune system to destroy pathogens, eliminate damaged cells, repair damaged tissues, and in this way, the cytokines protect against pathogens and inhibit the HPV virus.
Description
Technical Field
The invention relates to the technical field of female sanitary nursing products, in particular to a complexing agent with an HPV virus inhibiting function and a preparation method thereof.
Background
Human Papilloma Virus (HPV) is an epitheliophilic virus with high specificity. HPV are known to cause benign tumors and warts in humans, such as human common warts, condyloma acuminatum, and papillomas, which grow on the skin and mucous membranes in the vicinity of the reproductive organs.
HPV is a DNA virus, which is a collective term for a group of viruses, and can be clinically divided into a plurality of subtypes, and different subtypes can cause different diseases. HPV can be classified into low-risk type and high-risk type according to the risk of different types of HPV and tumor. Low risk types of HPV include HPV6, 11, 42, 43, 44, etc., which often cause benign lesions such as external genital condyloma, including cervical intraepithelial low grade lesions (CINI). High-risk types of HPV including HPV16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, etc. are associated with the occurrence of cervical cancer and cervical intraepithelial height lesions (CINII/III).
The most common types of genital tract infections with HPV are types 6, 11, 16 and 18, wherein HPV6 and 11 often infect vulva, anus, vagina and other parts; the research on the cervical cancer tissue specimens from various countries in the world discovers that the tumors caused by HPV16 and 18 types of infection are most common.
The current treatment technologies for gynecological diseases caused by HPV viruses are as follows: the injection is injected into muscles, orally taken medicines, operations and the like, and the methods have large side effect on human bodies; the other medicines are mainly administered through vagina, and the dosage forms of the common vagina administration preparations mainly comprise suppository, gel, effervescent tablets, vaginal tablets, lotion, capsules, film agents and the like, but the dosage forms have some defects more or less and influence the medicine to play the effect.
Disclosure of Invention
The invention aims to solve the defects of the prior art and provides a complexing agent with the function of inhibiting HPV virus and a preparation method thereof.
In order to achieve the purpose, the invention adopts the following technical scheme:
a complexing agent with the function of inhibiting HPV virus comprises the following specific components in percentage by mass: 20-35% of olive extract, 0.5-2% of cell factor freeze-dried powder and the balance of sterile water.
The olive extract is obtained by performing molecular modification and ion catalytic separation on high-purity olive oil.
The cytokine freeze-dried powder is prepared by a stem cell culture solution through a freeze-drying technology.
The preparation method of the compound agent with the function of inhibiting the HPV virus comprises the following specific steps:
s1 preparation of olive extract
The olive oil enters molecular modification equipment for reaction after being inspected and disinfected, meanwhile, a compressor is used for cooling, the olive oil enters membrane separation equipment after the reaction is finished, and more than 90% of high-effective components enter a mixing and stirring tank after being inspected;
s2 preparation of cytokine freeze-dried powder
Subculturing human mesenchymal stem cells to P3 generation, collecting sterile stem cell culture supernatant when the coverage rate of the stem cell culture supernatant is more than or equal to 80% after the stem cell culture supernatant is cultured in a cell culture medium of P3 generation, and storing at 4 ℃ or freezing at-20 ℃ for later use;
adding excipient into the collected sterile stem cell culture supernatant according to the proportion, and fully dissolving;
pre-freezing the dissolved culture solution at-20 deg.C for 8h, and freeze-drying with a freeze-drying machine to obtain cytokine lyophilized powder;
s3 preparation of complexing agent
The olive extract, the cell factor freeze-dried powder and the sterile water are uniformly stirred according to the proportion to obtain the complexing agent product.
In step S1, less than 10% of the low active ingredients in the membrane separation device operation are separated as raw materials for skin care products.
In step S2, the excipients added were mannitol and dextran.
In step S2, the culture conditions were as follows: the temperature was 37 ℃, the carbon dioxide concentration was 5%, and the humidity was 99%.
In step S2, the cell culture medium is ScienCell serum-free medium in usa.
The invention has the beneficial effects that: the compound preparation of the invention has no stimulation to vagina, safety and no side effect; the carried antivirus and antibacterial components can quickly permeate into the skin, protect mucosal cells from being invaded by viruses, quickly repair skin tissues and improve the healing and regenerating capability of wounds; the immunity of the organism can be improved, and a good antiviral effect can be achieved; in the course of viral infection, cytokines stimulate the human immune system to destroy pathogens, eliminate damaged cells, repair damaged tissues, and in this way, the cytokines protect against pathogens and inhibit the HPV virus. The compound preparation does not contain heavy metal, hormone, antibiotic and sex hormone, and olive bacteriostatic plant natural acid, and is easy to absorb and free of residue.
The effective rate of the complexing agent on low-risk HPV infection can reach 95.6 percent; the effective rate for high-risk HPV infection can reach 89.6 percent.
Detailed Description
The invention will be further illustrated with reference to specific examples:
a complexing agent with the function of inhibiting HPV virus comprises the following specific components in percentage by mass: 20-35% of olive extract, 0.5-2% of cell factor freeze-dried powder and the balance of sterile water.
The olive extract is obtained by performing molecular modification and ion catalytic separation on high-purity olive oil.
The cytokine freeze-dried powder is prepared by a stem cell culture solution through a freeze-drying technology.
The preparation method of the compound agent with the function of inhibiting the HPV virus comprises the following specific steps:
s1 preparation of olive extract
The olive oil enters molecular modification equipment for reaction after being inspected and disinfected, meanwhile, a compressor is used for cooling, the olive oil enters membrane separation equipment after the reaction is finished, and more than 90% of high-effective components enter a mixing and stirring tank after being inspected;
s2 preparation of cytokine freeze-dried powder
Subculturing human mesenchymal stem cells to P3 generation, collecting sterile stem cell culture supernatant when the coverage rate of the stem cell culture supernatant is more than or equal to 80% after the stem cell culture supernatant is cultured in a cell culture medium of P3 generation, and storing at 4 ℃ or freezing at-20 ℃ for later use;
adding excipient into the collected sterile stem cell culture supernatant according to the proportion, and fully dissolving;
pre-freezing the dissolved culture solution at-20 deg.C for 8h, and freeze-drying with a freeze-drying machine to obtain cytokine lyophilized powder;
s3 preparation of complexing agent
The olive extract, the cell factor freeze-dried powder and the sterile water are uniformly stirred according to the proportion to obtain the complexing agent product.
In step S1, less than 10% of the low active ingredients in the membrane separation device operation are separated as raw materials for skin care products.
In step S2, the excipients added were mannitol and dextran.
In step S2, the culture conditions were as follows: the temperature was 37 ℃, the carbon dioxide concentration was 5%, and the humidity was 99%.
In step S2, the cell culture medium is ScienCell serum-free medium in usa.
The olive extract in the invention is prepared by processing olive oil by PTF-9 beta molecular modification equipment which is independently developed, so that an important medicinal component Senoliv in the olive oil is extracted, and various original medicinal components of the olive are combined, so that the product has strong comprehensive advantages, has a sterilization rate of 99.99% to pathogenic bacteria such as escherichia coli, staphylococcus aureus, candida albicans and the like, has no drug resistance, has a remarkable effect on prevention of HPV (human papillomavirus) viruses, and has good applicability to women in climacteric period and menopausal period.
The cell factor freeze-dried powder is produced in the process of stem cell culture, the stem cell culture solution is prepared into the cell factor freeze-dried powder by a freeze-drying technology, the cell factor has anti-infection and anti-tumor effects, the cell factor has high affinity with a cell factor receptor, and the obvious biological effect can be exerted only by a trace amount; in addition, the medicine also has the functions of mediating and regulating immune response and inflammatory reaction, promoting proliferation and differentiation of target cells, stimulating hematopoiesis, promoting tissue repair and the like. During viral infection, cytokines stimulate the human immune system to destroy pathogens, eliminate damaged cells, and repair damaged tissues. In this way, the cytokine protects against pathogens and is able to inhibit the HPV virus.
The using method comprises the following steps: before use, the hands and the vulva are cleaned, the knees are bent to lie on the back, the push tube is gently sent into the vagina, after all the liquid is injected, the push tube is slowly drawn out and is discarded, and the patient stands still for three minutes.
Specific example 1:
a complexing agent with the function of inhibiting HPV virus comprises the following specific components in percentage by mass: 20% of olive extract, 0.5% of cytokine freeze-dried powder and the balance of sterile water.
The preparation method comprises the following steps:
s1: preparation of olive extract
The olive oil enters molecular modification equipment for reaction after being inspected and disinfected, meanwhile, a compressor is used for cooling, the olive oil enters membrane separation equipment after the reaction is finished, and more than 90% of high-effective components enter a mixing and stirring tank after being inspected.
S2: preparation of lyophilized powder of cytokine
Subculturing human mesenchymal stem cells to P3 generation, and collecting sterile stem cell culture supernatant after culturing in cell culture medium of P3 generation until the coverage rate is more than or equal to 80%, and storing at 4 deg.C;
adding excipients (mannitol and dextran) into the collected cell culture solution according to a ratio, and fully dissolving;
pre-freezing the dissolved culture solution at-20 ℃ for 8h, and then freeze-drying by a freeze dryer to obtain cell factor freeze-dried powder;
the cell culture medium is American ScienCell serum-free medium; the culture conditions were 37 ℃ temperature, 5% carbon dioxide concentration and 99% humidity.
S3 preparation of complexing agent
Mixing fructus Canarii albi extract, cytokine lyophilized powder, and sterile water at a certain proportion, and stirring.
Specific example 2:
a complexing agent with the function of inhibiting HPV virus comprises the following specific components in percentage by mass: 35% of olive extract, 2% of cytokine freeze-dried powder and the balance of sterile water.
The preparation method comprises the following steps:
s1: preparation of olive extract
The olive oil enters molecular modification equipment for reaction after being inspected and disinfected, meanwhile, a compressor is used for cooling, the olive oil enters membrane separation equipment after the reaction is finished, and more than 90% of high-effective components enter a mixing and stirring tank after being inspected.
S2: preparation of lyophilized powder of cytokine
Subculturing human mesenchymal stem cells to P3 generation, and collecting sterile stem cell culture supernatant and freezing at-20 ℃ for later use when the coverage rate of the stem cells is more than or equal to 80% after the stem cells are cultured in a cell culture medium of P3 generation;
adding excipients (mannitol and dextran) into the collected cell culture solution according to a ratio, and fully dissolving;
pre-freezing the dissolved culture solution at-20 ℃ for 8h, and then freeze-drying by a freeze dryer to obtain cell factor freeze-dried powder;
the cell culture medium is American ScienCell serum-free medium; the culture conditions were 37 ℃ temperature, 5% carbon dioxide concentration and 99% humidity.
S3 preparation of complexing agent
Mixing fructus Canarii albi extract, cytokine lyophilized powder, and sterile water at a certain proportion, and stirring.
Specific example 3:
a complexing agent with the function of inhibiting HPV virus comprises the following specific components in percentage by mass: 30% of olive extract, 1% of cytokine freeze-dried powder and the balance of sterile water.
The preparation method comprises the following steps:
s1: preparation of olive extract
The olive oil enters molecular modification equipment for reaction after being inspected and disinfected, meanwhile, a compressor is used for cooling, the olive oil enters membrane separation equipment after the reaction is finished, and more than 90% of high-effective components enter a mixing and stirring tank after being inspected.
S2: preparation of lyophilized powder of cytokine
Subculturing human mesenchymal stem cells to P3 generation, and collecting sterile stem cell culture supernatant and freezing at-20 ℃ for later use when the coverage rate of the stem cells is more than or equal to 80% after the stem cells are cultured in a cell culture medium of P3 generation;
adding excipients (mannitol and dextran) into the collected cell culture solution according to a ratio, and fully dissolving;
pre-freezing the dissolved culture solution at-20 ℃ for 8h, and then freeze-drying by a freeze dryer to obtain cell factor freeze-dried powder;
the cell culture medium is American ScienCell serum-free medium; the culture conditions were 37 ℃ temperature, 5% carbon dioxide concentration and 99% humidity.
S3 preparation of complexing agent
Mixing fructus Canarii albi extract, cytokine lyophilized powder, and sterile water at a certain proportion, and stirring.
Clinical trial
1. Test subjects: among 250 cases of HPV virus infections, 135 cases of low risk type HPV infections and 115 cases of high risk type HPV infections.
2. Age:
(1) of 135 patients with low risk HPV infection: minimum age 19 years, maximum age 47 years;
(2) among 115 patients with high risk HPV infection: the minimum age is 22 years, and the maximum age is 46 years.
3. The distribution of disease course:
(1) of 135 patients with low risk HPV infection: the shortest time is 2 months, and the longest time is 50 months;
(2) among 115 patients with high risk HPV infection: the shortest period is 1 month, and the longest period is 22 months.
4. Methods of observation and treatment:
the compound preparation for inhibiting HPV virus is used for 1 time a day, the hands and the pudendum are cleaned before use, one of the compound preparation is taken out, the cap of the pushing tube is removed, the knee is bent to lie on the back, the pushing tube is lightly sent into the vagina, liquid is injected into the deep part at one time, and the pushing tube is slowly drawn out and discarded.
The low-risk HPV infected patients are used for 2 months and then are subjected to the re-diagnosis; patients with high risk HPV infection are re-diagnosed after 3 months of use.
5. And (4) judging the standard:
(1) the effect is shown: HPV turns negative.
(2) Null, HPV did not turn negative.
6. The treatment results are as follows: see table 1.
TABLE 1 therapeutic results
| Group of | Number of people | Show effect | Invalidation | Total effective rate |
| Low risk type | 135 | 129 | 5 | 95.6％ |
| High risk type | 115 | 103 | 12 | 89.6％ |
Table 1 shows that: the effective rate of the gynecological gel prepared by the invention for low-risk HPV infection is 95.6 percent; the effective rate for high-risk HPV infection is 89.6%.
The compound preparation of the invention has no stimulation to vagina, safety and no side effect; the carried antivirus and antibacterial components can quickly permeate into the skin, protect mucosal cells from being invaded by viruses, quickly repair skin tissues and improve the healing and regenerating capability of wounds; the immunity of the organism can be improved, and a good antiviral effect can be achieved; in the course of viral infection, cytokines stimulate the human immune system to destroy pathogens, eliminate damaged cells, repair damaged tissues, and in this way, the cytokines protect against pathogens and inhibit the HPV virus. The compound preparation does not contain heavy metal, hormone, antibiotic and sex hormone, and olive bacteriostatic plant natural acid, and is easy to absorb and free of residue.
The present invention has been described in connection with the specific embodiments, and it is obvious that the specific implementation of the present invention is not limited by the above-mentioned manner, and it is within the protection scope of the present invention as long as various modifications are made by using the method concept and technical solution of the present invention, or the present invention is directly applied to other occasions without modification.
Claims (8)
1. The compound agent with the function of inhibiting HPV virus is characterized by comprising the following specific components in percentage by mass: 20-35% of olive extract, 0.5-2% of cell factor freeze-dried powder and the balance of sterile water.
2. The complexing agent for inhibiting HPV virus according to claim 1, wherein said olive extract is obtained by molecular modification and ion-catalyzed separation of high purity olive oil.
3. The composition with the function of inhibiting HPV virus according to claim 2, wherein the lyophilized cytokine powder is prepared from a stem cell culture solution by a freeze-drying technology.
4. The method for preparing the complexing agent with the function of inhibiting the HPV virus according to claim 3, which is characterized by comprising the following steps:
s1 preparation of olive extract
The olive oil enters molecular modification equipment for reaction after being inspected and disinfected, meanwhile, a compressor is used for cooling, the olive oil enters membrane separation equipment after the reaction is finished, and more than 90% of high-effective components enter a mixing and stirring tank after being inspected;
s2 preparation of cytokine freeze-dried powder
Subculturing human mesenchymal stem cells to P3 generation, collecting sterile stem cell culture supernatant when the coverage rate of the stem cell culture supernatant is more than or equal to 80% after the stem cell culture supernatant is cultured in a cell culture medium of P3 generation, and storing at 4 ℃ or freezing at-20 ℃ for later use;
adding excipient into the collected sterile stem cell culture supernatant according to the proportion, and fully dissolving;
pre-freezing the dissolved culture solution at-20 deg.C for 8h, and freeze-drying with a freeze-drying machine to obtain cytokine lyophilized powder;
s3 preparation of complexing agent
The olive extract, the cell factor freeze-dried powder and the sterile water are uniformly stirred according to the proportion to obtain the complexing agent product.
5. The method of claim 4, wherein in step S1, less than 10% of the effective components in membrane separation equipment are separated and used as raw materials for skin care products.
6. The method for preparing a complexing agent with HPV viral inhibiting effect according to claim 5 wherein in step S2, the excipients are mannitol and dextran.
7. The method for preparing a complexing agent with HPV viral inhibiting activity according to claim 6, wherein in step S2, the culture conditions are as follows: the temperature was 37 ℃, the carbon dioxide concentration was 5%, and the humidity was 99%.
8. The method for preparing a complexing agent with HPV viral inhibiting activity according to claim 7 wherein in step S2, the cell culture medium is ScienCell serum-free culture medium.
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| WO2024087356A1 (en) * | 2022-10-28 | 2024-05-02 | 上海市东方医院（同济大学附属东方医院） | Use of mesenchymal stem cells in preparation of drugs for treating persistent hpv infection |
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Application publication date: 20210326 |