CN108404111B - A bacteriostatic and antiviral preparation containing semen Phaseoli vulgaris phytohemagglutinin as main ingredient - Google Patents
A bacteriostatic and antiviral preparation containing semen Phaseoli vulgaris phytohemagglutinin as main ingredient Download PDFInfo
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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
- A61K38/168—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/13—Amines
- A61K31/155—Amidines (), e.g. guanidine (H2N—C(=NH)—NH2), isourea (N=C(OH)—NH2), isothiourea (—N=C(SH)—NH2)
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/702—Oligosaccharides, i.e. having three to five saccharide radicals attached to each other by glycosidic linkages
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/74—Synthetic polymeric materials
- A61K31/785—Polymers containing nitrogen
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/74—Synthetic polymeric materials
- A61K31/785—Polymers containing nitrogen
- A61K31/787—Polymers containing nitrogen containing heterocyclic rings having nitrogen as a ring hetero atom
- A61K31/79—Polymers of vinyl pyrrolidone
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/36—Polysaccharides; Derivatives thereof, e.g. gums, starch, alginate, dextrin, hyaluronic acid, chitosan, inulin, agar or pectin
- A61K47/38—Cellulose; Derivatives thereof
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/0012—Galenical forms characterised by the site of application
- A61K9/0034—Urogenital system, e.g. vagina, uterus, cervix, penis, scrotum, urethra, bladder; Personal lubricants
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/06—Ointments; Bases therefor; Other semi-solid forms, e.g. creams, sticks, gels
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/08—Solutions
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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
- A61P31/02—Local antiseptics
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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
- A61P31/04—Antibacterial agents
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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
- A61P31/10—Antimycotics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/02—Immunomodulators
- A61P37/04—Immunostimulants
Abstract
An antibacterial and antiviral preparation with kidney bean phytohemagglutinin as the main component, which belongs to the technical field of medicines. The relative weight share of each active component is 0.1-1.0 part of kidney bean phytohemagglutinin, 0.2-2.0 parts of one or two of stachyose and raffinose, and one or more than one of povidone iodine 0.1-0.5 part, polyhexamethylene biguanide 0.1-0.3 part and chlorhexidine 0.1-0.5 part; if a thickener is required, the thickener is hydroxyethyl cellulose. The invention has definite components, can be controlled quantitatively, can ensure the consistency and stability of the whole quality, can play a definite inhibiting role on bacteria, fungi and viruses, and has wider antibacterial and antiviral ranges; the product does not need to be stored at low temperature, has long storage time and is convenient to use.
Description
Technical Field
The present invention belongs to the field of medicine technology.
Background
At present, Sexually Transmitted Diseases (STDs) including HIV pose great threats to the physical and psychological health of people, and abuse of antibiotics also leads to the widespread emergence of drug-resistant bacteria. In the case of viral diseases, the problem is that the virus is of a large variety, and some viruses have a rapid variation, for example, because the HIV virus has a rapid variation, no ideal vaccine effective against HIV has been available for many years. There is an urgent need for a simple formulation with broad spectrum antibacterial and antiviral properties for daily protection and treatment. Common traumas such as burn, frostbite, incised wound, bruise, contusion and the like have good disinfection effect by iodophor. Compared with iodine tincture and alcohol, the iodine tincture has the advantages of slight irritation and pain caused by iodine, easy acceptance by patients, wide application range, and definite effect, and can be used as a substitute for skin mucosa disinfectant such as alcohol, mercuric oxide, iodine tincture, and gentian violet.
Polyhexamethylene guanidine has broad sterilization spectrum, low effective concentration, high action speed, stable property, easy water solubility, long-term bacteriostasis, no side effect, no corrosivity, no color, no odor, no toxicity, no combustion and explosion, safe use, moderate price and convenient transportation, and can be regarded as the best bactericide. In fact, the product is an environment-friendly multipurpose novel polymer and has extremely wide application in industry, agriculture, medical use and daily life. The product is a cationic polymer, and has antibacterial effect by dissolving lipid, changing permeability of bacterial cell membrane, and making metabolism in bacteria generate disorder. The wet tissue treated by the antibacterial agent has the characteristics of wide antibacterial spectrum, effectiveness on gram-positive bacteria, gram-negative bacteria and mold, no color, no smell, no irritation to skin, no toxicity and no corrosion, can be quickly sterilized, does not mildew, and has a shelf life of two years.
Chlorhexidine, also known as chlorhexidine, is chemically known as chlorhexidine, is a cationic surfactant, has quite strong broad-spectrum antibacterial and bactericidal effects, is a better bactericidal and disinfectant drug, and has stronger antibacterial effects on gram positive and negative bacteria than benzalkonium bromide. The medicine has positive charges, is adsorbed on plaque with negative charges and the surface of oral mucosa when the oral cavity gargles, then the adsorbed medicine is dispersed from the parts and gradually separated out to generate continuous action, the concentration of the medicine in saliva is reduced until 24 hours later, the medicine is adsorbed on a permeation barrier of a bacterial cell serosa to enable cell contents to leak out, the medicine has a bacteriostatic action at low concentration and a bactericidal action at high concentration, and the medicine is effective even in the presence of serum, blood and the like and has little local irritation and anaphylactic reaction. Chlorhexidine is a biguanide high-efficiency broad-spectrum bactericide, plays a role in sterilization by changing the permeability of a bacterial cytoplasmic membrane, has stronger sterilization effect than quaternary ammonium salt cationic surfactants, has stronger effect on gram-positive bacteria than negative bacteria, is also effective on pseudomonas aeruginosa and fungi, but has no effect on acid-resistant bacteria, spores and viruses.
The Chinese patent document with the application number of 2013100565888 discloses an application of kidney bean phytohemagglutinin in preparing human medicines and a pharmaceutical composition thereof, wherein the medicine takes kidney bean phytohemagglutinin, tremella polysaccharide, stachyose or/and raffinose and bletilla gum, or extracts of scutellaria baicalensis, berberis julianensis and sophora flavescens as active components, and the product does not need to be stored at low temperature, has long storage time and convenient use, but the antibacterial and antiviral abilities and the quality stability of the product need to be further improved.
Disclosure of Invention
Compared with the prior art, the bacteriostatic antiviral preparation which takes the kidney bean phytohemagglutinin as the main component has the advantages of no need of low-temperature storage, long storage time, convenient use, stronger antibacterial and antiviral abilities and easy quality control.
The preparation comprises active ingredients, by weight, 0.1-1.0 part of kidney bean phytohemagglutinin, 0.2-2.0 parts of stachyose and/or raffinose and one or more than one of povidone iodine 0.1-0.5 part, polyhexamethylene biguanide 0.1-0.3 part and chlorhexidine 0.1-0.5 part; if a thickener is required, the thickener is hydroxyethyl cellulose.
The kidney bean phytohemagglutinin can be white kidney bean phytohemagglutinin or red kidney bean phytohemagglutinin.
Because the chemical properties of the active ingredients in the preparation of the invention are different, the preparation is difficult to be matched with most thickeners, and through a large amount of screening and combination, the hydroxyethyl cellulose can be finally determined to be used together with the components, and the chemical properties of the active ingredients and the biological activity of the agglutinin are not changed.
The povidone iodine, the polyhexamethylene biguanide and the chlorhexidine are medicinal products.
The preparation of the invention can be water-soluble compound liquid or gel, and can also be solid preparation such as suppository, sustained release agent, disintegrating tablet and other pharmaceutical preparations suitable for external use, vagina and rectum.
The preparation of the invention is mainly used for external application, has the effects of preventing and treating bacterial, fungal and viral infection on the body surface or mucosa of a human body, can also be used for preventing and treating other diseases of the human body, such as treatment of wounds and the like, and can also be used as a contraceptive.
If the preparation is a gel liquid, 100ml of the gel liquid contains the following components:
0.5-1.0 g of kidney bean phytohemagglutinin,
0.5 to 2.0 g of stachyose and/or raffinose,
0.1 to 0.5g of povidone iodine,
0.1 to 0.3 g of polyhexamethylene biguanide,
0.1 to 0.5g of chlorhexidine,
0.8 to 0.9 g of NaCl,
the pH buffering agent is in an amount acceptable for a pharmaceutically gelled fluid,
the stabilizer is the amount acceptable by the gel liquid in pharmacy,
thickening agent: 1.2-2.0 g of hydroxyethyl cellulose.
If the preparation is liquid, 100ml of the preparation liquid contains the following components:
0.1-1.0 g of kidney bean phytohemagglutinin,
0.5 to 2.0 g of stachyose and/or raffinose,
0.1 to 0.5g of povidone iodine,
0.1 to 0.3 g of polyhexamethylene guanidine or biguanide,
0.1 to 0.5g of chlorhexidine,
0.8 to 0.9 g of NaCl,
the pH buffering agent is in an amount acceptable for a pharmaceutically gelled fluid,
the stabilizer is in an amount acceptable for pharmaceutical gel solutions.
The compound medicine is prepared by organically matching the components and the dosage according to the pathological state of an organism and is used for symptomatic treatment of multiple parts and different conditions.
The gel liquid can be used by cleaning wounds or the outside of the vagina and the vaginal orifice with liquid or warm water or normal saline according to the proportion of 1:10 or 1: 100-200, and then applying the gel liquid.
For external application, the composition can be in liquid form, or in gel form for vaginal or anal/rectal application.
The kidney bean agglutinin used in the experiments and examples below was a white kidney bean or red kidney bean phytoagglutinin. The lectin is extracted from white kidney bean and red kidney bean produced in Yunnan province by the patent applicant using the granted patent technology (application number is 200910094233.1).
The antibacterial action of the product is realized by selecting the combination of chemical bacteriostats of povidone iodine, polyhexamethylene biguanide and chlorhexidine which have definite antibacterial action, long-term use history and are proved to be effective and safe. The proportions of the various components can be adjusted within the concentration range used. For example, to prepare a less coloured gel with anti-bacteriostatic action, three bacteriostatic agents may be used simultaneously, at which time the minimum concentration of each of the three components is 0.1%, namely 0.1 g of povidone-iodine, 0.1 g of polyhexamethylene monoguanidine or biguanide, and 0.1 g of chlorhexidine, the sum of the three components being 0.3 g, and 100ml of a gel or liquid bacteriostatic solution may be prepared after being combined with the other components; if two components are adopted, each component is formed by combining two single components with each component being 0.15-0.2 g, namely 100ml of gel or liquid needs two bacteriostasis and the total amount is not less than 03-0.4 g; if only one bacteriostatic agent is used for preparation, the single bacteriostatic component required by 100ml of gel or liquid is 0.5g, and the antibacterial or bacteriostatic effect can reach 95-100%. The antiviral effect of the preparation is achieved by adding kidney bean phytohemagglutinin. The effects of directly resisting virus or enhancing immunity of organism can be achieved by direct action and indirect action of kidney bean phytohemagglutinin. In addition, because of the sugar binding property of lectin and the characteristics of agglutinable blood and sperm proved by previous research, the preparation has contraceptive effect when used in vagina. Other beneficial effects are also manifested in:
1. phytohemagglutinin is a mitogen which, at low concentrations, promotes cell division and thus wound healing.
2. The kidney bean phytohemagglutinin extracting solution also contains estrogen kidney bean isoflavone, and relevant researches at home and abroad show that the estrogen can reverse the atrophy change of the vaginal wall to relieve symptoms of senile vaginitis patients, the thickness of vaginal epithelium and the elasticity of the vaginal wall are increased by the supplement of the estrogen, the pH value of the vagina is reduced, and the self-cleaning effect of the vagina is recovered;
3. besides activating T lymphocytes, kidney bean agglutinin can stimulate B lymphocytes to produce interferon, interleukin and the like, and improve the humoral immunity level.
4. Our previous findings indicate that vaginal and rectal administration of kidney bean agglutinin can increase the conversion rate of T lymphocytes.
The product of the invention is suitable for skin or mucosa wound, bacterial or viral infection.
Compared with the prior art, the invention has the advantages that the components are clear, the quantitative control can be realized, the consistency and the stability of the whole quality can be ensured, the definite inhibition effect on bacteria, fungi and viruses can be realized, and the antibacterial and antiviral ranges are wider; the product does not need to be stored at low temperature, has long storage time and is convenient to use.
Experiment 7 below also shows that the agglutination time of the red blood cells by the preparation of the invention is much shorter than the agglutination time of the lectins alone, which is one of the unexpected benefits.
Experiment 1, extraction, purification and activity determination of lectins such as white kidney beans.
(1) The extraction and purification of lectins such as white kidney bean or red kidney bean are carried out by adopting the technology disclosed in Chinese patent document with the application number of 200910094233.1, and the protein content is measured by adopting a general double venation contraction method.
(2) The lectin activity was measured by the technique disclosed in chinese patent application No. 201110184061.4.
Experiment 2, inhibition effect of kidney bean agglutinin and chemical bacteriostatic agent combined gel on escherichia coli
First, equipment
1. The strain Escherichia coli 8099, 5 th to 6 th generations, is provided by the common microorganism center of China Committee for culture Collection of microorganisms.
2. The test sample is kidney bean agglutinin bacteriostatic lubricating gel, and the production date or batch number 2015 is 8 months and 15 days.
Second, method
1. The test is carried out according to the test method of the bacteriostatic performance of the dissolution antibacterial product in appendix C4 of GB15979-2002 hygienic Standard for Disposable sanitary articles.
2. Performing antibacterial lubrication on gel Escherichia coli stock solution with kidney bean agglutinin for 2min,5min, lOmin, and 20min, respectively, and repeating the experiment at 20s and 1 deg.C for 3 times.
Three, result in
Under the condition of 20 seconds and 1 ℃, the results of three repeated tests show that the kidney bean agglutinin bacteriostatic lubricating gel stock solution acts on escherichia coli for 2 minutes, the average bacteriostatic rate is 100.00 percent, and the results are shown in the following table:
bacteriostatic action of kidney bean agglutinin bacteriostatic lubricating gel stock solution on escherichia coli
Fourth, conclusion
The kidney bean agglutinin bacteriostatic lubricating gel stock solution acts on escherichia coli for 2min, the average bacteriostatic rate is 100.00%, and the product stock solution has a strong bacteriostatic action on the escherichia coli according to the evaluation standard of GB15979-2002 sanitation Standard for Disposable sanitary articles.
Experiment 3, inhibition effect of kidney bean agglutinin and chemical bacteriostatic agent combined gel on staphylococcus aureus
First, equipment
1. The strain is staphylococcus aureus ATCC6538, 5 th to 6 th generations, and is provided by the common microorganism center of China Committee for culture Collection of microorganisms.
2. The test sample is kidney bean agglutinin bacteriostatic lubricating gel, and the production date or batch number 2015 is 8 months and 15 days.
Second, method
1. The test is carried out according to the test method of the bacteriostatic property of the dissolution type anti-bacteria product in appendix C4 of GB15979-2002 hygienic Standard for Disposable sanitary articles.
2. Lubricating gel Staphylococcus aureus stock solution with kidney bean agglutinin antibacterial agent, respectively acting on Staphylococcus aureus for 2min,5min, 10min, and repeating the test at 20 + -10 deg.C for 3 times.
Three, result in
Under the condition of 20 seconds and 1 ℃, the results of three repeated tests show that the kidney bean agglutinin bacteriostatic lubricating gel stock solution acts on staphylococcus aureus for 2min, the average bacteriostatic rate is 100.00 percent, and the results are shown in the following table:
bacteriostatic action of kidney bean agglutinin bacteriostatic lubricating gel stock solution on staphylococcus aureus
Fourth, conclusion
The kidney bean agglutinin bacteriostatic lubricating gel stock solution acts on staphylococcus aureus for 2min, the average bacteriostatic rate is 100.00%, and the product stock solution has strong bacteriostatic action on the staphylococcus aureus according to the evaluation standard of GB15979-2002 hygienic standard for disposable sanitary articles.
Experiment 4, the inhibition effect of the kidney bean agglutinin and chemical bacteriostatic agent combined gel on candida albicans
First, equipment
1. The strain is Candida albicans ATCC10231, 5 th to 6 th generations, and is provided by the common microorganism center of China Committee for culture Collection of microorganisms.
2. The test sample is kidney bean agglutinin bacteriostatic lubricating gel, and the production date or batch number 2015 is 8 months and 15 days.
Second, method
1. The test is carried out according to the test method of the bacteriostatic performance of the dissolution antibacterial product in appendix C4 of GB15979-2002 hygienic Standard for Disposable sanitary articles.
2, using kidney bean agglutinin to inhibit bacteria and lubricate gel candida albicans stock solution, respectively acting on candida albicans for 2 min-5 min, lOmin for 20min, and repeating the test for 3 times at 20 ℃ and 1 ℃.
Three, result in
Under the condition of 20 seconds and 1 ℃, the results of three repeated tests show that the kidney bean agglutinin bacteriostatic lubricating gel stock solution acts on candida albicans for 2 minutes, the average bacteriostatic rate is 100.00 percent, and the results are shown in the following table:
bacteriostatic action of kidney bean agglutinin bacteriostatic lubricating gel stock solution on candida albicans
Fourth, conclusion
The kidney bean agglutinin bacteriostatic lubricating gel stock solution has an effect on candida albicans for 2min, the average bacteriostatic rate is 100.0096, and the product stock solution has a strong bacteriostatic effect on the candida albicans according to the evaluation standard of GB15979-2002 hygienic standard for disposable sanitary products.
Experiment 5, stability test of bacteriostasis of kidney bean agglutinin and chemical bacteriostat combined gel
The gel is stored for 90 days at 37 ℃, and the antibacterial effect is respectively tested by using the methods of experiments 2 to 4.
First, equipment
1. The strains are Escherichia coli 8099 (5 th-6 th generation), Staphylococcus aureus ATCC6538 (5 th-6 th generation) and Candida albicans ATCC10231 (5 th-6 th generation), which are provided by the common microorganism center of China Committee for culture Collection of microorganisms.
2. The test sample is kidney bean agglutinin bacteriostatic lubricating gel, and the production date or batch number 2015 is 8 months and 15 days.
Second, method
1. The test was carried out according to the stability test method of appendix C6 of GB15979-2002 hygienic Standard for Disposable sanitary articles.
2. And (3) stability test, namely storing the kidney bean agglutinin antibacterial lubricating gel at 37 ℃ for 90 days, respectively inhibiting escherichia coli, staphylococcus aureus and candida albicans for 2min,5min and lOmin for 20min by using stock solution, and repeating the test at 20 +/-10 ℃ for 3 times.
Three, result in
The results of three repeated tests at the temperature of 20 +/-1 ℃ show that after the kidney bean agglutinin antibacterial lubricating gel is stored for 90 days at the temperature of 37 ℃, stock solution of the kidney bean agglutinin antibacterial lubricating gel acts on escherichia coli, staphylococcus aureus and candida albicans for 2min,5min and lOmin and 20min, the average antibacterial rate is 100.00%, and the results are shown in tables 1-3 below.
TABLE 1 bacteriostatic action of kidney bean agglutinin bacteriostatic lubricating gel on escherichia coli after heat preservation
TABLE 2 bacteriostatic action of kidney bean agglutinin bacteriostatic lubricating gel on staphylococcus aureus after heat preservation
TABLE 3 bacteriostatic action of Kidney Bean agglutinin bacteriostatic lubricating gel stock solution on Candida albicans after incubation
Fourth, conclusion
The kidney bean agglutinin bacteriostatic lubricating gel is stored for 90 days at 37 ℃, the stock solution acts on escherichia coli, staphylococcus aureus and candida albicans for 2min,5min, l0min and 20min, the average bacteriostatic rate is 100.00%, and the bacteriostatic action of the product can be maintained for at least 2 years at room temperature.
Experiment 6, kidney bean agglutinin chemical bacteriostat combined gel for multiple times of vagina mucosa irritation test
1. Test materials and methods
(1) The test substance is kidney bean agglutinin bacteriostatic lubricating gel. Test group, sample as it is as the test object; control group was sterilized normal saline.
(2) Animals, i la rabbits 4, a common grade, provided by kunzhu technologies ltd, and animals, SCXK (yunnan) K2012-00020 feed, provided by kunzhu technologies ltd, and SCXK (yunnan) K2012-0003. Animals were examined in vivo for no abnormalities.
(3) The environmental conditions are that the temperature is 20-23 ℃ and the relative humidity is 40-60 percent. The facility certificate number is SYXK (Dian) K2015-0004.
(4) The test method is based on the vaginal mucosa irritation test of 2.3.5 in the Disinfection technical Specification of the Ministry of health (2002). The test was set with a test group and a negative control group, each group containing 3 animals. The animal is fixed on its back to expose perineum and vaginal opening. The catheter was wetted with the test solution and gently inserted into the vagina for 5cm, and 2mL of the test substance was slowly injected with a syringe. Control animals were treated with sterile saline as well. The infection is repeated every 24h for 5 days continuously. 24h after the last infection, the animals are killed by an air embolism method, the complete vagina is taken out after the laparotomy, the vagina is longitudinally cut open, whether congestion, edema and other phenomena exist or not is observed by naked eyes, the vagina is put into 10% formalin solution for fixation for more than 24h, tissues at two ends and 3 parts in the center of the vagina are selected for flaking, and histopathology examination is carried out after HE staining.
2. Test results
Gross dissection without obvious congestion, edema and erosion.
Glycopectin antibacterial lubricating gel vaginal mucosa irritation response score
Note that the average integral (stimulation response integral) is added and divided by the total number of observations (total number of observations x 3).
Stimulation index-average integral of test group-average integral of control group-5.20-0.56-4.64
3. Conclusion
Under the experimental condition, the kidney bean agglutinin combined gel is judged to be extremely light irritability according to the grading standard of vaginal mucosa irritation strength when grading the result of multiple vaginal mucosa irritation tests of the Yila rabbit.
Experiment 7, test of agglutination of erythrocyte by kidney bean agglutinin chemical bacteriostatic agent combined gel
The gel was diluted with a balanced salt solution (PBS) to a dilution containing 1mg/ml of lectin, and then diluted with PBS to a dilution of 1/256 in multiples, and then added to the first 8 wells of a 9-well plate, wherein the 9 th well of the 9-well plate was a PBS control group. Human or mouse blood was taken and washed with PBS solution and erythrocytes, and then prepared into 4% erythrocyte solution. Adding 4% of erythrocyte liquid into 1-9 holes of a 9-hole plate according to the volume ratio of 1: 1. Standing at the room temperature of 20-25 ℃ for 60-120 minutes, and observing the agglutination condition of each hole. The test result shows that the agglutination activity is between 1/128 and 1/256 mg. The combination has no influence on the agglutination activity, and a new phenomenon that the agglutination time is greatly shortened compared with the agglutination generated after the single lectin acts is found, the agglutination phenomenon generated after the single lectin acts is generally 60 minutes, and the time for the gel to cause the erythrocyte to gel can be generated 5 minutes after the reaction, namely the gel reaction is accelerated by more than 10 times. Sigma-Aldrich is the largest chemical reagent manufacturer worldwide and Sigma-Aldrich is a high-tech business leading biological/chemical reagents. The agglutination activity was determined according to the standard of Sigma-Aldrich, USA: (Agglutination activity is expressed in μ g/mL and is determined from the series of reactions of a 1mg/mL solution. this activity is the local convergence of the formation of the Agglutination of a 2% subset of human analytes in a phosphorus soluble salt, pH 6.8, after 1hr accumulation at 25. where http:// www.sigmaaldrich.com/ligand/product/sigma/l 2646. The gel compounded by the method greatly shortens the agglutination time. Such a result is extremely advantageous for the application of the combination gel, for example in contraception, by rapidly agglutinating sperm, increasing the contraceptive effect of the gel; in the aspect of wound hemostasis, because red blood cells are rapidly agglutinated, the bleeding of small wounds can be rapidly stopped.
Experiment 8, semen Phaseoli vulgaris agglutinin chemical bacteriostatic agent combined gel for killing sperm and agglutinating
Human and dog sperm were used for this experiment. Human sperm was collected by masturbation of a healthy human body, suspended in EF solution [ whole pine, Zhou Hai Width, mountain wild department, etc. ] and frozen in an electrolyte-free solution for fertilization ability of human sperm [ J ]. proceedings of first university of military medicine, 2002, 22(10):928-930 ], and stored at 4 ℃ for use within 1 hour. The dog sperm is artificially collected and stored and used by the same method as the above method.
According to the method of Sander-Cramer et al [ Bharitkar Y P, Banerjee M, Kumar S, et al, search for a needle microbial peptide from the isolates of Shorearobusta stress [ J ]. Contraception, 2013, 88(1):133-140 ], 1 volume of semen was taken and mixed thoroughly with 5 volumes of RKBL gels of different concentrations, respectively. That is, 100. mu.l of semen was mixed with 500. mu.l of xanthan gum gel containing 5, 2.5, 1, 0.5, 0.25, 0.1, 0.05, 0.025, 0.01, 0.005, 0.0025, 0.001(mg/ml) RKBL, respectively, immediately blown uniformly, 5. mu.l each was taken out within 20s, observed under a microscope of magnification X100, observed again after 5min, and if 100% of motile sperm were observed to be agglutinated after 5min, this was the final result, otherwise, observation was repeated every 20min thereafter, and observation was continued for 3 hours. The experiment was repeated 7 times.
The results of spermicidal and agglutination experiments show that the gel containing 5mg/ml kidney bean agglutinin can inactivate sperms within 20s, and RKBL concentration in the gel is higher than or equal to 1mg/ml, so that the sperms lose their activity within 20s, and brake and lethal effects are generated, but no agglutination phenomenon is generated. The gels containing 0.5mg/ml, 0.25mg/ml, 0.1mg/ml, 0.05mg/ml, 0.025mg/ml, 0.01mg/ml RKBL were all able to induce the agglutination of the sperm. At a concentration of 0.05mg/ml, the sperm aggregation exceeded 50%. The contraceptive function can be effectively exerted according to the sperm agglutination standard established by a WHO human sperm examination and treatment laboratory manual (2011, fifth edition).
Detailed Description
Example 1: and (5) formula A. The gel solution contains the following components in 100 ml:
0.4-0.5 g of white kidney bean phytohemagglutinin,
0.5g of oligosaccharide (the total amount of stachyose and raffinose),
chemical bacteriostatic agent: 0.2 to 0.5g of povidone iodine,
chemical bacteriostatic agent: 0.1 to 0.3 g of polyhexamethylene guanidine,
chemical bacteriostatic agent: 0.1 to 0.3 g of chlorhexidine acetate,
0.85 g of NaCl is added to the solution,
pH buffer is acetic acid-sodium acetate buffer (pH4.5, taking 1.8 g of sodium acetate, adding glacial acetic acid 0.98m1,
then adding water to dilute to 100ml, thus obtaining the product,
the stabilizing agent is PEG 40000.5-2.0 g,
1.4-2.0 g of thickening agent hydroxyethyl cellulose,
the pH value is 4.3-4.8.
Example 2: and (5) preparing a formula B. The gel solution contains the following components in 100 ml:
0.4-0.5 g of phytohemagglutinin of red kidney bean,
0.2-0.5 g of oligosaccharide (total amount of stachyose and raffinose),
chemical bacteriostatic agent: 0.2 to 0.5g of povidone iodine,
chemical bacteriostatic agent: 0.1 to 0.3 g of polyhexamethylene guanidine,
0.7 to 0.85 g of NaCl,
pH buffer is acetic acid-sodium acetate buffer (pH4.5, taking 1.8 g of sodium acetate, adding glacial acetic acid 0.98m1, adding water and diluting to 100ml,
the stabilizing agent is PEG 40000.5-2.0 g,
1.2-2.0 g of thickening agent hydroxyethyl cellulose
The pH value is 4.3-4.8
Example 3: the thickening agent is subtracted from the formulas A and B to obtain the kidney bean agglutinin chemical bacteriostatic (washing) solution.
Phytohemagglutinin 0.4-0.5 g,
0.3 to 0.5g of oligosaccharide,
chemical bacteriostatic agent: 0.2 to 0.5g of povidone iodine,
chemical bacteriostatic agent: 0.1 to 0.3 g of polyhexamethylene guanidine.
Example 4, detection of bacteriostatic action of combinations of two or three bacteriostatic agents and lectin at different concentrations the strains and methods of experiments 2,3 and 4 were used, treatment time was 5min, and detection was performed at 37.0 ± 1 ℃. The results are shown in tables A, B and C below. The results show that the bacteriostasis rate of the bacteriostat with different combinations can reach more than 95 percent under the concentration of 0.2 percent of one or two of 0.1 percent PHA, stachyose and raffinose.
Inhibition of Escherichia coli by a bacteriostatic wetting solution containing 0.1% of kidney bean agglutinin and other bacteriostatic components
B.0.1% kidney bean agglutinin and other bacteriostatic component combined bacteriostatic wetting liquid for inhibiting Staphylococcus aureus
C.0.1% kidney bean agglutinin and other bacteriostatic component combined bacteriostatic moistening liquid for inhibiting Candida albicans
Example 5: can be used for treating female vaginal pruritus and abnormal odor, or daily cleaning and caring male and female vulva. The bacteriostatic solution of the embodiment 3 is firstly directly sprayed or smeared on the vulva and the vaginal orifice, or warm water is added according to the proportion of 1:00 or 1:200 for even blending, and the vulva is cleaned. And then 2-5 ml of the gel of the formula A or B is pushed into the vagina. The traditional Chinese medicine composition is used for 1-2 times every day, so that abnormal leucorrhea can be reduced, and the effect of eliminating bad symptoms such as peculiar smell and pruritus is obvious. The gel has definite effect after being actually used by tens of people.
The bacteriostatic solution prepared in the embodiment 3 is used and mixed with warm water according to the proportion of 1:00 or 1:200, and can be used for daily cleaning and nursing male pudendum and female pudendum.
Example 6: lubrication and contraceptive effect. Because the gel has the functions of killing erythrocytes and sperms at high concentration and agglutinating at low concentration, the gel A and the gel B can play good roles in lubrication and contraception. Experiments prove that the concentration of the agglutinin gel is 1mg/ml to 5 mg/ml. Spraying the vaginal opening with a pressing bottle or a gynecological pusher of 4.5-5.0 g before sexual life or pushing the vaginal opening with the gynecological pusher of 4.5-5.0 g. From 9 months 2015 to 9 months 2017, over 100 people have used this new gel for lubrication and contraception, with no 1 failure.
Example 7: anti-herpes virus effect
In 2017, in 9 months, a single example of the pudendum herpes simplex virus is treated. In the treatment of certain female, a local clustered small blister appears at perineum part, the small blister is broken by a 5# sterilization injection needle, liquid is squeezed out by soft pressure by a disinfection cotton swab as much as possible, then gel is smeared, the medicine is continuously applied for 7 days in the morning and at night every day, and then the herpes part is healed.
Example 8: therapeutic effect on bovine and porcine foot-and-mouth disease virus
Therapeutic observations of bovine and porcine foot and mouth disease in mile city and yilian county, respectively, in 2016 were made, summarized as follows:
oral treatment: the oral cavity is washed by 0.1 to 0.2 percent potassium permanganate solution or 2 to 3 percent acetic acid or vinegar, then gel is coated on the ulcerated surface and is coated once every other day, and the ulcerated surface is diluted by 1:100 and normal saline and then added into drinking water to be taken by sick pigs and cattle, so that the oral liquid can achieve obvious effect after 8 to 14 days. The total effective rate is about 85%.
Hoof treatment: soaking the hooves in 3 to 5 percent copper sulfate solution. After drying, the composition is smeared with gel once a day or every other day, diluted by adopting 1:100 and normal saline, added into drinking water to be taken by sick pigs and cattle, and obvious effect can be seen after continuous use for 10-15 days. The total effective rate is about 80%.
Treatment of breast: milking is done by conventional sterilization, rinsing with warm water, and then applying the gel once a day or every other day. 8-12 can see obvious effect. The total effective rate is about 80%. The number of cattle treated by the method is about 30, and the number of pigs treated by the method is about 50, and the result shows that the effective rate of the gel treatment is improved by more than 30 percent compared with the traditional treatment method.
Claims (1)
1. An antibacterial and antiviral preparation with kidney bean phytohemagglutinin as main component is characterized by being gel liquid, wherein 100ml of the gel liquid contains the following components:
0.5-1.0 g of kidney bean phytohemagglutinin,
0.5 to 2.0 g of stachyose and/or raffinose,
0.1 to 0.5g of povidone iodine,
0.1 to 0.3 g of polyhexamethylene biguanide,
0.1 to 0.5g of chlorhexidine,
0.8 to 0.9 g of NaCl,
the pH buffering agent is in an amount acceptable for a pharmaceutically gelled fluid,
the stabilizer is the amount acceptable by the gel liquid in pharmacy,
thickening agent: 1.2-2.0 g of hydroxyethyl cellulose.
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| CN111450235A (en) * | 2020-06-05 | 2020-07-28 | 杨正东 | Contraceptive gel jelly |
| CN113230205B (en) * | 2021-05-22 | 2023-07-07 | 绍兴鸿瑞生物科技有限公司 | Gynecological gel preparation and preparation method and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101508730A (en) * | 2009-03-18 | 2009-08-19 | 王敏康 | Extract method for lectin of leguminous plants |
| CN103157095A (en) * | 2013-02-22 | 2013-06-19 | 云南康洲生物科技有限公司 | Kidney bean phytolectin applications in preparation of human drugs and drug composition thereof |
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101508730A (en) * | 2009-03-18 | 2009-08-19 | 王敏康 | Extract method for lectin of leguminous plants |
| CN103157095A (en) * | 2013-02-22 | 2013-06-19 | 云南康洲生物科技有限公司 | Kidney bean phytolectin applications in preparation of human drugs and drug composition thereof |
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| 芸豆植物凝集素的提取及血凝效果研究;郭春生等;《中国兽药杂志》;20080131;第42卷(第1期);第49-52页 * |
| 阴道或直肠注入芸豆植物凝集素的亚慢性实验研究;王昌梅等;《中国预防医学杂志》;20110430(第4期);第299-302页 * |
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