CN103751105A - Application of ginkgolic acid nanometer liposome in treatment of gynecological diseases and venereal diseases - Google Patents
Application of ginkgolic acid nanometer liposome in treatment of gynecological diseases and venereal diseases Download PDFInfo
- Publication number
- CN103751105A CN103751105A CN201310408906.2A CN201310408906A CN103751105A CN 103751105 A CN103751105 A CN 103751105A CN 201310408906 A CN201310408906 A CN 201310408906A CN 103751105 A CN103751105 A CN 103751105A
- Authority
- CN
- China
- Prior art keywords
- nanometer liposome
- gingkolic acid
- acid nanometer
- preparation
- gingkolic
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Landscapes
- Medicinal Preparation (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Cosmetics (AREA)
Abstract
The invention relates to a use of ginkgolic acid nanometer liposome in preparation of external application preparations for treatment of gynecological diseases and venereal diseases. According to the present invention, the ginkgolic acid can be obtained from extracts of the exopleura part of gingko through a chemical separation method; the ginkgolic acid nanometer liposome is prepared by using a film ultrasound-high pressure homogenization method; the gynecological diseases are infectious and inflammatory genitalia diseases; the drug or the disinfectant external application preparation containing the ginkgolic acid nanometer liposome and provided for treatment of gynecological diseases and venereal diseases contains 0.001-80% of the ginkgolic acid nanometer liposome and a pharmaceutically and commonly used external application preparation substrate or an acceptable carrier; the preparation is the any one external application formulation in the pharmacy; and results of pharmacology studies and clinical trials show that the ginkgolic acid nanometer liposome provides prominent treatment effects for a variety of gynecological diseases and venereal diseases, wherein the effective rate is 100%, and characteristics of safe use and no side effect are provided.
Description
Technical field
The invention belongs to biological medicine technology field, be specifically related to the purposes of gingkolic acid nanometer liposome in the external preparation of preparation treatment gynaecopathia and sexually transmitted disease (STD).
Background technology
Gynaecopathia and sexually transmitted disease (STD) are one of clinical modal diseases, and in various places, have the trend rising year by year.Because this type of disease persistent period is long, easily outbreak repeatedly, brings misery to patient, has a strong impact on patient's physical and mental health.This type of disease is many with genitals and to reside pathogenic infection around relevant, as infection such as gonococcus, herpesvirus, human papillomavirus (HPV), treponema pallidum, pubic louses, cause that multiple sexually transmitted disease (STD) is as gonorrhea, genital herpes, condyloma acuminatum, syphilis, pubic louse etc.; Gynaecopathia is as trichomonal vaginitis, colpitis mycotica, bacterial vaginitis, pudendal pruritus, cervicitis, cervical erosion etc.At present this type of disease for the treatment of mostly is antimicrobial drug, antiviral agents or antifungal agent, and corticosteroid hormone and immunosuppressant, though these medicines have effect for the moment, and easily recurrence after slow, the drug withdrawal of taking effect; And prolonged application can cause many side effect.For fear of the generation of this type of bad phenomenon, market demand develop a kind of derive from natural, effect significantly, without the treatment gynaecopathia of obvious adverse reaction and the external preparation of sexually transmitted disease (STD), to alleviate patient suffering.These type of kinds of Diseases are many and patient is numerous, and therefore the external preparation product of this type of disease of exploitation treatment has good potential applicability in clinical practice, to patient, is very useful and necessary.
Semen Ginkgo is the medicinal plants of China's preciousness, and leaf, fruit and the episperm etc. of Semen Ginkgo all have medicinal exploitation and are worth.According to < < Compendium of Materia Medica > >, record, Semen Ginkgo property is sweet, bitter, flat, puckery; Have that relieving asthma and cough, leukorrhagia stopping are turbid, the merit of pain relieving; Cure mainly cough of cold type phlegm dyspnea, asthma, frequent micturition, leucorrhea with red and white discharge, rhagadia manus et pedis etc.Particularly it is found that in recent decades in Folium Ginkgo and contain plurality of active ingredients, main effective ingredient is flavonoid and terpene lactone compound, has the function of control cardiovascular and cerebrovascular disease.China has gingko episperm soak is used as pesticide morning among the people, and the insects such as aphid, rice borer are had to killing action.
Gingkolic acid is colourless or flaxen oily or powdered substance, is the component that is mainly present in a class in gingko episperm and has important physiologically active.Gingkolic acid involved in the present invention is alkyl or alkenyl phenol acid, according to whether being connected with carboxyl on phenyl ring in molecule, is divided into ginkgoic acid and the large class of ginkgol two.Ginkgoic acid is the series compound that salicylic acid molecule has saturated or unsaturated chain alkyl on phenyl ring C6 position, by chemical constitution, can be divided into 5 kinds, and side chain R is C
13h
27, C
15h
29, C
15h
3l, C
17h
31and C
17h
33, corresponding molecular weight is respectively 320,346,348,372,374.Gingkolic acid comprises ginkgolic acid, hydroginkgoic acid, Semen Ginkgo eo-acid, the sub-acid of Semen Ginkgo, hydroginkgolinic acid, anacardic acid, ginkgol, Semen Ginkgo diphenol, Ginnol etc.Gingkolic acid (ginkgoic acid, ginkgol, Semen Ginkgo diphenol) structure is as follows.
the structure of gingkolic acid
Nanometer liposome is the ultra micro spheroid carrier preparation that drug encapsulation is formed in lipoids bilayer.Liposome, as a kind of novel pharmaceutical carrier, has obtained development rapidly in preparation for external application to skin research and application aspect in recent years.Nanometer liposome as the feature of percutaneous drug delivery carrier is: compatibility, can strengthen the transdermal effect of medicine; Skin histology targeting; Slow-releasing; Liposomal encapsulated drug solubility and the stability of improving.The advantage such as that nanometer liposome external preparation has is efficient, long-acting, safety, skin targeting are strong.Exploitation external preparation for skin nano liposome preparations is to improve enepidermic trend, has become the direction of external used medicine and cosmetics research and development.
Have not yet to see the report of the application of gingkolic acid nanometer liposome in treatment gynaecopathia and sexually transmitted disease (STD).Therefore, about gingkolic acid nanometer liposome external preparation is prevented and treated the research and development of gynaecopathia and sexually transmitted disease (STD) newtype drug, cosmetics, disinfectant, and the abundant exploitation to gingko resource, will produce huge society and economic benefit.
Summary of the invention
The object of this invention is to provide the new purposes of gingkolic acid nanometer liposome, gingkolic acid nanometer liposome is prevented and treated the purposes in the external preparation of gynaecopathia and sexually transmitted disease (STD) in preparation.It has evident in efficacy, use safe feature.
The object of the present invention is achieved like this:
The purposes of gingkolic acid nanometer liposome in the external preparation of preparation treatment gynaecopathia and sexually transmitted disease (STD) is provided.
Described gingkolic acid comprises ginkgoic acid, ginkgol, Semen Ginkgo diphenol, can be by Chemical Decomposition method, from the extract at the positions such as the episperm of Semen Ginkgo, obtain; Or gingko episperm is extracted to elaboration products and in the extraction course of processing, in remaining waste material, again reclaim; Or obtain the each several part extract that contains this compounds from other plant; Or obtain by the method for chemosynthesis.The materials such as claimed this compounds of the present invention and pharmaceutically acceptable salt thereof, ester, solvate, crystal habit.
Described gingkolic acid nanometer liposome, by adopting the preparation of thin-film ultrasonic-high pressure homogenization method.
Described sexually transmitted disease (STD) is infectious and struvite genital diseases, as multiple sexually transmitted disease (STD)s such as gonorrhea, syphilis, condyloma acuminatum, genital herpes, human papillomavirus (HPV) infection, pubic louses.Described gynaecopathia is trichomonal vaginitis, colpitis mycotica, bacterial vaginitis, pudendal pruritus, cervicitis, cervical erosion etc.
The treatment gynaecopathia that contains gingkolic acid nanometer liposome and medicine, the disinfectant external preparation of sexually transmitted disease (STD) are also provided, contain gingkolic acid nanometer liposome 0.001-80%, and conventional external preparation substrate or external preparation acceptable carrier pharmaceutically.
Further, contain gingkolic acid nanometer liposome 0.001-30%.
Further, contain gingkolic acid nanometer liposome 0.5-20%.
By above-mentioned weight ratio scope get principal agent gingkolic acid nanometer liposome, pharmaceutically conventional external preparation substrate or acceptable carrier prepares gingkolic acid nanometer liposome external preparation; And comprise medicine, the disinfectant external preparation that gingkolic acid nanometer liposome and any combinations of substances are used for the treatment of gynaecopathia and sexually transmitted disease (STD).Described preparation is said exterior-applied formulation on any pharmaceutics, is liquid preparation, cream cream, gel, membrane, lotion, powder, aerosol spray, suppository, transdermal absorption formulation.
The invention has the beneficial effects as follows, pharmacology test research shows, gingkolic acid nanometer liposome external preparation of the present invention has significant therapeutic effect to rat bacterial vaginitis, colpitis mycotica, cervicitis model tool, and can suppress the propagation of human papillomavirus (HPV) infection cell, also there is the remarkable effect of killing the resident pathogen (as gonococcus, trichomonal vaginitis, herpesvirus, treponema pallidum, pubic louse) of genitals.What is more important, the effect of gingkolic acid nanometer liposome is significantly better than gingkolic acid matched group.Local irritation test research shows safety of the present invention, without untoward reaction such as skin irritation, allergy.Sick and the std patient result on trial of clinical multiple gynecological shows, the present invention has significant therapeutic effect to this type of disease tool, and effective percentage reaches 100%, uses safe having no side effect.
The specific embodiment
Below in conjunction with embodiment, the present invention is further described.
Embodiment 1:
In ginkgo nut episperm, gingkolic acid extraction process is as follows: ginkgo nut episperm is stripped down, dry, pulverized with pulverizer.Accurately take a certain amount of gingko episperm through pulverizing, adding solid-liquid ratio is 1:6(W/V) 80% ethanol, at 60 ℃ of bath temperatures, stirring and leaching 2 times, each 4 hours extraction times.After lixiviate finishes, use nutsch filter sucking filtration, collect filtrate.Through D101 macroporous resin adsorption purification, the 100% ethanol desorbing of 10BV, then through purification by silica gel column chromatography, petroleum ether: ethyl acetate (90:10, V/V) eluting, use vacuum rotary evaporator concentrate eluant, condition is 55 ℃ of temperature, pressure 0.008-0.009MPa, concentrating under reduced pressure.In 60 ℃ of vacuum dryings, obtain gingkolic acid extractum, character is: light brown extractum, micro-smelly, be dissolved in alcohol, water insoluble; Weigh, and measure gingkolic acid content (mg/g).
Gingkolic acid assay: adopt HPLC method to measure.Gingkolic acid standard substance are purchased from Yuan Ye bio tech ltd, Shanghai.The standard solution of gingkolic acid is between 100-1200 μ g/ml, and respective peaks area and gingkolic acid mass concentration have significant linear relationship, have worked out standard curve.With dehydrated alcohol (chromatographically pure), by the dissolving of gingkolic acid extractum, filter.At column temperature: room temperature; Mobile phase: methanol-water (9:1, v/v); Flow velocity 1ml/min; Sample size: 20 μ l; Detect wavelength 310nm; Analysis time: measure gingkolic acid content in extractum under 25min condition.According to the retention time of standard specimen, carry out qualitatively, calculate the content of gingkolic acid by external standard method.In extractum, gingkolic acid content is 912.49mg/g.
Gingkolic acid extraction process and content assaying method are public method.
Soybean phospholipid ethanol solution, cholesterol ethanol solution, filter; Add above-mentioned gingkolic acid extractum, vitamin E, jolting is even, puts in round-bottomed flask, 30 ℃ of constant temperature water bath rotary evaporations, decompression film forming, makes film material on bottle wall, form uniform class membrane of lipoprotein, adds at 50 ℃ of PBS buffer fully vibration aquation, come off completely to wall upper film, immerse 50 ℃ of stirred in water bath, ultrasonic 5min under 25 ℃ of water bath condition, the each 5s of 200Hz, intermittently 5s, to forming the translucent turbid liquid that shows slightly opalescence.High pressure homogenize, circulates 3 times under 100 Mpa pressure, crosses microporous filter membrane, obtains light yellow transparent colloid shape gingkolic acid nanometer liposome suspension.
The preparation of gel: carbopol 10g is sprinkled upon to 500ml distilled water surface, swelling spending the night.Take glycerol 60g, propylene glycol 120ml, is added to respectively in swelling good carbopol, grinds evenly rapidly, obtains transparent gel-type vehicle.Get above-mentioned gingkolic acid nanometer liposome suspension 40ml, be dissolved in 100ml 95% alcoholic solution, be slowly added in gel-type vehicle, stir.Slowly add triethanolamine 15ml again, mend distilled water to enough 1000 g, obtain faint yellow gingkolic acid nanometer liposome gel 1000 g, gingkolic acid nano-lipid body burden is 4%.Be sub-packed in 50 tubules every 20 g.
Gel preparation technology is public method.
Embodiment 2:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 924.51mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
The preparation of ointment: get above-mentioned gingkolic acid nanometer liposome suspension 5ml, be dissolved in 100ml 95% alcoholic solution, then add wherein triethanolamine 50ml, heat to 70 ℃.Paraffin 300g, stearic acid 300g, propylene glycol 100ml are added respectively to stir and melt, mend distilled water to enough 1000 g, stir to paste, obtain faint yellow gingkolic acid nanometer liposome ointment 1000 g, gingkolic acid nano-lipid body burden is 0.5%.Be sub-packed in 50 tubules every 20 g.
Ointment preparation technology is public method.
Embodiment 3:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 945.07mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
The preparation of tincture: get above-mentioned gingkolic acid nanometer liposome suspension 100ml, be dissolved in 600ml 95% alcoholic solution, add respectively wherein again triethanolamine 50ml, propylene glycol 200ml stirs, mend 95% alcoholic solution to enough 1000 ml, obtain faint yellow gingkolic acid nanometer liposome tincture 1000 ml, gingkolic acid nanometer liposome suspension content is 10%.Be sub-packed in 50 tubules every 20 ml.
Tincture preparation technology is public method.
Embodiment 4:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 922.16mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
Get gingkolic acid nanometer liposome suspension 10ml, by embodiment 1 gel preparation technology, prepare gingkolic acid nanometer liposome gel 1000g, gingkolic acid nano-lipid body burden is 1%.Be sub-packed in 50 tubules every 20 g.
Embodiment 5:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 940.23mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
Get gingkolic acid nanometer liposome suspension 200ml, by embodiment 2 ointment preparation technologies, prepare gingkolic acid nanometer liposome ointment 1000g, gingkolic acid nano-lipid body burden is 20%.Be sub-packed in 50 tubules every 20 g.
Embodiment 6:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 909.79mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
Get gingkolic acid nanometer liposome suspension 400ml, by embodiment 3 tincture preparation technologies, prepare gingkolic acid nanometer liposome tincture 1000ml, gingkolic acid nano-lipid body burden is 40%.Be sub-packed in 50 tubules every 20 ml.
Embodiment 7:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 923.61mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
Get gingkolic acid nanometer liposome suspension 1ml, by embodiment 3 tincture preparation technologies, prepare gingkolic acid tincture 1000ml, gingkolic acid content is 0.1%.Be sub-packed in 50 tubules every 20 ml.
Embodiment 8:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 923.61mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
Get gingkolic acid nanometer liposome suspension 0.1ml, by embodiment 1 gel preparation technology, prepare gingkolic acid gel 1000g, gingkolic acid content is 0.01%.Be sub-packed in 50 tubules every 20 g.
Embodiment 9:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 950.02mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
Get gingkolic acid nanometer liposome suspension 300ml, by embodiment 3 tincture preparation technologies, prepare gingkolic acid nanometer liposome tincture 1000ml, gingkolic acid nano-lipid body burden is 30%.Be sub-packed in 50 tubules every 20 ml.
Embodiment 10:
In ginkgo nut episperm, gingkolic acid extraction process and content assaying method, with embodiment 1, obtain gingkolic acid extractum, and gingkolic acid content is 913.42mg/g.
By embodiment 1 gingkolic acid nanometer liposome suspension preparation technology, prepare gingkolic acid nanometer liposome suspension.
Get gingkolic acid nanometer liposome suspension 20ml, by embodiment 2 ointment preparation technologies, prepare gingkolic acid nanometer liposome ointment 1000g, gingkolic acid nano-lipid body burden is 2%.Be sub-packed in 50 tubules every 20 g.
The gingkolic acid nanometer liposome external preparation of making in order to top method, carries out relevant pharmacology pharmacodynamic test, the experiment of local irritation safety evaluatio and clinic trial test.Experimental data of the present invention below:
To the female rats of the therapeutical effect of rat bacterial vaginitis: body weight 200-250g, from vagina, inject staphylococcus aureus (3 × 10
5cFU/ml) and escherichia coli (1.6 × 10
6cFU/ml) mixed bacteria liquid, bacterium liquid injection rate is 0.05ml/100g body weight, the next day, inoculates 1 time.Within the 5th day and the 7th day, get respectively vaginal secretions and do microscopy, antibacterial culturing, and isolate respectively staphylococcus aureus and escherichia coli, then be inoculated in respectively in culture medium, after cultivating, do identification of strains.Microscopy and identification of strains are positive for infecting successful rat.Get the successful rat of infection and be divided at random 4 groups, model control group, gingkolic acid matched group, gingkolic acid nanometer liposome group of the present invention, ofloxacin ointment group, 10 every group.In each rat vagina of administration group, inject respectively relative medicine, saline injection in each rat vagina of model control group, every day 1 time, continuous 5 days.Within after drug withdrawal the 1st day and the 4th day, get rat vagina secretions and identify, all negative being decided to be turns out cloudy to identify pathogen for 2 times.Within after drug withdrawal the 5th day, rat is dissected, got vagina specimen and carry out perusal; Vagina specimen is marked by hyperemia, edema, hemorrhage 3 indexs: without hyperemia, edema, hemorrhagely comment 0 point; Vaginal mucosa still smoothly has mild hyperaemia, is chosen as 1 point; Vaginal mucosa is level and smooth moderate hyperemia still, and visible point-like petechia, is chosen as 2 points; Vaginal mucosa is unsmooth, and severe hyperemia is bled profusely a little as seen, organizes Mild edema, is chosen as 3 points.With the score value of model control group, be decided to be 100% (degree of inflammation), medicine group and model control group comparison, can draw medicine group inflammation suppression ratio.With t check, inflammation is marked and added up, use χ
2check is added up negative conversion rate.Result shows (in Table 1), and gingkolic acid nanometer liposome gel of the present invention has significant therapeutic effect to rat bacterial vaginitis tool, and effect is significantly better than ofloxacin and gingkolic acid external preparation matched group.
The therapeutical effect of table 1. to rat bacterial vaginitis
X ± s. n=10. compared with model control group, *
p<0.05, * *
p<0.01. compared with gingkolic acid group,
# p<0.05,
## p<0.01.
To the female rats of the therapeutical effect of rat colpitis mycotica: body weight 200-250g, the 35 mg/kg anesthesia of lumbar injection pentobarbital sodium, spay.Perform the operation after 10 days, rat skin lower injection estradiol benzoate 10mg/kg, continuous 5 days, vaginal smear under microscope, visible a large amount of keratinocytes, shape is large and irregular, illustrates that rat is in false rutting period.Every 1 estradiol benzoate of injection on the 3rd, make rat maintain false rutting period later.To, in the estrous rat of vacation, from vaginal injection, enter 2.5 × 10
6the Candida albicans 0.1ml of CFU/ml, block vaginal orifice with aseptic cotton balls, prevent that bacterium liquid stream from going out, microscopy vaginal secretions after 10 days, visible rat pudendum is obviously red and swollen, and has a large amount of white secretions, and secretions is inoculated in sabouraud culture medium, after cultivating, microscopy is positive and is the successful rat of infection.Get the successful rat of infection and be divided at random 4 groups, model control group, gingkolic acid matched group, gingkolic acid nanometer liposome group of the present invention, Miconazole Nitrate Cream group, 10 every group.In each rat vagina of administration group, inject respectively relative medicine, saline injection in each rat vagina of model control group, every day 1 time, continuous 7 days.Before administration in the 4th day and get rat vagina secretions the 8th day morning and cultivate in sabouraud culture medium 48 hours, as without albicans growth for curing, otherwise, do not cure.Use χ
2check is added up cure rate.Result shows (in Table 2), and gingkolic acid nanometer liposome ointment of the present invention has significant therapeutic effect to rat colpitis mycotica tool, and effect is significantly better than miconazole nitrate and gingkolic acid external preparation matched group.
The therapeutical effect of table 2. to rat colpitis mycotica
X ± s. n=10. compared with model control group, *
p<0.05, * *
p<0.01. compared with gingkolic acid group,
# p<0.05,
## p<0.01.
To the therapeutical effect of rat cervicitis: set up rat cervicitis model with phenol rubber cement vaginal injection.Phenol adding distil water, Radix Acaciae senegalis are made 25% rubber cement, extract the injection of the capable vagina of 0.15ml deep during modeling, and give self-control medical aseptic cotton silver blocking vaginal orifice, prevent that medicinal liquid from flowing out, and 1 time on the every 3rd, totally 4 times, 12 days.Get the successful rat of modeling and be divided at random 4 groups, model control group, gingkolic acid matched group, gingkolic acid nanometer liposome group of the present invention, the safe rubber cement group of two azoles, 10 every group.In each rat vagina of administration group, inject respectively relative medicine, saline injection in each rat vagina of model control group, every day 1 time, continuous 7 days.Perusal rat pudendum part and secretions change, and whether vaginal orifice is dry, and mucosa has or not redness, hyperemia, and vagina has or not flow liquid to bleed etc.The pathological change of light Microscopic observation cervical tissue cellularity.Result shows, perusal model group pudendum turns white, and cervix uteri hyperemia, redness have individually dense sexual secretion; The above-mentioned Non Apparent Abnormality of organizing for the treatment of group.The necrosis of micro-Microscopic observation model group cervical mucosal, comes off, and forms ulcer, visible a large amount of cell infiltration under cervical mucosal, little vasodilation, hyperemia, interstitial edema, the visible columnar epithelium squamous metaplasia of squama column intersection; Gingkolic acid matched group, the necrosis of the safe group of two azoles mucosa, come off, and vasodilation, hyperemia, edema alleviate, and inflammatory cell is less; The above-mentioned performance of gingkolic acid nanometer liposome gel group of the present invention obviously alleviates.Illustrate that gingkolic acid nanometer liposome gel of the present invention has significant therapeutic effect to rat cervicitis tool, can promote wound tissue healing, curative effect is better than the two calm and peaceful gingkolic acid external preparation of azoles matched groups.
The therapeutical effect that human papillomavirus (HPV) is infected: HPV infects positive epithelial cells from male patients with condyloma acuminata patient foreskin position disease damage and women's patients of cervical ruin cervix uteri port part disease damage.Cell culture: trypsin digestion obtains HPV and infects epithelial cells, with 1 × 10
5cell number is inoculated in the 100ml Tissue Culture Flask of coating Mus tail collagen, and condition of culture is 5%CO
2, 37 ℃; Culture medium is RPMI1640 culture fluid, containing 10% calf serum and antibiotics.Use 2.5gL
-1the epithelial cells of trypsinization the 3rd generation monolayer culture, makes single cell suspension, every hole 4 × 10 with culture fluid
4individual cell is inoculated in 24 orifice plates, and every hole adds 1ml culture fluid cellar culture, within 3 days, changes liquid 1 time, continuously 24d.Centrifugal collecting cell, observation of cell form under microscope, and adopt tetramethyl azo azoles salt (MTT) colorimetric determination cell proliferation vigor.Cell is inoculated in to 96 orifice plates, and (every porocyte number is 2 × 10
3), establish gingkolic acid matched group, gingkolic acid nanometer liposome group of the present invention, interferon group, establish 5 dosage groups for every group, add respectively relative medicine 0,20,40,60,80,100 μ l, each dosage group is established 12 multiple holes; Cultivate after 3 days, sucking-off culture fluid, in every hole, add MTT solution 100 μ l in 37 ℃ of reactions 4 hours, discard MTT, every hole adds 100 μ l DMSO and vibrates 10 minutes, abundant dissolving crystallized thing, 490nm place measures absorbance A value, and experimental result represents (experimental group A value/control group A value × lOO) with cell proliferation rate.Result shows (in Table 3), and compared with blank group, gingkolic acid nanometer liposome tincture of the present invention can suppress the propagation of HPV infection cell; The cell that the visible HPV of micro-Microscopic observation infects recovers Normocellular form gradually, illustrate that the present invention has the effect that anti-HPV infects, and effect is significantly better than interferon and gingkolic acid external preparation matched group.
The impact of table 3. on the HPV infection cell rate of increase (%)
X ± s. n=12. compared with blank,
* p<0.01. compared with gingkolic acid group,
# p<0.05,
## p<0.01.
To gonococcal bacteriostatic activity: use agar dilution gonococcus to be carried out to the mensuration of Mlc.Collect bacterial strain: bacterial strain is that STD clinics is collected and separated and obtains, and is inoculated in immediately the defiber Sanguis caprae seu ovis culture medium that contains 10% after specimen obtains, 37 ℃ of CO2 gas incubator cultivation 48h.Through morphologic dyeing, oxidase test, growth qualification test and sugar fermentating test, identify
After, with skim milk, be stored in-70 ℃ of refrigerators standby.Antibacterial culturing: experimental strain is inoculated in to GC cultivation and hatches after 24h based on 37 ℃ of incubators, observe the form of bacterium colony, picking list bacterium colony is through identifying, standby.The bacteria containing amount of bacterial concentration is 1.5 X 10
8cFU/mL.MIC measures: the liquid diluting method providing according to the NCCLS of National Committee of Clinical Laboratory Standards is carried out the MIC value of determination experiment bacterial strain, with 2 times of dilution method dilution medicines, the concentration of gingkolic acid nanometer liposome tincture and gingkolic acid tincture is respectively 0.1g/mL, 0.05 g/mL, 0.025 g/mL, 0.0125 g/mL, 0.00625 g/mL, adds 1.5 X 10 in every pipe
8the antibacterial 10 μ L of CFU/mL.Use inoculating loop picking bacterium liquid, streak inoculation, on flat board, is also inoculated in the not flat board containing medicinal liquid simultaneously, and the bacterium liquid of inoculation same amount is contrast, and 37 ℃ of incubators are hatched observed result after 24h.Experiment repeats 3 times, take 3 times or 2 identical persons as report foundation.Experimental result shows, the antibacterial activity in vitro of gingkolic acid nanometer liposome tincture of the present invention is stronger, and MIC is 0.006 g/mL, and the MIC of gingkolic acid tincture is 0.02g/mL.Illustrate that the present invention has significant bacteriostatic activity to gonococcus; And effect is better than gingkolic acid external preparation matched group.
To the therapeutical effect of genital herpes guinea pig model: genital herpes virus is taken from the bleb liquid of clinical patients with genital herpes, after identifying, Virus culture and PCR turn out to be HSV-II.Experiment is 10 with virus virulence
-7tCID
50.Get 40 of healthy guinea pigs, body weight 300 ± 20g, gets in advance vaginal secretions and does HSV-II nucleic acid qualitative detection, gets rid of HSV-II and infects.The intravaginal of genital herpes virus liquid is injected after modeling success, is divided at random 4 groups, i.e. model control group, gingkolic acid matched group, gingkolic acid nanometer liposome group of the present invention, acyclovir group, 10 every group.In each rat vagina of administration group, inject respectively relative medicine, saline injection in each rat vagina of model control group, every day 1 time, continuous 10 days.Carry out therapeutic evaluation by pudendum symptom score (the 10th day skin lesion integrated value) and skin lesion regression time.Result shows (in Table 4), and gingkolic acid nanometer liposome ointment of the present invention has significant therapeutic effect to genital herpes guinea pig model tool, and effect is significantly better than acyclovir and gingkolic acid external preparation matched group.
The therapeutical effect of table 4. to genital herpes guinea pig model
| Grouping | Skin lesion regression time (my god) | The 10th day skin lesion integrated value |
| Model contrast | 0 | 5.07±1.45 |
| Gingkolic acid | 6.25±1.09 ** | 1.16±0.32 ** |
| Gingkolic acid nanometer liposome | 3.37±0.83 **## | ?0.29±0.07 **## |
| Acyclovir | 7.07±1.26 ** | 1.39±0.44 ** |
X ± s. n=10. compared with model control group, *
p<0.05, * *
p<0.01. compared with gingkolic acid group,
# p<0.05,
## p<0.01.
To the killing action of trichomonal vaginitis: experiment worm kind is picked up from Out-patient Clinic of Department of Gynecology trichomonal vaginitis patient.Get patient's posterior fornix of vagina secretions, be inoculated in liver and soak in soup culture medium, 37 ℃ of aseptic anaerobism are cultivated, and within every 3 days, go down to posterity once, pass altogether 3 times, and the infusorian that obtains not cloning aseptic culture is tested.If blank group, gingkolic acid matched group, gingkolic acid nanometer liposome group of the present invention, metronidazole group, drug dose is 2mmol/L, establishes 10 multiple holes for every group; After cultivation 2,4,8,24h, get small part and adopt trypan blue staining to observe polypide survival condition.Get appropriate mixed liquor and be put on blood cell counting plate, in high power Microscopic observation polypide active state; Under low power lens, according to the result of Trypan Blue, count respectively dead or living worm body, and calculate polypide survival rate.After cultivating 24h, without the remaining part of culture tube of the worm that lives, remake normal transferred species cultivation, get nearly bottom culture transferred species to new culture medium, after .48h, adopt above-mentioned Trypan Blue observational method, if without infusorian survival, can judge that survival rate is zero.Experimental result shows (in Table 5), and gingkolic acid nanometer liposome tincture of the present invention is external has killing action significantly to trichomonal vaginitis, and effect is significantly better than metronidazole and gingkolic acid external preparation matched group.
The killing in vitro effect of table 5. to trichomonal vaginitis
X ± s. n=10. compared with blank,
* p<0.01. compared with gingkolic acid group,
# p<0.05,
## p<0.01.
To the killing action of papovavirus: from the condyloma acuminatum specimen of clinical criteria, after cutting, wash away blood with normal saline. under aseptic condition, specimen is shredded, grind to form homogenate standby.Get Condyloma Acuminata homogenate 0.6ml, add respectively gingkolic acid tincture, gingkolic acid nanometer liposome tincture 2.4ml of the present invention.Control tube adds dehydrated alcohol 2.4ml, puts in 4 ℃ of refrigerators, extracts DNA on time.Fluorescence quantitative polymerase chain reaction (FQ-PCR) reactions steps: adopt conventional alkaline lysis method of extracting DNA 0.2ml to enter in thin-walled reaction tube, add certain density primer, F-PROBE, dNTP, archaeal dna polymerase and buffer etc. simultaneously, put into PE5700 fluorescent PCR instrument, 93 ℃ of 2min give after degeneration, by 93 ℃ of 45s, 55 ℃ of 120s, carry out 40 circulations repeatedly, and quantitative result is calculated the quantitative result of initial copy number by computer software automatic analysis.Experimental result shows (in Table 6), and gingkolic acid nanometer liposome tincture of the present invention is external has killing action significantly to papovavirus, and effect is significantly better than gingkolic acid external preparation matched group.
The killing action of table 6. to papovavirus
Local irritation test: get 8 of adult rabbit, male and female half and half, every animal sub-cage rearing, first raises 3-4 days at laboratory condition.Then rabbit spinal column both sides dorsal body setae is shaved off to every lateral area 5 × 10cm with electric shaver-for women
2.Carefully check that whether depilation district has redness and damage, if any abandoning it, can not test.Adopt the test method of multiple dosing, in depilation, gingkolic acid nanometer liposome external preparation of the present invention is smeared in district, every day 2 times, smears continuously 7 days.Whether last is smeared after 24 hours and is washed away smear with warm water, observes local skin reaction respectively at l, after 24,48,72 hours, as erythema, edema, desquamation, incrustation etc., have petechia, skin peptide coarse or poor etc., records time of origin and regression time.Experimental result shows, is smearing continuously gingkolic acid nanometer liposome external preparation of the present invention after 7 days, and each rabbit skin does not all show obvious skin irritation and anaphylaxis.Show product safety of the present invention, have no adverse reaction.
Clinic trial test: the gingkolic acid nanometer liposome external preparation of making in order to top method, on probation through multiple gynecological disease and std patient, concrete case and the course for the treatment of are in Table 7.All patients are coated with disease damage place outward with the present invention, every day 1 time.Patient does not all use any other external used medicine during treating.Criterion of therapeutical effect: cure: clinical symptoms performance all disappears, and pathogen microscopy is negative; Effective: clinical symptoms performance is basic to disappear, and pathogen microscopy is the weak positive; Effective: clinical symptoms performance part disappears, and pathogen microscopy is the weak positive; Invalid: clinical symptoms shows constant or increases the weight of, and pathogen microscopy is positive.Cure rate=healing number/total case X100%; Effective percentage=(healing number+effective number+efficiently individual quantity)/total case X100%.The results are shown in Table 7.Clinic trial result shows, multiple gynecological is sick and std patient use gingkolic acid nanometer liposome external preparation of the present invention is all effective, and effective percentage reaches 100%; Cure rate all exceedes 85%.During application the present invention, all there is not the untoward reaction such as skin irritation, allergy in all patients.Therefore the present invention is the external preparation of a kind of desirable treatment multiple gynecological disease and sexually transmitted disease (STD), evident in efficacy, safe and reliable.
The clinical effectiveness of table 7 gingkolic acid nanometer liposome of the present invention external preparation treatment gynaecopathia and sexually transmitted disease (STD)
The present invention is compared with prior art: the present invention derives from natural precious medicinal plants Semen Ginkgo, from positions such as gingko episperms, extract and separate gingkolic acid, take gingkolic acid as effective ingredient, make nanometer liposome external preparation, there is the remarkable effect of killing genitals and residing pathogen and antiinflammatory around.To multiple gynecological, sick and sexually transmitted disease (STD) tool has significant therapeutic effect, and curative effect is better than common antibiotics or immunosuppressant, without the side effect due to life-time service immunosuppressant, antibiotic.Therefore gingkolic acid nanometer liposome external preparation of the present invention is used safety, and aspect treatment multiple gynecological disease and sexually transmitted disease (STD), curative effect is outstanding, has good potential applicability in clinical practice.
Claims (7)
1. gingkolic acid nanometer liposome is prevented and treated the purposes in the external preparation of gynaecopathia and sexually transmitted disease (STD) in preparation.
2. purposes as claimed in claim 1, described gynaecopathia is trichomonal vaginitis, colpitis mycotica, bacterial vaginitis, pudendal pruritus, cervicitis, cervical erosion; Described sexually transmitted disease (STD) is infectious and struvite genital diseases, as gonorrhea, syphilis, condyloma acuminatum, genital herpes, human papillomavirus (HPV) infect, the multiple sexually transmitted disease (STD) of pubic louse.
3. gingkolic acid nanometer liposome as claimed in claim 1, described gingkolic acid is by Chemical Decomposition method, from the extract at the episperm position of Semen Ginkgo, obtains; Or gingko episperm is extracted to elaboration products and in the extraction course of processing, in remaining waste material, again reclaim; Or obtain the each several part extract that contains this compounds from other plant; Or obtain by the method for chemosynthesis; And the material of this compounds pharmaceutically acceptable salt, ester, solvate, crystal habit; Described gingkolic acid nanometer liposome, by adopting the preparation of thin-film ultrasonic-high pressure homogenization method.
4. external preparation as claimed in claim 1 is the treatment gynaecopathia that contains gingkolic acid nanometer liposome and medicine, cosmetics or the disinfectant external preparation of sexually transmitted disease (STD); And comprise medicine, cosmetics or the disinfectant external preparation that gingkolic acid nanometer liposome and any combinations of substances are used for the treatment of gynaecopathia and sexually transmitted disease (STD); It is characterized in that, described preparation is said exterior-applied formulation on any pharmaceutics, is liquid preparation, cream cream, gel, powder, membrane, lotion, aerosol spray, transdermal absorption formulation, subcutaneous injection agent.
5. medicine as claimed in claim 4, cosmetics or disinfectant external preparation, is characterized in that, contains gingkolic acid nanometer liposome 0.001-80%.
6. medicine as claimed in claim 4, cosmetics or disinfectant external preparation, is characterized in that, contains gingkolic acid nanometer liposome 0.001-30%.
7. medicine as claimed in claim 4, cosmetics or disinfectant external preparation, is characterized in that, contains gingkolic acid nanometer liposome 0.5-20%.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310408906.2A CN103751105A (en) | 2013-09-11 | 2013-09-11 | Application of ginkgolic acid nanometer liposome in treatment of gynecological diseases and venereal diseases |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310408906.2A CN103751105A (en) | 2013-09-11 | 2013-09-11 | Application of ginkgolic acid nanometer liposome in treatment of gynecological diseases and venereal diseases |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN103751105A true CN103751105A (en) | 2014-04-30 |
Family
ID=50518542
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310408906.2A Pending CN103751105A (en) | 2013-09-11 | 2013-09-11 | Application of ginkgolic acid nanometer liposome in treatment of gynecological diseases and venereal diseases |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103751105A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106177475A (en) * | 2016-09-26 | 2016-12-07 | 贵州柏泰合生物科技有限公司 | A kind of Chinese medicine composition for gynaecopathia |
| PL423496A1 (en) * | 2017-11-18 | 2019-05-20 | Inst Immunologii I Terapii Doswiadczalnej Polskiej Akademii Nauk | Application of maidenhair-tree Ginkgo biloba leaves extract |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102349935A (en) * | 2011-08-18 | 2012-02-15 | 王青 | Purpose of ginkgolic acid in preparing external preparation for treating venereal disease, gynopathy and perianal disease |
-
2013
- 2013-09-11 CN CN201310408906.2A patent/CN103751105A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102349935A (en) * | 2011-08-18 | 2012-02-15 | 王青 | Purpose of ginkgolic acid in preparing external preparation for treating venereal disease, gynopathy and perianal disease |
Non-Patent Citations (1)
| Title |
|---|
| 陈桐楷等: "柔性纳米脂质体经皮给药新载体的研究进展", 《药学进展》, vol. 12, no. 6, 31 December 2009 (2009-12-31) * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106177475A (en) * | 2016-09-26 | 2016-12-07 | 贵州柏泰合生物科技有限公司 | A kind of Chinese medicine composition for gynaecopathia |
| PL423496A1 (en) * | 2017-11-18 | 2019-05-20 | Inst Immunologii I Terapii Doswiadczalnej Polskiej Akademii Nauk | Application of maidenhair-tree Ginkgo biloba leaves extract |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| KR101796423B1 (en) | Composition and ladies genital area cleanser composition comprising composition for inducing Antiviral and antibacterial cause vaginitis and method for manufacturing the same | |
| CN102085248B (en) | Traditional Chinese medicinal composition for treating cervix diseases, method for preparing same and method for detecting same | |
| CN107184911A (en) | One kind is used to prevent cervical carcinoma and treatment papillomavirus（HPV）Infection and the product and preparation method of colpitis | |
| CN104888058A (en) | Antibacterial and anti-inflammation external preparation and method for preparing same | |
| CN103751105A (en) | Application of ginkgolic acid nanometer liposome in treatment of gynecological diseases and venereal diseases | |
| CN102349935A (en) | Purpose of ginkgolic acid in preparing external preparation for treating venereal disease, gynopathy and perianal disease | |
| CN103372076B (en) | Treat the medicine of disease and the application of Radix Sanguisorbae and/or Radix Sangusorbae extract that are caused by propionibacterium acnes | |
| CN102319272B (en) | Application of ginkgolic acid in preparation of external preparation for treating dermatopathy | |
| CN103251636B (en) | Medicament for treating Candida infections and diseases caused by Candida, and preparation method thereof | |
| CN102743432B (en) | Application of patchouli oil in preparation of medicines used for treating colpitis | |
| CN1321655C (en) | Gel of Chinese traditional medicine for treating cervicitis, colpitis and preparing method | |
| CN103751230A (en) | Application of gingko extract nanometer liposome in gynecological disease treatment | |
| CN105233144A (en) | Traditional Chinese medicine preparation for treating dermatomycoses of animals and preparation method of traditional Chinese medicine preparation | |
| CN106177900B (en) | A kind of gel of anti-human papilloma virus (anti-HPV) and preparation method thereof | |
| CN105147827B (en) | A kind of antibacterial medicines and preparation method thereof | |
| CN102688300B (en) | Pharmaceutical composition and usesthereof | |
| CN102652773B (en) | Medicinal composition and preparation method thereof | |
| CN103751231A (en) | Application of gingko extract nanometer liposome in acne removing and dermatosis treatment | |
| CN1970020B (en) | A pharmaceutical composition, preparation process and application thereof | |
| CN101757140B (en) | Medicament with spermicidal effect in vitro and bacteriostatic action, and preparation method and application thereof | |
| CN109820901A (en) | A kind of Traditional Chinese medicinal gel preparation with sterilization | |
| CN1293893C (en) | Gel of Chinese traditional medicine and preparation method | |
| CN110368463A (en) | A kind of Chinese medicine composition and the preparation method and application thereof for treating human papilloma virus HPV infection | |
| CN107362310A (en) | It is a kind of to be used to repair Baicao Fuyanqing preparation of vagina environment and preparation method thereof | |
| CN109364148A (en) | A kind of FUKE QIANJIN PIAN and preparation method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20140430 |