CN1663585A - Pharmaceutical composition for resisting herpes simplex virus and its preparing process - Google Patents
Pharmaceutical composition for resisting herpes simplex virus and its preparing process Download PDFInfo
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Abstract
The invention discloses a pharmaceutical composition for resisting herpes simplex virus and its preparing process, wherein the composition is prepared from coptis root, gentian root, pipewort, baras camphor by a predetermined weight ratio, the obtained active constituent has the actions of reducing fever, purging the intense heat and detoxicating. The composition can be prepared into eye drops, nose spray and electuary.
Description
Technical field
The present invention relates to pharmaceutical composition of a kind of anti-herpes simplex virus and preparation method thereof, belong to Chinese medicine medicine and manufacturing technology field.
Background technology
Herpes simplex virus (herpers simplex vinus, be called for short HSV) through the oral cavity, number of ways such as respiratory tract, reproductive tract mucosa and skin injury invade body, can and hide in the trigeminal ganglion ganglion cell, in the keratocyte or skin, mucomembranous surface.Infect caused herpes simplex keratitis (herpers simplex keratitis by HSV, be called for short HSK) be to endanger one of serious viral infection oculopathy in the world today, polymorphic type, easily recurrence, course of disease progress has virus, multipaths such as immunity, too many levels factor interaction cause transparency cornea to be replaced by a large amount of new vesselses, scar tissue, and secondary uveitis, glaucoma, antibacterial or fungal infection facilitate that sb.'s illness took a turn for the worse, the serious harm visual function, even cause blind.The blind rate of HSK accounts for first of corneal blindness, be extensive use of along with antibiotic, hormone, immunosuppressant in recent years and the increase of acquired immune deficiency syndrome (AIDS), HSK is high especially, is second diseases causing blindness except that cataract, the serious harm human health is worldwide ophthalmology thorny problem.
Still there is not at present effectively this sick medicine of control, see drug resistance when Western medicine acycloguanosine, idoxuridine, Sorivudine, vidarabine, trifluridine, acyclovir treatment repeatly, can not effectively control recurrence, even after corneal transplantation, send out again, and there be a lot of bad sending out to answer, class cholesterol application of cortical hormone causes immunosuppressant, reduces the interferon generation, promotes virus breeding.See Tao Peizhen for details at " anti-herpesvirus medicament progress " (figure classification number among Chinese Journal of New Drugs in 2003 the 12nd volume fourth phase P253-P257[] R978.7[article numbering] 1003-3734 (2003) 04-0253-05).
Extraction has anti-bacteria and anti-virus from plant, the immune isoreactivity composition of adjusting has advantages such as little, the difficult generation drug resistance of toxic and side effects.Therefore, from the Chinese medicine traditional Chinese medical science, excavate the important content that effective Therapeutic Method and medicine have become Chinese anti-HSV-I active component research.
Chinese herbal medicine such as existing report Radix Et Rhizoma Rhei, Galla Chinensis, Flos Caryophylli have certain anti-HSV-I activity; The side selects the Decoction of Gentiana for Purging the Liver-fire plus-minus in calendar year 2001 the 19th volume the 7th phase P65-66 " oral medicinal herb the is main treatment herpes simplex keratitis 20 examples " literary composition, side's medicine: rush down in Radix Gentianae, grassy marsh, and Caulis Akebiae, Radix Bupleuri, Radix Scutellariae, the Radix Rehmanniae, Fructus Gardeniae, Radix Arnebiae (Radix Lithospermi), Herba Houttuyniae, Radix Glycyrrhizae, decocting divide the Chinese medicine decoction treatment of decoction being taken warmly early;
Surpass atomizing eye-douche method, intravenous drip, the existing clinical or mechanism research of sub-conjunctival injection treatment herpes simplex keratitis with SHUANGHUANGLIAN FENZHENJI, the shuanghuanglian powder injection pure Chinese medicinal preparation, the aseptic injection of being processed into for medicines such as Flos Lonicerae, Fructus Forsythiae, Radix Scutellariaes, but at this polymorphic type of HSK, easy oculopathy repeatedly,, can not effectively not control the further deterioration of focus to eye mucosa and the direct administration of nasal mucosa.
Rhizoma Coptidis toxic materials clearing away decoction is the classic prescriptions in the heat-clearing and toxic substances removing side, and this side is made up of Rhizoma Coptidis (nine grams), Radix Scutellariae (six grams), Cortex Phellodendri (six grams), Fructus Gardeniae (nine grams) four medicines, has the effect of clearing away heat-fire detoxifcation, cures mainly the card of all excess-heat fire-toxin, three warmers intenseness of heat.Medicinal liquid to the Rhizoma Coptidis toxic materials clearing away decoction active component carries out the inhibitory action that extracorporeal antivirus effect experiment discovery has herpesvirus.
But above-mentioned Chinese medicine decoction all has the shortcoming that takes effect slowly, easily affects the state of an illness adversely.And the still not Chinese medicine preparation of the anti-HSV that comes into the market of effective treatment means of approval and active drug so far, as eye drop preparation etc.
According to the Chinese medicine theory, with identifying and correcting and the bonded thinking understanding of differential diagnosis of diseases HSK: black eyeball (cornea) belongs to wind wheel (liver) in " five take turns theory ".Primary disease mainly is outer wind-engaging pyretic toxicity heresy, and the endogenous cause of ill Liver Channel is intimately gone up and stopped up, and the wind-fire pyretic toxicity is fought with in black eyeball; Or damp-heat accumulation, folder ailment said due to cold or exposure pyretic toxicity is tied in black eyeball; Or cloudy depletion declines, wind-heat invading the lung, pathogenic heat passes liver, follow through on attack and cause " dendritic keratitis " in black eyeball.Its pathogenesis development law is that deficiency and excess folds, the sign characteristics are to be revealed in outer real image (bulbar conjunctiva hyperemia, herpetic keratitis sample pathological changes, corneal epithelial cell generation apoptosis) to coexist in lying concealed in interior virtual image (immunologic function disorder or decline), and interact, malpractice can cause " keratomalacia ", treatment is when " symptomatic treatment in acute condition, radical treatment in chronic case ".So must originally rule together by table.
The key medicine Rhizoma Coptidis of Chinese medicine clearing away heat-damp and expelling toxic material pathogenic fire purging, " property of medicine opinion " " poison reduces phlegm and internal heat for Rhizoma Coptidis point acute conjunctivitis dusk pain, town liver ", the modern pharmacological research Rhizoma Coptidis influences antibacterial, mycete, virus, chlamydia trachomatis by the alkaloid number of ways and reaches the antibiosis effect; Have remarkable anti-inflammatory activity and make macrophage activation, engulf digestive function and improve, and make activated macrophage be directed chemotactic to move, arrive site of complement activation and be the chemokine-like effect; Berberine has killing, suppresses multiple pathogenic microorganism, antibiotic, antiviral and antiinflammatory action in the Rhizoma Coptidis, also has the phagocytic activity that strengthens reticuloendothelial system, the effect that improves immunity.
Radix Gentianae removing fire from the liver, clearing away damp-heat are gone into Liver Channel, " medical science opens former ": " Radix Gentianae is controlled the swollen inflammatory exophthalmos of two conjunctival congestions, control in the eye disease must medication also." modern pharmacological research shows that the Radix Gentianae decoct has inhibitory action to various bacteria, mycetes such as bacillus pyocyaneus, golden Portugal bacterium, achorion gypseums; Its contained erythricine antiinflammatory action is strong, can obviously promote inflammatory cell phagocytic function, macrophage phagocytic function and lymphocyte transformation.
The Flos Eriocauli acrid in the mouth is returned Liver Channel, wind-dispelling heat-dissipating, and improving acuity of vision and removing nebula utilized its antibacterial action to multiple microorganisms such as bacillus pyocyaneus modern age, to the inhibitory action of fungus; Strengthen with the Radix Gentianae compatibility that dispelling wind purges heat, the merit of liver heat removing and eyesight improving; Compendium of Material Medica is thought function: " curing mainly blindness nebula film ", " book on Chinese herbal medicine justice " thinks its " merit of improving acuity of vision and removing nebula, on Flos Chrysanthemi also ", and the Flos Eriocauli medical material is outside the certified products divided by Flos Eriocauli, the also equal people's medicine of the head inflorescence of the wounded in the battle stem of congener match Flos Eriocauli and white pearl Flos Eriocauli
The Borneolum Syntheticum heat radiation of having one's ideas straightened out, pain relieving makes eye bright, and among the peoplely often fries in shallow oil water with Rhizoma Coptidis and endures cream, go into a little eye dripping of Borneolum Syntheticum, its acrid in the mouth hardship of Compendium of Material Medica " can walk and can loose, make the tonneau that is jammed; the meridians bar reaches and the fever (of children) accompanied by fear self-balancing, sore can go out ", cold nature, perfume (or spice) are scurried and are apt to away, nowhere less than, the stagnated fire of loosing, logical all keys, its feature is " sickly use it in the depths; can priming is deeply sick locate ", the traditional Chinese medical science is thought Borneolum Syntheticum acrid in the mouth, hardship, and is cool in nature, GUIXIN, spleen, lung meridian have the merit of the refreshment of having one's ideas straightened out, clearing away heat to alleviate pain, granulation promoting.Amplification on Canon of Materia Medica is thought Borneolum Syntheticum " a little less than the gesture of walking alone then, assistant makes then meritorious ".Recently studies confirm that Borneolum Syntheticum can promote eye drop to pass cornea to enter the anterior chamber to have the short effect of oozing.That Borneolum Syntheticum has is antibiotic, antiinflammatory, analgesic effect, and local external use is very light to sensorineural stimulation, the protective action of pain relieving gentleness.Borneolum Syntheticum also has the blood brain barrier of seeing through, promotes drug transdermal to absorb, promote that medicine enters blood brain barrier, impels the medicine bulk concentration to increase, improve the effect of bioavailability.
Summary of the invention
The objective of the invention is pathogenesis and sign characteristics, utilize the Chinese crude drug performance, through the modern pharmaceutical processes at HSK, obtain drug effect obviously, the pharmaceutical composition of the anti-herpes simplex virus of processing science.
The technical solution adopted for the present invention to solve the technical problems is:
A kind of pharmaceutical composition of anti-herpes simplex virus, it is characterized in that by following mainly be that component is formed originally:
A. Rhizoma Coptidis 40～80 weight portions, b. Radix Gentianae 40～80 weight portions, c. Flos Eriocauli 30～70 weight portions, d. Borneolum Syntheticum 0.5～2 weight portion
Above-mentioned raw materials is preferably a. Rhizoma Coptidis 55～65 weight portions, Radix Gentianae 55～65 weight portions, c. Flos Eriocauli 45～55 weight portions, d. Borneolum Syntheticum 0.9～1.1 weight portion; More preferably a. Rhizoma Coptidis 60 weight portions, b. Radix Gentianae 60 weight portions, c. Flos Eriocauli 50 weight portions, d. Borneolum Syntheticum 1.0 weight portions
In order to reach curative effect preferably, in the described primary raw material c Flos Eriocauli can also for one of following or both and both more than mixture replacing:
Periostracum Cicadae, Herba Equiseti Hiemalis, Fructus Gardeniae, Flos Chrysanthemi Indici, Herba Houttuyniae, Flos Chrysanthemi, Flos Buddlejae.
The b Radix Gentianae can also substitute with Radix Et Rhizoma Rhei in the described primary raw material.
Alternative raw material comprises following on its pharmacology of described primary raw material:
(1) Fructus Gardeniae (2) Flos Chrysanthemi Indici (3) Herba Taraxaci (4) Herba Houttuyniae (6) Flos Buddlejae (7) Radix Scutellariae (8) Herba Equiseti Hiemalis (9) Radix Astragali (10) Radix Et Rhizoma Rhei (11) Radix Arnebiae (Radix Lithospermi) (12) Periostracum Cicadae (13) Flos Chrysanthemi (14) Folium Isatidis.The preparation of drug combination method of above-mentioned anti-herpes simplex virus, it comprises the following steps:
(a) take by weighing raw material by proportional quantity:
Rhizoma Coptidis, Radix Gentianae, Flos Eriocauli, Borneolum Syntheticum;
(b) Rhizoma Coptidis of proportional quantity is added 4～8 times 0.1%～2% hydrochloric acid solution, after the immersion, decocted 1.5 hours, leach 0.1%～2% hydrochloric acid solution that decocting liquid adds 4～8 times again and decoct for the second time, decocting time is 1 hour; Merge decocting liquid twice, it is 1.12 that filtration gained filtrate decompression is concentrated into proportion, makes extractum A;
The Radix Gentianae and the Flos Eriocauli of proportional quantity are decocted with water twice, add water and be not advisable to have powder, decocting time was respectively 1.5 hours, 1 hour, merged decocting liquid twice, filtered to get filtrate, and it is 1.12 that filtrate decompression is concentrated into proportion, makes extractum B;
(c) to add ethanol to the alcohol amount of containing be 65～75% for extractum A that obtains in the combining step (b) and extractum B, left standstill cold preservation 24～72 hours.Draw supernatant, decompression recycling ethanol adds purified water to there not being the alcohol flavor, adjusts pH value to 7.0, and supernatant is drawn in cold preservation 12～48 hours;
(d) Borneolum Syntheticum with proportional quantity adds 1/3 scale surface-active agent for grinding, and adds purified water again and stirs evenly, and joins the supernatant that step (c) obtains, and stirs, and makes active ingredient in pharmaceutical of the present invention;
(e) active component that step (d) is obtained adds conventional additives, makes all ingredients.
Hydrochloric acid described in the step (b): concentration range is 0.1～2%; Preferred 0.5%, Rhizoma Coptidis can also be 1%～8% acetic acid extraction with concentration; Or 0.2%～1% sulphuric acid extraction.Radix Gentianae, amount of water was advisable with the submergence powder when Flos Eriocauli decocted with water, and drew by orthogonal experiment: best decocting time is that 1.5 hours, best for the second time decocting time are 1 hour for the first time.
The precipitate with ethanol consumption is 8～12 times in the step (c), and the best is 10 times, and to contain the alcohol amount to the mixed solution be 65%～75%, more preferably 70%.
Step (d)) surfactant is a surfactant conventional in the Chinese medicine preparation in, in order to increase the dissolubility of Borneolum Syntheticum, preferred tween 80.
The active component that above-mentioned raw materials is produced through processing, required various conventional adjuvant in the time of can adding the different preparation of preparation according to different needs is prepared into various dosage forms as surfactant etc. with the method for Chinese medicinal of routine, as eye drop, nasal spray, unguentum etc.
Eye drop is the main dosage form of the present composition, and it can adopt the following step preparation:
Add purified water to amount of preparation with obtaining active component in the step (e), add 0.0005g with every ml amount of preparation and add ethyl hydroxybenzoate, regulate pH value to 6～7 with hydrochloric acid, and to regulate osmotic pressure with sodium chloride solution be 7.4, to cross the microporous filter membrane fine straining twice of 0.8 μ m, embedding is in the 250ml saline bottle, and 100 ℃ of flowing steams were sterilized 0.5～1 hour, the cooling back is aseptic subpackaged in eye-drop liquid bottle
The present invention is a primary raw material with Rhizoma Coptidis, Radix Gentianae, Flos Eriocauli, Borneolum Syntheticum, effect with heat clearing and damp drying, eliminating fire and detoxication, bacterial diseasees such as mucocutaneous pathological changes, carbuncle sore tumefacting virus, the internal strength of treatment poison, the knowledge of also can be applicable to have excessive noxious heat, stopping up syndrome in damp and hot swells and ache, pharyngitis.
Pharmaceutical composition of the present invention has the broad-spectrum antivirus action, and experiment showed, also have certain anti-SARS virus and the infected cell activity effect of protection through the SARS resisting medicine screening active ingredients.
The eye drop of pharmaceutical composition of the present invention has the infringement that better inhibition HSK-I corneal causes, and promotes the reparation of cornea focus, helps having shortened the course of disease, alleviates the formation of corneal nebula.Clinically metamorphosis such as vernal conjunctivitis, blepharitis, phlyctenular conjunctivitis immunity, anaphylactic disease are had obvious curative effects, and toxicity is low and other antiviral drugs does not have crossing drug resistant, can be used as the Comprehensive Treatment medicine of herpes simplex keratitis
The specific embodiment
Below in conjunction with embodiment the present invention is further described.
Embodiment 1
(a) take by weighing the following traditional Chinese medicines material as raw material:
Rhizoma Coptidis 6kg, Radix Gentianae 6kg, Flos Eriocauli 5kg, Borneolum Syntheticum 0.1kg.
(b) Rhizoma Coptidis 6kg is added 36L, 0.5% hydrochloric acid solution decocts twice, for the first time decocting time is that 1.5 hours, medicinal residues add 30L again, 0.5% hydrochloric acid solution decocts for the second time, and decocting time is 1 hour, and decocting liquid filters, filtrate decompression is condensed into the extractum A that relative density is about 1.12 (20 ℃)
Radix Gentianae 6kg, Flos Eriocauli 5kg two flavor medicines are decocted with water twice, decocting time is 1 hour for the first time, leach decocting liquid, it is 0.5 hour that medicinal residues add the water decocting time second time again, add water was not advisable to have powder at every turn, decocting liquid filters, and filtrate decompression is condensed into the extractum B that relative density is about 1.12 (20 ℃).
(c) extractum A of combining step (b) and extractum B add 10 times of amount ethanol, and the alcohol amount that contains to solution reaches 70%, leaves standstill cold preservation 48 hours.Absorption supernatant, decompression recycling ethanol be not to there being the alcohol flavor, and adding an amount of purified water adjustment pH value is 7.0, and supernatant is drawn in cold preservation 24 hours.
(d) get Borneolum Syntheticum 0.1kg and add the 0.04L tween 80, add the fresh purified water of 10L and stir evenly, add the supernatant of (c), stir, add fresh purified water to 100L for grinding; The ethyl hydroxybenzoate that adds 50g, regulating PH with hydrochloric acid is 6～7, and sodium chloride solution is regulated osmotic pressure and is about 7.4 atmospheric pressure, and being adjusted to the solution that sodium chloride content reaches about 0.9 is isosmotic solution; Fine straining twice, embedding in the 250mL saline bottle, 100 ℃ of flowing steams sterilization 30 minutes, after the cooling, through fine straining (by the microporous filter membrane of 0.8 μ m) once after, aseptic subpackaged to eye-drop liquid bottle.
Embodiment 2
(a) take by weighing raw material: Rhizoma Coptidis 4kg, Radix Gentianae 4kg, Borneolum Syntheticum 0.05kg, Herba Houttuyniae 4kg Flos Chrysanthemi 4kg
(b) Herba Houttuyniae 4kg, Flos Chrysanthemi 4kg extract Aromatic water, and the amount of getting Aromatic water is 20L.
Rhizoma Coptidis 4kg decocts with the hydrochloric acid solution of 0.5%24L, and decocting time is 1.5 hours for the first time, leaches decocting liquid, and the hydrochloric acid solution that medicinal residues add 20L again decocted 1 hour for the second time, and decocting liquid filters, and relative density was about 1.12 extractum A when filtrate decompression was condensed into 20 ℃.
Get Radix Gentianae 4kg and decoct with water twice, amount of water is advisable with the submergence powder, and first decocting time is 1 hour, leaches decocting liquid, and medicinal residues decocted 0.5 hour for the second time, and decocting liquid filters, and when filtrate decompression was concentrated into 20 ℃, relative density was about 1.12 extractum B.
(c) get the extractum A of step (b) gained and extractum B and add ethanol to the alcohol amount that contains of solution and reach 70%, left standstill cold preservation 48 hours.Draw supernatant, decompression recycling ethanol adds an amount of purified water to there not being the alcohol flavor, adjusts pH value to 7.0, and supernatant is drawn in cold preservation 24 hours.
(d) get the 0.05kg Borneolum Syntheticum and add the 0.017L tween 80 for grinding, adding the fresh purified water of 5L stirs evenly, add the supernatant of (c) and Aromatic water (b), stir, add fresh purified water to 100L, the ethyl hydroxybenzoate that adds 50g, hydrochloric acid regulate that pH value is 6～7, sodium chloride solution is regulated osmotic pressure and is about 7.4 atmospheric pressure, and being adjusted to the solution that sodium chloride content reaches about 0.9 is isosmotic solution; Fine straining twice, embedding in the 250ml saline bottle, 100 ℃ of flowing steams sterilization 30 minutes, after the cooling, through fine straining (by the microporous filter membrane of 0.8 μ m) once after, aseptic subpackaged to eye-drop liquid bottle.
Embodiment 3
(a) take by weighing following Chinese crude drug and make raw material:
Rhizoma Coptidis 8kg Radix Gentianae 8kg Borneolum Syntheticum 0.2kg Fructus Gardeniae 8kg Flos Chrysanthemi Indici 8kg Periostracum Cicadae 8kg
(b) Flos Chrysanthemi Indici 8kg extracts Aromatic water, the amount 25L of Aromatic water.
Getting Rhizoma Coptidis 8kg is that 6% acetic acid decocts with 48L concentration, and decocting time is 1.5 hours for the first time, leaches decocting liquid, and medicinal residues decocted 1 hour for the second time, and filtrate decompression is condensed into, and relative density is about 1.12 (20 ℃) equivalent extract.
Radix Gentianae 8kg, Fructus Gardeniae 8kg, Periostracum Cicadae 8kg three flavor medicines decoct with water, each amount of water is advisable for the submergence powder, and decocting time is 1 hour, leaches decocting liquid that medicinal residues add water and decocted for the second time 0.5 hour, decocting liquid filters, and filtrate decompression is condensed into the extractum A that relative density is about 1.12 (20 ℃).
(c) step (b) gained extractum A and extractum B are added ethanol, make the alcohol amount that contains in the solution reach 75%, left standstill cold preservation 48 hours.Draw supernatant, decompression recycling ethanol adds an amount of purified water to there not being the alcohol flavor, adjusts pH value to 7.0, and supernatant is drawn in cold preservation 24 hours.
(d) Borneolum Syntheticum 0.2kg is added the tween 80 of 0.067L for grinding, adding the fresh purified water of 10L stirs evenly, add the supernatant of (c) and Aromatic water (b), stir, add fresh purified water to 100L, the ethyl hydroxybenzoate that adds 50g, hydrochloric acid regulate that pH value is 6～7, sodium chloride solution is regulated osmotic pressure and is about 7.4 atmospheric pressure, and being adjusted to the solution that sodium chloride content reaches about 0.9 is isosmotic solution; Fine straining twice, embedding in the 250ml saline bottle, 100 ℃ of flowing steams sterilization 30 minutes, after the cooling, through fine straining (by the microporous filter membrane of 0.8 μ m) once after, aseptic subpackaged to eye-drop liquid bottle.
Embodiment 4:
(a) take by weighing the following traditional Chinese medicines material as raw material:
Rhizoma Coptidis 5.5kg, Radix Gentianae 5.5kg, Borneolum Syntheticum 0.09kg, Flos Buddlejae 5.5kg, Flos Chrysanthemi 5.5kg
(b) Flos Chrysanthemi is extracted Aromatic water, gets the amount 22L of Aromatic water.
Rhizoma Coptidis 5.5kg is decocted with the 27L0.2% sulfuric acid solution, and decocting time is 1.5 hours for the first time, leaches decocting liquid, and medicinal residues decocted 1 hour for the second time with the 25L sulfuric acid solution, and decocting liquid filters, and filtrate decompression is condensed into the equivalent extract A that relative density is about 1.12 (20 ℃).
Get Radix Gentianae 5.5kg, Flos Buddlejae 5.5kg decocts with water, and adds water and is not advisable to have powder, for the first time decocting time is 1 hour, leach decocting liquid, medicinal residues add the water decocting time second time 0.5 hour, and decocting liquid filters, and filtrate decompression is condensed into the equivalent extract B that relative density is about 1.12 (20 ℃).
(c) gained extractum A among the step b and extractum B are added ethanol, make to contain alcohol amount in the solution and reach 70%, left standstill cold preservation 48 hours.Draw supernatant, decompression recycling ethanol adds an amount of purified water to there not being the alcohol flavor, adjusts pH value to 7.0, and supernatant is drawn in cold preservation 24 hours.
(d) get Borneolum Syntheticum 0.09kg and add the 0.03L tween 80 for grinding, adding the fresh purified water of 10L stirs evenly, add the supernatant of step c and the Aromatic water of step b, stir, add fresh purified water to 100L, the ethyl hydroxybenzoate that adds 50g, hydrochloric acid regulate that pH value is 6～7, sodium chloride solution is regulated osmotic pressure and is about 7.4 atmospheric pressure, and being adjusted to the solution that sodium chloride content reaches about 0.9 is isosmotic solution; Fine straining twice, embedding in the 250ml saline bottle, 100 ℃ of flowing steams sterilization 30 minutes, after the cooling, through fine straining (by the microporous filter membrane of 0.8 μ m) once after, aseptic subpackaged to eye-drop liquid bottle.
Embodiment 5
(a) take by weighing the following traditional Chinese medicines material as raw material:
Rhizoma Coptidis 6.5kg, Flos Eriocauli 6.5kg, Borneolum Syntheticum 0.11kg, Radix Et Rhizoma Rhei 6.5kg.
(b) Radix Et Rhizoma Rhei 6.5kg is 60% ethanol percolation with 52L concentration, and percolate is deposited in addition.
The hydrochloric acid solution that Rhizoma Coptidis 6.5kg adds 39L0.1% decocts, decocting time is 1.5 hours for the first time, leaches decocting liquid, and the hydrochloric acid solution that medicinal residues add 35L again decocted 1 hour for the second time, decocting liquid filters, and filtrate decompression is condensed into the extractum A that relative density is about 1.12 (20 ℃).
Flos Eriocauli 6.5kg decocts with water twice, adds water and is not advisable to have powder, and decocting time is 1 hour for the first time, leaches decocting liquid, adds water and decocts for the second time 0.5 hour, and decocting liquid filters, and it is about 1.12 (20 ℃) extractum B that filtrate decompression is condensed into relative density.
(c) combining step b gained extractum A and extractum B add ethanol and make the alcohol amount that contains of solution reach 65%, leave standstill cold preservation 48 hours.Draw supernatant, add the percolate mixing of step b, decompression recycling ethanol adds an amount of purified water to there not being the alcohol flavor, adjusts pH value to 7.0, and supernatant is drawn in cold preservation 24 hours.
(d) get Borneolum Syntheticum 0.11kg and add the 0.04kg tween 80, add the fresh purified water of 20L and stir evenly, add the supernatant of step c for grinding, stir, add fresh purified water to 100L, add the ethyl hydroxybenzoate of 50g, regulate PH and osmotic pressure with hydrochloric acid, sodium chloride respectively, twice of fine straining, embedding is in the 250ml saline bottle, and 100 ℃ of flowing steams were sterilized 30 minutes, after the cooling, through fine straining (by the microporous filter membrane of 0.8 μ m) once after, aseptic subpackaged to eye-drop liquid bottle.
Embodiment 6
(a) take by weighing the following traditional Chinese medicines material and make raw material:
Rhizoma Coptidis 6.5kg, Radix Gentianae 5.5kg, Flos Eriocauli 5.5kg Borneolum Syntheticum 0.1kg
(b) Rhizoma Coptidis 6.5kg decocts with the hydrochloric acid solution of 36L 0.5%, and decocting time is 1.5 hours for the first time, leaches decocting liquid, the hydrochloric acid solution that medicinal residues add 30L decocted 1 hour for the second time, decocting liquid filters, and filtrate decompression is condensed into relative density and is about 1.12 (20 ℃) extractum A, and is standby.
5.5kg Radix Gentianae, 5.5kg Flos Eriocauli two flavor medicines are decocted with water twice, add water at every turn and be advisable with the submergence powder, decocting time is 1 hour for the first time, leaches decocting liquid, and medicinal residues add water and decocted for the second time 0.5 hour; Decocting liquid filters, and filtrate decompression is condensed into the extractum B that relative density is about 1.12 (20 ℃), and is standby.
(c) extractum A of combining step b gained and extractum B add ethanol to the alcohol amount of containing of solution and reach 70%, leave standstill cold preservation 48 hours.Draw supernatant, decompression recycling ethanol adds an amount of purified water to there not being the alcohol flavor, adjusts pH value to 7.0, and supernatant is drawn in cold preservation 24 hours.
(d) Borneolum Syntheticum 0.1kg adds the 0.03L tween 80 for grinding, add the fresh purified water of 10L and stir evenly, add the supernatant of step (c), stir, add fresh purified water to 100L, the ethyl hydroxybenzoate that adds 50g, fine straining twice, embedding is in the 250ml saline bottle, 100 ℃ of flowing steams were sterilized 30 minutes, after the cooling, through fine straining (by the microporous filter membrane of 0.8 μ m) once after, aseptic subpackaged to the nasal spray bottle.
Below being the test example of pharmaceutical preparation of the present invention, is example with the eye drop.
Experimental example 1: the acute toxicity test of eye drop of the present invention
1. experiment material:
Animal: ICR mice, 20, male and female half and half, body weight 20～22g.Male and female sub-cage rearing, the animal quality certification: No. (96) 2001, the accurate word of Zhejiang laboratory animal are provided by the experimental center, Zhejiang Province.
Medicine: medicine eye drop of the present invention, brown liquid, specification 10ml are equivalent to crude drug 1.74g.
2. experimental technique:
20 fasting of ICR mice are after 16 hours, irritate stomach and give medicine eye drop 40ml/kg body weight of the present invention, same dosage gastric infusion was 1 time every 5 hours, totally 3 times, accumulated dose is every kg body weight 120ml, be equivalent to every kg crude drug 208.8g, the toxic reaction of mice comprises after the observation administration: general symptom, outward appearance, hair color, defecation, the situation of ingesting, autonomic activities etc. and the death condition in 7 days, and administration was weighed on the 7th day.
3. experimental result:
Autonomic activities, hair color, outward appearance after the mice administration, ingest all normally, first administration is visible stool color burn about 5 hours, recovers normal after 24 hours, other no abnormality seen reaction; Not seeing in seven days observation period has dead mouse, and the body weight of male and female mice all has growth, sees Table 1.
The body weight change of table 1 eye drop maximum tolerance determination of the present invention mice (X ± SD)
| Grouping | Quantity (only) | Before the administration (gram) | After the administration (gram) |
| Male | ????10 | ????21.0±0.86 | ????24.0±1.54 |
| Female | ????10 | ????21.1±0.79 | ????22.9±1.47 |
Learnt that by experiment the oral eye drop of the present invention of ICR mice 40ml/kg3 time does not have tangible toxic reaction, the oral maximum tolerated dose of eye drop mice is 120ml/kg, is equivalent to crude drug 208.8g/kg.
The clear collyrium eye irritation test of experimental example 2 pyretic toxicity
1. experiment material
Animal: rabbit, New Zealand's kind, male, body weight 2.1-2.8kg, eyes are normal, available from Zhejiang Province's Experimental Animal Center.
Medicine: eye drop of the present invention, the TCM Preparation Room of Hangzhou Chinese Medicinal Hospital provide.
2. experimental technique
4 of rabbit splash into eye drop 0.1ml respectively at a branch hole eyeball, passive closed 10 seconds, splash into normal saline 0.1ml in contrast at opposite side, morning and afternoon every day each 1 time, continuous 7 days.Test and rose in the 2nd day the irritant reaction of observing positions such as cornea, conjunctiva before administration every day, 1 week after drug withdrawal.
Mark by " eye stimulates standards of grading " in the chief editor of Ministry of Public Health pharmaceutical control and administration department " study of tcm new drug guide ", and be converted into the irritant reaction grade:
3. experimental result
Tangible irritant reaction does not appear in 4 lagophthalmos that give the clear collyrium of pyretic toxicity: conjunctiva does not have hyperemia, no edema, no secretions; Cornea does not have muddiness; Iris is normal.Be 0 grade according to " study of tcm new drug guide " reaction classification.
The clear collyrium of pyretic toxicity every day twice, each 0.1ml gives the rabbit eye drip, and continuous 7 days, the irritation that do not go out to lose face, it is 0 minute that eye stimulates a comprehensive mean scores.As seen the clear collyrium multiple dosing of pyretic toxicity does not have tangible irritant reaction.
Experimental example 3 eye drop treatment herpes simplex keratitis of the present invention clinical observations
1, physical data:
Accept outpatient service 108 examples for medical treatment, 2 examples of being in hospital, totally 110 examples (170 eyes) HSK patient,
Random packet medicine eye drop of the present invention nebulae inhalation group, eye drop eye drip group and acyclovir (acycloguanosine) eye drip matched group.
Wherein 50 examples (79 eyes) are organized in the eye drop atomizing: male's 15 examples, women's 35 examples; Age is at 12～84 years old, average (41.91 ± 15.54) year; Before this experiment, the shortest 2d of the course of disease, the longest 20 years.
Eye drop eye drip group 30 examples (46 eyes): male's 9 examples, women's 21 examples; 13～76 years old age, average (37.07 ± 15.57) year; Before this experiment, the shortest 3d of the course of disease, the longest 10 years.
Acyclovir (acycloguanosine) eye drip matched group 30 examples (47 eyes): male's 10 examples, women's 20 examples; 11～69 years old age, average (36.73 ± 15.62) year; The shortest 1d of the course of disease, the longest 12 years.
2, diagnostic criteria
Dialectical and the Western medicine diagnose standard of Chinese medicine, according to Ministry of Public Health " new drug (Chinese medicine) clinical research guideline ", " traditional Chinese medical science disease controversial issue criterion of therapeutical effect " and " ophthalmology of Chinese medicine " (the 5th edition teaching material of traditional Chinese medical science institution of higher learning) and " practical ophthalmology " (People's Health Publisher), mainly rely on medical history and examination of eyes definite to the diagnosis of herpes simplex keratitis.
Tcm diagnosis: a. eye symptom: 1. main symptom: black eyeball consor punctate keratitis, it is greyish white that color is; 2. accompanied symptoms: the puckery discomfort of conscious ophthalmic gritty; Photophobia is shed tears; Hyperemia of the bulbar conjunctiva or ciliary hyperemia.
B. whole body accompanied symptoms: 1. fever with aversion to wind, pharyngalgia; 2. yellow and thin fur or Huang; 3. wiry and frequent pulse or floating number.
Doctor trained in Western medicine herpes simplex keratitis diagnostic criteria: 1. skin bleb infringement or simple property keratitis medical history are arranged; 2. there are specific go up sense, heating etc. to recur factors in the recent period; 3. the course of disease is delayed more, and antibiotic therapy is invalid, and 17-hydroxy-11-dehydrocorticosterone makes that sb.'s illness took a turn for the worse; Specific forms such as dendroid, point-like, starlike, map shape appear in 4. cornea fluorescein sodium stained positive, focus; 5. corneal sensation is gone down.
3, experimental technique
1. eye drop atomizing group of the present invention: 20ml adds 402 ultrasonic atomizatio instrument nebulae inhalations, each 15min, 2 times/d
Eye drop eye drip group of the present invention: 6 times/d, each 2.
Matched group: 0.1% acyclovir collyrium eye drip, 6 times/d, each 2.
2. three groups of equal whole bodies are used polyinosini injection 5d, 2ml/d, intramuscular injection;
3. the course of treatment with follow up a case by regular visits to: 10d is a course of treatment, and the 3d further consultation is once treated 1～2 course of treatment, this laboratory observation 15d; Every group 50% case was followed up a case by regular visits to 4～12 months.
4, curative effect determinate standard:
With reference to " practical ophthalmology ", " traditional Chinese medical science disease diagnosis criterion of therapeutical effect "
1. recovery from illness: irritation disappears, conjunctival congestion (-), and cornea focus edema is soaked into and is absorbed point-like, dendroid, map shape ulcer healing, fluorescent staining (-).
2. produce effects: irritation disappears substantially, conjunctival congestion (-), and the cornea focus is soaked into and disappeared, and ulcer healing, fluorescent staining reduce and reach 2/3, but slight edema is arranged, and thickens to be no more than cornea complete thick 1/5～1/4.
3. effective: irritation alleviates, and focus is dwindled, and ulcer surface dwindles, and edema is soaked into and taken a turn for the better.
4. invalid: every sign of symptomatology and keratitis does not have improvement, even increases the weight of.
Traditional Chinese medical science disease efficacy evaluation:
1. clinical recovery: the integration of symptomatology and sign disappears to 0.
2. produce effects: symptomatology and sign are significantly improved, and integration descends 〉=2/3.
3. effective: symptomatology and sign all take a favorable turn, and integration descends between 1/3～2/3.
4. invalid: symptomatology and sign do not take a turn for the better, and integration descends＜1/3 or do not have decline, even increases the weight of.
5. health giving quality index: compare with the variation before and after the treatment.
5, experimental result
1. total effects
See Table 2
The treatment of table 2 group is compared (example) to the total effective rate of herpes simplex keratitis
| Group | The example number | Cure | Produce effects | Effectively | Invalid | Total effective rate (%) | Cure rate (%) |
| Eye drop atomizing group of the present invention eye drip group of the present invention matched group | ?50 ?50 ?50 | ?40 ?16 ?9 | ?6 ?7 ?6 | ??3 ??4 ??9 | ???1 ???3 ???7 | ??98.00 ??90.00 ??76.67 | ??80.00 ??53.33 ??30.00 |
Ridit analyzes: eye drop atomizing group of the present invention is compared with matched group :=3.39, and P＜0.01; Eye drip group of the present invention is compared with matched group :=2.07, and P＜0.05.
2. the cardinal symptom curative effect relatively sees Table 3
Table 3
| Symptom | Group | ????n | Difference | ????F | ????P |
| Blurred vision | Eye drop atomizing group of the present invention eye drip group of the present invention matched group | ????50 ????30 ????30 | ?3.80±1.16 ?3.06±1.36 ?2.33±1.1 | ????13.86 | ????＜0.01 |
| The puckery uncomfortable photophobia of intraocular gritty shed tears the conjunctival congestion opacity of the cornea fever with aversion to wind thin Huang of pharyngalgia tongue or yellow superficial and rapid pulse or string number | Eye drops atomizing group eye drip group of the present invention control group eye drops atomizing of the present invention group eye drip group of the present invention control group eye drops atomizing of the present invention group eye drip group of the present invention control group eye drops atomizing of the present invention group eye drip group of the present invention control group eye drops atomizing of the present invention group eye drip group of the present invention control group eye drops atomizing of the present invention group eye drip group of the present invention control group eye drops atomizing of the present invention group of the present invention eye drip group of the present invention control group | ??50??4.08±1.34 ??30??3.40±1.50 ??30??2.53±1.74 ??50??3.96±1.11 ??30??3.27±1.78 ??30??2.33±1.58 ??50??4.04±1.24 ??30??3.40±1.50 ??30??2.40±1.43 ??50??2.96±1.23 ??30??2.46±1.14 ??30??1.86±1.38 ??41??2.54±1.27 ??26??2.54±1.56 ??27??1.93±1.17 ??50??2.58±1.05 ??30??2.40±1.22 ??30??2.00±1.44 ??50??2.28±1.29 ??30??2.10±1.40 ??30??1.50±1.53 | ????10.02 ? ? ????11.85 ? ? ????13.49 ? ? ????10.46 ? ? ????2.04 ? ? ????2.93 ? ? ????3.01 | ??＜0.01 ? ? ??＜0.01 ? ? ??＜0.01 ? ? ??＜0.01 ? ? ??＜0.01 ? ? ??＜0.01 ? ? ??＜0.01 |
3. follow up a case by regular visits to situation
15 groups of effective cases were followed up a case by regular visits to by 4,8,12 months, the results are shown in following table:
Table 4 is followed up a case by regular visits to situation after respectively organizing drug withdrawal
| Group eye drop atomizing of the present invention group eye drip group of the present invention matched group | 4 months | 8 months | 12 months | ||||||
| Example several 29 18 16 | Recur 002 | There is not recurrence 29 18 14 | Example several 27 17 14 | Recur 114 | There is not recurrence 26 16 10 | Example several 25 14 11 | Recur 225 | There is not recurrence 23 12 6 | |
Followed up a case by regular visits to 4 months, the relapse rate of eye drop atomizing group of the present invention and eye drip group of the present invention is 0, and the matched group relapse rate is 12.90%; Followed up a case by regular visits to 8 months, eye drop atomizing group relapse rate of the present invention is 3.70%, and the relapse rate of eye drop of the present invention is 14.29%, and the relapse rate of matched group is 28.57%; Followed up a case by regular visits to 12 months, eye drop atomizing group relapse rate of the present invention is 8.00%, and eye drop group relapse rate of the present invention is 14.29%, and the matched group relapse rate is 45.45%.
4. untoward reaction
All are observed case and are not seen that untoward reaction is arranged, and select 9 routine patients to carry out relevant index observings such as hepatic and renal function, blood glucose at random, find no harmful effect.
Experimental example 4: eye drop treatment allergic conjunctivitis of the present invention clinical observation
1, physical data
Accept outpatient service 86 examples for medical treatment, spring catarrb conjunctivitis 53 examples, phlyctenular conjunctivitis 33 examples
Random packet medicine eye drop of the present invention nebulae inhalation group and matched group.
Wherein 52 examples are organized in the eye drop atomizing: vernal conjunctivitis 31 examples, phlyctenular conjunctivitis 21 examples; Male's 37 examples, women's 15 examples; Age is at 7～32 years old, average (12.62 ± 5.54) year; Before this experiment, the vernal conjunctivitis course of disease is the shortest 21 days, and is the longest 4 years, and phlyctenular conjunctivitis is the shortest 1 day, and is the longest 19 days.
Matched group 34 examples: vernal conjunctivitis 22 examples, phlyctenular conjunctivitis 12 examples; Male's 24 examples, women's 10 examples; 6～31 years old age, average (12.97 ± 6.22) year; The Spring Festival, the catarrhal conjunctivitis course of disease was the shortest 19 days, and the longest 4 is sticking, and phlyctenular conjunctivitis is the shortest 1 day, and is the longest 20 days.
2, diagnostic criteria
Vernal conjunctivitis, make a definite diagnosis according to dependent diagnostic standard in " traditional Chinese medical science disease diagnosis curative effect " of the promulgation of national traditional chinese medical science administration: eyes are very itched, and photophobia is shed tears; Big and flat nipple and near the conjunctiva glue sample hypertrophy of cornea, the visible cornea point-like of severe patient muddiness appear in palpebral conjunctiva; Secretions is sticking, and the conjunctiva scraping blade has a large amount of eosinophilic granulocytes; Seasonal periodical attack; Divide palpebral conjunctiva type, limbus of corneae type and mixed type clinically.
Phlyctenular conjunctivitis: be decided to be according to dependent diagnostic standard in " ophthalmology pandect ": occur nodositas infringement (cellular infiltration) under conjunctiva, the limbal epithelium; When being positioned at cornea central authorities, just cause city infringement in various degree; Be divided into phlyctenular conjunctivitis, phlyctenular keratitis, eczematous keratoconjunctivitis according to sick position difference; Can seasonal alternate repetition outbreak.
3, Therapeutic Method
1. eye drop atomizing group of the present invention: eye drop 10ml of the present invention adds 402 ultrasonic atomizatio instrument and cooperates the disposable eyeshield of self-control, each 15 minutes, every day 1 time.
Matched group: dexamethasone injection 80,000 adds 402 ultrasonic atomizatio instrument equally and cooperates the disposable eyeshield of self-control, each 15 minutes, each 1 time.
2. course of treatment: 10 days is a course of treatment, and 2 each course of treatment of treatment, further consultation was 1 time in per at least 5 days.Once a day.
3. follow up a case by regular visits to: every group of about 40% case of each sick kind followed up a case by regular visits to, and vernal conjunctivitis was followed up a case by regular visits to 3 years, and phlyctenular conjunctivitis was followed up a case by regular visits to 1 year.
4, curative effect judgment criteria
1. the Spring Festival catarrhal conjunctivitis: with reference to " traditional Chinese medical science disease diagnosis criterion of therapeutical effect " be: cure: the angle conjunctive disorder disappears, and ophthalmic pruritus is eliminated.Take a turn for the better: the angle conjunctive disorder is alleviated, and ophthalmic pruritus alleviates.Do not heal: the angle conjunctive disorder, ophthalmic pruritus does not have improvement.
2. phlyctenular conjunctivitis: cure: conjunctival epithelium inferior thyroid tubercle sample pathological changes disappears, the congested absorption.Take a turn for the better: conjunctival epithelium inferior thyroid tubercle sample pathological changes is alleviated, and hyperemia alleviates.Do not heal: conjunctival epithelium inferior thyroid tubercle sample pathological changes, congested nothing take a turn for the better
5, experimental result
See Table 5, table 6, table 7, table 8, measurement data is checked with t, enumeration data is checked with x2
The total effects of table 5 eye drop nebulae inhalation of the present invention allergic conjunctivitis
Group example number is cured the more cure rate % improvement rate % that takes a turn for the better not
Treatment organizes 52 37 13 2 71.15 96.15
Matched group 34 16 15 3 47.06 91.18
Add up to 86 53 28 5
Cure rate: x
2=4.08 p＜0.05
The curative effect of table 6 eye drop nebulae inhalation of the present invention vernal conjunctivitis and phlyctenular conjunctivitis
| Group | Vernal conjunctivitis | Phlyctenular conjunctivitis | ||||||
| Treatment group matched group adds up to | Example several 31 22 53 | Cure 17 4 21 | Take a turn for the better 12 15 27 | Do not heal 235 | Example several 21 12 33 | Cure 20 12 32 | Take a turn for the better 101 | Do not heal 000 |
The Spring Festival catarrhal conjunctivitis cure rate: x
2=5.78 p＜0.05
Phlyctenular conjunctivitis cure rate: x
2=0.83 p＞0.05
Table 7 liang group patient healing time relatively
| Group | Vernal conjunctivitis | Phlyctenular conjunctivitis | ||||||
| Treatment group matched group | Example several 17 4 | Healing time 16.88 ± 1.73 19.00 ± 1.41 | ???t ?2.27 | ????P ??＜0.05 | Example several 20 12 | Healing time 14.15 ± 3.86 16.83 ± 3.01 | ????t ???2.06 | ????p ?＜0.05 |
Table 8 catarrhal conjunctivitis in the Spring Festival situation of following up a case by regular visits to that heals the sick
| Group | The example number | Recurrence | Not recurrence |
| Treatment group matched group adds up to | ????9 ????3 ????12 | ????3 ????2 ????5 | ????6 ????1 ????7 |
x
2＝4.11?p＜0.05
The observation of curative effect of experimental example 5 eye drop external curing eye herpes zoster facialis of the present invention
1, physical data
Select ward eye, facial area herpes zoster (HZ) patient 75 examples at random, be divided into 2 groups: treatment group and matched group
Wherein: the treatment group: 37 examples; Male's 19 examples, women's 18 examples; Age is at 28～82 years old, average (56.32 ± 15.84) year; The shortest 1～the 9d of the course of disease, average 4.41 ± 1.64d; Right eye 23 examples, left eye 14 examples; Concurrent keratitis 22 examples, concurrent iritis 16 examples, concurrent scleritis 3 examples.
Matched group: 38 examples; Male's 19 examples, women's 19 examples; Age is at 26～80 years old, average (55.03 ± 16.61) year; The shortest 1～the 9d of the course of disease, average 4.50 ± 1.61d; Right eye 19 examples, left eye 19 examples; Concurrent keratitis 21 examples, concurrent iritis 15 examples, concurrent scleritis 4 examples.
2, diagnostic criteria
According to " clinical dermatology ": become family's vesicle, distribute, be arranged in band shape, one-sided property and characteristics such as tangible neuralgia are arranged along neural.Concurrent keratitis; Bulbar conjunctiva hyperemia, edema, corneal epithelium muddiness, herpes, ulcer.Concurrent iritis; Kp+, anterior chamber tyn+, contracted pupil behind the cornea, light reflex is blunt or disappear.
3, Therapeutic Method
1. treatment group: use eye drop eye drip of the present invention: per 2 hours 1 time, each 1; Eyelid, facial area skin soak eye drop soak of the present invention with 5 layers of antiseptic gauzes, and each 0.5 hour, keep gauze moistening, every d6 time;
Matched group: with 0.1% acyclovir eyedrop eye drip, per 2 hours 1 time, each 1, skin is coated with outward with JIDESHENGSHEYAO, and every day repeatedly.Secondary cases iritis or scleritis add the U.S. pupil collyrium of 0.02% fluorine, and every day, eye drip was 4 times, each one; Secondary glaucoma adds with 0.25% timolol collyrium, and every d2 time, each 1.All use the intravenous drip of Chinese medicine SHUANGHUANGLIAN ZHUSHEYE for two groups, every d3g adds in the 500ml normal saline.
2. observational technique: respectively after treatment the 1st, 2,3,4,5,6,7,8,9,10,14,21d observes 1 time.Observation index:
Erythra: stop to occur the new dermexanthesis time; The all dry time of erythra; Erythra all forms a scab the time; Complete analgesic time; Have or not post herpetic neuralgia (erythra incrustation back pain belongs to post herpetic neuralgia).
Eye symptom: photophobia, shed tears, twinge etc. stimulates the disease extinction time; Epithelium keratitis or interstitial keratitis, disciform keratitis extinction time; The iritis extinction time.
4, criterion of therapeutical effect
Recovery from illness: in the medication 8d, lose subsequently outside the neuralgia, facial area, eyelid herpes, concurrent keratitis, iritis symptom, sign all disappear;
Effectively: in the medication 8d, except that post herpetic neuralgia, facial area, the eyelid herpes, concurrent keratitis takes a turn for the better, and iritis alleviates;
Invalid: in the medication 8d, except that post herpetic neuralgia, facial area, eyelid herpes, concurrent keratitis, iritis symptom, sign do not have and are clearly better.
5, experimental result
See Table 9, table 10, table 11, table 12, table 13
Table 9 liang group total effects
The n invalid routine number of the effective routine number of routine number of fully recovering
Treatment organizes 37 30 70
Matched group 38 21 17 0
x
2＝4.62?*p＜0.05
Table 10 eye drop of the present invention and treatment of control group eyelid skin of face HZ curative effect are relatively
The new rash of example number is stopped paying out erythra incrustation of dry time of time erythra complete analgesic time of time
Treatment organizes 37 1.19 ± 0.52 1.97 ± 0.60 2.89 ± 0.52 5.43 ± 3.36
Matched group 38 2.24 ± 0.91 3.21 ± 0.70 4.84 ± 1.10 11.32 ± 4.92
T value 6.114 8.227 9.77 6.04
P value＜0.01＜0.01＜0.01＜0.01
Table 11 eye drop of the present invention and treatment of control group HZ keratitis curative effect are relatively
Example several numbers (only) irritation extinction time keratitis sign extinction time
Treatment organizes 22 22 5.20 ± 1.36 5.66 ± 1.86
Matched group 21 21 7.27 ± 1.35 8.35 ± 1.84
T value 6.27 6.44
P value＜0.01＜0.01
Table 12 eye drop of the present invention and treatment of control group HZ secondary iritis curative effect are relatively
Example several numbers (only) irritation extinction time iritis sign extinction time
Treatment organizes 16 16 5.06 ± 1.06 6.13 ± 0.96
Matched group 15 15 6.46 ± 1.88 7.20 ± 1.66
T value 2.57 2.21
P value＜0.05＜0.05
Table 13 eye drop of the present invention and treatment of control group eye face
Grouping n post herpetic neuralgia is lost neuralgia without male offspring
Treatment organizes 37 1 36
Matched group 38 3 35
*P＞0.05
Experimental example 6: the external anti-I type herpes simplex virus of eye drop of the present invention effect experiment
1, general material:
1. herpes simplex virus lyophilizing HSV-I 168 strains derive from China Sickness Prevention Control Center Virus Disease Prevention Control Institute.
2. medicine
Eye drop of the present invention contains raw medicinal herbs 174mg for every milliliter, abandons precipitation behind the centrifugal 20min during use, gets supernatant.
Control drug: aciclovir eye drop Wuhan People's Armed Police pharmaceutical factory preparation
3. cell Ren sus domestica passage cell strain (IBRS-2), Zhejiang Academy of Medical Sciences bio-engineering research institute Viral Laboratory preserves.Adding after trypsinization when going down to posterity that growth-promoting media piping and druming disperses to make cell number is 5 * 10
-5Individual/ml, be incorporated in 96 hole micro plates, every hole 100 l put 5%～7%CO
2Incubator, 37 ℃ leave standstill, and 24～48h grows up to monolayer.
4. culture fluid is made the IBRS-2 cell culture fluid with Eagla`s and the LH that contains 10% calf serum, and virus is kept liquid and contained 2% calf serum.
2, experimental technique
1. the virosis kind goes down to posterity and the mensuration of virus virulence
Strain is inoculated in the IBRS-2 cell through serum-free Eagla`s liquid dissolving, and the viral liquid behind the rejuvenation three generations partly places liquid nitrogen to irritate, balance-30 ℃.Going down to posterity in cell culture during use increases poison, treats that collecting Virus culture after virus virulence is stablized is for experiment.Virus virulence is collected Virus culture after stable be for experiment with keeping liquid.With keeping liquid with virus 10
-1～10
-6Be diluted to 6 kinds of concentration, add respectively in 96 well culture plates, each concentration is inoculated 3 holes, the 100l/ hole, and absorption 30min washes 3 times, adds culture fluid, 37 ℃ of 5%～7%CO
2Hatch, observe pathological changes caused by virus effect (CPE) day by day,, and measure half sensibility reciprocal (TCID for 6 days
50).If cell contrast.
2. drug cell toxicity test
Get eye drop of the present invention with keeping 2 times of liquid be diluted to 1: 10 (17000mg/L), 1: 20 (8500mg/L), 1: 40 (4000mg/L), 1: 80 (2000mg/L), 1: 160 (1000mg/L), 1: 320 (500mg/L) 6 kinds of dilution factors, be incorporated in respectively in the micro plate that has grown up to cell monolayer, every hole 100l, 37 ℃ of 5%～7%CO
2Cultivate 7d, microscopy day by day, observation of cell pathological changes.Measure medicine half toxic concentration (TC
50) and maximal non-toxic concentration (TC
0).If cell contrast.
3. control drug aciclovir eye drop toxicity test
Get aciclovir eye drop with keeping liquid be diluted to 1: 10 (100000g/L), 1: 100 (10000g/L), 1: 200 (5000g/L), 1: 1000 (1000g/L) 4 kinds of dilution factors, below operation is with 2.
4. the anti-HSV-I effect of eye drop of the present invention is with reference to carrying out about the detection method of antivirus action in " drug study methodology ".
Maximal non-toxic concentration eye drop suppresses the CPE effect that causes of the HSV-I virus liquid of variable concentrations: with 100TCID
50(10
-3) and TCID
50(10
-4) HSV-I virus liquid infects on the IBRS-2 passage cell that has grown up to monolayer on the 96 hole micro plates 37 ℃ of 5%～7%CO respectively
2Absorption 30min, sucking-off virus is washed 3 times, discards venom; Add the maximal non-toxic concentration liquid, each viral dilution hole respectively adds 4 holes.Set up virus, cell, medicine, acyclovir matched group separately.37 ℃ of 5%～7%CO
2Hatched 3 days, microscopy is observed CPE day by day.
The eye drop of the present invention of variable concentrations causes the inhibitory action of CPE to HSV-I:
Diluted medicine to be measured adds 10TCID
50The HSV-I venom is as test group, and each drug level respectively adds 4 holes; If medicine (drug level is consistent with test group), HSV-I, cell matched group.Measure the ID of eye drop of the present invention at last
50
Detection method and cytopathy criterion: behind the cell counteracting toxic substances (or adding medicine), observe CPE day by day.Normal IBRS-2 cellular morphology is fusiformis, and the degeneration variation appears in cell behind the counteracting toxic substances; Swelling, change circle are seen giant cell and fused cell.Branch Pyatyi: 1. 0 grade: (-) 4 holes all do not have CPE, and score value is 8 minutes; 2. 1 grade: CPE 〉=2 holes appear in (+) 25% following cell, score value be 6 minutes 3. 2 grades: CPE 〉=2 holes appear in (++) 5%-50% cell, score value 4 minutes; 4. 3 grades: score value 2 minutes CPE 〉=2 holes appear, in (+++) 50%-75% cell; 5. 4 grades: score value 0 minute CPE 〉=2 holes appear, in (++ ++) 75% above cell.
3, experimental result
1. the virulence of herpesvirus I (HSV-I)
HSV-I causes the IBRS-2 cytopathic effect: do not produce under the cytopathy condition at cell maintenance medium: 10
-1～10
-3Even three kinds of viral dilution degree a large amount of specific lesions occur in first day cell of inoculation: cellular swelling, change circle, refractive power increase, cavity form, the target shape; 10
-4The HSV-I dilute liquid medicine pathological changes occurs in back first day part cell of inoculation; 10
-1～10
-5Five kinds of dilute liquid medicines all occurred the typical cells specific lesions in second day in inoculation; And 10
-6Medicinal liquid is avirulence.
50% tissue infection dosage HSV-I TCID appears in the high dilution of virus titer
50Be 5.5.
2. eye drop medicine of the present invention is to the toxicity of IBRS-2 cell
Observe 4d, the eye drop pair cell of the present invention that is higher than 4000000g/L concentration has toxicity in various degree.Its half toxic concentration (TC
50) be 4000000g/L.Maximal non-toxic concentration (TC
0) 2000000g/L.
3. positive control drug aciclovir eye drop toxicity
Observe 4d result and show, acyclovir concentration 100000g/L (1: 10), 10000g/L (1: 100) pair cell are toxic, and 5000g/L is the maximal non-toxic concentration of this medicine.
4. eye drop of the present invention is anti-HSV-I effect in cell culture
A. eye drop of the present invention suppresses the CPE effect that causes of the HSV-I virus liquid of variable concentrations
See Table 14, eye drop of the present invention (2000000g/L crude drug) is to infecting 100TCID
50, 10TCID
50The IBRS-2 cell of HSV-I virus liquid causes CPE and has very strong inhibitory action, and any specific cell pathological changes does not appear in cell after the dosing, and statistical analysis is compared difference with acyclovir do not have significance (P＞0.05).
Table 14 10TCID
50With 100TCID
50Dosing experiment behind the HSV-I virus attack cell
Test method 10TCID
50HSV-I organizes 100TCID
50The HSV-I group
Observe natural law and observe natural law
1??????2??????3??????1??????2??????3
Medicine+HSV-I------
The HSV-I contrast+++ ++ +++++ ++
Acyclovir+HSV-I------
The medicine contrast------
The acyclovir contrast------
The cell contrast-----
B. variable concentrations eye drop of the present invention causes the CPE inhibitory action to HSV-I:
See Table 15, eye drop 4000000～63000g/L of the present invention causes CPE to HSV-I and has very effective inhibitory action, the drug effect that is higher than 2000000g/L, 1000000g/L drug level, identical with the effect of the medicine group pair cell that does not add virus, both do not have significance at difference, curative effect and concentration are negative correlation in 1: 40～1: 1280 matched group dilution range, and this cytotoxicity with medicine is relevant.
The medicine of table 15 different pharmaceutical concentration causes the CPF inhibitory action to HSV-I
Medicine is dense
Observe natural lawIntegration t value p value
Group degree mg/l 1234567
Test group 4000--+++ +++++ ++ ++ ++ 4.00 ± 3.46
Medicine to (1: 40)--+++ +++++ ++ ++ ++ 4.00 ± 3.46 0＞0.05
According to group
HSV-I???????????????-????+++??++++?++++?++++?++++?++++??1.42±2.99??1.49?＞0.05
Contrast
Test group 2000----++++++ +++5.71 ± 3.35
Medicine to (1: 80)----+++ ++ ++ 6.00 ± 3.05 0.17＞0.05
According to group
HSV-I???????????????-????+++??++++?++++?-+++?++++?++++??1.42±2.99??2.53?＜0.05
Contrast
Test group 1000-----++ ++ ++ 6.28 ± 3.15
Medicine to (1: 160----++ ++ 6.86 ± 1.95 0.41＞0.05
According to group)
HSV-I????????????-??+++??++++?+++??++++?++++?++++??1.42±2.99????2.96???＜0.05
Contrast
Test group 500-----+++ 6.86 ± 2.27
Medicine to (1: 320------++ 7.43 ± 1.51 0.55＞0.05
According to group)
HSV-I????????????-??+++??++++?++++?++++?++++?++++??1.42±2.99????3.83???＜0.01
Contrast
Test group 125-----+++ 7.14 ± 1.57
Medicine was to (1: 640-------8.00 ± 0.00 0.67＞0.05
According to group)
HSV-I????????????-??+++??++++?++++?++++?++++?++++??1.42±2.99????4.48???＜0.01
Contrast
Test group 63-------7.43 ± 1.51
Medicine was to (1: 128-------8.00 ± 0.00 0.10＞0.05
According to group 0)
SV-I?????????????-??+++??++++?++++?+++??++++?++++??1.42±2.99????4.75???＜0.01
Contrast
Cell is right-------
According to
The ID of eye drop of the present invention
50≤ 63000g/L, eye drop of the present invention might directly kill HSV-I or stop the adhesion and the intrusion of HSV-I pair cell, duplicate effect.And toxicity is lower; Maximal non-toxic concentration is 2000000g/L, and acyclovir is 5000g/L.
Experimental example 7: eye drop anti-SARS virus screening active ingredients experiment of the present invention
1, detect raw material:
Eye drop of the present invention: national new drug brush is chosen heart sample number: S00842
Strain: BJ-01 (source: microorganism epidemic research institute of Beijing Academy of Military Sciences)
2, detection method:
Test sample is handled: sample is dissolved in culture fluid or DMSO, is made into suitable initial concentration, 5 times of dilutions, 3 dilution factors
The Vero-E6 cell inoculation in 96 well culture plates, is put 37 ℃, 5%CO
2Cultivate, behind the SARS virus infection cell, add the sample of different diluted concentrations respectively, establish virus control, cell contrast and sample contrast.Observed result under the every day mirror, record CPE, and measure OD value with dimethyl diaminophenazine chloride dyeing, carries out the calculating and the evaluation of sample anti-SARS virus active function with reference to contrasting.
3, testing result
Testing result sees Table 16, when 100g/ml concentration, see certain anti-SARS virus is arranged, the protection infected cell activity effect.
Table 16
Ultimate density (dilution rate) CPE cytoprotective rate (EC_) % cytotoxicity rate (CC_) %
10％??????????????+++????????27?????????????????27
2％???????????????++++???????0??????????????????0
0.4％?????????????++++???????0??????????????????0
Illustrate: as virus host cell (permissive cell), test the effect of eye drop opposing virus infected cell of the present invention with the Vero-E6 cell, detecting index is cytopathy reaction (CPE) and infection cell protection
Cytopathy political reform (CPE): with "-" expression cell attachment complete form, not seeing has tangible disengaging, but cell quantity what do not compare; With "+" expression cytopathy degree,＜25%+, 25%～50%++, 50%～75%+++,＞75%++++.
Infected cytoprotective rate: by comparing the OD value of virus control, cell contrast and sample contrast, calculation sample is to the protection activity of virus infected cell, protective rate＞EC
20Think that tentatively eye drop of the present invention to being had certain protective role by the cell of viral infection, has antiviral activity.
Claims (7)
1, a kind of pharmaceutical composition of anti-herpes simplex virus is characterized in that mainly being made up of the raw material of following weight portion:
A. 40～80 parts of Rhizoma Coptidis, 40～80 parts of b. Radix Gentianaes, 30～70 parts of c. Flos Eriocaulis, 0.5～2 part of d. Borneolum Syntheticum.
2, the pharmaceutical composition of anti-herpes simplex virus as claimed in claim 1, feature are that described raw material preferable amount is: a. Rhizoma Coptidis 55～65 weight portions, b. Radix Gentianae 55～65 weight portions, c. Flos Eriocauli 45～55 weight portions, d. Borneolum Syntheticum 0.9～1.1 weight portion.
3, a kind of preparation of drug combination method of a kind of anti-herpes simplex virus as claimed in claim 1 or 2, it comprises the following steps:
(a) take by weighing Rhizoma Coptidis, Radix Gentianae, Flos Eriocauli, Borneolum Syntheticum, standby;
(b) Rhizoma Coptidis decocts twice with 0.1～0.8% hydrochloric acid solution of 4～8 times of amounts, and each decocting time is 1.0～1.5 hours, and collecting decoction filters to get filtrate; Filtrate concentrates, and makes extractum A;
Radix Gentianae and Flos Eriocauli decoct with water twice, and each decocting time is 1.0～1.5 hours, merges decocting liquid twice, filters to get filtrate; Filtrate concentrates, and makes extractum B;
(c) merge extractum A and extractum B, adding ethanol is 65～75% to containing the alcohol amount; Leave standstill cold preservation 24～72 hours, and drew supernatant, reclaim ethanol; Adjust pH value to 7.0, supernatant is drawn in cold preservation 12～48 hours;
(d) add surfactant in the Borneolum Syntheticum, stir evenly with purified water then; Borneolum Syntheticum solution joins the supernatant that step (c) obtains, and mixes thoroughly, makes active component;
(e) in the active component that step (d) makes, add conventional additives, make preparation.
4, the preparation of drug combination method of a kind of anti-herpes simplex virus as claimed in claim 3 is characterized in that in the step (b) that concentration of hydrochloric acid solution is 0.5% for good.
5, the preparation of drug combination method of a kind of anti-herpes simplex virus as claimed in claim 3 is characterized in that adding in the step (c) ethanol to the alcohol amount of containing and is 70% for good.
6, the preparation of drug combination method of a kind of anti-herpes simplex virus as claimed in claim 3 is characterized in that the surfactant in the step (d) is selected tween 80 for use.
7, a kind of preparation of drug combination method as claimed in claim 3, feature are to produce the active component that obtains in the step (d), to add ethyl hydroxybenzoate, purified water preparation, hydrochloric acid is regulated PH, and sodium chloride solution is regulated osmotic pressure, through the microporous filter membrane fine straining, conventional preparation eye drop.
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| CNB2004100168028A CN100341546C (en) | 2004-03-05 | 2004-03-05 | Pharmaceutical composition for resisting herpes simplex virus and its preparing process |
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| CNB2004100168028A CN100341546C (en) | 2004-03-05 | 2004-03-05 | Pharmaceutical composition for resisting herpes simplex virus and its preparing process |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101019967B (en) * | 2007-02-09 | 2012-07-18 | 杭州市中医院 | Process of preparing antiviral medicine composition |
| CN102579637A (en) * | 2012-03-14 | 2012-07-18 | 广西强寿药业集团有限公司 | Chinese patent drug--herpes virus tablet for treating herpes simplex virus as well as preparation method of same |
| CN105748868A (en) * | 2016-04-15 | 2016-07-13 | 王鑫磊 | Traditional Chinese medicine for treating herpes simplex viral keratitis and preparation method thereof |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1246029C (en) * | 2002-05-28 | 2006-03-22 | 杨毓龙 | Medicine for treating herpes zoster |
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2004
- 2004-03-05 CN CNB2004100168028A patent/CN100341546C/en not_active Expired - Fee Related
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101019967B (en) * | 2007-02-09 | 2012-07-18 | 杭州市中医院 | Process of preparing antiviral medicine composition |
| CN102579637A (en) * | 2012-03-14 | 2012-07-18 | 广西强寿药业集团有限公司 | Chinese patent drug--herpes virus tablet for treating herpes simplex virus as well as preparation method of same |
| CN105748868A (en) * | 2016-04-15 | 2016-07-13 | 王鑫磊 | Traditional Chinese medicine for treating herpes simplex viral keratitis and preparation method thereof |
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| CN100341546C (en) | 2007-10-10 |
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