CN101926858B

CN101926858B - External medicinal composition for treating eczema and dermatitis and preparation method thereof

Info

Publication number: CN101926858B
Application number: CN2009100597897A
Authority: CN
Inventors: 占堆
Original assignee: HOSPITAL OF TRADITIONAL TIBETAN MEDICINE TIBET AUTONOMOUS REGION
Current assignee: Tibet Cheezheng Tibetan Medicine Co Ltd
Priority date: 2009-06-26
Filing date: 2009-06-26
Publication date: 2012-07-25
Anticipated expiration: 2029-06-26
Also published as: CN101926858A

Abstract

The invention provides an external medicinal composition for treating eczema and dermatitis. The composition is an external medicament prepared from the following raw material medicaments in part by weight: 27 to 81 parts of baical skullcap root, 27 to 81 parts of whin, 19 to 57 parts of rhubarb and 27 to 81 parts of medicine terminalia fruit. The invention also provides a preparation method and application of the external medicinal composition. The composition has good curative effect on the eczema and the dermatitis including contact dermatitis, anaphylactic dermatitis, self-sensitive dermatitis, infectious eczematoid dermatitis and the like.

Description

A kind of externally-applied medicinal composition of treating eczematous dermatitis and preparation method thereof

Technical field

The present invention relates to a kind of externally-applied medicinal composition of treating eczematous dermatitis and preparation method thereof, belong to drug world.

Background technology

Eczema and dermatitis are clinical common, multiple dermatosiss, account for the 15%-30% of dermatological department outpatient service amount, wherein see with acute eczema, dermatitis especially more, are one of emphasis problems of the clinical and basic research of domestic and international Dermatology.Because primary disease often has intensive pruritus, exudate is many, easy relapse, and general of especially prolonged and repeated outbreak and skin lesion are more painful, are one of emphasis problems of the clinical and basic research of domestic and international dermatosis.Modern medicine proves that the pathological change of eczema all is intercellular edema (sponge formation) and intracellular edema to occur at epidermis, and high dermis has telangiectasis, edema and monocyte infiltration.From seeing clinically, eczema often is difficult for finding clear and definite reason.In recent years think that eczema is tardy paraphilia reactive disorder, is caused by multiple internal and external factor, and is relevant with inherited genetic factors.A lot of so-called universal eczemas, disseminated neurodermatitis etc. in fact all belong to atopic dermatitis (or eczema).

Application number: 200610104407, denomination of invention: a kind of ointment that is used for alleviating pain and detumescence and preparation method thereof, the present invention provides and relates to a kind of ointment that is used for alleviating pain and detumescence and preparation method thereof.It is to be raw material by Herba Oxytropis Kansuensis 10～15%, sub-rhubarb extract 4～8%, Fructus Chebulae's (enucleation) 10～15%, Radix aconiti szechenyiani 8-12%, Fructus Terminaliae Billericae 10～15%, Fructus Phyllanthi 10～15%, Benzoinum 2～6%, Herba Lycopodii 15～25%, Moschus or artificial Moschus 1～5%, is that substrate is prepared from the emulsifying product of surfactant, oil phase, water.Ointment of the present invention adopts modern advanced to produce, and keeps active substance and the effective ingredient of medical material in the medical material effectively, has improved the dissolution of crude drug composition again greatly; Improve bioavailability simultaneously; Effectively improve therapeutic effect, overcome the deficiency of the former ointment of Tibetan medicine simultaneously, the product free from extraneous odour; Have good stability, easy to use, carry and preserve; Simultaneously of the present invention simple for process, raw material and adjuvant steady sources are easy to industrialization.Application number: 200710188509.3; Denomination of invention: the alcohol extraction method for preparing of Tibetan medicine QINGPENG GAOJI; The invention provides the new method for preparing of a kind of Tibetan medicine QINGPENG GAOJI, this method is the basis with the prescription of former Tibetan medicine QINGPENG GAOJI, and QINGPENG GAOJI is to be the preparation that feedstock production forms by Herba Oxytropis Kansuensis, sub-rhubarb extract, Radix aconiti szechenyiani, Fructus Chebulae, Fructus Terminaliae Billericae, Fructus Phyllanthi, Benzoinum, Herba Lycopodii, artificial Moschus or Moschus; The active substance in the reservation medical material and the active ingredient of medical material; Improve the dissolution of crude drug composition, improved bioavailability simultaneously, effectively improved therapeutic effect.

Summary of the invention

Technical scheme of the present invention has provided a kind of externally-applied medicinal composition of treating eczematous dermatitis, and another technical scheme of the present invention has provided this preparation of drug combination method and purposes.

The invention provides a kind of externally-applied medicinal composition of treating eczematous dermatitis, it is the topical agent that is prepared from following raw materials by weight proportions:

Radix Scutellariae 27-81 part, Herba Oxytropis Kansuensis 27-81 part, Radix Et Rhizoma Rhei 19-57 part, Fructus Chebulae 27-81 part.

Further preferably, it is the topical agent that is prepared from following raw materials by weight proportions:

54 parts of Radix Scutellariaes, 54 parts of Herba Oxytropis Kansuensiss, 38 parts of Radix Et Rhizoma Rhei, 54 parts of Fructus Chebulaes.

Pharmaceutical composition of the present invention is to be active component by Radix Scutellariae, Herba Oxytropis Kansuensis, Radix Et Rhizoma Rhei, Fructus Chebulae's water or extractive with organic solvent, adds the topical agent that acceptable accessories or complementary composition are prepared from.

Wherein, described topical agent is: cream, plaster, rubber-emplastrum, external spraying agent, liniment, emplastrum, paste or cataplasma.

Wherein, the weight percentage that every preparation unit contains baicalin in the described topical agent is not less than 0.2%.

Wherein, the substrate in the described cream is oil-in-water type substrate.

The invention provides a kind of method for preparing the externally-applied medicinal composition of described treatment eczematous dermatitis, it comprises the steps:

A, take by weighing following raw materials by weight proportions:

Radix Scutellariae 27-81 part, Herba Oxytropis Kansuensis 27-81 part, Radix Et Rhizoma Rhei 19-57 part, Fructus Chebulae 27-81 part;

B, soak, heated and boiled is got decoction liquor; Medicinal residues are decocte with water again, merges decoction liquor, filters, and filtrating is put cold;

C, will filtrate through the macroporous adsorptive resins of having handled well, and discard effusive water liquid, water washing cylinder discards cleaning mixture, does not detect to there being flavones ingredient with the ethanol elution of 60-90%; Merge ethanol elution, concentrating under reduced pressure, drying adds acceptable accessories or complementary composition again and is prepared into external preparation pharmaceutically commonly used.

Wherein, described macroporous adsorbent resin is a D101 type macroporous adsorbent resin.

Wherein, the described ethanol elution of c step is 70% ethanol elution.

The present invention also provides the purposes of described externally-applied medicinal composition in the medicine of acute, the subacute eczema of preparation, erythra that the chronic eczema acute attack causes and pruritus.

Department of Pharmacy of the present invention develops externally used compound preparation according to the SDA relevant requirements on the basis of Hospital of Traditional Tibetan Medicine's clinical proved recipe preparation; Its modern study document of forming medicine is very abundant, confirms that fully it has the active substance and the pharmacological action of many-sided treatment eczema and dermatitis.Radix Scutellariae has effects such as antiinflammatory, analgesic, antiallergic action, inhibition pathogenic microorganism in the side; Herba Oxytropis Kansuensis bitter in the mouth, cold in nature, little poison have heat-clearing and toxic substances removing, and anti-inflammation hemostasia cures mainly wound hemorrhage, open sore part inflammation, but external reducing swelling and alleviating pain; Radix Et Rhizoma Rhei bitter in the mouth, cold in nature.Rush down scorchingly hot, internal organs heat, the clots absorbing eliminate indigestion, anti-inflammation is used for jaundice due to damp-heat; This three medicine is a principal agent.Fructus Chebulae's bitter in the mouth, puckery, warm in nature.It is sick to control wind, gallbladder, expectorant, blood, and this medicine is an accessory drugs, with the Herba Oxytropis Kansuensis compatibility, can reduce its toxicity.Cold febrile disease is controlled; Cold the controlling of calentura is the more important of Tibetan medicine's determination of treatment based on pathogenesis obtained through differentiation of symptoms and signs, and dermatosis belongs to calentura, more than in the Tibetan medicine bitter and puckery flavor; Cold; So to eczema, erythra and pruritus that especially acute, subacute eczema or chronic eczema acute attack cause have better curative effect, are worth carrying out multiple center clinical study to investigate its curative effect.

Medicine of the present invention is made up of the middle Tibetan medicine of clearing away heat-damp and promoting diuresis, anti-inflammatory anti-itch function promptly according to above-mentioned pathogenesis, aims at the skin limitation inflammation that eczema causes and establishes.Radix Scutellariae has effects such as antiinflammatory, analgesic, antiallergic action, inhibition pathogenic microorganism in the raw material prescription; Herba Oxytropis Kansuensis bitter in the mouth, cold in nature, little poison have heat-clearing and toxic substances removing, and anti-inflammation hemostasia cures mainly wound hemorrhage, open sore part inflammation, but external reducing swelling and alleviating pain; Radix Et Rhizoma Rhei bitter in the mouth, cold in nature.Rush down scorchingly hot, internal organs heat, the clots absorbing eliminate indigestion, anti-inflammation is used for jaundice due to damp-heat; This three medicine is a principal agent.Fructus Chebulae's bitter in the mouth, puckery, warm in nature.It is sick to control wind, gallbladder, expectorant, blood, and this medicine is an accessory drugs, with the Herba Oxytropis Kansuensis compatibility, can reduce its toxicity.Cold febrile disease is controlled; Cold the controlling of calentura is the more important of Tibetan medicine's determination of treatment based on pathogenesis obtained through differentiation of symptoms and signs, and dermatosis belongs to calentura, more than in the Tibetan medicine bitter and puckery flavor; Cold, eczema and dermatitis (comprising contact dermatitis, allergic dermatitis, autosensitization dermatitis, infectious eczematoid dermatitis etc.) are had better curative effect.

The specific embodiment

The preparation of embodiment 1 medicinal frost formulation of the present invention

[prescription] Radix Scutellariae 54g Herba Oxytropis Kansuensis 54g Radix Et Rhizoma Rhei 38g Fructus Chebulae 54g

[method for making] above four flavors add 12 times of water logging bubbles 1 hour, and heated and boiled 2 hours is got decoction liquor; Medicinal residues add the water of 10 times of amounts to be fried in shallow oil 1 time again, merges decoction liquor, filters; Filtrating is put cold, through the D101 macroporous adsorptive resins of having handled well, discards effusive water liquid; Water washing cylinder with 4 times of amounts of medical material amount discards cleaning mixture, and the ethanol elution with 70% does not detect (amount of ethanol is about 4-5 times of medical material amount) to there being flavones ingredient.Merge ethanol elution, concentrating under reduced pressure, the dry check of vacuum (or spraying), adding blank oil-in-water type emulsifiable paste matrix ad pond om is 1000g, stirs, and is filled into ointment tube and promptly gets.

The preparation of embodiment 2 medicine patch of the present invention

Get Radix Scutellariae 81g Herba Oxytropis Kansuensis 27g Radix Et Rhizoma Rhei 19g Fructus Chebulae 27g

Press the method for embodiment 1 and extract medicine, behind the ethanol elution concentrate drying, add the medical pressure-sensitive adhesive that contains additive and Percutaneous absorption enhancer and process in the substrate, process coating, be coated with cream, drying, cutting, the lid lining is cut into small pieces, and cuts 1000 altogether, promptly gets.

The preparation of embodiment 3 medicament spraying agents of the present invention

Get Radix Scutellariae 27g Herba Oxytropis Kansuensis 81g Radix Et Rhizoma Rhei 57g Fructus Chebulae 81g, press the method for embodiment 1 and extract material medicine, add spray adjuvant commonly used again, be prepared into the spray of 1000ml.

The preparation technology's of embodiment 4 medicines of the present invention parameter is selected test

1, the screening of process conditions

1.1 the screening of material medicine process conditions

1.1.1 the screening of decocting condition

1.1.1.1 the medical material water absorption rate is measured and got the prescription medical material, mixing adds 5 times of water gagings and soaks, and soaks into until medical material.Leach unabsorbed moisture, calculate, the water absorption rate of trying to achieve the prescription medical material is 150%.

1.1.1.2 the selection of decocting condition is with amount of water (A), decocting time (B) decocts 4 factors that number of times (C) and soak time (D) decoct recovery rate for influence and decoct quality, and every factor 3 levels are with L9 (3 ⁴) orthogonal table makes an experiment, with must measuring of baicalin in the 100g prescription drug as evaluation criterion.

The design of table 1 decocting condition orthogonal test gauge outfit

Table 2 water extraction process orthogonal test table

Tested number NO	Quantity of solvent (A)	Decocting time (B)	Decoct number of times (C)	Soak time (D)	Yield (%)	Content of baicalin (%)	Total baicalin amount (gram) *
								1	1 (5 times)	1 (1 hour)	1 (1 time)	1 (1 hour)	12.8	5.94	0.76
2	1	2 (2 hours)	2 (2 times)	2 (2 hours)	27.2	4.91	1.33
								3	1	3 (3 hours)	3 (3 times)	3 (3 hours)	28.1	2.48	0.70
4	2 (10 times)	1	2	3	29.6	3.05	0.90
								5	2	2	3	1	37.2	5.52	2.05
6	2	3	1	2	16.0	5.85	0.93
								7	3 (15 times)	1	3	2	31.9	2.86	0.91
8	3	2	1	3	13.8	2.58	0.36
								9	3	3	2	1	36.0	5.40	1.94
I _j	2.79	2.57	2.05	4.75	?	?	?
								II _j	3.88	3.74	4.17	3.17	?	?	?
III _j	3.21	3.57	3.66	1.96	?	?	?
								I _j	0.93	0.86	0.68	1.58	?	?	?
II _j	1.29	1.25	1.39	1.06	?	?	?
								III _j	1.07	1.19	1.22	0.65	?	?	?
R	0.36	0.39	0.71	0.93	?	?	?

In visible 4 factors of The above results:

D＞C＞B＞A

Wherein: A2＞A3＞A1; B2＞B3＞B1;

C2＞C3＞C1；D1＞D2＞D3。

According to orthogonal experiments, optimum extraction process is:

A ₂B ₂C ₂D ₁

* content of baicalin assay method:

[assay] measured according to HPLC (appendix VID)

Chromatographic condition and system suitability test use octadecylsilane chemically bonded silica to be filler; Methanol-3.5% acetic acid (43: 57) is mobile phase; Detect wavelength 278nm, theoretical cam curve is pressed the baicalin peak and is calculated, and should be not less than 1500.

The preparation precision of reference substance solution takes by weighing at 4 hours baicalin reference substance of 60 ℃ of vacuum dryings an amount of, adds 70% ethanol and processes the solution that every 1ml contains 200 μ g, as reference substance solution.

About 1 gram of these article is got in the preparation of need testing solution, and accurate the title decides, and adds 70% ethanol 30ml; Put in the water-bath reflux 3 hours, and put coldly, filter; Filtrating is put in the 50ml measuring bottle, and with a small amount of 70% ethanol gradation washing container and residue, washing liquid is filtered in the same measuring bottle; Add 70% ethanol dilution to scale, shake up.

Accurate respectively reference substance solution 15 μ l and the need testing solution 5 μ l of drawing of algoscopy inject chromatograph of liquid, measure, and promptly get.

Pair of orthogonal test optimum condition has carried out repetition and scale-up, and the result lists following table in:

Table 3 repeats and scale-up

* content of baicalin is 8.33% in the radix scutellariae medicinal materials.

1.1.2 the selection of impurities removing method

1.1.2.1 the purpose of removing of mechanicalness impurity is deposition and other mechanicalness impurity of removing in the water decoction, through repetition test, proves in the commercial production filter method commonly used, can achieve the goal like filter press etc.

1.1.2.2 impurity property composition is removed

Get prescription medical material 5kg, decoct, merge decoction liquor by above-mentioned best decoction scheme, mixing, 5 parts of five equilibriums, handle with following method respectively:

A, decoction liquor is concentrated into dried (water-boiling method);

B, decoction liquor adds ethanol after concentrating, and extremely pure content reaches 60% (decocting in water alcohol deposition method);

C, decoction liquor is passed through the D101 macroporous adsorbent resin, with 50% ethanol 6L eluting, eluent decompression recycling ethanol and evaporate to dryness (Amberlyst process-1)

D, same C is with 70% ethanol 6L eluting (Amberlyst process-2);

E, same C is with 85% ethanol 6L eluting (Amberlyst process-3);

Get the above 5 kinds of extractum that are equivalent to 1g prescription medical material respectively, add 70% ethanol 80ml, heating in water bath refluxed 2 hours, put coldly, filtered, and filtrating is put in the 100ml measuring bottle, adds 70% ethanol to scale, shakes up, as sample solution.Other gets the about 2mg of baicalin reference substance, and accurate the title decides, and puts in the 10ml measuring bottle, adds 70% ethanol to scale, shakes up, as reference substance solution.Accurate reference substance and each 5 μ l of different sample solution of drawing; Inject high performance liquid chromatograph; Carry out the mensuration of baicalin according to the content assaying method of regulation in " quality standard draft and draft explanation ", calculate content of baicalin in the distinct methods gained extractum, result of the test is summarized in table 4.

The comparison of table 4 water decoction impurities removing method

Method	The product character	The thin layer chromatography inspection	Yield (%)	Content of baicalin (%)	Baicalin conversion ratio (%)
						A	The strong moisture absorption bitter in the mouth of dark-brown extractum good water solubility	Detect Herba Oxytropis Kansuensis, Radix Et Rhizoma Rhei	36.20	4.88	78.54
B	Pale brown color extractum moisture absorption bitter in the mouth water solublity is better	Detect Herba Oxytropis Kansuensis, Radix Et Rhizoma Rhei	20.34	5.71	51.64
						C	The yellow pulverizing extractum that is prone to is difficult for moisture absorption bitter in the mouth * *	Detect Herba Oxytropis Kansuensis, Radix Et Rhizoma Rhei	9.65	11.96	51.13
D	The yellow pulverizing extractum that is prone to is difficult for moisture absorption, bitter in the mouth	Detect Herba Oxytropis Kansuensis, Radix Et Rhizoma Rhei	12.00	12.95	69.09
						E	The yellow pulverizing extractum that is prone to is difficult for moisture absorption, the bitter in the mouth poorly water-soluble	Detect Herba Oxytropis Kansuensis, Radix Et Rhizoma Rhei	10.85	12.55	60.54

*, content of baicalin is 8.33% in the medical material.

*, the inspection of baicalin and other flavones ingredient in C, D, the discarded water liquid of the E three therapeutic methods of traditional Chinese medicine:

(1), get C, D, depleted each 100ml of water liquid of the E three therapeutic methods of traditional Chinese medicine, be concentrated into driedly, residue adds methanol 10ml, dissolving is filtered, and gets filtrating 1ml, adds a small amount of and hydrochloric acid 1ml of magnesium powder, is not the color reaction of flavone compound;

(2), the methanol solution of the methanol solution of above-mentioned C, D, E three therapeutic methods of traditional Chinese medicine gained sample, baicalin and control medicinal material and garbage (water liquid concentrate and get) is carried out thin layer chromatography result relatively, do not see the corresponding speckle appearance that baicalin, rhubarb anthraquinone and Herba Oxytropis Kansuensis, Fructus Chebulae's medical material are arranged in the garbage chromatograph

The eluting terminal point determining: get macroporous resin 70% ethanol elution 20ml, water bath method, residue are used the 5ml70% dissolve with ethanol, filter, and filtrating adding magnesium powder is an amount of, and hydrochloric acid 3-4 drips, and compare with the retinue blank, should not show the flavonoid color reaction.

In addition, still available thin layer chromatography is judged the eluting terminal point whether to detect flavone compound, and method is following:

The preparation of reference substance solution takes by weighing at 4 hours baicalin reference substance of 60 ℃ of vacuum dryings an amount of, adds methanol and processes the solution that every 1ml contains 2mg, as reference substance solution.

Macroporous resin 70% ethanol elution 20ml is got in the preparation of need testing solution, and water bath method, residue are used the 2ml dissolve with methanol, as need testing solution.

Test according to thin layer chromatography (appendix IVB of Chinese Pharmacopoeia version in 2000); Draw need testing solution, each 10 μ l of reference substance solution, put respectively in same be on the silica gel g thin-layer plate of binding agent with the sodium carboxymethyl cellulose, be developing solvent with n-butyl alcohol-glacial acetic acid-water (6: 1.5: 2.5); Launch; Take out, dry, spray 1%FeCl ₃The ethanol test solution, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on, must not show identical dirty-green speckle.

Visible by testing and go up in the table:

1, water decoction with B, C, D, the removal of impurity of E method after, the representative compound content of baicalin all has loss in various degree in the effective ingredient;

2, the extractum content of A, B two method gained is high, but content of baicalin is low, so consumption is big, and because of the extractum hygroscopicity is stronger, preparation need add a large amount of adjuvants, is its significant disadvantages;

3, C, D, E three therapeutic methods of traditional Chinese medicine products obtained therefrom character are roughly the same, but C method recovery rate with the equivalent solvent elution time is lower, and the conversion ratio of baicalin is also low.As increase the eluting solvent consumption, and though can improve recovery rate, elution time is long, solvent recovering rate is low, is its shortcoming;

4, the product of E method gained and other method products obtained therefrom compare, and water solubility is relatively poor, because used concentration of alcohol is higher, cost is also higher;

5, to compare with C, E method, the D method has product recovery rate and the equal advantage of higher of baicalin conversion ratio;

6, all do not detect the main component in the prescription medical material in the discarded water liquid of C, D, the E three therapeutic methods of traditional Chinese medicine.

Main component is impurity property compositions such as sugar, inorganic salt, pigment, macromolecular tannin polymer in the waste liquid through analyzing.

Above-mentioned evidence: with the water decoction of macroreticular resin absorbing method processing prescription medical material, both can remove hygroscopicity composition wherein, and can make the effective ingredient enrichment in the medical material again, and think that therefore with this method removal of impurity be feasible.

1.1.3 macroporous adsorbent resin process conditions screening

1.1.3.1 the selection of macroporous adsorbent resin type is according to big this characteristic of chemical compound polarity such as effective ingredient flavonoid, phenolic acids and anthraquinone glycoside of prescription medical material; Select for use nonpolar adsorption resin D101, Diaion HP-20 and low pole adsorbent resin D201 to make an experiment; The result proves that D101, two kinds of resins of Diaion HP-20 have similar adsorption effect; Resulting product has essentially identical thin layer chromatography; But D101 resin system home products, price is cheap far beyond Diaion HP-20, is fit to commercial production and China's national situation.The D201 resin is lower to the absorption affinity of phenol, sour isopolarity composition, and adsorbance is little, and resin demand is big, and elution time is long, and solvent consumption is many, so be not suitable for effective ingredient enrichment in the prescription medical material.According to result of the test, select the D101 macroporous adsorbent resin for use.The removal of impurity technology that this resin of evidence is used for the preparation of this prescription material medicine is feasible.

1.1.3.2JD-1 (WLD) type macroporous adsorbent resin data

JD-1 (WLD) type macroporous adsorbent resin belongs to D101 type macroporous adsorbent resin, and its related standards and parameter are following:

1, JD-(WLD) type macroporous adsorbent resin company standard code: Q/7144991-(B513902209)

2, JD-1 (WLD) type macroporous adsorbent resin enterprise specification standards

Title: macroporous adsorbent resin

Model: JD-1 (WLD)

Structure: styrene type EVA

Cross-linking agent: divinylbenzene

Porogen: toluene

Outward appearance: milky or faint yellow pure pearl body

Abnormal smells from the patient: free from extraneous odour

Polarity: nonpolar

Physical parameter:

Water content (%)≤70

Granularity (0.23-1.25mm, %) >=95

Bulk density (dry state, g/ml) 0.23-0.32

Wet apparent density (g/ml) 0.62-0.68

Skeletal density (g/ml) 1.00-1.20

Specific surface area (m ²/ g) 350-450

Average pore size (nm) 35-65

Porosity (%) 60.0-68.0

Pore volume (ml/g) 1.70-2.20

Resin residue thing amount:

A. it is not muddy that 3 times of amounts of muddy test adsorbent resin alcohol leaching liquid adds the equivalent distilled water.

B. toluene, xylene＜50ppm

C. divinylbenzene and diethylbenzene class＜40ppm

D. alkane＜1ppm

E. styrene＜1ppm

F. phenol＜1ppm

G. acrylonitrile＜1ppm

Saturated extent of adsorption >=40mg/g

Other character chemical property is stable, acid and alkali resistance, water insoluble and common organic solvents is all stable to general oxidation, Reducing agent and heat (＜150 ℃).

1.1.3.3D101 the pass of absorption with macroporous adsorbent resin amount and temperature ties up to the relation of having investigated resin absorption amount and temperature under the different temperatures, three times result of the test is listed following table in:

The relation of table 5 temperature and adsorbance ℃

Visible by last table, with the rising of fluid temperature, the D101 macroporous adsorbent resin reduces the absorbability of composition in the prescription medical material water decoction.Therefore must be noted that the temperature of water decoction and the consumption of resin, in order to avoid cause damage because of leaking to inhale.Usually decoction liquor is placed or is adjusted to below 25 ℃ to well.

1.1.3.4 5 parts of water lotion consumption water intaking decoction liquor; Every part of 1L is respectively through 5 identical macroporous resin adsorption posts, respectively with the water washing of 1,2,3,4,5 times of medical material amounts; Continue with 70% ethanol elution, carry out thin layer chromatography behind the ethanol elution concentrating under reduced pressure of each post relatively.Evidence with the water washing adsorption column of 4 times of amounts after resulting eluate promptly have stable thin layer chromatography, therefore think that the water washing adsorption column with 4 times of medical material amounts gets final product.

1.1.3.570% the ethanol elution consumption carries out the eluting terminal point determining with thin layer chromatography, proves the eluting that can accomplish effective site with 70% ethanol of 6 times of amounts, and alcoholic acid losing quantity is about 25-30%.

1.1.3.6 elution speed is got prescription medical material 10kg, decocts by best decocting condition, merges decoction liquor; 5 parts of five equilibriums; Through 4kg D101 macroporous resin absorption post is housed, after cylinder is used the 8L water washing respectively, use the friction speed eluting respectively with 70% ethanol; And judge the eluting terminal point to detect baicalin with thin layer chromatography, result of the test is listed following table in:

The influence of the different elution speeds of table 6

Visible by last watch: more complete because of desorption when flow velocity is slow, the eluant consumption is less, and along with the quickening of elution speed, because of desorption is incomplete, required quantity of solvent also increases.Therefore, elution speed with every kg resin minute 0.02-0.04L for well.In middle trial production, press the 100kg macroporous resin and calculate, when elution speed is per minute 2-4L usually, in 90 minutes, can finish by eluting.Also find in test that because of desorption needs the regular hour elution speed can not infinitely increase, generally flow velocity can slow down automatically behind the beginning eluting.

1.1.3.7 the pretreatment of resin and regeneration

1.1.3.7.1 the new resin of the pretreatment of resin carries out pretreatment by conventional method usually, promptly with acetone or the soak with ethanol that contains 5% hydrochloric acid after one week, water is washed till neutrality and can uses.Once sour water on probation or aqueous alkali replace acetone or sour ethanol to do the pretreatment of resin, and the absorbability of resin is poor as a result, removal of impurity poor effect.

1.1.3.7.2 regeneration of resin has been carried out the comparison of different renovation process.Get 6 of glass columns, pretreatment D packs in the every post ₁₀₁Macroporous adsorbent resin 100g; And add prescription medical material water decoction and make the resin state (having the sucking-off of leakage existing) that reaches capacity with thin layer chromatography and test-tube reaction inspection; Every post with the 100ml water washing after, the 1-5 post is handled with different regeneration methods, the adsorption column after the regeneration carries out the absorption of prescription drug water decoction once more; 70% ethanol carries out eluting, and eluent concentrates, weigh after the drying.No. 6 post is not regenerated and absorption for the second time, and as contrast, result of the test is listed following table in eluate first time weight:

The comparison of table 7 macroporous resin renovation process

Visible by last table:

1, sour water can not be as regenerated solvent;

2, aqueous alkali can partly be removed remaining impurities on the post;

3, sour ethanol effectively resin regenerate;

4, reuse acid Ethanol Treatment can not only be regenerated to resin effectively after alkali liquor was removed partial impurities, and can reduce sour amount of ethanol;

5, acetone can partly be removed remaining impurities on the post, but regeneration effect is not so good as sour ethanol, and cost is higher.

In sum, visible renovation process is best with alkali liquor+sour alcoholic acid method, and repeatedly middle practice of trial production proves that also this method is feasible.

1.1.4 concentrated and dry owing to adopt macroporous resin removal of impurity technology, water decoction need not concentrate; Ethanol elution then adopts the method for concentrating under reduced pressure to concentrate by routine.

Vacuum drying and two kinds of methods of spray drying are compared, and the result lists following table in:

Table 8 vacuum drying and spray-dired comparison

Visible by last table: the result of two kinds of drying means gained is basic identical, and the drying means that all can be used as these article is described.Vacuum dryer is comparatively simple, and the general pharmaceutical factory of traditional Chinese medicine all possesses this condition, but the required time is longer, and need pulverize after the drying, and product color is darker, is its shortcoming.Spray drying products obtained therefrom quality need not pulverized for well than vacuum is dry, and powder size is thin, is easy to stir during the preparation emulsifiable paste, and the finished product outward appearance is even, fine and smooth, color is shallow, attractive in appearance, is its advantage.So, advocate and adopt spray drying method to carry out drying in pharmaceutical factory with good conditionsi.

Because whether stablizing of material medicine is normal relevant with the content of its moisture, in order to reduce its moisture, spray-dired condition is studied, the result lists following table in:

The comparison of table 9 spray drying condition

Visible by last table: during as evaluation criterion, the product water content of test 2,3 is all lower with water content, but tests 3 higherly because of relative density, and spraying is difficulty, and the quality of product is than 1,2 for poor, so think that to test 2 condition better.

1.1.5 middle trial production by above-mentioned decoction, the removal of impurity, concentrate, technology such as drying carries out trial production in three batches, the result lists following table in:

Produce result's (intermediate products) in table 10 material medicine as a trial

* the medical material content of baicalin that is used for this test is 8.33%.

1.2 preparations shaping technical study

1.2.1 the selection of dosage form

Medicine medicated powder of the present invention through decocting, the removal of impurity, concentrate, technology such as drying makes, the effective ingredient water solublity is good, and stable in properties.Medicine of the present invention is mainly used in dermatosiss such as treatment eczema, contact dermatitis, neurodermatitis, is widely used in the Tibetan area.Former cream uses vaseline to be substrate, and water absorption is poor, and greasy is big, is difficult for washing, and influences penetrating and absorbing of medicine.To effect, purposes and the medicinal property of medicine of the present invention, plan does not change pharmaceutical dosage form, and the substrate of former dosage form has been carried out screening again.

1.2.2 the selection of matrix species

Oil, water solublity, oil-in-water (O/W), Water-In-Oil (W/O) emulsion-type substrate are prepared ointment and screens.

1, greasing base

Prescription: material medicine 1g

Liquid paraffin is an amount of

Vaseline adds to 20g

Method for making: get material medicine and put in the mortar, add appropriate amount of fluid paraffin and be ground into pasty state, gradation adds vaseline, grinds well promptly to get.

2, water-soluble base

Prescription: material medicine 1.0g

Glycerol 0.4g

Gelatin 4.0g

Distilled water 15.5ml

Method for making: get glycerol, gelatin, the dissolving of water mixing post-heating, must freeze the shape thing after the cooling, in mortar, add material medicine, grind well promptly and get.

3, water-in-oil type (W/O) emulsifiable paste matrix

Prescription: material medicine 1.00g glyceryl monostearate 2.00g

Paraffin 2.00g white vaseline 1.00g

Liquid paraffin 10.00g span 40 0.10g

Polyoxyethylene nonylphenol ether 0.10g ethylparaben 0.02g

Distilled water 5.00ml

Method for making: glyceryl monostearate, white vaseline, liquid paraffin, span 40, polyoxyethylene nonylphenol ether are heated to 80 ℃ make fusing; Ethylparaben is soluble in water; The water heating also remains on 80 ℃, and fused oil phase is slowly added water, and the limit edged is stirred to condensation; Promptly get water-in-oil emulsion type substrate, the adding material medicine grinds well promptly and gets in mortar.

4, oil-in-water (W/O) type emulsifiable paste matrix

Prescription: material medicine 1.00g glyceryl monostearate 0.40g

White vaseline 2.40g octadecanol 2.40

Glycerol 1.40g sodium lauryl sulphate 0.30g

Vitamin C 0.20g ethyl hydroxybenzoate 0.04g

Distilled water 10.00ml

Method for making: white vaseline, octadecanol, glyceryl monostearate are heated to 70-80 ℃ make the substrate fusing; Sodium lauryl sulphate, glycerol, vitamin C, ethyl hydroxybenzoate are dissolved in water and are heated to 70-80 ℃; Under constantly stirring, water is slowly added oil phase (70-80 ℃); Continue to be stirred to condensation, promptly get O/W type emulsion-type substrate, the adding medicine grinds well promptly and gets in mortar.

Result to above four kinds of substrate test shows that greasing base ointment greasy is strong, washing inconvenience, and be prone to dirty medicated clothing; It is strong that water-soluble base ointment has viscosity, is difficult for the shortcoming of coating; Water-in-oil type emulsifiable paste matrix ointment also has greasy strong, is difficult for washing, is prone to dirty medicated clothing, and medicine is difficult for shortcomings such as release and absorption; Oil-in-water type emulsifiable paste matrix ointment non-greasy is prone to washing, and feel is good; Help medicine in dermal osmosis and absorption; So think that oil-in-water emulsion type substrate obviously is superior to other three kinds of substrate, therefore, confirm to adopt oil-in-water (O/W) type emulsifiable paste matrix at last as ointment base.

1.2.3 the comparison of medicine adding method

Adding method to material medicine is studied; Find that different adding methods has tangible influence to the quality of ointment; Result of the test proves: material medicine forms initial stage (70-80 ℃) adding at emulsifiable paste; Be stirred to condensation, emulsifiable paste has little water to analyze after placing, and the color and luster of emulsifiable paste is also even inadequately; Material medicine adds in condensed emulsifiable paste, and the ointment color and luster is even, places the no moisture in back and oozes out.Its reason possibly be that material medicine extractum absorbs moisture at the higher temperature that emulsifiable paste forms the initial stage, has destroyed the emulsion layer of emulsifiable paste, thereby has influenced the stability of emulsifiable paste.Therefore, the blank substrate of the suitable preparation earlier of the method for preparing of medicine of the present invention adds the material medicine mixing then.

1.2.4 prescription is formed and method for preparing is passed through the comparative study to different substrates, selects the substrate of oil-in-water type emulsifiable paste matrix as prescription drug for use, preparation prescription composition and method for preparing are following:

1), prescription is formed

Oil phase: white vaseline 150g

Octadecanol 150g

Glyceryl monostearate 20g

Water: sodium lauryl sulphate 15g

Glycerol 75g

Vitamin C 10g

Ethyl hydroxybenzoate 1g

Distilled water 630ml*

Material medicine 20g**

*, distilled water has part because of evaporation loss in heating process.

*, material medicine calculates with recovery rate 10%.

Sodium lauryl sulphate is an emulsifying agent in the substrate, and glycerol is wetting agent, and vitamin C is an antioxidant, and ethyl hydroxybenzoate is an antibacterial.

2), method for preparing is heated to 70-80 ℃ with oil phase makes substrate fusing, water is slowly adding oil phase (70-80 ℃) with water in 70-80 ℃ of heating for dissolving under constantly stirring; Continue to be stirred to substrate and solidify (temperature is about 30-40 ℃), stop to stir, a small amount of blank substrate is put in the ointment slab; Adding material medicine grinds well; Progressively add blank substrate, grind well, promptly get.

2, method for preparing

2.1 technological process

The prescription medical material adds 12 times of water logging bubbles 1 hour, and heated and boiled 2 hours is got decoction liquor; Medicinal residues add the water of 10 times of amounts to be fried in shallow oil 1 time again, merges decoction liquor, filters; Filtrating is put cold, through the D101 macroporous adsorptive resins of having handled well, discards effusive water liquid; Water washing adsorbent resin cylinder with 4 times of amounts of medical material amount discards cleaning mixture, and the ethanol elution with 70% does not detect (amount of ethanol is about 4-5 times of medical material amount) to there being flavones ingredient.Merge ethanol elution, concentrating under reduced pressure, vacuum (or spraying) drying, check adds blank oil-in-water type emulsifiable paste matrix, stirs, and packing promptly gets.

2.2 middle trial production

Manufactured experimently 3 batches " medicines of the present invention " according to clinical trial continuously with the technology of sample preparation, every batch of inventory is 3.325kg, and 3 batches of pilot product experimental results are seen table

Produce the result in the table 11 as a trial

2.3 the quality examination of pilot product

The 3 batches of pilot products by " prescription under an appendix IR of the Chinese pharmacopoeia current edition ointment item, by " drug standard draft " inspection, and carry out health examination *, the result sees the following form.

Table 12 quality control

Below prove beneficial effect of the present invention through pharmacodynamics test.

Below through four aspects such as antiinflammatory, antibacterial, antipruritic, immunomodulating, select its pharmacological action of test model objective evaluation of generally acknowledging both at home and abroad, when carrying out clinical research the pharmacology is provided reference material for the clinicist.

Medicine and reagent

Medicine of the present invention: substrate, white cream, not drug.10% medicine of the present invention (by embodiment 1 preparation), faint yellow cream, every 100g frost contains crude drug in whole 10g.20% medicine of the present invention, the yellow cream of light brown, every g contains crude drug in whole 20g.40% medicine of the present invention, isabelline cream, every 100g frost contains crude drug in whole 40g.By the medicine chamber Luo Zeyuan researcher of this institute lot number 990906 is provided.

Fluocinonide ointment: the accurate word (1981) of medicine is defended No. 001463 in Tianjin.Specification: 10g:25mg/ props up.Usage: external is applied to the affected part, 2-3 time/day.Tianjin pharmaceutcal corporation, Ltd produces, lot number 980807.

Ice the happy ointment of yellow skin: (98) are defended the accurate word Z-031 of medicine number.Specification: 15g/ props up.Main component: compositions such as Radix Et Rhizoma Rhei, Rhizoma Curcumae Longae, sulfur, Borneolum Syntheticum.Have effects such as dispeiling pathogenic wind and removing dampness, heat-clearing and toxic substances removing, killing parasites for relieving itching, be used for the skin pruritus due to neurodermatitis, eczema, tinea pedis and the psoriasis etc.Usage: external is applied to the affected part, 3 times/day.Tibet Zhizhi Pharmacy Group Co., Ltd. produces, lot number 980616.

JIEERYIN XIYE: (93) are defended the accurate word Z-32 of medicine number.The secret kind of country's Chinese patent medicine.Prescription is formed Fructus Cnidii, Radix Sophorae Flavescentis, Rhizoma Atractylodis, Cortex Phellodendri etc.Specification 60ml/ bottle.Heat clearing and damp drying, killing parasites for relieving itching.(1) cures mainly women's damp-heat vaginal discharge.Disease sees that pudendal pruritus is red and swollen, profuse leukorrhea, yellow skin or like the bean dregs shape, bitter taste xerostomia, yellowish urine constipation, red tongue with yellowish and greasy fur, wiry and frequent pulse.Be applicable to mycotic, infusorian property and nonspecific vaginitis.(2) be used for following dermatosis: acute eczema (damp-heat type), contact dermatitis (pyretic toxicity folder wet type), tinea corporis and cruris (rheumatism pattern of fever).Chengdu En Wei pharmaceutical Co. Ltd produces, lot number 990813.

Ergamisole: the accurate word (1990) of medicine is defended No. 005517 in the river, specification 25mg5100 sheet, and Zi Gong the 3rd pharmaceutical factory produces, lot number: 970305.

Cyclophosphamide sheet: the accurate word of Su Wei medicine (82) 2358-1 number.Specification: 50mg/ sheet, 100 slices/bottle.Nantong second pharmaceutical factory produces, lot number 980201.

Pentobarbital sodium: analytical pure, the 25g/ bottle meets BP 1958 editions, Denmark's import packing, China Drug Co.'s Shanghai chemical reagent purchasing and supply station, lot number 970213.

Benzylpenicillin sodium for injection: specification 800,000 u/ bottles, Ji are defended the accurate word 1995 of medicine No. 000004, North China pharmaceutical Co. Ltd product, lot number 980725T.

ZHUSHEYONG LIUSUAN LIANMEISU: specification 1,000,000 u/ bottles, Ji are defended the accurate word 1995 of medicine No. 000010, North China pharmaceutical Co. Ltd product, lot number 9809114.

2.4-dinitrochlorobenzene: CP level, 25g/ bottle, Shanghai reagent one factory's product, lot number 981001.

Sodium sulfide: analytical pure, the 500g/ bottle, chemical reagent factory in Chengdu produces, lot number 980923.The Dextran 40 glucose injection: the accurate word (1981) of medicine is defended No. 000747 in the river, Sichuan hundred news pharmaceutical Co. Ltds, lot number 990402.

Xylene: the 500ml/ bottle, analytical pure, chemical reagent factory in Chengdu produces, lot number 980416.

Carrageenin: specification 10g/ bottle, Shenyang Pharmacy College provides, lot number 980801.

India ink: the biological dye, specification 50ml/ bottle, the chemical plant produces in the west, Beijing, lot number 980301.

Alsever liquid (glucose 2.05g, citrin sodium 0.8g, sodium chloride 0.42g, adding distil water is to 100ml.9 pounds of sterilizations in 10 minutes are subsequent use).

Hemoglobin conversional solution (Shanghai [G/95] defends medicine and give birth to the card word No. 20 for Dou Shi reagent, HiCH method, and the glad biotechnology research of Shanghai section provides, lot number 990702).

Sodium carbonate: analytical pure, 500g/ bottle, Chongqing chemical reagent factory product, lot number 971218.

Animal and bacterial strain

The SD rat, one-level is qualified, and No. the 90th, the real moving pipe in river is provided by institute of antibiotics, Sichuan Province Experimental Animal Center.

Kunming mouse, one-level is qualified, and No. the 92nd, the real moving pipe in river is provided by institute of antibiotics, Sichuan Province Experimental Animal Center.

The NIH mice, one-level is qualified, and No. the 117th, the real moving pipe in river is provided by Chengdu Biological Products Inst., Ministry of Public Health's Experimental Animal Center.

Cavia porcellus, one-level is qualified, the real moving pipe in river 98-2 number, laboratory animal room of Blood Transfusion Inst., Chinese Academy of Medical Sciences provides.

Bacterial strain: select the reference culture and clinical isolates strain 16 strains of dermatosis common pathogen.Wherein staphylococcus aureus ATCC6538, escherichia coli ATCC25922, bacillus pyocyaneus ATCC27853, Candida albicans ATCC10231, beta hemolytic streptococcus ATCC19615, Cray Bai Shi bacillus ATCC13833 are internationally recognized quality control standard bacterial strain, are provided by Sichuan Province epidemic prevention station antibacterial section.All the other clinical strains derive from the separation recently of People's Hospital, Sichuan Prov. or Sichuan Province's epidemic prevention station and preserve bacterial strain.

Experimental apparatus

Day island proper Tianjin EB-3200D precise electronic balance (precision 0.01g).

The JN-B of Shanghai Second Balance Factory type precision torsion balance (0.5 precision mg).

The Chongqing CS213 of testing equipment factory electric heating incubator.

The LD5-2A of Beijing Medical Centrifugal Machine Factory centrifuge.

Shanghai the 3rd analytical tool factory 722 type grating spectrophotometers.

Shanghai City second hardware factory 501 type thermostatic water-circulator bath casees.

Day island proper Tianjin UV-730 semi-automatic biochemical analyzer.

Rat foot volume determination appearance.

Time clock etc.

The test of Test Example 1 medicine antiinflammatory action of the present invention

1.1 medicine xylol of the present invention causes the influence of mice ear

(1) purpose: cause chmice acute ear swelling model with xylene, cause the antiinflammatory action that scorching front and back mice ear degree is observed medicine of the present invention through mensuration.

(2) method: 70 of Kunming mouses, male, body weight 18 ~ 22g is divided into 7 groups at random by body weight, 10 every group.If normal control group (blank), model control group (substrate) and positive controls (Western medicine fluocinonide ointment, Chinese medicine are iced the happy ointment of yellow skin); Receive 3 dose groups of reagent thing (10%, 20%, 40% medicine of the present invention; Promptly 10,20,40g crude drug in whole/100g frost, be equivalent to clinical plan 0.5,1,2 times respectively with concentration).20 μ l drip in mouse right ear with xylene, smear in mouse right ear behind the 1min to be administered once, and weight such as employing not isoconcentration ointment are smeared the auris dextra administration, dosage 0.2g frost/only.Wipe medicine gently away and put to death animal with cotton balls behind the 20min, with card punch (diameter 9mm) in the left and right sides ear same area beat and get auricle, weigh respectively, with two auricle weight differences as auricular concha swelling degree.Respectively organize swelling degree and inhibitory rate of intumesce according to computes,, compare each group difference significance, inspection level α=0.05 with t or t ' check.

Swelling degree=auris dextra sheet heavy (mg)-left auricle heavy (mg)

(3) result: visible by table 1, medicine of the present invention tries 3 dose groups mice auricular concha swelling degree and all significantly is lower than and model control group, and p＜0.05 or 0.01 shows that medicine xylol of the present invention causes the chmice acute ear swelling inhibitory action is arranged.

The influence of the acute ear swelling of table 1 medicine xylol of the present invention induced mice (x ± S)

Compare (t check or t ' check) with model control group: * P＞0.05; * P＜0.05; * * P＜0.01.

1.2 medicine on Carrageenan of the present invention is brought out the influence of rat foot claw swelling

(1) purpose: be chosen in the effect proinflammatory agent carrageenin local injection that injects the 2-4 hour swelling peaking in back and bring out rat foot claw swelling, the characteristics of this model are to cause the synthetic increase of scorching local PG, and bring out edema with vaso-active substance and kassinin kinin class.Cause the antiinflammatory action that medicine of the present invention is observed in the volumetrical variation of scorching front and back rat paw edema through measurement.

(2) method: 70 of SD rats, male, body weight 120 ~ 150g is divided into 7 groups at random by body weight, 10 every group.If normal control group (blank), model control group (substrate) and positive controls (Western medicine fluocinonide ointment, Chinese medicine are iced the happy ointment of yellow skin); Receive 3 dose groups of reagent thing (10%, 20%, 40% medicine of the present invention; Promptly 10,20,40g crude drug in whole/100g frost, be equivalent to clinical plan 0.5,1,2 times respectively with concentration).Weight such as employing not isoconcentration ointment are smeared the left hind administration, and dosage 0.5g frost/only, administration 2 times (at a distance from 4 hours) between the first day causes inflammation for morning next day the left back sufficient plantar aponeurosis injected 1% carrageenin 0.1ml of rat, smear immediately then to be administered once.Adopt rat foot claw volume determination appearance to measure to cause scorching before with cause scorching back different time (at interval 1 ~ 2 hour once, continuous 5 times) rat foot claw volume, and according to computes inhibitory rate of intumesce (%).The result compares each group difference significance, inspection level α=0.05 with the t check.

(3) result: can know by table 2; When the carrageenin local injection brings out rat sufficient pawl swelling in 1 ~ 8 hour peak; 3 dose groups rat foot claws of medicine of the present invention swelling degree is starkly lower than matched group; Have significant difference significance (P＜0.05 or＜0.01), show that medicine on Carrageenan local injection of the present invention brings out rat foot claw swelling inhibitory action is arranged.

Table 2 medicine on Carrageenan of the present invention is brought out the influence (x ± S) of rat foot claw swelling

Annotate: () interior numerical value is inhibitory rate of intumesce (%)

Compare (t check or t ' check) with model control group: ^*P＞0.05; ^*P＜0.05; ^{* *}P＜0.01.

5.1.3 the influence that medicine of the present invention forms the mice granuloma induced by implantation of cotton pellets

(1) purpose: cotton balls is implanted in the mice body can cause connective tissue proliferation, and this model can be observed medicine of the present invention to anti-inflammatory later stage connective tissue proliferation function.

(2) method: 62 of Kunming mouses, male, body weight 23 ~ 27g is divided into 6 groups at random by body weight, and every group of 10-11 is only.If model contrast (substrate) and positive drug matched group (Western medicine fluocinonide ointment, Chinese medicine are iced the happy ointment of yellow skin); Receive 3 dose groups of reagent thing (10%, 20%, 40% medicine of the present invention; Promptly 10,20,40g crude drug in whole/100g frost, be equivalent to clinical plan 0.5,1,2 times respectively with concentration).Lumbar injection pentobarbital sodium 30mg/kg anesthetized mice; It is fixing to lie on the back, and iodine tincture, alcohol disinfecting hypogastric region skin are cut off along ventrimeson and to be about the 0.5cm otch; With 2 sterilization cotton balls (heavy 10mg of each cotton balls; Autoclaving drips each 0.05ml of 800u/ml penicillin and 650u/ml respectively, 50 ℃ of oven dry) to implant the left and right strange portion of mice respectively subcutaneous.Postoperative began to adopt the same day isometric(al) not isoconcentration ointment smear strange portion administration (matched group is smeared substrate), dosage 0.2g frost/only, every day 2 times, continuous 8 days.Animal is put to death in 1h cervical vertebra displacement after the last administration, and cotton balls is taken out together with connective tissue on every side, rejects fatty tissue; Place 55 ℃ of baking oven 24hr oven dry, weigh after the cooling, deduct the raw cotton ball weight; Promptly get granuloma weight, granuloma weight is with mg (granuloma)/100g (body weight) expression.The result compares each group difference significance, inspection level α=0.05 with the t check.

The influence that table 3 medicine of the present invention forms the mice granuloma induced by implantation of cotton pellets (x ± S)

Compare (t check or t ' check) with model group: ^*P＞0.05; ^*P＜0.05; ^{* *}P＜0.01.

(3) result: visible by table 3; Medicine of the present invention tries three dose groups granuloma induced by implantation of cotton pellets and weighs less than and matched group granuloma weight; The significant difference highly significant; And have certain dose-effect difference trend, can think that this medicine can suppress the formation of cotton balls granulation tissue, has antiinflammatory action to this inflammatory model.

The test of Test Example 2 medicine bacteriostasis of the present invention

2.1 medicine of the present invention is to the influence of skin common pathogen growth

(1) purpose: select the reference culture and the clinical isolates strain of dermatosis common pathogen, observe the bacteriostasis of the contained extractum of medicine of the present invention with test tube method.

(2) test method: accurately take by weighing drug extract powder 2.0g of the present invention (every gram is equivalent to crude drug in whole 10.6g), adding distil water is settled to 15.62ml, and making into concentration is the suspendible medicinal liquid of 128mg/ml, boiling water bath heating 15 minutes, and 4 ℃ of cold preservations are subsequent use.Get suspension 0.5ml during test with the meat soup dilution of opposing doubly, make that concentration is respectively 64,32,16,8,4,2,1mg/ml.Test organisms is inoculated in the plain agar flat board by nutritional requirement or blood agar is dull and stereotyped, cultivates 18 hours for 37 ℃, scrapes the antibacterial tongue that takes a morsel and is emulsifiable in meat soup (streptococcus is used serum broth), than its concentration of turbid correction, remakes 100,000 times of dilutions, makes final concentration be equivalent to 10 ⁴Cfu/ml meat soup.Minimal inhibitory concentration (MIC) measure to adopt test tube liquid doubling dilution, the every pipe dosage of test organisms liquid 0.5ml, former medicine final concentration 32,16,8,4,2,1,0.5mg/ml.Because the Chinese medicine color is darker and muddy, can't direct observation, after 37 ℃ of overnight incubation, every pipe is got an inoculating loop changes kind of an agar plate (streptococcus changes blood plate), to observe the growing state of every arm.Establish contrast of bacterium liquid meat soup and former medicine blank, positive drug (JIEERYIN XIYE) contrast simultaneously.

(3) visible by table 4; Medicine of the present invention all has the obvious suppression effect to dermatosis common pathogen staphylococcus aureus, staphylococcus epidermidis, beta hemolytic streptococcus, Candida albicans, escherichia coli, klebsiella, bacillus pyocyaneus, bacteriostasis intensity order (descending): klebsiella＞Staphylococcus aureus=bacillus pyocyaneus=staphylococcus epidermidis＞Candida albicans＞beta hemolytic streptococcus＞escherichia coli.

The external minimal inhibitory concentration of table 4 medicine of the present invention (MIC) is measured the result

Conclusion (of pressure testing):

Medicine local skin of the present invention administration can significantly suppress rat paw edema due to acute ear swelling of xylene induced mice and the carrageenin, and the mice granuloma induced by implantation of cotton pellets is formed with inhibitory action.Clinical and reference culture all has the obvious suppression effect that the obvious suppression effect is all arranged to medicine of the present invention to 16 strains such as dermatosis common pathogen staphylococcus aureus, staphylococcus epidermidis, beta hemolytic streptococcus, Candida albicans, escherichia coli, klebsiella, bacillus pyocyaneus, bacteriostasis intensity order (descending): klebsiella＞Staphylococcus aureus=bacillus pyocyaneus=staphylococcus epidermidis＞Candida albicans＞beta hemolytic streptococcus＞escherichia coli.Medicine of the present invention can significantly can also cause the certain inhibitory action trend of having of mice whole body pruritus to dextran to the local pruritus of guinea pig skin due to the antihistamine.In addition, medicine of the present invention also has significant antagonism to the auricular concha swelling that DNCB induced mice skin delayed hypersensitivity causes, and can improve the normal mouse mononuclear-macrophage phagocytic function, but the normal mouse hemolytic antibody is generated obviously influence of nothing.

Test Example 3 clinical drug trials of the present invention

One, clinical drug trial number of the present invention: 439 examples

Two, examine the material standard:

1, Western medicine diagnose standard

Subacute eczema:

(1) medical history:

1. often be that the untimely or malpractice of acute eczema treatment causes a disease due to the journey delay.

2. the conscious pruritus that has in various degree.

(2) sign:

1. boundary's stage between acute and chronic eczema.

2. the skin lesion inflammation is lighter, with pimple, and incrustation, squama is main, only has a small amount of papulovesicle or phlysis or slight erosion to ooze out.

2, Chinese medical discrimination standard

Damp-heat syndrome

Primary symptom: skin infiltration, ooze out, incrustation, erythema, with pruritus, scorching hot;

Inferior disease: vexed, thirsty, yellowish urine, big dry stool;

Tongue arteries and veins: red tongue, yellow and greasy fur, rolling pulse.

Possess primary symptom, doublely see in time disease can dialectically to be assertive evidence with reference to the tongue arteries and veins more than 1 or 1.

3, skin lesion pruritus degree

0 minute: no pruritus.

2 minutes: idol had pruritus, does not influence daily life.

4 minutes: the paroxysmal pruritus, the time light when heavy, the influence sleep.

6 minutes: violent pruritus had a strong impact on sleep and work.

4, skin lesion form

Erythema degree: 0 minute: do not have.

1 minute: slight, may be seen indistinctly.

2 minutes: moderate, apparent in view, color was redder.

3 minutes: severe, very obvious, color was scarlet.

Pimple degree: 0 minute: do not have.

1 minute: slight, be dispersed in, omit projection, color is little red.

2 minutes: moderate, comparatively dense, obvious projection, color is red.

3 minutes: severe, very dense is noncountable, obvious projection, color is scarlet.

Vesicle degree: 0 minute: do not have.

1 minute: slight, be dispersed in, may be seen indistinctly.

2 minutes: moderate, comparatively dense, or merge mutually.

3 minutes: severe, very dense merged in flakes.

Ooze out degree: 0 minute: do not have.

1 minute: slight, point-like was dispersed in, and mays be seen indistinctly.

2 minutes: moderate, many, see through napkin easily.

3 minutes: severe, a lot, be globule shape.

Rotten to the corn degree: 0 minute: do not have.

1 minute: slight, point-like was dispersed in.

2 minutes: moderate, the some lamellar, part merges mutually.

3 minutes: severe, very significantly large stretch of rotten to the corn face.

Incrustation degree: 0 minute: do not have.

1 minute: slight, be dispersed in tiny thin crust.

2 minutes: moderate, more faint yellow crust.

3 minutes: severe, the thicker yellow crusts of more mutual fusion.

5, target skin lesion area

Greatest length * the Breadth Maximum (cm2) of target skin lesion area=target skin lesion

0 minute: skin lesion disappeared fully.

1 minute: the skin lesion area reduced >=75%.

2 minutes: >=30% skin lesion area reduced＜75%.

3 minutes: the skin lesion area do not reduce or＜30% (treatment front target skin lesion area be 3 fens).

Annotate: can not be with facial eczema as the target skin lesion.

6, tcm syndrome standards of grading

Primary symptom: skin symptom/sign: with reference to above-mentioned standards of grading;

Inferior disease:

Vexed: 0 minute: as not have;

1 minute: have.

Thirsty: 0 minute: as not have;

1 minute: have.

Yellowish urine: 0 minute: do not have;

1 minute: have.

Big dry stool: 0 minute: do not have;

1 minute: have.

The tongue arteries and veins is not scored.

7, inclusion criteria

1. meet the subacute eczema diagnosis;

2. Chinese medical discrimination is a damp-heat syndrome;

3. skin lesion limits to or area≤body surface area 20%;

Three, process of the test: test group: the medicinal frost formulation of the present invention of Test Example 1 preparation, external, scumbling affected part, 3 times/day.

The clinical trial validity evaluation index:

1, skin lesion overall score (skin lesion pruritus degree+skin lesion form+target skin lesion area): before the medication, medication 7 days, medication evaluated in 14 days.

2, tcm syndrome curative effect: before the medication, medication 7 days, medication carried out the evaluation of tcm syndrome integration in 14 days.

3, skin each item symptom, sign change: before the medication, medication 7 days, medication carried out skin each item symptom in 14 days, sign changes evaluation.

4, take a picture in target skin lesion position: before the medication, medication carried out target skin lesion position in 14 days and take a picture.

Wherein, FAS (Full Analysis Set) refers to: the complete analysis collection; PPS (Per ProtocolSet) refers to: meet the scheme data set.

Brief summary:

Subacute eczema skin lesion overall score situation of change, medicinal frost formulation of the present invention can be treated the subacute eczema skin lesion, and its effect is not inferior to the happy ointment of the yellow skin of ice, FAS, PPS conclusion unanimity.FAS result: treat that subacute eczema skin lesion overall score test group on average descended 4.38 ± 3.40 fens after 7 days, matched group descended 4.23 ± 3.72 fens, and the variation of two groups of relative baselines all has statistical significance (P＜0.05).The comparison of two groups of treatments subacute eczema skin lesion overall score and baseline difference after 7 days, difference do not have statistics anticipate (P＞0.05).Treat that subacute eczema skin lesion overall score test group on average descended 8.37 ± 5.11 fens after 14 days, matched group descended 7.75 ± 5.60 fens, and each variation of organizing relative baseline all has statistical significance (P＜0.05).The least square mean of treating two groups of relative baseline variation differences of subacute eczema skin lesion overall score (test group-matched group) after 14 days is-0.954 (1.976,0.068).The covariance analysis result who considers treatment, center, baseline effect shows, the comparison of the two groups of treatments relative baseline variation of subacute eczema skin lesion overall score difference after 14 days, difference not statistically significant (P＞0.05).Consider that treatment, center, baseline, treatment and the interactive covariance analysis result in center show, the interaction item not statistically significant (P＞0.10) at treatment and center.PPS result: treat that subacute eczema skin lesion overall score test group on average descended 4.67 ± 3.27 fens after 7 days, matched group descended 4.77 ± 3.43 fens, and the variation of two groups of relative baselines all has statistical significance (P＜0.05).The comparison of two groups of treatments subacute eczema skin lesion overall score and baseline difference after 7 days, difference do not have statistics anticipate (P＞0.05).Treat that subacute eczema skin lesion overall score test group on average descended 8.88 ± 4.79 fens after 14 days, matched group descended 8.71 ± 5.01 fens, and each variation of organizing relative baseline all has statistical significance (P＜0.05).The least square mean of treating two groups of relative baseline variation differences of subacute eczema skin lesion overall score (test group-matched group) after 14 days is-0.548 (1.509,0.413).The covariance analysis result who considers treatment, center, baseline effect shows, the comparison of the two groups of treatments relative baseline variation of subacute eczema skin lesion overall score difference after 14 days, difference not statistically significant (P＞0.05).Consider that treatment, center, baseline, treatment and the interactive covariance analysis result in center show, the interaction item not statistically significant (P＞0.10) at treatment and center.PPS is similar with FAS result.

The tcm syndrome curative effect, medicinal frost formulation of the present invention has the effect that improves the subacute eczema damp-heat syndrome, and its curative effect is not inferior to the happy ointment of the yellow skin of ice, FAS, PPS conclusion unanimity.FAS result: treat the clinical recovery rate 16.3% of test group after 14 days, obvious effective rate 27.3%, effective percentage 40.5%, inefficiency 16.0%; The clinical recovery rate 9.3% of matched group, obvious effective rate 26.2%, effective percentage 42.1%, inefficiency 22.4%.In the deduction in the heart the CMH X 2 test of effect show that test group and matched group tcm syndrome curative effect grade comprehensively compare, difference not statistically significant (P＞0.05).PPS result: treat the clinical recovery rate 17.3% of test group after 14 days, obvious effective rate 29.0%, effective percentage 42.3%, inefficiency 11.4%; The clinical recovery rate 10.4% of matched group, obvious effective rate 29.2%, effective percentage 45.8%, inefficiency 14.6%.In the deduction in the heart the CMH X 2 test of effect show that test group and matched group tcm syndrome curative effect grade comprehensively compare, difference not statistically significant (P＞0.05).PPS is similar with FAS result.

Individual event skin symptom/sign curative effect: medicinal frost formulation of the present invention has the effect of treatment subacute eczema each individual event skin symptom/sign (pruritus degree, erythema degree, pimple degree, vesicle degree, ooze out degree, rotten to the corn degree, incrustation degree, target erythra area); Wherein, The effect of medicinal frost formulation treatment target skin lesion area of the present invention is superior to icing the happy ointment of yellow skin, and FAS is consistent with the PPS conclusion.The effect of treating other each item individual event skin symptom/signs is not inferior to the happy ointment of the yellow skin of ice.1. pruritus degree: FAS result: treat after 14 days, the disappearance rate 39.6% of test group, improvement rate 44.2%, variability 16.3%; The disappearance rate 33.6% of matched group, improvement rate 43.0%, variability 23.4%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS result: treat after 14 days, the disappearance rate 42.0% of test group, improvement rate 46.3%, variability 11.7%; The disappearance rate 37.5% of matched group, improvement rate 46.9%, variability 15.6%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS is similar with FAS result.2. erythema degree: FAS result: treat after 14 days, the disappearance rate 27.6% of test group, improvement rate 46.0%, variability 26.4%; The disappearance rate 19.6% of matched group, improvement rate 47.7%, variability 32.7%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS result: treat after 14 days, the disappearance rate 29.3% of test group, improvement rate 48.5%, variability 22.1%; The disappearance rate 21.9% of matched group, improvement rate 52.1%, variability 26.0%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS is similar with FAS result.3. pimple degree: FAS result: treat after 14 days, the disappearance rate 40.8% of test group, improvement rate 33.1%, variability 26.1%; The disappearance rate 33.3% of matched group, improvement rate 32.4%, variability 34.3%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS result: treat after 14 days, the disappearance rate 42.9% of test group, improvement rate 34.1%, variability 23.0%; The disappearance rate 37.2% of matched group, improvement rate 36.2%, variability 26.6%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS is similar with FAS result.4. vesicle degree: FAS result: treat after 14 days, the disappearance rate 69.0% of test group, improvement rate 13.2%, variability 17.8%; The disappearance rate 58.2% of matched group, improvement rate 11.9%, variability 29.9%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS result: treat after 14 days, the disappearance rate 72.1% of test group, improvement rate 13.3%, variability 14.5%; The disappearance rate 67.2% of matched group, improvement rate 13.8%, variability 19.0%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS is similar with FAS result.5. ooze out degree: FAS result: treat after 14 days, the disappearance rate 73.3% of test group, improvement rate 10.5%, variability 15.8%; The disappearance rate 61.4% of matched group, improvement rate 11.4%, variability 27.1%.Two groups of grades comprehensively compare difference has statistical significance (P＜0.05).PPS result: treat after 14 days, the disappearance rate 76.2% of test group, improvement rate 10.5%, variability 13.3%; The disappearance rate 66.7% of matched group, improvement rate 12.7%, variability 20.6%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).6. rotten to the corn degree: FAS result: treat after 14 days, the disappearance rate 70.1% of test group, improvement rate 14.9%, variability 14.9%; The disappearance rate 59.7% of matched group, improvement rate 12.9%, variability 27.4%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS result: treat after 14 days, the disappearance rate 73.3% of test group, improvement rate 15.8%, variability 11.0%; The disappearance rate 65.5% of matched group, improvement rate 14.5%, variability 20.0%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS is similar with FAS result.7. degree forms a scab: FAS result: after treating 14 days, and the disappearance rate 41.0% of test group, improvement rate 21.7%, variability 37.3%; The disappearance rate 32.5% of matched group, improvement rate 21.7%, variability 45.8%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).At PPS: after treating 14 days, the disappearance rate 43.2% of test group, improvement rate 23.1%, variability 33.8%; The disappearance rate 37.5% of matched group, improvement rate 25.0%, variability 37.5%.Two groups of grades comprehensively compare difference not statistically significant (P＞0.05).PPS is similar with FAS result.8. target erythra area: FAS result: treat after 14 days, the disappearance rate 19.9% of test group, improvement rate 50.0%, variability 30.1%; The disappearance rate 10.3% of matched group, improvement rate 49.5%, variability 40.2%.Two groups of grades comprehensively compare difference has statistical significance (P＜0.05).PPS result: treat after 14 days, the disappearance rate 21.2% of test group, improvement rate 52.8%, variability 26.1%; The disappearance rate 11.5% of matched group, improvement rate 54.2%, variability 34.4%.Two groups of grades comprehensively compare difference has statistical significance (P＜0.05).PPS is similar with FAS result.

Comparison shows that of pruritus onset time, extinction time, aspect treatment subacute eczema pruritus symptom, medicinal frost formulation of the present invention is compared with the happy ointment of the yellow skin of ice, has the characteristics rapid-action, that transference cure is fast.1. treat 14 days pruritus onset times: FAS result: test group average out to 4.84 ± 2.72 days, matched group average out to 6.11 ± 3.12 days.PPS result: test group average out to 4.84 ± 2.72 days, matched group average out to 6.15 ± 3.12 days.Two groups of difference have statistical significance (P＜0.05), and PPS is similar with FAS result.Survival analysis relatively, test group meta pruritus onset time is 4 days, matched group is 6 days, two groups of difference have statistical significance (P＜0.05).2. treat 14 days pruritus extinction times: FAS result: test group average out to 8.45 ± 3.23 days, matched group average out to 9.95 ± 3.27 days.Two groups of difference have statistical significance (P＜0.05), and PPS is consistent with FAS result.Survival analysis compares, and test group meta pruritus extinction time is 8 days, and matched group is 10 days, and two groups of difference have statistical significance (P＜0.05).

Above result shows; Medicine of the present invention has antiinflammatory, antibacterial, antipruritic and certain multiple effects that help eczema such as immunomodulating; Can be used for treating erythra and pruritus that acute, subacute eczema, chronic eczema acute attack cause, drug effect is obvious, steady quality; Controllability is strong, for clinical a kind of new selection is provided.

Description of drawings: accompanying drawing is a pharmaceutical preparation flow chart of the present invention

Claims

1. method for preparing the externally-applied medicinal composition of treating eczematous dermatitis, it is characterized in that: it comprises the steps:

A, take by weighing following raw materials by weight proportions:

2. the method for preparing of externally-applied medicinal composition according to claim 1, it is characterized in that: the weight proportion of the described crude drug of a step is:

3. the method for preparing of externally-applied medicinal composition according to claim 1, it is characterized in that: described macroporous adsorbent resin is a D101 type macroporous adsorbent resin.

4. the method for preparing of externally-applied medicinal composition according to claim 1, it is characterized in that: the described ethanol elution of c step is 70% ethanol elution.

5. the method for preparing of externally-applied medicinal composition according to claim 1, it is characterized in that: described external preparation is: cream, unguentum, external spraying agent, liniment or paste.

6. the method for preparing of externally-applied medicinal composition according to claim 5, it is characterized in that: described unguentum is rubber-emplastrum, emplastrum, cataplasma.

7. the method for preparing of externally-applied medicinal composition according to claim 5 is characterized in that: the weight percentage that every preparation unit contains baicalin in the described external preparation is not less than 0.2%.

8. the method for preparing of externally-applied medicinal composition according to claim 5, it is characterized in that: the substrate in the described cream is oil-in-water type substrate.