CN101417038A

CN101417038A - Traditional Chinese medicine composition for treating male infertility, preparation method and quality control method thereof

Info

Publication number: CN101417038A
Application number: CNA2007101762267A
Authority: CN
Inventors: 付立家; 付建家
Original assignee: Beijing Asia East Bio Pharmaceutical Co Ltd
Current assignee: Beijing Asia East Bio Pharmaceutical Co Ltd
Priority date: 2007-10-23
Filing date: 2007-10-23
Publication date: 2009-04-29
Anticipated expiration: 2027-10-23
Also published as: CN101417038B

Abstract

The invention relates to a traditional Chinese medicine composition used for treating male sterility, a preparation method and a quality control method thereof. The traditional medicine composition contains crude drugs such as fruits of Chinese wolfberry, dodder (fried), raspberry, fructus schizandrae (steamed), plantago seeds (fried with salt) and the like. The preparation method comprises an ultrafine grinding technology and the like. The quality control method of the medicine composition comprises a thin-layer identification method for fruits of Chinese wolfberry, dodder, raspberry, fructus schizandrae and plantago seeds as well as a method for measuring the content of schizandrin B in the plantago seeds. The medicine composition has obvious therapeutic effect in treating renal deficiency and lumbago, post-micturition dribble, spermatorrhea, premature ejaculation, impotence and sterility.

Description

Chinese medicine composition of treatment male infertility and preparation method thereof and method of quality control

Technical field

The present invention relates to a kind of Chinese medicine composition and preparation method thereof and method of quality control, relate in particular to a kind of Chinese medicine composition for the treatment of male infertility and preparation method thereof and method of quality control, belong to technical field of Chinese medicines.

Background technology

Male sterility is not single specific diseases, but by the male genital organ that Different types of etiopathogenises causes, external genitalia and gonad axis varying level is unusual in comprising, finally shows as the syndrome that fertility descends or loses.Just because of this, can not cure male infertility due to the various different causes of disease with a kind of specific medicine or method for male infertility.Treatment, immunization therapy, surgical intervention, the artificial insemination of endocrine therapy, urogenital tract infection are arranged at the clinical Therapeutic Method that certain curative effect arranged at present, still have many agnogenio, the male infertility that acatalepsia is true, the Therapeutic Method of this kind situation is concluded to be had: medicines such as hormone, antibiotic, thyroxine, vitamin, trace element, aminoacid, be aided with simultaneously and advise that patient avoids tobacco and wine, prevent psychentonia, the overtired and living habit etc. that keeps normal nutrition and rule, but many curative effects are not certainly.

The traditional Chinese medical science early has understanding to male sterility, and tcm clinical efficacy is remarkable than doctor trained in Western medicine, and side effect seldom.The traditional Chinese medical science is thought kidney controlling essence storage, for reproduction this, so treat, accumulated many experiences based on the kidney invigorating.The leaf sky scholar of the Qing Dynasty is very deep to the understanding of male sterility, " disease about the pregnant person of tire, the man is then in essence, the woman is then at blood, and is not enough nothing but and right.All men's deficiency then has smart cunning, seminal plasma, cold sperm, or it is not hard to face thing, or stream and not penetrating, or the nocturnal emission frequency, or turbid urine pouring drop.Or be fond of women, so that the deficiency of YIN, then the waist nephralgia is exhausted for the deficiency of YIN.Or good male wind, so that anode, the then high and exhaustion of YIN of anode.Or too strong, strong then victory or defeat is not in harmony.Or the plain scrotal hernia of suffering from, scrotal hernia then Liver and kidney well-behaved from.Decline the sun then much cold that declines in addition or with sun.Or with the deficiency of YIN, the deficiency of YIN is frequent fever then, is diseases of man all, and the married woman that must not use up Venezuelan also.If its source and cure it, then thing does not have not good." he is rare excessively from the sexual dysfunction to the seminal fluid, unable to genital disease from ejaculating, and excessively causes the hernia of suffering from a deficiency of the kidney from sexual life.For male infertility, as do not have special disease, the traditional Chinese medical science is emphasized the kidney invigorating.Because of kidney controlling essence storage, deficiency of the kidney is then smart closes admittedly, often shows as seminal emission, spermatorrhea or premature ejaculation.Deficiency of the kidney must cause deficiency of both QI and blood, so the tcm treatment method of male infertility is nothing more than the kidney invigorating, two replenishing QI and blood.The speciality of performance Chinese medicine provides a kind of Chinese medicine composition of effective treatment male infertility being very important at present.

Summary of the invention

The object of the present invention is to provide a kind of Chinese medicine composition that is used for the treatment of male infertility;

The present invention also aims to provide a kind of preparation method of this Chinese medicine composition;

The present invention also aims to provide the method for quality control of this Chinese medicine composition.

The present invention seeks to be achieved through the following technical solutions:

The Chinese medicine composition of treatment male infertility of the present invention is to be made by the crude drug of following weight ratio:

Fructus Lycii 15～75 weight portion Semen Astragali Complanatis 20～60 weight portion Fructus Rubies 10～40 weight portions

Fructus Schisandrae Chinensis 1～8 weight portion Semen Plantaginis (salt stir-fry) 4～15 weight portions

Traditional Chinese medicinal composition raw materials optimum ratio of the present invention can for:

Fructus Lycii 56 weight portion Semen Astragali Complanatis 25 weight portion Fructus Rubies 20 weight portions

Fructus Schisandrae Chinensis 5 weight portion Semen Plantaginiss (salt stir-fry) 7 weight portions

The Chinese medicine composition of treatment male infertility of the present invention can be made by the crude drug of following weight ratio:

Fructus Lycii 15～75 weight portion Semen Cuscutae 20～60 weight portion Fructus Rubies 10～40 weight portions

Chinese medicine composition raw material optimum ratio of the present invention is:

Fructus Lycii 40～65 weight portion Semen Cuscutae (stir-fry) 20～30 weight portion Fructus Rubies 15～30 weight portions

Fructus Schisandrae Chinensis (steaming) 4～7 weight portion Semen Plantaginiss (salt stir-fry) 5～9 weight portions

Traditional Chinese medicinal composition raw materials optimum ratio of the present invention can also for:

Fructus Lycii 56 weight portion Semen Cuscutae (stir-fry) 25 weight portion Fructus Rubies 20 weight portions

Fructus Schisandrae Chinensis (steaming) 5 weight portion Semen Plantaginiss (salt stir-fry) 7 weight portions

Male infertility is many by deficiency of kidney-QI, due to the deficiency of YIN-essence.Fructus Lycii, Semen Astragali Complanati the kidney invigorating essence in the pharmaceutical composition prescription of the present invention, the tonifying YANG road helps spirit; Fructus Rubi is supported Kidney-Yin, and controlling nocturnal emission with astringent drugs closes, and plays sexual impotence; Fructus Schisandrae Chinensis the kidney invigorating water, lung benefiting gas ends spermatorrhea; Semen Plantaginis diuresis, " Mingyi Bielu " claim that it can " nourishing the liver reinforcing YIN-essence benefit smart ", match with said medicine, and the invigorating middle warmer residence rushes down, and mend and oiliness.The merit that full side plays the kidney invigorating and essence nourishing altogether is applicable to lumbago due to renal deficiency, dribble of urine, and the seminal emission premature ejaculation, sexual impotence is sterile.

Compositions of the present invention technology adding adjuvant is routinely made clinical acceptable forms such as tablet, capsule, drop pill; Described adjuvant comprises solvent, disintegrating agent, correctives, antiseptic, coloring agent, binding agent, lubricant, substrate etc.

The preparation method of Chinese medicine composition pill of the present invention is:

The above five tastes adopt superfine communication technique, are ground into micropowder, mixing; 90-110 weight portion Mel is in the time of 116-118 ℃, and refining density is 1.37, promptly gets refined honey; Per 100 weight portion powder add the water of refined honey 35-50 weight portion and 100-160 weight portion, general ball, and 60-80 ℃ of drying 8～12 hours, promptly.

Pharmaceutical composition method of quality control of the present invention comprises one or more in following discrimination method and/or the assay:

Differentiate:

(1) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3～4g, porphyrize, the 40～60ml that adds diethyl ether, supersound process 15～30 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets Fructus Lycii control medicinal material 1～2g, grinds, and the 20～30ml that adds diethyl ether shines medical material solution in pairs with legal system.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15～25:15～25:0.1～0.5) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(2) get pharmaceutical preparation of the present invention and be equivalent to crude drug 7～8g, porphyrize adds petroleum ether (30～60 ℃) 25～35ml, supersound process 20～40 minutes, filter, discard petroleum ether liquid, medicinal residues are waved most petroleum ether, add methanol 30～40ml supersound process 15～30 minutes, filter, filtrate evaporate to dryness, residue add ethanol 5～6ml to be made it to disperse, and gets supernatant as need testing solution.Other gets Semen Cuscutae control medicinal material 2g, and porphyrize shines medical material solution in pairs with the method preparation.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with toluene-ethyl acetate-formic acid (8～12:4～6:3～5) is developing solvent, launches, and takes out, dry, spray 1% aluminum chloride alcoholic solution, airing is put under the ultra-violet lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3～4g, porphyrize, the 40～60ml that adds diethyl ether, supersound process 15～30 minutes, filter, filtrate volatilizes, and residue adds chloroform 30～50ml, supersound process 15～25 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets the deoxyschizandrin reference substance, adds ethyl acetate and makes the solution that every 1ml contains 1mg, in contrast product solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, reference substance solution 2 μ l put in same silica gel G F respectively ₂₅₄On the lamellae, be developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect with the upper solution of petroleum ether (30～60 ℃)-Ethyl formate-formic acid (10～20:3～7:1～2).In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(4) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3～4g, porphyrize adds 40～60mL petroleum ether, and Soxhlet is extracted 2～3h defat, and residue volatilizes petroleum ether, adds 40～60mL methanol eddy and extracts 2～4h, filters, and filtrate is concentrated into 1～2mL, as need testing solution; Other gets the Fructus Rubi control medicinal material, shines medical material solution in pairs with legal system; According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ L of above-mentioned two kinds of solution, put respectively on same silica gel G plate with ethyl acetate-methanol-water (4～7: 1～2: 1～2) be developing solvent, launch, take out, dry, spray 0.5%AlCl ₃Methanol solution is put under the uviol lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.

(5) get pharmaceutical preparation of the present invention and be equivalent to crude drug 1～2g, porphyrize adds methanol 15～30ml, soaks 2～4h, filters, and filtrate evaporate to dryness, residue add methanol 1～2ml makes dissolving, as need testing solution.Get Semen Plantaginis control medicinal material 1～2g, methanol 10～30ml, heating and refluxing extraction 0.5～1.5h filters, and filtrate volatilizes, and residue adds 1～2ml dissolve with methanol, in contrast medical material solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution, each 20 μ l of control medicinal material solution, put respectively on same silica gel g thin-layer plate, with chloroform-methanol-ammonia (10～20: 1～2: 0.5～1) be developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 100 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the speckle of same color.

Assay:

Measure according to high performance liquid chromatography (Chinese Pharmacopoeia version appendix in 2005 VID)

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; With acetonitrile-water-acetic acid (65:40:0.1) is mobile phase; The detection wavelength is 254nm.

It is an amount of that the schisandrin B reference substance is got in the preparation of reference substance solution, and accurate the title decides, and adds methanol and makes the solution that every 1ml contains 30 μ g, promptly.

The preparation of need testing solution is got pharmaceutical preparation of the present invention and is equivalent to crude drug 3～4g, porphyrize, and precision takes by weighing about 4～6g, put in the apparatus,Soxhlet's, it is an amount of to add normal hexane, and soaked overnight was in 80～85 ℃ of reflux, extract, 5～8 hours, extracting solution low temperature evaporate to dryness, residue adds the methanol slight fever makes dissolving, moves in the 25ml measuring bottle, adds methanol to scale, shake up, promptly.

Accurate respectively reference substance solution and each 5～15 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.Pharmaceutical preparation of the present invention is equivalent to crude drug 0.5～1g and contains Fructus Schisandrae Chinensis with schisandrin B (C ₂₃H ₂₈O ₆) meter, must not be less than 0.05～0.08mg.

Pharmaceutical composition method of quality control of the present invention is preferably as follows one or more in discrimination method and/or the assay:

Differentiate:

(1) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize, the 50ml that adds diethyl ether, supersound process 20 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets Fructus Lycii control medicinal material 1g, grinds, and the 20ml that adds diethyl ether shines medical material solution in pairs with legal system.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-Ethyl formate-formic acid (20:20:0.1) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(2) get this product 10g, porphyrize adds petroleum ether (30～60 ℃) 30ml, supersound process 30 minutes, filter, discard petroleum ether liquid, medicinal residues are waved most petroleum ether, add methanol 30ml supersound process 20 minutes, filter, filtrate evaporate to dryness, residue add ethanol 5ml to be made it to disperse, and gets supernatant as need testing solution.Other gets Semen Cuscutae control medicinal material 2g, and porphyrize shines medical material solution in pairs with the method preparation.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with toluene-ethyl acetate-formic acid (10:5:4) is developing solvent, launches, and takes out, dry, spray 1% aluminum chloride alcoholic solution, airing is put under the ultra-violet lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize, the 50ml that adds diethyl ether, supersound process 20 minutes, filter, filtrate volatilizes, and residue adds chloroform 40ml, supersound process 20 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets the deoxyschizandrin reference substance, adds ethyl acetate and makes the solution that every 1ml contains 1mg, in contrast product solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, reference substance solution 2 μ l put in same silica gel G F respectively ₂₅₄On the lamellae, be developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect with the upper solution of petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15:5:1).In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

(4) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize adds the 50mL petroleum ether, and Soxhlet is extracted the 2.5h defat, and residue volatilizes petroleum ether, adds the 50mL methanol eddy and extracts 3h, filters, and filtrate is concentrated into 2mL, as need testing solution; Other gets the Fructus Rubi control medicinal material, shines medical material solution in pairs with legal system; According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ L of above-mentioned two kinds of solution, put respectively on same silica gel G plate with ethyl acetate-methanol-water (5: 1: 1), launch, take out, dry spray 0.5%AlCl ₃Methanol solution is put under the uviol lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.

(5) get pharmaceutical preparation of the present invention and be equivalent to crude drug 1.8g, porphyrize adds methanol 20ml, soaks 3h, filters, and filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, as need testing solution.Get Semen Plantaginis control medicinal material 1g, methanol 20ml, heating and refluxing extraction 1h filters, and filtrate volatilizes, and residue adds the 1ml dissolve with methanol, in contrast medical material solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, control medicinal material solution 20 μ l, put respectively on same silica gel g thin-layer plate, with chloroform-methanol-ammonia (15: 1.5: 0.5) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 100 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the speckle of same color.

Assay:

The preparation of need testing solution is got pharmaceutical preparation of the present invention and is equivalent to crude drug 4g, porphyrize, and precision takes by weighing about 5g, put in the apparatus,Soxhlet's, it is an amount of to add normal hexane, and soaked overnight was in 80～85 ℃ of reflux, extract, 6 hours, extracting solution low temperature evaporate to dryness, residue adds the methanol slight fever makes dissolving, moves in the 25ml measuring bottle, adds methanol to scale, shake up, promptly.

Accurate respectively reference substance solution and each 5～15 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.Pharmaceutical preparation of the present invention is equivalent to crude drug 0.719g and contains Fructus Schisandrae Chinensis with schisandrin B (C ₂₃H ₂₈O ₆) meter, must not be less than 0.07mg.

The method of quality control of Chinese medicine composition provided by the present invention, be by obtaining behind the creative experiment sieving of big measuring, pass through screening in the discrimination method to sample treatment, the selection of developing solvent, make and differentiate that specificity is fine, and method is economic and practical, the result is quick, and can both use different lamellaes.Carried out the methodological study of assay in the assay, show that this method is stable, accurate, can effectivelyly carry out quality control, and will compare more stable that product that additive method measures shows on pharmacological effect with the product that this method is measured to product.

Following experimental example and embodiment are used to further specify but are not limited to the present invention.

The research of test example 1 preparation method

1, smashing fineness test

Pharmaceutical composition of the present invention is analyzed the effective ingredient of each flavor medical material when research design preparation technology, in order to keep the effective ingredient in the medicine preferably, the direct pulverizing of medical material is used as medicine, and makes pill.Get totally 6 parts of Fructus Lycii 56g, Semen Astragali Complanati 25g, Fructus Rubi 20g, Fructus Schisandrae Chinensis 5g, Semen Plantaginis (salt stir-fry) 7g, be divided into two groups, adopt different preparation methoies to make pill respectively, measure the content of the schisandrin B in the preparation, the results are shown in following table 1:

The selection of table 1 breaking method

Preparation method	Schisandrin B average content (mg/g)
Preparation method	Schisandrin B average content (mg/g)	The above five tastes adopt superfine communication technique, are ground into micropowder, mixing.Per 100 weight portion powder add an amount of water pill with refined honey 35～50 weight portions, and drying is made water-honeyed pill, promptly.	0.18
The above five tastes are ground into fine powder, sieve mixing.Every 100g powder adds an amount of water pill with refined honey 35～50g, and drying is made water-honeyed pill, promptly.	0.12		0.18

As can be seen from the above table, its active ingredient amount of recording is significantly higher than general pulverizing after the medicine micronizing, illustrates that micronizing can significantly improve the dissolution of active ingredient in the medicine, and this also is the main cause that curative effect of medication is significantly improved.

2. amount of water test

Press five parts of medical materials of recipe quantity configuration, carry out micronizing by technological requirement, divide four groups to do experiment respectively: water honey ratio is respectively: 2:1, and 2.5:1,3:1,3.5:1 is that index is determined amount of water with pill viscosity, pill outward appearance, drying time respectively.The results are shown in Table 2,

Table 2: amount of water result of the test

Above result shows: water honey ratio is 3:1, and when drying time was 11h, every index was better, is 3:1 so select water honey ratio in producing for use.

Test example 2 quality standard researches

1, the Fructus Lycii thin layer is differentiated

(1) investigation of need testing solution supersound extraction time

Get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize, the 50ml that adds diethyl ether, supersound process filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets Fructus Lycii control medicinal material 1g, grinds, and the 20ml that adds diethyl ether shines medical material solution in pairs with legal system.Drawing each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, is developing solvent with petroleum ether (30～60 ℃)-Ethyl formate-formic acid (20:20:0.1), launches, and takes out, and dries, and puts under the ultra-violet lamp (365nm) and inspects.Compare test sample and the color developing effect of control medicinal material on lamellae, the results are shown in following table:

The investigation of table 3 need testing solution supersound extraction time

Extraction time

5min

10min

20min

40min

Color developing effect

The colour developing speckle does not all appear in test sample and control medicinal material on lamellae.

The test sample speckle is very shallow, and the control medicinal material speckle is unclear.

Test sample and control medicinal material are clear in the colour developing of relevant position speckle.

As can be seen from the above table, more than the supersound extraction 20min, gained need testing solution and control medicinal material solution, after the expansion, on lamellae relevant position speckle colour developing clear, so Pass Test requirement is supersound extraction 20min.

(2) proportioning of developing solvent

According to the thin layer chromatography test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developing solvent with petroleum ether (30～60 ℃)-Ethyl formate-formic acid, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.Under the more different proportioning developing solvents, the unfolded effect of each speckle in test sample and the control medicinal material chromatograph the results are shown in following table:

The proportioning test result of table 4 developing solvent

The developing solvent proportioning	10:30:0.1	20:30:0.1	30:10:0.1	20:20:0.1
The developing solvent proportioning	10:30:0.1	20:30:0.1	30:10:0.1	20:20:0.1	Launch effect	Each speckle of test sample and control medicinal material separates unclear, launches poor effect.	Each speckle of test sample and control medicinal material separates unclear, launches weak effect.	Each speckle of test sample and control medicinal material has the tail of taking off phenomenon, launches weak effect.	Each speckle of test sample and control medicinal material separates clear, launches effective.

As can be seen from the above table, when the developing solvent proportioning was 20:20:0.1, the speckle that respectively develops the color in test sample and the control medicinal material chromatograph separated clear, launch effective, the Pass Test requirement.

2, the thin layer of Semen Cuscutae is differentiated

The preparation of need testing solution: get pharmaceutical preparation of the present invention and be equivalent to crude drug 7.2g, porphyrize adds petroleum ether (30～60 ℃) 30ml, supersound process 30 minutes, filter, discard petroleum ether liquid, medicinal residues are waved most petroleum ether, add methanol 30ml supersound process 20 minutes, filter, filtrate evaporate to dryness, residue add ethanol 5ml to be made it to disperse, and gets supernatant as need testing solution.

The preparation of control medicinal material solution: get Semen Cuscutae control medicinal material 2g, porphyrize shines medical material solution in pairs with the method preparation.

The selection test of developing solvent:

According to the thin layer chromatography test, draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, be developing solvent with toluene-ethyl acetate-formic acid (10:5:4), launch, take out, dry, spray 1% aluminum chloride alcoholic solution, airing is put under the ultra-violet lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.More different developing solvents, the expansion effect of each speckle in test sample and the control medicinal material chromatograph the results are shown in following table:

The selection result of the test of table 5 developing solvent

Developing solvent	Toluene-ethyl acetate-formic acid (5:5:3)	Toluene-ethyl acetate-formic acid (10:5:4)	Chloroform-ethyl acetate-formic acid (10:5:4)	Toluene-n-butyl alcohol-formic acid (10:5:4)
Developing solvent	Toluene-ethyl acetate-formic acid (5:5:3)	Toluene-ethyl acetate-formic acid (10:5:4)	Chloroform-ethyl acetate-formic acid (10:5:4)	Toluene-n-butyl alcohol-formic acid (10:5:4)	Launch effect	Test sample and reference substance	Test sample and reference substance	Test sample and reference substance	Test sample and reference substance

The chromatograph relevant position shows the speckle of same color, but each speckle separation is unclear, negative noiseless.

The corresponding position of chromatograph shows the speckle of same color, launches effective, negative noiseless.

The corresponding position of chromatograph shows the speckle of same color, but separating effect is very poor, and feminine gender has interference.

The corresponding position of chromatograph shows the speckle of same color, and the tail of taking off phenomenon is arranged, and feminine gender has interference.

As can be seen from the above table, be developing solvent with toluene-ethyl acetate-formic acid (10:5:4), launch effectively, and speckle colour developing is clear, and is negative noiseless, favorable reproducibility.

3, the thin layer of Fructus Schisandrae Chinensis is differentiated

The preparation of need testing solution:

Method one: get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize adds chloroform 50ml, and reflux 0.5 hour filters, and filtrate evaporate to dryness, residue add chloroform 1ml makes dissolving, as need testing solution.

Method two: get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize is put in the flask, adds normal hexane 50ml, and merceration spends the night, and in 80～85 ℃ of reflux, extract, 2 hours, filters, and filtrate low temperature evaporate to dryness, residue add ethyl acetate 2ml makes dissolving, as need testing solution.

Method three: get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize adds kieselguhr 5g, grind well the 50ml that adds diethyl ether, supersound process 20 minutes, filter, filtrate volatilizes, and residue adds chloroform 40ml, supersound process 20 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.

The preparation of reference substance solution: get the deoxyschizandrin reference substance, add ethyl acetate and make the solution that every 1ml contains 1mg, promptly.According to thin layer chromatography (appendix VI B) test, draw need testing solution 10 μ l, reference substance solution 2 μ l put in same silica gel G F respectively ₂₅₄On the lamellae, be developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect with the upper solution of petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15:5:1).Compare test sample chromatograph and corresponding reference substance chromatograph position, the unfolded effect of speckle

The preparation method result of the test of table 6 need testing solution

The need testing solution preparation method	Method one	Method two	Method three
The need testing solution preparation method	Method one	Method two	Method three	Launch effect	Need testing solution with the reference substance solution relevant position on identical colour developing speckle is arranged, but feminine gender has interference.	Need testing solution with the reference substance solution relevant position on identical colour developing speckle is arranged, but feminine gender has interference.	Need testing solution with the reference substance solution relevant position on identical colour developing speckle is arranged, negative noiseless.

As can be seen from the above table, the need testing solution of application process three preparations launches effective, and is negative noiseless.So system of selection three preparation need testing solutions.

4, the thin layer of Fructus Rubi is differentiated

The preparation of need testing solution: get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, add the 50mL petroleum ether, Soxhlet is extracted the 2.5h defat, and residue volatilizes petroleum ether, adds the 50mL methanol eddy and extracts 3h, filters, and filtrate is concentrated into 2mL, as need testing solution;

The preparation of control medicinal material solution: get the Fructus Rubi control medicinal material, shine medical material solution in pairs with legal system;

According to thin layer chromatography test, draw above-mentioned two kinds of solution, put respectively on same silica gel G plate with ethyl acetate-methanol-water, launch, take out, dry spray 0.5%AlCl ₃Methanol solution is put under the uviol lamp (365nm) and is inspected.Under more different point sample amounts, the different developing solvent proportioning, the expansion effect of need testing solution and reference substance solution the results are shown in following table:

The comparative test of table 7 developing solvent proportioning and test sample point sample amount

As can be seen from the above table, the proportioning of developing solvent ethyl acetate-methanol-water is 5: 1: 1, when the point sample amount is 5 μ l, test sample and control medicinal material chromatograph relevant position show same color fluorescence speckle, and each speckle good separating effect, and it is clear to develop the color, negative noiseless, the Pass Test requirement.

5, the thin layer of Semen Plantaginis is differentiated

The preparation of need testing solution: get pharmaceutical preparation of the present invention and be equivalent to crude drug 1.8g and add methanol 20ml, soak 3h, filter, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, as need testing solution.

The preparation of control medicinal material solution: get Semen Plantaginis control medicinal material 1g, methanol 20ml, heating and refluxing extraction 1h filters, and filtrate volatilizes, and residue adds the 1ml dissolve with methanol, in contrast medical material solution.

Drawing need testing solution, each 20 μ l of control medicinal material solution, put respectively on same silica gel g thin-layer plate, is developing solvent with chloroform-methanol-ammonia, launches, and takes out, and dries, and spray is with 10% ethanol solution of sulfuric acid, and 100 ℃ to be heated to the speckle colour developing clear.More different proportioning developing solvents, test sample and the control medicinal material expansion effect on thin layer chromatography the results are shown in following table:

The selection test of table 8 developing solvent proportioning

The developing solvent proportioning	15: 4: 0.5	15: 3: 0.5	15: 2: 0.5	15: 1.5: 0.5
The developing solvent proportioning	15: 4: 0.5	15: 3: 0.5	15: 2: 0.5	15: 1.5: 0.5	Launch effect	In test sample and the control medicinal material chromatograph, each speckle separates very unclear, disturbs big.	In test sample and the control medicinal material chromatograph, each speckle separates very unclear, disturbs big.	In test sample and the control medicinal material chromatograph, each speckle separates unclear, and interference is arranged.	In test sample and the control medicinal material chromatograph, each speckle good separating effect, it is clear to develop the color.

As can be seen from the above table, be developing solvent with 15: 1.5: 0.5 ratio of chloroform-methanol-ammonia, it is best to launch effect, and Pass Test requirement, and favorable reproducibility are negative noiseless.

6, the assay of schisandrin B

Detecting instrument: Tianjin, the island SPD-10ATvp of company type high performance liquid chromatograph

Chromatographic column: Di Ma company (Zorbax C18 4.6 * 150mm, 5 μ m)

Mobile phase: acetonitrile-water-acetic acid (65:40:0.1)

Detect wavelength: 254nm flow velocity: 1.000ml/min column temperature: room temperature

Reference substance: schisandrin B is purchased lot number: the 110765-200205 in Nat'l Pharmaceutical ﹠ Biological Products Control Institute

Assay method:

The preparation of need testing solution is got pill by embodiment 2 methods preparations and is equivalent to crude drug 4.5g and pulverizes, and precision takes by weighing about 5g, and the accurate 50ml methanol that adds takes by weighing weight, and supersound process 30 minutes is put coldly, supplies weight, shake up, and filtration, promptly.

The blank sample that the preparation of embodiment 2 methods lacks Fructus Schisandrae Chinensis is pressed in the preparation of negative controls, the preparation negative controls.

Filter with microporous filter membrane (0.45 μ m).The accurate respectively negative controls of drawing, each 5～10 μ l of reference substance solution and need testing solution inject chromatograph of liquid, measure, promptly.

1. content assaying method is investigated:

(1) stability test is got reference substance solution (34 μ g/ml), respectively at preparing the back 0,2,4,6,12,24 hour, measures in accordance with the law, and the result shows that it is basicly stable in 24 hours, the results are shown in following table:

Table 9 stability test result

(2) linear relationship is investigated and to be got reference substance solution (35.52 μ g/ml) and shake up, accurate respectively 1,5,10,15, the 19 μ l of absorption inject high performance liquid chromatograph, the results are shown in following table, and drawing standard curve, show that schisandrin B is linear between 0.03552 μ g-0.67488 μ g, its regression equation is: y=2E+06x+4938.3 (r=0.9998)

Table 10 linear relationship is investigated the result

(3) precision is tested accurate reference substance solution, repeats sample introduction 5 times, tries to achieve relative standard deviation＜2.5%, the results are shown in following table:

Table 11 Precision test result

(4) the text method is pressed in repeatability test, gets with 5 parts of a collection of pharmaceutical preparatioies of the present invention, and every part is measured, and tries to achieve relative standard deviation＜2%, the results are shown in following table:

Table 12 reproducible test results

(5) the recovery test precision takes by weighing accurate again schisandrin B reference substance (the 35.52 μ g/ml) 8ml that adds of pharmaceutical preparation 2.5g same a collection of of the present invention of known content, press the preparation method operation of text need testing solution, measure its content, and calculate its response rate, measurement result sees the following form:

Table 13 recovery test result

2. measurement result:

Three crowdes of assay results of table 14

Lot number	Content (mg/ gram)
Lot number	Content (mg/ gram)	11201	0.115
11302	0.116	11201	0.115
11302	0.116	11403	0.115

According to above experimental result, medicine assay method of the present invention is stable, meets the requirements.

Test example 3 pharmacodynamic studies

1 material

Medicine and reagent:

Medicine I of the present invention: according to the pharmaceutical preparation of embodiment 8 preparations

Medicine II of the present invention: according to the pharmaceutical preparation of embodiment 2 preparations

Medicine III of the present invention: according to the pharmaceutical preparation of embodiment 1 preparation

Positive control medicine: SHENGJING JIAONANG commercially available product

Pentobarbital sodium: with the normal saline preparation, for lumbar injection usefulness, Shanghai chemical reagent purchasing and supply station packing.

Scotcil: with the water for injection preparation, for lumbar injection usefulness, North China Pharmaceutical Factory produces.

The hydrocortisone sodium succinate injection: crude drug pharmaceutical factory in Tianjin produces.

Testosterone propionate test kit, Tianjin get the outlet of general DDC company.

Animal: the SD rat (80～100g), kunming mice (20～22g), Cavia porcellus (750～850g), be male, provide by Henan Province's Experimental Animal Center.

Instrument: threshold of pain analyzer (being used for electricity irritation), Haidian, Beijing electron medical treatment factory produces.γ radioimmunity enumerator, Xi'an automatization institute is produced.

2 methods and result

2.1 influence to the castrated rats syndrome of deficiency of kidney-YANG

Method: get 48 male rats, through intraperitoneal anesthesia, behind the sterilization local skin, select 8 animals to make blank (sham-operation) group at random with pentobarbital sodium 40mg/kg, surplus animal is all excised bilateral testes and intramuscular injection scotcil 20,000 Ukg ^-1Neuter was divided into 5 groups at random in the 11st day, 8 every group, and the beginning medication, continuous 12 days.Be blank group (sham operated rats), model control group: give 015%CMC2Na10mlkg ^-1Ig; Positive controls (SHENGJING JIAONANG group): 4.0g crude drug kg ^-1Medicine I of the present invention, II, III group: be respectively 2.8g crude drug kg ^-1Current intensity was 4mA, observed erection incubation period and persistent period with Wa29E analyzer stimulation animal penis part in 1 hour after the last medication.Put to death animal next day, get preputial glands, seminal fluid capsule-prostate and levator ani and weigh, and calculate acropetal coefficient (mg/100g).Result: see Table 15.

Table 15 five-seed Yanzong concentrated pill to the influence of castrated rats syndrome of deficiency of kidney-YANG (x ± s, n=8)

Annotate: compare △ △ P＜0.01 with the blank group; Compare * P＜0.05 with model control group, * * P＜0.01; Compare ★ P＜0.05 with positive controls, ★ ★ P＜0.01.

The erection of positive controls and medicine group of the present invention is significantly shorter than model group incubation period, the erection persistent period is higher than model group (P＜0.05 or 0.01), accessory sex organ's coefficient of positive controls and medicine group of the present invention (preputial glands, seminal fluid capsule-prostate and levator ani) is all apparently higher than model group (P＜0.05 or 0.01), present certain dose-effect relationship, the weight of medicine group levator ani of the present invention is apparently higher than positive control medicine group.Medicine III wherein of the present invention is the most remarkable to the influence of castrated rats syndrome of deficiency of kidney-YANG, and effect is best.

2.2 to the influence of fertility of insufficiency of kidney-YANG mice

Method: get 60 male mices, be divided into 6 groups at random, 10 every group (grouping and dosage see Table 2).Except that the blank group, all the other animals are used hydrocortisone 0.5mlkg ^-1Continuous 9 days of ip; Begin to irritate stomach simultaneously in modeling, continuous 12 days.1h takes a blood sample through eye socket after the last administration, presses the explanation of ria-determination test kit, surveys blood plasma testosterone content; And after each Mus testis, epididymis weighed, epididymis is put into the plate that fills ringer solution, sperm suspension is made in grinding, 37 ℃ of water-bath 15min, press the contained sperm count of the every g epididymis of erythrometry observed and recorded, and above-mentioned suspension dripped on sheet glass, optical microscope is the percentage rate of motile sperm in 200 sperms of record down.Result: see Table 16.

Table 16 five-seed Yanzong concentrated pill is to the yang deficiency mice influence of fertility (x ± s)

Annotate: compare △ P＜0.05 with the blank group, △ △ P＜0.01; Compare * P＜0.05 with model control group, * * P＜0.01; Compare ★ P＜0.05 with positive controls, ★ ★ P＜0.01.

The blood plasma testosterone content of positive drug matched group and medicine group of the present invention is apparently higher than model group (P＜0.01); The testis of each medication group, epididymis weight and model group do not have obvious difference (P〉0.05); The sperm number of positive drug matched group and medicine group of the present invention is higher than model group (P＜0.01 or 0.05); The motility of sperm of medicine group of the present invention is apparently higher than model group (P＜0.01).Medicine of the present invention all is significantly higher than the positive control medicine to the raising of every indexs such as yang deficiency mice plasma testosterone concentration, sperm count, motility of sperm, and the effect of medicine III especially of the present invention is best.

The influence of 3 pairs of male mice mating abilities

Get 60 of male Kunming kind white mice, be divided into 6 groups at random, 10 every group.Normal control group distilled water is irritated stomach (20ml/kg), the plain group of propanoic acid highland ball subcutaneous injection administration (＜0.025mg/ is only). SHENGJING JIAONANG group gastric infusion 4.0g/kg.Medicine I of the present invention, II, III gastric infusion 2.8g/kg.More than each group, irritate stomach every day 1 time (propanoic acid is started the plain group of ball and is the subcutaneous injection administration), 10d continuously, 5d begins every male Mus and 5 female Mus are fed with cage, 6d injects estradiol benzoate (200 μ g/kg) down for every female Corium Mus.Female Mus vaginal orifice is respectively organized in the every morning inspection or intravaginal has or not white thing to occur with beginning next day behind the cage, and adularescent thing person takes out it for cloudy bolt occurring from cage; Calculate cloudy bolt occurrence rate.Knot sees the following form 17:

Table 17 shakes the shadow noon of male exhibition gesture pellet to the male mice mating ability

Group	Dosage (g/kg)	Number of animals (n)	The Mus number (n) that cloudy bolt occurs	Cloudy bolt occurrence rate (%)
Group	Dosage (g/kg)	Number of animals (n)	The Mus number (n) that cloudy bolt occurs	Cloudy bolt occurrence rate (%)	The normal control group	Distilled water 20ml	50	10	20
The plain group of propanoic acid highland ball	--	50	31	62 ^**	The normal control group	Distilled water 20ml	50	10	20
The plain group of propanoic acid highland ball	--	50	31	62 ^**	The SHENGJING JIAONANG group	4.0	50	25	50 ^**
Medicine I group of the present invention	2.8	50	32	64 ^**	The SHENGJING JIAONANG group	4.0	50	25	50 ^**
Medicine I group of the present invention	2.8	50	32	64 ^**	Medicine II group of the present invention	2.8	50	35	70 ^**★
Medicine III group of the present invention	2.8	50	39	78 ^**★★	Medicine II group of the present invention	2.8	50	35	70 ^**★

Annotate: compare * P＜0.05 with the normal control group, * * P＜0.01; Compare ★ P＜0.05 with the SHENGJING JIAONANG group, ★ ★ P＜0.01.

As seen from the above table, medicine group of the present invention and positive drug matched group (plain group of propanoic acid highland ball and SHENGJING JIAONANG group) strengthen mice mating ability (P＜0.01) more significantly with the normal control group, the effect of medicine of the present invention is significantly higher than SHENGJING JIAONANG, and the effect of medicine III wherein of the present invention is best.

Following embodiment all can realize the described effect of above-mentioned experimental example

Embodiment 1

Fructus Lycii 56g Semen Astragali Complanati 25g Fructus Rubi 20g

Fructus Schisandrae Chinensis 5g Semen Plantaginis (salt stir-fry) 7g

Take by weighing above raw medicinal material, the above five tastes adopt superfine communication technique, are ground into micropowder, mixing; 90-110g Mel is in the time of 116-118 ℃, and refining density is 1.37, promptly gets refined honey; Every 100g powder adds the water of refined honey 40g and 110g, general ball, and 60 ℃ of dryings 11 hours are made 100g, promptly.

(1) get this product 5g, porphyrize, the 50ml that adds diethyl ether, supersound process 20 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets Fructus Lycii control medicinal material 1g, grinds, and the 20ml that adds diethyl ether shines medical material solution in pairs with legal system.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-Ethyl formate-formic acid (20:20:0.1) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(2) get this product 5g, porphyrize adds kieselguhr 5g, grinds well, and 50ml adds diethyl ether, supersound process 20 minutes filters, and filtrate volatilizes, and residue adds chloroform 40ml, supersound process 20 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets the deoxyschizandrin reference substance, adds ethyl acetate and makes the solution that every 1ml contains 1mg, in contrast product solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, reference substance solution 2 μ l put in same silica gel G F respectively ₂₅₄On the lamellae, be developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect with the upper solution of petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15:5:1).In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

Assay:

The preparation of need testing solution is got this product and is pulverized, and precision takes by weighing about 5g, puts in the apparatus,Soxhlet's, and it is an amount of to add normal hexane, soaked overnight, in 80～85 ℃ of reflux, extract, 6 hours, extracting solution low temperature evaporate to dryness, residue add the methanol slight fever made dissolving, move in the 25ml measuring bottle, add methanol, shake up, promptly to scale.

Accurate respectively reference substance solution and each 5～15 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.The every gram of this product contains Fructus Schisandrae Chinensis with schisandrin B (C ₂₃H ₂₈O ₆) meter, must not be less than 0.07mg.

Embodiment 2

Fructus Lycii 56g Semen Cuscutae (stir-fry) 25g Fructus Rubi 20g

Fructus Schisandrae Chinensis (steaming) 5g Semen Plantaginis (salt stir-fry) 7g

(3) get this product 5g, porphyrize adds kieselguhr 5g, grinds well, and 50ml adds diethyl ether, supersound process 20 minutes filters, and filtrate volatilizes, and residue adds chloroform 40ml, supersound process 20 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets the deoxyschizandrin reference substance, adds ethyl acetate and makes the solution that every 1ml contains 1mg, in contrast product solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, reference substance solution 2 μ l put in same silica gel G F respectively ₂₅₄On the lamellae, be developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect with the upper solution of petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15:5:1).In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

Assay:

Embodiment 3

Fructus Lycii 15g Semen Cuscutae (stir-fry) 20g Fructus Rubi 10g

Fructus Schisandrae Chinensis 1g Semen Plantaginis (salt stir-fry) 4g

Take by weighing above raw medicinal material, the above five tastes adopt superfine communication technique, are ground into micropowder, mixing; 90-110g Mel is in the time of 116-118 ℃, and refining density is 1.37, promptly gets refined honey; Every 100g powder adds the water of refined honey 40g and 110g, general ball, and 60 ℃ of dryings 11 hours, promptly.

(4) get this product powder 5g, add the 50mL petroleum ether, Soxhlet is extracted the 2.5h defat, and residue volatilizes petroleum ether, adds the 50mL methanol eddy and extracts 3h, filters, and filtrate is concentrated into 2mL, as need testing solution; Other gets the Fructus Rubi control medicinal material, shines medical material solution in pairs with legal system; According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ L of above-mentioned two kinds of solution, put respectively on same silica gel G plate with ethyl acetate-methanol-water (5: 1: 1), launch, take out, dry, spray 0.5%AlCl3, methanol solution is put under the uviol lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color.

(5) get this product content 1.5g and add methanol 20ml, soak 3h, filter, filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, as need testing solution.Get Semen Plantaginis control medicinal material 1g, methanol 20ml, heating and refluxing extraction 1h filters, and filtrate volatilizes, and residue adds the 1ml dissolve with methanol, in contrast medical material solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, control medicinal material solution 20 μ l, put respectively on same silica gel g thin-layer plate, with chloroform-methanol-ammonia (15: 1.5: 0.5) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 100 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the speckle of same color.

Embodiment 4

Fructus Lycii 25g Semen Astragali Complanati 25g Fructus Rubi 20g

Fructus Schisandrae Chinensis 5g Semen Plantaginis (salt stir-fry) 7g

Embodiment 5

Fructus Lycii 75g Semen Cuscutae 60g Fructus Rubi 40g

Fructus Schisandrae Chinensis 8g Semen Plantaginis (salt stir-fry) 15g

Take by weighing above raw medicinal material, adopt superfine communication technique, pulverize separately becomes micropowder, and mixing is even according to the mixed of weight ratio 3:7 with Macrogol 4000, splashes in the refrigerative liquid paraffin, makes drop pill, promptly.

Embodiment 6

Fructus Lycii 56g Semen Cuscutae (stir-fry) 25g Fructus Rubi 20g

Take by weighing above raw medicinal material, be crushed to 80 orders, mixing incapsulates, promptly.

Embodiment 7

Fructus Lycii 56g Semen Cuscutae (stir-fry) 25g Fructus Rubi 20g

Take by weighing above raw medicinal material, be crushed to 80 orders, mixing is that 8% starch slurry is granulated with concentration, drying, and granulate, tabletting, coating is made 350, promptly.

Embodiment 8

Fructus Lycii 25g Semen Cuscutae (stir-fry) 25g Fructus Rubi 12.5g

Fructus Schisandrae Chinensis (steaming) 3.125g Semen Plantaginis (salt stir-fry) 6.25g

[method for making] above five tastes are ground into fine powder, sieve mixing; 90-110g Mel is in the time of 116-118 ℃, and refining density is 1.37, promptly gets refined honey; Every 100g powder adds an amount of water pill with refined honey 35～50g, and drying is made 100g, promptly.

[character] this product is tan water-honeyed pill; Sweet, sour, the little hardship of distinguishing the flavor of.

This product is got in [discriminating] (1), puts microscopically and observes: plant skin stone cell surface and see irregular polygon, and wall thickness, wavy bending, laminated striation is clear.It is fallow to plant skin epidermis stone cell, and the class polygon is seen on the surface, and wall is thicker, and the hole ditch is fine and closely woven, and cell contains the burgundy thing.Plant skin palisade cells 2 row, the outer row of interior row are long, and light line is arranged.Plant the Intradermal epidermal surface and see rectangle like, wall is wavy slightly, is one group with several cells, and zyklopisch is arranged slightly.Nonglandular hair is unicellular, wall thickness, and lignify, the back vestiges that come off are like the stone cell shape.

(2) get this product 5g, porphyrize, the 50ml that adds diethyl ether, supersound process 20 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets Fructus Lycii control medicinal material 1g, grinds, and the 20ml that adds diethyl ether shines medical material solution in pairs with legal system.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-Ethyl formate-formic acid (20:20:0.1) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(3) get this product 10g, porphyrize adds petroleum ether (30～60 ℃) 30ml, supersound process 30 minutes, filter, discard petroleum ether liquid, medicinal residues are waved most petroleum ether, add methanol 30ml supersound process 20 minutes, filter, filtrate evaporate to dryness, residue add ethanol 5ml to be made it to disperse, and gets supernatant as need testing solution.Other gets Semen Cuscutae control medicinal material 2g, and porphyrize shines medical material solution in pairs with the method preparation.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with toluene-ethyl acetate-formic acid (10:5:4) is developing solvent, launches, and takes out, dry, spray 1% aluminum chloride alcoholic solution, airing is put under the ultra-violet lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.

(4) get this product 5g, porphyrize adds kieselguhr 5g, grinds well, and 50ml adds diethyl ether, supersound process 20 minutes filters, and filtrate volatilizes, and residue adds chloroform 40ml, supersound process 20 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets the deoxyschizandrin reference substance, adds ethyl acetate and makes the solution that every 1ml contains 1mg, in contrast product solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, reference substance solution 2 μ l put in same silica gel G F respectively ₂₅₄On the lamellae, be developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect with the upper solution of petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15:5:1).In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.

[inspection] should meet every regulation relevant under the pill item (appendix I A).

[assay] measured according to high performance liquid chromatography (Chinese Pharmacopoeia version appendix in 2005 VI D)

The preparation of need testing solution is got this product and is pulverized, and precision takes by weighing about 5g, and the accurate 50ml methanol that adds takes by weighing weight, and supersound process 30 minutes is put coldly, supplies weight, shake up, and filtration, promptly.

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

The every gram of this product contains Fructus Schisandrae Chinensis with schisandrin B (C ₂₃H ₂₈O ₆) meter, must not be less than 0.07mg.

[function with cure mainly] the kidney invigorating and essence nourishing.Be used for lumbago due to renal deficiency, dribble of urine, the seminal emission premature ejaculation, sexual impotence is sterile.

[usage and consumption] is oral.A 6g, 2 times on the one.

The heavy 10g of [specification] per 100 balls

[storage] sealing.

Claims

1, a kind of Chinese medicine composition for the treatment of male infertility is characterized in that it being that crude drug by following weight ratio is made:

2, Chinese medicine composition as claimed in claim 1 is characterized in that this Chinese medicine composition can be made by the crude drug of following weight ratio:

3, Chinese medicine composition as claimed in claim 1 is characterized in that this Chinese medicine composition can be made by the crude drug of following weight ratio:

4, Chinese medicine composition as claimed in claim 3 is characterized in that this Chinese medicine composition can be made by the crude drug of following weight ratio:

5, Chinese medicine composition as claimed in claim 4 is characterized in that this Chinese medicine composition can be made by the crude drug of following weight ratio:

6,, it is characterized in that technology adding adjuvant is made clinical acceptable forms such as tablet, capsule, pill, drop pill routinely as any described Chinese medicine composition of claim 1-5; Described adjuvant comprises solvent, disintegrating agent, correctives, antiseptic, coloring agent, binding agent, lubricant, substrate etc.

7, Chinese medicine composition as claimed in claim 6 is characterized in that the preparation method of this Chinese medicine composition pill is:

8, as claim 1-5 any one or 7 described method of quality control, it is characterized in that this method of quality control comprises one or more in following discrimination method and/or the assay:

Differentiate:

(1) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3～4g, porphyrize, the 40～60ml that adds diethyl ether, supersound process 15～30 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets Fructus Lycii control medicinal material 1～2g, grinds, and the 20～30ml that adds diethyl ether shines medical material solution in pairs with legal system.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15～25:15～25:0.1～0.5) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

(2) get pharmaceutical preparation of the present invention and be equivalent to crude drug 7～8g, porphyrize adds petroleum ether (30～60 ℃) 25～35ml, supersound process 20～40 minutes, filter, discard petroleum ether liquid, medicinal residues are waved most petroleum ether, add methanol 30～40ml supersound process 15～30 minutes, filter, filtrate evaporate to dryness, residue add ethanol 5～6ml to be made it to disperse, and gets supernatant as need testing solution.Other gets Semen Cuscutae control medicinal material 2g, and porphyrize shines medical material solution in pairs with the method preparation.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with toluene-ethyl acetate-formic acid (8～12:4～6:3～5) is developing solvent, launches, and takes out, dry, spray 1% aluminum chloride alcoholic solution, airing is put under the ultra-violet lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

(3) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3～4g, porphyrize, the 40～60ml that adds diethyl ether, supersound process 15～30 minutes, filter, filtrate volatilizes, and residue adds chloroform 30～50ml, supersound process 15～25 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets the deoxyschizandrin reference substance, adds ethyl acetate and makes the solution that every 1ml contains 1mg, in contrast product solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, reference substance solution 2 μ l put in same silica gel G F respectively ₂₅₄On the lamellae, be developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect with the upper solution of petroleum ether (30～60 ℃)-Ethyl formate-formic acid (10～20:3～7:1～2).In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

(4) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3～4g, porphyrize adds 40～60mL petroleum ether, and Soxhlet is extracted 2～3h defat, and residue volatilizes petroleum ether, adds 40～60mL methanol eddy and extracts 2～4h, filters, and filtrate is concentrated into 1～2mL, as need testing solution; Other gets the Fructus Rubi control medicinal material, shines medical material solution in pairs with legal system; According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VIB) test, draw each 5 μ L of above-mentioned two kinds of solution, put respectively on same silica gel G plate with ethyl acetate-methanol-water (4～7:1～2:1～2), launch, take out, dry, spray 0.5%AlCl3, methanol solution is put under the uviol lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color;

(5) get pharmaceutical preparation of the present invention and be equivalent to crude drug 1～2g, porphyrize adds methanol 15～30ml, soaks 2～4h, filters, and filtrate evaporate to dryness, residue add methanol 1～2ml makes dissolving, as need testing solution.Get Semen Plantaginis control medicinal material 1～2g, methanol 10～30ml, heating and refluxing extraction 0.5～1.5h filters, and filtrate volatilizes, and residue adds 1～2ml dissolve with methanol, in contrast medical material solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution, each 20 μ l of control medicinal material solution, put respectively on same silica gel g thin-layer plate, with chloroform-methanol-ammonia (10～20:1～2:0.5～1) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 100 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the speckle of same color;

Assay:

Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; With acetonitrile-water-acetic acid (65:40:0.1) is mobile phase; The detection wavelength is 254nm;

It is an amount of that the schisandrin B reference substance is got in the preparation of reference substance solution, and accurate the title decides, and adds methanol and makes the solution that every 1ml contains 30 μ g, promptly;

The preparation of need testing solution is got pharmaceutical preparation of the present invention and is equivalent to crude drug 3～4g, porphyrize, and precision takes by weighing about 4～6g, put in the apparatus,Soxhlet's, it is an amount of to add normal hexane, and soaked overnight was in 80～85 ℃ of reflux, extract, 5～8 hours, extracting solution low temperature evaporate to dryness, residue adds the methanol slight fever makes dissolving, moves in the 25ml measuring bottle, adds methanol to scale, shake up, promptly;

Accurate respectively reference substance solution and each 5～15 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly; Pharmaceutical preparation of the present invention is equivalent to crude drug 0.5～1g and contains Fructus Schisandrae Chinensis with schisandrin B (C ₂₃H ₂₈O ₆) meter, must not be less than 0.05～0.08mg.

9, as the method for quality control of Chinese medicine composition as described in the claim 8, it is characterized in that this method of quality control comprises one or more in following discrimination method and/or the assay:

Differentiate:

(1) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize, the 50ml that adds diethyl ether, supersound process 20 minutes filters, and filtrate volatilizes, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets Fructus Lycii control medicinal material 1g, grinds, and the 20ml that adds diethyl ether shines medical material solution in pairs with legal system.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with petroleum ether (30～60 ℃)-Ethyl formate-formic acid (20:20:0.1) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

(2) get this product 10g, porphyrize adds petroleum ether (30～60 ℃) 30ml, supersound process 30 minutes, filter, discard petroleum ether liquid, medicinal residues are waved most petroleum ether, add methanol 30ml supersound process 20 minutes, filter, filtrate evaporate to dryness, residue add ethanol 5ml to be made it to disperse, and gets supernatant as need testing solution.Other gets Semen Cuscutae control medicinal material 2g, and porphyrize shines medical material solution in pairs with the method preparation.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 2 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with toluene-ethyl acetate-formic acid (10:5:4) is developing solvent, launches, and takes out, dry, spray 1% aluminum chloride alcoholic solution, airing is put under the ultra-violet lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color;

(3) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize, the 50ml that adds diethyl ether, supersound process 20 minutes, filter, filtrate volatilizes, and residue adds chloroform 40ml, supersound process 20 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds ethyl acetate 1ml makes dissolving, as need testing solution.Other gets the deoxyschizandrin reference substance, adds ethyl acetate and makes the solution that every 1ml contains 1mg, in contrast product solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, reference substance solution 2 μ l put in same silica gel G F respectively ₂₅₄On the lamellae, be developing solvent, launch, take out, dry, put under the ultra-violet lamp (254nm) and inspect with the upper solution of petroleum ether (30～60 ℃)-Ethyl formate-formic acid (15:5:1).In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color;

(4) get pharmaceutical preparation of the present invention and be equivalent to crude drug 3.6g, porphyrize adds the 50mL petroleum ether, and Soxhlet is extracted the 2.5h defat, and residue volatilizes petroleum ether, adds the 50mL methanol eddy and extracts 3h, filters, and filtrate is concentrated into 2mL, as need testing solution; Other gets the Fructus Rubi control medicinal material, shines medical material solution in pairs with legal system; According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw each 5 μ L of above-mentioned two kinds of solution, put respectively on same silica gel G plate with ethyl acetate-methanol-water (5: 1: 1), launch, take out, dry, spray 0.5%AlCl3, methanol solution is put under the uviol lamp (365nm) and is inspected.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color;

(5) get pharmaceutical preparation of the present invention and be equivalent to crude drug 1.8g, porphyrize adds methanol 20ml, soaks 3h, filters, and filtrate evaporate to dryness, residue add methanol 1ml makes dissolving, as need testing solution.Get Semen Plantaginis control medicinal material 1g, methanol 20ml, heating and refluxing extraction 1h filters, and filtrate volatilizes, and residue adds the 1ml dissolve with methanol, in contrast medical material solution.According to thin layer chromatography (" Chinese pharmacopoeia version appendix in 2005 VI B) test, draw need testing solution 10 μ l, control medicinal material solution 20 μ l, put respectively on same silica gel g thin-layer plate, with chloroform-methanol-ammonia (15: 1.5: 0.5) is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and 100 ℃ to be heated to the speckle colour developing clear.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the speckle of same color;

Assay:

The preparation of need testing solution is got pharmaceutical preparation of the present invention and is equivalent to crude drug 4g, porphyrize, and precision takes by weighing about 5g, put in the apparatus,Soxhlet's, it is an amount of to add normal hexane, and soaked overnight was in 80～85 ℃ of reflux, extract, 6 hours, extracting solution low temperature evaporate to dryness, residue adds the methanol slight fever makes dissolving, moves in the 25ml measuring bottle, adds methanol to scale, shake up, promptly;

Accurate respectively reference substance solution and each 5～15 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly; Pharmaceutical preparation of the present invention is equivalent to crude drug 0.719g and contains Fructus Schisandrae Chinensis with schisandrin B (C ₂₃H ₂₈O ₆) meter, must not be less than 0.07mg.