CN101991636B

CN101991636B - Detection method of Aidi injection preparation

Info

Publication number: CN101991636B
Application number: CN2010105310881A
Authority: CN
Inventors: 窦啟玲; 叶湘武; 汤琼
Original assignee: Guizhou Yibai Pharmaceutical Co Ltd
Current assignee: Guizhou Yibai Pharmaceutical Co Ltd
Priority date: 2007-11-19
Filing date: 2007-11-19
Publication date: 2012-04-25
Anticipated expiration: 2027-11-19
Also published as: CN101991636A

Abstract

The invention provides a detection method of an Aidi injection preparation, which comprises the items of character, discrimination, inspection and content determination, wherein the discrimination comprises alpha-naphthol chromogenic reaction and thin-layer chromatography discrimination on ginseng and radix astragali in the preparation by taking ginsenoside Re, Rg1, Rb and astragaloside as references; the content determination comprises content determination of ginsenoside Re contained in the ginseng and content determination of the cantharidin contained in cantharis; and the inspection comprises inspection of safety indicators of pH value, heavy metal, pyrogen, protein, tannin and the like. Compared with the prior art, the quality control method of the invention can effectively control the product quality, thereby ensuring the clinical curative effect.

Description

The detection method of Aidi injection preparation

The application's name that to be the applicant submit on November 19th, 2007 is called: " method of quality control of Aidi injection preparation ", application number are: the dividing an application of the application for a patent for invention of 200710078008.X.

Technical field: the present invention relates to the detection method of Aidi injection preparation, belong to technical field of medicaments.

Background technology: cancer is one big type of disease of serious threat human health; Aidi injection preparation mainly is made up of Chinese blister beetle, genseng, the Radix Astragali and wilsonii, has effect clearing heat and detoxicating, the powder for dispersing blood stasis knot, is mainly used in primary carcinoma of liver; Lung cancer; The carcinoma of the rectum, malignant lymphoma, diseases such as gynecologic malignant tumor.Wherein " ad pro injection " publication is the 20th of Chinese medicine ministerial standard.But development along with society; Requirements for quality control for medicine is increasingly high; And it is also more and more urgent for the demand of seeking efficient and convenient controlled detection method; The inventor is through discovering, in the existing Aidi injection preparation quality control standard, the description of product proterties is definite inadequately; In the thin layer discriminating to genseng, the Radix Astragali, the preparation method of need testing solution is more loaded down with trivial details, and the method for operating time of upper prop wash-out is longer; The selection proportioning of developping agent is not ideal enough, be difficult to each effective ingredient particularly the ginsenoside Rg1 separate fully with Astragaloside IV, developping agent has the restriction of configuration condition simultaneously, these all are prone to cause in discrimination process degree of separation bad, it is clear inadequately that spot develops the color; Thin layer plate thickness is not stipulated in the primary standard in addition; But show that through a large amount of the realization genseng is very tight to the requirement of thin layer plate, if thin layer plate is thin excessively; The ginsenoside Rg1 is difficult to separate with Astragaloside IV; Must slab (more than the 500 μ m) could be with the two effective separation, humidity is also very big to its influence simultaneously, will strict controlled humidity in expansion process; So existing method of quality control can not effectively be controlled the quality of Aidi injection preparation, thereby will influence the clinical efficacy of said preparation.

Summary of the invention:

The objective of the invention is to: a kind of method of quality control of Aidi injection preparation is provided, and this ejection preparation comprises freeze drying powder injection, infusion solution and liquid drugs injection.The present invention is directed to the deficiency of prior art; Method of quality control to this Aidi injection preparation is studied; Content assaying method and the genseng of cantharidin, the thin layer discrimination method of the Radix Astragali have been improved; And increased the safety indexes inspection item of this Aidi preparation, improve the quality control standard of Aidi injection preparation, thereby guaranteed the clinical efficacy of said preparation.

Aidi injection preparation according to the invention is to constitute like this: calculate according to composition by weight: it mainly is prepared from Chinese blister beetle 0.5-5, genseng 20-80, Radix Astragali 200-600, wilsonii 100-200.Its preparation method is: above four flavors, and the genseng section is extracted 1-5 time with the 20-85% alcohol heating reflux, merges extract, filters, and reclaims ethanol, and soup is subsequent use; Three flavor boilings such as the dregs of a decoction and all the other Chinese blister beetles 1-6 time, collecting decoction filters; Filtrating merges with Ginseng extract; With stone sulphur method precipitation process 1-3 time, the gained supernatant adds ethanol to be made and contains the alcohol amount and reach 50-99%, hold over night; Get the supernatant decompression recycling ethanol to there not being the alcohol flavor, process freeze drying powder injection, infusion solution or liquid drugs injection respectively according to conventional method again.

Method of quality control according to the invention mainly comprise in proterties, discriminating, inspection and the assay project partly or entirely; Wherein discriminating comprises the alpha-Naphthol chromogenic reaction, serves as the thin-layer chromatography discriminating that contrasts the genseng in the preparation and the Radix Astragali with ginsenoside Re, Rg1, Rb and Astragaloside IV; Assay is the assay to the contained ginsenoside Re of genseng in the preparation, the contained cantharidin of Chinese blister beetle.

The discrimination method of the genseng and the Radix Astragali is to be contrast with ginsenoside Re, Rg1, Rb and Astragaloside IV, and with methenyl choloride: methyl alcohol: water=10-50: 1-9: 0.5-5 is the thin layer chromatography of developping agent.

The discrimination method of the genseng and the Radix Astragali can also be to be contrast with ginsenoside Re, Rg1, Rb and Astragaloside IV, and with methenyl choloride: ethyl acetate: methyl alcohol: water=1-10: 2-15: 1-10: 1-10 is the thin layer chromatography of developping agent.

Discrimination method comprises the part or all of of following project:

(1) gets freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, add ethanol and stir, filter; Sediment is used dissolved in distilled water with washing with alcohol 1-5 time with sediment, gets 0.5-3ml and puts in the test tube; Add 2-10% alpha-Naphthol ethanolic solution after the low-grade fever; Shake up, slowly add the concentrated sulphuric acid along tube wall again, at two liquid junction place displaing amaranth rings;

(2) method one: get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, put in the separating funnel, extract with the methenyl choloride jolting; Discard methenyl choloride liquid, water layer extracts 1-4 time with water-saturated n-butanol, merges normal butyl alcohol liquid; With ammonia solution washing 1-4 time, discard ammonia solution, normal butyl alcohol liquid evaporate to dryness in water-bath; Residue adds methyl alcohol makes dissolving, as need testing solution.Other gets ginsenoside Re, Rg ₁, Rb ₁And the Astragaloside IV reference substance, add dissolve with methanol, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With methenyl choloride: methyl alcohol: water=10-50: 1-9: 0.5-5 is a developping agent, launches, and takes out; Dry; Spray is with 10% ethanol solution of sulfuric acid, and heating is put under the ultraviolet lamp 200-500nm after the placement again and inspected; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

Method two: get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, put in the separating funnel, extract 1-4 time the merging extract with water saturated normal butyl alcohol; Put evaporate to dryness in the water-bath, residue is used dissolved in distilled water, adds on the DA-201 resin column for preparing in advance; Use water washing, use 20-80% methanol solution wash-out again, collect eluent; Evaporate to dryness, residue add methyl alcohol makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add dissolve with methanol, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=1-10: 2-15: 1-10: 1-10,10 ℃ of lower floor's solution with held are developping agent, launch; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

Concrete discrimination method comprises the part or all of of following project:

(1) getting freeze-dried powder 400-1000mg adds injection water 5-20ml and makes dissolving or get that transfusion 200-500ml is concentrated into 5-20ml or water intaking injection 20-100ml is concentrated into 5-20ml; Get above-mentioned medicine respectively, add ethanol 20-100ml, stir; Filter, sediment dissolves sediment with washing with alcohol 1-4 time with distilled water 2-10ml; Get 0.5-3ml and put in the test tube, add 2-10% alpha-Naphthol ethanolic solution 2-10 after the low-grade fever and drip, shake up; Slowly add concentrated sulphuric acid 0.2-2ml along tube wall again, at two liquid junction place displaing amaranth rings;

(2) method one: get freeze-dried powder 400-1000mg and add injection water 20-100ml and make dissolving or get transfusion 200-500ml and be concentrated into 20-100ml or water intaking injection 20-100ml; Put in the separating funnel, extract, discard methenyl choloride liquid with the jolting of 10-50ml methenyl choloride; Water layer extracts 1-4 time with water-saturated n-butanol, and each 40-60ml merges normal butyl alcohol liquid; With ammonia solution washing 1-4 time, each 10-50ml discards ammonia solution; Normal butyl alcohol liquid evaporate to dryness in water-bath, residue add methyl alcohol 0.5-3ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb ₁And the Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 0.5-3mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 3-20 μ l of above-mentioned two kinds of solution, put on same silica gel g thin-layer plate thick 500 μ m and more than the 500 μ m respectively; With methenyl choloride: methyl alcohol: water=10-50: 1-9: 0.5-5 is a developping agent, and controlled humidity launches where necessary; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put again under the ultraviolet lamp 200-500nm after placing more than 1 hour and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

Method two: get freeze-dried powder 400-1000mg and add injection water 20-100ml and make dissolving or get transfusion 200-500ml and be concentrated into 20-100ml or water intaking injection 20-100ml; Put in the separating funnel, extract 1-4 time, each 10-60ml with water saturated normal butyl alcohol; Merge extract, put evaporate to dryness in the water-bath, residue dissolves with distilled water 1-10ml; Add on the DA-201 resin column for preparing in advance internal diameter 0.5～2cm, long 10-20cm; After filling the DA-201 resin of 10-15cm, recharge the 1-5g neutral alumina; 20-80% methanol solution wash-out is used in water 50-200ml washing again, collects eluent, and evaporate to dryness, residue add methyl alcohol 0.5-5ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 0.5-3mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 3-20 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=1-10: 2-15: 1-10: 1-10,10 ℃ of lower floor's solution with held are developping agent, launch; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp 200-500nm and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color.

Ginsenoside Re's content assaying method is to be the AAS of contrast with ginsenoside Re's reference substance in the preparation.

The content assaying method of cantharidin is to be contrast with the cantharidin reference substance in the preparation, is the vapor-phase chromatography of immobile liquid with OV-17.

Content assaying method comprises:

(1) ginsenoside Re

The preparation of reference substance solution: precision takes by weighing ginsenoside Re's reference substance, adds dissolve with methanol, shakes up, and promptly gets;

The preparation of typical curve: precision is measured reference substance solution 0,20,40,60,80,100 μ l, puts respectively in the 10ml tool plug test tube, puts the water-bath Back stroke and desolvates, and takes out immediately; Put coldly, add 2-10% vanillic aldehyde glacial acetic acid solution, perchloric acid, shake up, put in the water-bath and heat; Take out, use water cooling, add glacial acetic acid, shake up; According to the Chinese Pharmacopoeia AAS, measure absorbance log, be that ordinate, concentration are horizontal ordinate with the absorbance log, the drawing standard curve;

The preparation of need testing solution: get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, put in the separating funnel, with chloroform extraction 1-5 time, the combined chloroform extract washs 1-4 time with distilled water; Discard chloroform solution, washing lotion and above-mentioned water layer merge, and put in the separating funnel, extract 1-5 time with water saturated normal butyl alcohol, merge n-butanol extracting liquid; Add anhydrous sodium sulfate, stir, put, normal butyl alcohol liquid is moved in the evaporating dish to clarification; With a small amount of normal butyl alcohol washing anhydrous sodium sulfate, washing lotion is incorporated in the evaporating dish, and evaporate to dryness is with a small amount of dissolved in distilled water residue; Be added on the macroporous absorbent resin DA-201 post, treat that liquid level is near cotton layer after, use the distilled water wash-out, discard washing lotion; Use the 40-90% ethanol elution, collect eluent in evaporating dish, put evaporate to dryness in the water-bath, use the dissolve with methanol residue; Move in the 5ml measuring bottle, add methyl alcohol, shake up, promptly get to scale;

Determination method: precision is measured need testing solution 100 μ l, the sighting target directrix curve prepare the method under the item, measure absorbance log from " putting in the 10ml tool plug test tube " in accordance with the law.Read ginsenoside Re's the need testing solution weight μ g from typical curve, calculate; In this ejection preparation, contain genseng in the freeze drying powder injection, must not be less than 1% in the ginsenoside Re; Contain genseng in the infusion solution in the ginsenoside Re, must not be less than 0.01mg/ml; Contain genseng in the liquid drugs injection in the ginsenoside Re, must not be less than 0.1mg/ml.

(2) cantharidin

Method one: according to the Chinese Pharmacopoeia gas chromatography determination.

Chromatographic condition and system suitability test: glass column 1-2m * 3mm id is an immobile liquid with OV-17, and OV-17 is made up of 50% methyl, 50% phenyl polysiloxane, and being coated with the stain amount is 0.5-3%, carrier Shimalite W (AW-DMCS) 60-120 order.Column temperature is 160 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and adds the chloroform dissolving, shakes up, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, add the 1-3mol/L sulfuric acid solution, the accurate again chloroform that adds, jolting is extracted, and standing demix is centrifugal in case of necessity, gets subnatant and promptly gets;

Determination method: precision is measured reference substance solution and need testing solution respectively, and inject gas chromatograph calculates; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml.

Method two: according to the Chinese Pharmacopoeia gas chromatography determination.

Chromatographic condition and system suitability test: with carbowax-20M and methyl silicone rubber SE-30 is immobile liquid, and coating concentration is respectively 5-20% and 1-10%, 0.5-2: 0.5-2 mixes the dress post.Column temperature is 180 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500.

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and adds the chloroform dissolving, as reference substance solution.

The preparation of need testing solution: get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, add the 1-3mol/L sulfuric acid solution, extract 1-5 time with the chloroform jolting, combined chloroform liquid with the concentrated constant volume of K-D concentrator, promptly gets.

Determination method precision is respectively measured reference substance solution and need testing solution, and inject gas chromatograph calculates, and promptly gets; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml.

Content assaying method comprises more specifically:

(1) ginsenoside Re

The preparation of reference substance solution: precision takes by weighing ginsenoside Re's reference substance, puts in the measuring bottle, adds dissolve with methanol and makes and contain ginsenoside Re 1-2mg among every 1ml, shakes up, and promptly gets;

The preparation of typical curve: precision is measured reference substance solution 0,20,40,60,80,100 μ l, puts respectively in the 10ml tool plug test tube, puts the water-bath Back stroke and desolvates, and takes out immediately; Put coldly, add 2-10% vanillic aldehyde glacial acetic acid solution 0.05-1ml, perchloric acid 0.5-1.5ml, shake up; Put in the 40-80 ℃ of water-bath and heated 8-25 minute, take out, with water cooling 1-3 minute; Add glacial acetic acid 2-10ml, shake up, according to the Chinese Pharmacopoeia AAS; Measuring absorbance log in the 400-700nm wavelength, is that ordinate, concentration are horizontal ordinate with the absorbance log, the drawing standard curve;

The preparation of need testing solution: get freeze-dried powder 200-400mg and add injection water 10-30ml and make dissolving or get transfusion 50-200ml and be concentrated into 10-30ml or water intaking injection 10-30ml; Precision is measured above-mentioned medicine respectively, put in the separating funnel, and with chloroform extraction 1-5 time, each 10-30ml, combined chloroform extract; With distilled water washing 1-4 time, each 2-10ml discards chloroform solution, and washing lotion and above-mentioned water layer merge, and put in the separating funnel; Extract 1-5 time with water saturated normal butyl alcohol, each 40-60ml merges n-butanol extracting liquid, adds anhydrous sodium sulfate 1-5g, stirs; Put to clarification, normal butyl alcohol liquid is moved in the evaporating dish, with a small amount of normal butyl alcohol washing anhydrous sodium sulfate, washing lotion is incorporated in the evaporating dish, evaporate to dryness; With a small amount of dissolved in distilled water residue, be added on the macroporous absorbent resin DA-201 post, treat that liquid level is near cotton layer after, with distilled water 100ml wash-out, the control flow velocity is 0.3-0.6ml/min; Discard washing lotion,, collect eluent in evaporating dish, put evaporate to dryness in the water-bath, use the dissolve with methanol residue with 40-90% ethanol 70-100ml wash-out; Move in the 5ml measuring bottle, add methyl alcohol, shake up, promptly get to scale;

(2) cantharidin

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and chlorination is copied into the solution that every 1ml contains 0.005-0.02mg, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder 30-100g and add injection water 5-20ml and make dissolving or get that transfusion 30-100ml is concentrated into about 5-20ml or the injection 5-20ml that fetches water; Precision is measured above-mentioned medicine respectively, adds 1-3mol/L sulfuric acid solution 0.5-2ml, the accurate again 0.5-2ml chloroform that adds, and jolting is extracted, and standing demix is centrifugal in case of necessity, gets subnatant and promptly gets;

Determination method: precision is measured reference substance solution and each 2-12 μ l of need testing solution respectively, and inject gas chromatograph calculates; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml.

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and chlorination is copied into the solution that every 1ml contains 0.05-0.2mg, as reference substance solution.

The preparation of need testing solution: get freeze-dried powder 400-1000mg and add injection water 20-100ml and make dissolving or get transfusion 200-500ml and be concentrated into 20-100ml or water intaking injection 20-100ml; Add 1-3mol/L sulfuric acid solution 2-10ml; Extract 1-5 time each 10-100ml, combined chloroform liquid with the chloroform jolting; Be settled to 2-8ml with the K-D concentrator is concentrated, promptly get.

Inspection item comprises the part or all of of following project:

PH value: should be 3.0-6.5;

Heavy metal: get freeze-dried powder 20-40mg and add injection water 2ml and make dissolving or get transfusion 8-20ml and be concentrated into 2ml or water intaking injection 2ml; Put in the crucible, evaporate to dryness in the water-bath according to the inspection of Chinese Pharmacopoeia heavy metal inspection technique, contains heavy metal and must not cross 5/1000000ths;

Pyrogen: get freeze-dried powder 300-600mg and add injection water 30ml and make dissolving or get transfusion 100-300ml and be concentrated into 30ml or water intaking injection 30ml; Be diluted to 100ml with sodium chloride injection or 5～10% glucose injections, by the dosage of rabbit body weight 1Kg injection 10ml, according to the inspection of Chinese Pharmacopoeia pyrogen test, should be up to specification;

Protein: get freeze-dried powder 10-20mg and add injection water 1ml and make dissolving or get transfusion 4-10ml and be concentrated into 1ml or water intaking injection 1ml; The 30% sulfosalicylic acid solution 1ml that adds new preparation, mixing was placed 5 minutes, muddiness must not occur;

Tannin: get freeze-dried powder 10-20mg and add injection water 1ml and make dissolving or get transfusion 4-10ml and be concentrated into 1ml or water intaking injection 1ml; The physiological sodium chloride solution 5ml that contains 1% Ovum Gallus domesticus album that adds new preparation in case of necessity, filters with the 0.45um miillpore filter, places 10 minutes, muddiness or deposition must not occur;

Resin: get freeze-dried powder 50-100mg and add injection water 5ml and make dissolving or get transfusion 20-50ml and be concentrated into 5ml or water intaking injection 5ml; Add 1 of hydrochloric acid, placed 30 minutes, deposition must not occur;

Oxalates: get freeze-dried powder 20-40mg and add injection water 2ml and make dissolving or get transfusion 8-20ml and be concentrated into 2ml or water intaking injection 2ml; Regulate pH value to 1-2 with watery hydrochloric acid, filter, filtrating is regulated pH value to 5-6, adds 3% calcium chloride solution 2-3 and drips, and places 10 minutes, muddiness or deposition must not occur;

Potassium ion: get freeze-dried powder 20-40mg and add injection water 2ml and make dissolving or get transfusion 8-20ml and be concentrated into 2ml or water intaking injection 2ml; Evaporate to dryness is measured according to Chinese Pharmacopoeia potassium ion determination method, should be up to specification;

Arsenic salt: get freeze-dried powder 10-20mg and add injection water 1ml and make dissolving or get transfusion 4-10ml and be concentrated into 1ml or water intaking injection 1ml; Precision is measured, and evaporate to dryness in crucible adds 2% magnesium nitrate ethanolic solution 3ml, lights; In the afterburnt, burning with little heated earlier makes charing, blazing to ashing fully at 500-600 ℃ again; Put coldly, add hydrochloric acid 5ml and water 21ml makes dissolving, according to the inspection of Chinese Pharmacopoeia arsenic salt inspection technique; Arsenic content must not cross 2/1000000ths in every 1ml finished product;

Haemolysis and cohesion: rabbit blood or sheep blood number milliliter are got in the preparation of 2% red blood cell suspension, put into the conical flask that fills beaded glass, jolting 10 minutes; Remove fibrinogen; Make to become to take off fine blood, the physiological sodium chloride solution that adds about 10 times of amounts washs 2-3 time, till the apparent redness of supernatant; The gained red blood cell is made into 2% suspension with physiological sodium chloride solution, promptly gets;

The preparation of need testing solution: get freeze-dried powder 100-200mg and add injection water 10ml and make dissolving or get transfusion 40-100ml and be concentrated into 10ml or water intaking injection 10ml; Add physiological sodium chloride solution and be diluted to 50ml, mixing promptly gets;

Test method: get 6 in test tube, proportional quantity adds 2% red blood cell suspension and physiological sodium chloride solution successively in the according to the form below, behind the mixing; In 37 ℃ of constant temperature ovens, placed 30 minutes, add the need testing solution of different amounts respectively, the 6th pipe is control tube; After shaking up, put in 37 ℃ ± 0.5 ℃ constant temperature oven, beginning was whenever observed once at a distance from 15 minutes; After 1 hour, whenever once, observed altogether 3 hours at a distance from 1 h observation;

By last method inspection, be as the criterion with the 3rd pipe, this product haemolysis and red blood cell condensation must not occur in 3 hours;

Undue toxicity: get freeze-dried powder add the injection water make the dissolving or get the transfusion or the water intaking injection; By every 0.5ml of mouse, according to the inspection of Chinese Pharmacopoeia undue toxicity inspection technique;

Allergin inspection: get freeze-dried powder and add the injection water and make dissolving or get transfusion or the water intaking injection; With former concentration, be 0.5ml/ by the cavy priming dose, booster dose is 1.0ml/; According to the inspection of Chinese Pharmacopoeia allergin inspection technique, should be up to specification;

The material that boosts inspection: get freeze-dried powder and add the injection water and make dissolving or get transfusion or the water intaking injection; With former concentration, press the dosage of rat body weight 1kg injection 10ml, according to the Chinese Pharmacopoeia material inspection technique inspection of boosting, should be up to specification;

Depressor inspection: get freeze-dried powder and add the injection water and make dissolving or get transfusion or the water intaking injection; With former concentration, by the dosage of cat body weight 1kg injection 1ml, according to the inspection of Chinese Pharmacopoeia test of depressor substance, should be up to specification;

Other: should meet each item regulation relevant under the Chinese Pharmacopoeia injection item;

Method of quality control according to the invention is:

Proterties: for freeze drying powder injection, product is light yellow loose block to light brown;

For infusion solution, product is light yellow clear liquid to light brown;

For liquid drugs injection, product is light yellow clear liquid to light brown;

Differentiate:

(2) get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, put in the separating funnel, extract with the methenyl choloride jolting; Discard methenyl choloride liquid, water layer extracts 1-4 time with water-saturated n-butanol, merges normal butyl alcohol liquid; With ammonia solution washing 1-4 time, discard ammonia solution, normal butyl alcohol liquid evaporate to dryness in water-bath; Residue adds methyl alcohol makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb ₁And the Astragaloside IV reference substance, add dissolve with methanol, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With methenyl choloride: methyl alcohol: water=10-50: 1-9: 0.5-5 is a developping agent, launches, and takes out; Dry; Spray is with 10% ethanol solution of sulfuric acid, and heating is put under the ultraviolet lamp 200-500nm after the placement again and inspected; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

Determination method: precision is measured need testing solution 100 μ l, the sighting target directrix curve prepare the method under the item, measure absorbance log from " putting in the 10ml tool plug test tube " in accordance with the law; Read ginsenoside Re's the need testing solution weight μ g from typical curve, calculate; In this ejection preparation, contain genseng in the freeze drying powder injection, must not be less than 1% in the ginsenoside Re; Contain genseng in the infusion solution in the ginsenoside Re, must not be less than 0.01mg/ml; Contain genseng in the liquid drugs injection in the ginsenoside Re, must not be less than 0.1mg/ml;

(2) cantharidin is according to the Chinese Pharmacopoeia gas chromatography determination;

Chromatographic condition and system suitability test: with carbowax-20M and methyl silicone rubber SE-30 is immobile liquid, and coating concentration is respectively 5-20% and 1-10%, 0.5-2: 0.5-2 mixes the dress post; Column temperature is 180 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and adds the chloroform dissolving, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, add the 1-3mol/L sulfuric acid solution, extract 1-5 time with the chloroform jolting, combined chloroform liquid with the concentrated constant volume of K-D concentrator, promptly gets;

Determination method: precision is measured reference substance solution and need testing solution respectively, and inject gas chromatograph calculates, and promptly gets; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml.

Concrete method of quality control is:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(2) getting freeze-dried powder 400-1000mg adds injection water 20-100ml and makes dissolving or get transfusion 200-500ml and be concentrated into 20-100ml or water intaking injection 20-100ml; Put in the separating funnel, extract, discard methenyl choloride liquid with the jolting of 10-50ml methenyl choloride; Water layer extracts 1-4 time with water-saturated n-butanol, and each 40-60ml merges normal butyl alcohol liquid; With ammonia solution washing 1-4 time, each 10-50ml discards ammonia solution; Normal butyl alcohol liquid evaporate to dryness in water-bath, residue add methyl alcohol 0.5-3ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb ₁And the Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 0.5-3mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 3-20 μ l of above-mentioned two kinds of solution, put on same silica gel g thin-layer plate thick 500 μ m and more than the 500 μ m respectively; With methenyl choloride: methyl alcohol: water=10-50: 1-9: 0.5-5 is a developping agent, and controlled humidity launches where necessary; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put again under the ultraviolet lamp 200-500nm after placing more than 1 hour and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and chlorination is copied into the solution that every 1ml contains 0.05-0.2mg, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder 400-1000mg and add injection water 20-100ml and make dissolving or get transfusion 200-500ml and be concentrated into 20-100ml or water intaking injection 20-100ml; Add 1-3mol/L sulfuric acid solution 2-10ml; Extract 1-5 time each 10-100ml, combined chloroform liquid with the chloroform jolting; Be settled to 2-8ml with the K-D concentrator is concentrated, promptly get;

Method of quality control according to the invention also can be:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(2) get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, put in the separating funnel, extract 1-4 time the merging extract with water saturated normal butyl alcohol; Put evaporate to dryness in the water-bath, residue is used dissolved in distilled water, adds on the DA-201 resin column for preparing in advance; Use water washing, use 20-80% methanol solution wash-out again, collect eluent; Evaporate to dryness, residue add methyl alcohol makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add dissolve with methanol, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=1-10: 2-15: 1-10: 1-10,10 ℃ of lower floor's solution with held are developping agent, launch; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Tannin: get freeze-dried powder 10-20mg and add injection water 1ml and make dissolving or get transfusion 4-10ml and be concentrated into 1ml or water intaking injection 1ml; The physiological sodium chloride solution 5ml that contains 1% Ovum Gallus domesticus album that adds new preparation in case of necessity, filters with the 045um miillpore filter, places 10 minutes, muddiness or deposition must not occur;

Assay:

(1) ginsenoside Re

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and adds the chloroform dissolving, as the brilliant solution of contrast;

Determination method precision is respectively measured reference substance solution and need testing solution, and inject gas chromatograph calculates, and promptly gets; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 000001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml.

Concrete method of quality control is:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(2) getting freeze-dried powder 400-1000mg adds injection water 20-100ml and makes dissolving or get transfusion 200-500ml and be concentrated into 20-100ml or water intaking injection 20-100ml; Put in the separating funnel, extract 1-4 time, each 10-60ml with water saturated normal butyl alcohol; Merge extract, put evaporate to dryness in the water-bath, residue dissolves with distilled water 1-10ml; Add on the DA-201 resin column for preparing in advance internal diameter 0.5～2cm, long 10-20cm; After filling the DA-201 resin of 10-15cm, recharge the 1-5g neutral alumina; 20-80% methanol solution wash-out is used in water 50-200ml washing again, collects eluent, and evaporate to dryness, residue add methyl alcohol 0.5-5ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 0.5-3mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 3-20 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=1-10: 2-15: 1-10: 1-10,10 ℃ of lower floor's solution with held are developping agent, launch; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp 200-500nm and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

The preparation of need testing solution: get freeze-dried powder 400-1000mg and add injection water 20-100ml and make dissolving or get transfusion 200-500ml and be concentrated into 20-100ml or water intaking injection 20-100ml; Add 1-3mol/L sulfuric acid solution 2-10ml; Extract 1-5 time each 10-100ml, combined chloroform liquid with the chloroform jolting; Be settled to 2-8ml with the K-D concentrator is concentrated, promptly get; Determination method precision is respectively measured reference substance solution and need testing solution, and inject gas chromatograph calculates, and promptly gets; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml.

Method of quality control according to the invention also can be:

Proterties: for freeze drying powder injection, product is the loose block of light brown;

For infusion solution, product is the clear liquid of light brown;

For liquid drugs injection, product is the clear liquid of light brown;

Differentiate:

Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

Chromatographic condition and system suitability test: glass column 1-2m * 3mm id is an immobile liquid with OV-17, and OV-17 is made up of 50% methyl, 50% phenyl polysiloxane, and being coated with the stain amount is 0.5-3%, carrier Shimalite W (AW-DMCS) 60-120 order; Column temperature is 160 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

Concrete method of quality control is:

For infusion solution, product is the clear liquid of light brown;

For liquid drugs injection, product is the clear liquid of light brown;

Differentiate:

(2) getting freeze-dried powder 400-1000mg adds injection water 20-100ml and makes dissolving or get transfusion 200-500ml and be concentrated into 20-100ml or water intaking injection 20-100ml; Put in the separating funnel, extract 1-4 time, each 10-60ml with water saturated normal butyl alcohol; Merge extract, put evaporate to dryness in the water-bath, residue dissolves with distilled water 1-10ml; Add and make on each good DA-201 resin column internal diameter 0.5～2cm, long 10-20cm in advance; After filling the DA-201 resin of 10-15cm, recharge the 1-5g neutral alumina; 20-80% methanol solution wash-out is used in water 50-200ml washing again, collects eluent, and evaporate to dryness, residue add methyl alcohol 0.5-5ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 0.5-3mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 3-20 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=1-10: 2-15: 1-10: 1-10,10 ℃ of lower floor's solution with held are developping agent, launch; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp 200-500nm and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

The preparation of reference substance solution: it is an amount of to get the cantharidin reference substance, and accurate the title decides, and chlorination is copied into the solution that every 1ml contains 0.005-0.02mg, as reference substance solution;

Method of quality control of the present invention is the preferred plan that obtains through a large amount of shaker tests, and following experimental study is a preferred process of the present invention:

One, cantharidin content assaying method research

Cantharidin is an effective ingredient anticancer in the Aidi injection preparation; Cantharidin content assaying method two of the present invention is a method used in original standard, be with its content of gas chromatography determination, but its content assaying method all is restricted all the time; And the K-D concentrator is bought comparatively difficulty; This causes obstacle to measuring this project, and in order to address this problem, we study the methodology of cantharidin assay.

1. Tianjin, instrument island GC-14C gas chromatograph, chromatographic column: glass column 1-2m * 3mm id, the stain amount that is coated with of OV-17 (50% methyl, 50% phenyl polysiloxane) immobile liquid is 0.5-3%, carrier Shimalite W (AW-DMCS) 60-120 order; Detecting device: FID flame ionization ditector, ultrapure nitrogen (80KPa), hydrogen (60KPa), air (50KPa); Column temperature: 160 ℃, injector temperature: 240 ℃, detector temperature: 240 ℃, Weil-McLain jade for asking rain workstation; Microsyringe (10 μ l).

2. it is pure that reagent sulfuric acid, chloroform are analysis; Supply assay to provide by Nat'l Pharmaceutical & Biological Products Control Institute with the cantharidin reference substance.

3. chromatographic condition glass column 1-2m * 3mm id, the stain amount that is coated with of OV-17 (50% methyl, 50% phenyl polysiloxane) immobile liquid is 0.5-3%, carrier Shimalite W (AW-DMCS) 60～120 orders.Column temperature is 160 ± 10 ℃; Number of theoretical plate should be not less than 1500 by the cantharidin peak.

Because of used stationary phase chromatographic peak profile in the method two bad, so the stain amount that is coated with that changes stationary phase into OV-17 (50% methyl, 50% phenyl polysiloxane) immobile liquid is 0.5-3%, carrier Shimalite W (AW-DMCS) 60～120 orders.Detect the back at the chromatographic condition according to primary standard and find that sample goes out the peak at 4.07min, from the chromatogram that obtains, can find out that this main peak does not drop on the baseline place, the front also has a small peak not separate fully with it.Through test, when being reduced to 160 ℃ to column temperature, the sample separation degree is greater than 1.5, so change column temperature into 160 ℃.

4. system suitability test

Get the negative test sample liquid 4 μ l inject gas chromatographs of cantharidin reference substance, test sample liquid, scarce cantharidin respectively; Cantharidin reference substance retention time is 8.33 minutes, and negative sample did not have absorption in the time of 8 minutes, and sample is in this time degree of separation 2.16; Greater than 1.5; Theoretical cam curve 1904 is calculated with the cantharidin peak so confirm theoretical cam curve, should be not less than 1500.

5. it is an amount of that the preparation precision of reference substance solution takes by weighing the cantharidin reference substance, and chlorination is copied into the solution that every 1ml contains 0.01mg, as reference substance solution.

6. the preparation precision of need testing solution is measured these article 10ml, puts in the tool plug test tube, adds 1.8mol/L sulfuric acid solution 1ml, the accurate again 1ml chloroform that adds, and close plug, jolting 3 minutes, standing demix, centrifugal in case of necessity, get subnatant and promptly get.

7, precision test

1) precision is measured reference substance solution (C=0.01088mg/ml) 4ul, repeats sample introduction 5 times, and the result sees the following form:

Can find out that from the result reference substance repeats sample introduction 5 times, RSD% is 1.4%, explains that precision is good.

2) replica test: get same lot number medicine, handle 6 parts of test samples simultaneously by the test sample disposal route, the result sees the following form:

Can find out that from the result 6 parts of repeated sample size mean values are that 0.010945mg/ props up, RSD% is 2.2%, explains that this method repeatability is good.

8, stability test

Get this preparation, become test sample liquid by the test sample disposal methods, the result sees the following form:

Can find out that from the result 8 hours internal stabilities of this product are good.

9, accuracy test

This product accuracy test is pressed average recovery and is calculated:

Sample size is measured: precision is measured these article liquid drugs injection 10ml, gets 2 parts altogether, puts in the tool plug test tube, adds 1.8mol/L sulfuric acid solution 1ml respectively, the accurate respectively again 1ml chloroform that adds, and close plug, jolting 3 minutes, standing demix, centrifugal, get subnatant and promptly get.Precision is measured reference substance solution and each 4 μ l of need testing solution respectively again, and inject gas chromatograph calculates, and promptly gets.

Reference substance is table as a result:

Concentration	Peak area 1	Peak area 2	Mean value	RSD％
					0.01088mg/ml	348618	344274	346446	0.9

Test sample is table as a result:

Determination of recovery rates: precision is measured these article liquid drugs injection 5ml, gets 6 parts altogether, puts in the tool plug test tube, adds 1.8mol/L sulfuric acid solution 1ml respectively; Accurate respectively again adding 1ml cantharidin reference substance solution (C=0.01088mg/ml), close plug, jolting 3 minutes; Standing demix, centrifugal, get subnatant and promptly get.Precision is measured reference substance solution and each 4 μ l of need testing solution respectively again, and inject gas chromatograph calculates, and promptly gets.The result sees the following form:

Can find out that from the result average recovery rate is 100.4%, RSD% is 1.0%, explains that this method recovery is good, and accuracy is high.

10. scope is investigated

The content limit of cantharidin in the preparation according to the present invention, its content are less than 1%, and the scope of investigation is ± 50%.Get same batch sample, precision is measured 6 parts respectively, and two parts of 5ml, two parts of 10ml, two parts of 15ml become test sample liquid by the test sample disposal methods, the results are shown in following table:

Can find out that from the result 6 duplicate samples cantharidin assay mean values are that 0.01149mg/ props up, RSD is 1.2%, and the declared range test is good.

11. serviceability test

Get preparation of the present invention, become test sample liquid by the test sample disposal methods, through changing column temperature and detector temperature, the result sees the following form:

As can be seen from the above table, column temperature is bigger to the retention time influence, and detector temperature does not have influence to retention time, but change column temperature and detector temperature can both guarantee that sample size is constant basically, and this method good tolerance is described.

Can be known by the said method result of study, adopt the content assaying method of cantharidin according to the invention, accuracy is high, and test is accurate, stable, durable, and the method favorable reproducibility is the desirable content assaying method of Aidi preparation.

Two, genseng, Radix Astragali thin layer Study on Identification

With ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance differentiate genseng, Milkvetch Root in the preparation:

Need testing solution preparation method one: the article of getting it filled, put in the separating funnel, extract 2 times with water saturated normal butyl alcohol, merge extract; Put evaporate to dryness in the water-bath, residue is used dissolved in distilled water, adds on the DA-201 resin column for preparing in advance; Use water washing, use 40% methanol solution wash-out again, collect eluent; Evaporate to dryness, residue add methyl alcohol makes dissolving, as need testing solution; The negative test liquid that is equipped with shortage of staff's ginseng, the Radix Astragali with legal system.

Need testing solution preparation method two: the article of getting it filled, put in the separating funnel, extract with the methenyl choloride jolting; Discard methenyl choloride liquid, water layer extracts 2 times with water-saturated n-butanol, merges normal butyl alcohol liquid; Use the ammonia solution washed twice, discard ammonia solution, normal butyl alcohol liquid evaporate to dryness in water-bath; Residue adds methyl alcohol makes dissolving, as need testing solution; The negative test liquid that is equipped with shortage of staff's ginseng, the Radix Astragali with legal system.

Need testing solution preparation method three: the article of getting it filled, put in the separating funnel, extract with the ether jolting; Discard ether solution, water layer extracts 2 times with water-saturated n-butanol, merges normal butyl alcohol liquid; Use the ammonia solution washed twice, discard ammonia solution, normal butyl alcohol liquid evaporate to dryness in water-bath; Residue adds methyl alcohol makes dissolving, as need testing solution; The negative test liquid that is equipped with shortage of staff's ginseng, the Radix Astragali with legal system.

Developping agent is selected: respectively with the mixed solution of chloroform, ethyl acetate, first alcohol and water different proportion; The mixed solution of cyclohexane, ethyl acetate different proportion; The mixed solution of methenyl choloride, first alcohol and water different proportion; The mixed solution of ethyl acetate, acetone, water different proportion is a developping agent.

Discover, adopt method two to prepare need testing solution, on the basis of water-saturated n-butanol extraction, passed through the removal of impurities process twice, can effectively remove impurity, simple to operate, time saving and energy saving; With methenyl choloride: methyl alcohol: water is developping agent, has reduced by a component ethyl acetate on the one hand, on the other hand not need 10 ℃ with held, make simple to operateization and effective ingredient separated fully; Used thickness is the above thin layer plates of 500 μ m, and controlled humidity simultaneously reaches the purpose that ginsenoside Rg1 and Astragaloside IV effectively separate; Be directly to put under the ultraviolet lamp to inspect after the colour developing in the primary standard, but find through test of many times that thin layer plate is put under the uviol lamp after placement the preceding paragraph time (at least one hour) again and inspected behind spray developer baking spotting out, sample point is more clear.Through test of many times, adopt thin layer discrimination method favorable reproducibility of the present invention, specificity is strong, and is negative noiseless.

The result: adopt method two to prepare need testing solution, with methenyl choloride: methyl alcohol: water=10-50: 1-9: 0.5-5 is a developping agent, and its degree of separation is good; The spot colour developing is clear; Negative control is noiseless, and the method reappearance is best, and best developping agent is: methenyl choloride: methyl alcohol: water=15: 6: 1.

Three, check item confirming of safety indexes down

Improve the requirement of traditional Chinese medicine quality standard for cooperating country; Simultaneously also be the quality of watching preparation attentively, below we increase Aidi injection preparation: the inspection of projects such as protein, tannin, resin, oxalates, potassium ion, arsenic salt, haemolysis and cohesion, allergic reaction material, depressor, the material that boosts, undue toxicity, osmotic pressure for this Ai Di of better control.

1. the inspection of protein: make an experiment according to Chinese Pharmacopoeia.

Method 1: get Ai Di product liquid drugs injection 1ml, add 30% sulfosalicylic acid solution 1ml of new preparation, mixing was placed observations 5 minutes;

Method 2: get Ai Di product liquid drugs injection 1ml, add 1～3 of tannic acid test solution, observations.

Through adopting above two kinds of methods to make an experiment to 10 lot sample article, the result does not all occur muddy, so with method 1 income end product quality standard.

2. the inspection of tannin: make an experiment according to Chinese Pharmacopoeia.

Method 1: get Ai Di product liquid drugs injection 1ml, add the physiological sodium chloride solution 5ml that contains 1% Ovum Gallus domesticus album (in case of necessity, filtering) of new preparation), placed observations 10 minutes with miillpore filter (0.45um).

Method 2: get Ai Di product liquid drugs injection 1ml, add 1 of spirit of vinegar, add 4～5 of sodium chloride gelatin test solutions again, observations.

Through adopting above two kinds of methods to make an experiment to 10 lot sample article, muddy or deposition does not all appear in the result, so with method 1 income end product quality standard.

3. the inspection of resin: make an experiment according to Chinese Pharmacopoeia.

Method 1: get Ai Di product liquid drugs injection 5ml, add 1 of hydrochloric acid, placed observations 30 minutes.

Method 2: get Ai Di product liquid drugs injection 5ml, add methenyl choloride 10ml jolting and extract, obtain methenyl choloride liquid, put evaporate to dryness in the water-bath, residue adds glacial acetic acid 2ml makes dissolving, puts in the tool plug test tube, adds water 3ml, and mixing was placed observations 30 minutes.

Through adopting above two kinds of methods to make an experiment to 10 lot sample article, deposition does not all appear in the result, so with method 1 income end product quality standard.

4. the inspection of oxalates: make an experiment according to Chinese Pharmacopoeia.Get Ai Di product liquid drugs injection 2ml, regulate pH value to 1～2, filter with watery hydrochloric acid; Filtrating is regulated pH value to 5～6; Add 2～3 of 3% calcium chloride solutions, placed 10 minutes, through 10 lot sample article are made an experiment; Muddy or deposition does not all appear in the result, so include the inspection of oxalates in the end product quality standard.

5. the inspection of potassium ion: make an experiment according to Chinese Pharmacopoeia.Get Ai Di product liquid drugs injection 2ml, evaporate to dryness burns to charing with little heated earlier, 500～600 ℃ of ashing extremely fully, adds spirit of vinegar 2ml and makes dissolving again, puts in the 25ml measuring bottle, and thin up is to scale, and mixing is as need testing solution.Get two of 10ml nessler colorimetric tubes, accurate adding standard potassium ion solution 0.8ml adds alkaline formaldehyde solution 0.6ml, 2 of 3% Calcium Disodium Versenate solution, 3% sodium tetraphenylborate solution 0.5ml in the first pipe, and thin up becomes 10ml; The accurate need testing solution 1ml that adds in the second pipe with the operation in accordance with the law simultaneously of first pipe, shakes up.First, second two pipes are from up to down had an X-rayed with putting on the black paper.Through 10 lot sample article are made an experiment, test sample pipe turbidity all is shallower than standard pipe as a result, so include the potassium ion inspection in the end product quality standard.

6. arsenic salt inspection: Aidi injection preparation contains organism and possibly exist function influences such as absorption, complexing to detect arsenic salt, therefore in experiment, adds the magnesium nitrate ethanolic solution earlier these article are carried out organic destruction, makes an experiment according to Chinese Pharmacopoeia.Inspection regulation according to " annex 2: the content of quality standard and project demand " the arsenic salt of " traditional Chinese medicine research technical requirement ": " by " inspection of Chinese pharmacopoeia current edition method must not fix tentatively 2/1000000ths.", and " preparation about standard arsenic spot in the Chinese pharmacopoeia appendix IX F arsenic salt inspection technique is: taking concentration is the standard arsenic solution 2ml of 1ug/ml.Our sampling amount that can be calculated as follows sample is 1ml thus:

\frac{2 \times 10^{- 6} g / ml}{1 μg / ml \times 2 ml} = 1 ml

Concrete operation method is: get Ai Di product liquid drugs injection 1ml in crucible, evaporate to dryness adds 2% magnesium nitrate ethanolic solution 3ml; Light, in the afterburnt, burning with little heated earlier makes charing; Again 500～600 ℃ blazing to fully ashing, put coldly, add hydrochloric acid 5ml and water 21ml makes dissolving; Add 5 in the inferior tin of potassium iodide test solution 5ml and acid chlorization again, placed 10 minutes, add zinc granule 2g according to the inspection of pharmacopeia appendix arsenic salt inspection technique first method in room temperature.Finished product with 10 lot numbers makes an experiment by last method, the arsenic spot all do not occur.So include the inspection of arsenic salt in the end product quality standard.

7. haemolysis is checked with cohesion: panaxitriol and oleanolic acid are that the ginsenoside of aglycon has significant haemocylolysis; For improving the security of medicine, Aidi injection preparation is done haemolysis and cohesion inspection according to pharmacopeia appendix traditional Chinese medicine security inspection method application directs principle.Get 6 in test tube, proportional quantity adds 2% red blood cell suspension and physiological sodium chloride solution successively in the according to the form below, behind the mixing; In 37 ℃ of constant temperature ovens, placed 30 minutes, the need testing solution that adds different amounts respectively is (according to ad pro injection usage and dosage " 50～100ml that is grown up a time adds sodium chloride injection 400～450ml iv drip of 0.9% "; So select these article of getting 10ml for use; Add physiological sodium chloride solution and be diluted to 50ml, mixing is as need testing solution; The 6th pipe is for control tube), after shaking up, to put in 37 ℃ ± 0.5 ℃ constant temperature oven, beginning is every to be observed once at a distance from 15 minutes, after 1 hour, whenever at a distance from 1 h observation once, observed altogether 3 hours.

Finished product with 10 lot numbers makes an experiment by last method, haemolysis and coacervation all do not occur.So include haemolysis and cohesion inspection in the end product quality standard.

8. undue toxicity inspection, allergin inspection, the material that boosts inspection, depressor inspection: through experimental study; The result shows that the inspection of ad pro injection undue toxicity, allergin inspection, the material that boosts inspection, depressor inspection all meet the requirements, so include its detection method in quality standard.Concrete detection method is:

The undue toxicity inspection: get Aidi injection preparation, by every 0.5ml of mouse, according to the inspection of Chinese Pharmacopoeia method, should be up to specification.

The allergin inspection: getting Aidi injection preparation, with former concentration, is 0.5ml/ by the cavy priming dose, and booster dose is 1.0ml/; According to the inspection of Chinese Pharmacopoeia method, should be up to specification.

The material that boosts inspection: get Aidi injection preparation,, press the dosage of rat body weight 1kg injection 10ml with former concentration, according to the inspection of Chinese Pharmacopoeia method, should be up to specification.

The depressor inspection: get Aidi injection preparation, with former concentration, by the dosage of cat body weight 1kg injection 1ml, according to the inspection of Chinese Pharmacopoeia method, should be up to specification.

9. the inspection of osmotic pressure: fail to accomplish because of the osmotic pressure project lacks the instrument and equipment that detects.

Show that through the safety indexes result security of these article is reliable; Development test shows that each related substance inspection also is feasible.So list protein, tannin, resin, oxalates, potassium ion, arsenic salt, haemolysis and cohesion, undue toxicity inspection, allergic reaction material, depressor, the material item that boosts in quality standard; And osmotic pressure for want of instrument and equipment fail to accomplish, so fail to include osmotic pressure in quality standard.

Compared with prior art; Method of quality control of the present invention provides the alpha-Naphthol chromogenic reaction, serve as that contrast is differentiated the thin-layer chromatography of the genseng in the preparation and the Radix Astragali with ginsenoside Re, Rg1, Rb and Astragaloside IV, to the content assaying method of contained ginsenoside Re and cantharidin in the preparation; And to the inspection of safety indexes such as pH value, heavy metal, pyrogen, protein, tannin; Above method of quality control can effectively be controlled the quality of Aidi injection preparation, thereby guarantees its clinical efficacy.

Embodiment: followingly further specify the present invention through embodiment, but not as limitation of the present invention.

The embodiment of the invention 1:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(1) getting freeze-dried powder 1000mg adds injection water 20ml and makes dissolving or get that transfusion 500ml is concentrated into 20ml or water intaking injection 100ml is concentrated into 20ml; Get above-mentioned medicine respectively, add ethanol 20ml, stir; Filter, sediment dissolves sediment with washing with alcohol 1 time with distilled water 10ml; Get 0.5ml and put in the test tube, add 2 of 2% alpha-Naphthol ethanolic solutions after the low-grade fever, shake up; Slowly add concentrated sulphuric acid 2ml along tube wall again, at two liquid junction place displaing amaranth rings;

(2) getting freeze-dried powder 1000mg adds injection water 100ml and makes dissolving or get transfusion 500ml and be concentrated into 100ml or water intaking injection 100ml; Put in the separating funnel, extract, discard methenyl choloride liquid with the jolting of 50ml methenyl choloride; Water layer extracts 1 time with water-saturated n-butanol, and each 60ml merges normal butyl alcohol liquid; With ammonia solution washing 4 times, each 10ml discards ammonia solution; Normal butyl alcohol liquid evaporate to dryness in water-bath, residue add methyl alcohol 3ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb ₁And the Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 3mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 3 μ l of above-mentioned two kinds of solution, put on same silica gel g thin-layer plate thick 800 μ m respectively; With methenyl choloride: methyl alcohol: water=10: 9: 5 is developping agent, and controlled humidity launches where necessary, takes out, and dries, and spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating several minutes, puts under the ultraviolet lamp 500nm after placing more than 1 hour again and inspects; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Heavy metal: get freeze-dried powder 40mg and add injection water 2ml and make dissolving or get transfusion 20ml and be concentrated into 2ml or water intaking injection 2ml; Put in the crucible, evaporate to dryness in the water-bath according to the inspection of Chinese Pharmacopoeia heavy metal inspection technique, contains heavy metal and must not cross 5/1000000ths;

Pyrogen: get freeze-dried powder 300mg and add injection water 30ml and make dissolving or get transfusion 100ml and be concentrated into 30ml or water intaking injection 30ml; Be diluted to 100ml with sodium chloride injection or 5～10% glucose injections, by the dosage of rabbit body weight 1kg injection 10ml, according to the inspection of Chinese Pharmacopoeia pyrogen test, should be up to specification;

Assay:

(1) ginsenoside Re

The preparation of reference substance solution: precision takes by weighing ginsenoside Re's reference substance, puts in the measuring bottle, adds dissolve with methanol and makes and contain ginsenoside Re 1mg among every 1ml, shakes up, and promptly gets;

The preparation of typical curve: precision is measured reference substance solution 0,20,40,60,80,100 μ l, puts respectively in the 10ml tool plug test tube, puts the water-bath Back stroke and desolvates, and takes out immediately; Put coldly, add 10% vanillic aldehyde glacial acetic acid solution 0.05ml, perchloric acid 0.5ml, shake up; Put in 80 ℃ of water-baths and heated 8 minutes, take out, with water cooling 1 minute; Add glacial acetic acid 2ml, shake up, according to the Chinese Pharmacopoeia AAS; Measuring absorbance log in the 700nm wavelength, is that ordinate, concentration are horizontal ordinate with the absorbance log, the drawing standard curve;

The preparation of need testing solution: get freeze-dried powder 400mg and add injection water 10ml and make dissolving or get transfusion 200ml and be concentrated into 10ml or water intaking injection 10ml; Precision is measured above-mentioned medicine respectively, put in the separating funnel, and with chloroform extraction 1 time, each 30ml, combined chloroform extract; With distilled water washing 1 time, each 2ml discards chloroform solution, and washing lotion and above-mentioned water layer merge, and put in the separating funnel; Extract 5 times with water saturated normal butyl alcohol, each 40ml merges n-butanol extracting liquid, adds anhydrous sodium sulfate 5g, stirs; Put to clarification, normal butyl alcohol liquid is moved in the evaporating dish, with a small amount of normal butyl alcohol washing anhydrous sodium sulfate, washing lotion is incorporated in the evaporating dish, evaporate to dryness; With a small amount of dissolved in distilled water residue, be added on the macroporous absorbent resin DA-201 post, treat that liquid level is near cotton layer after, with distilled water 100ml wash-out, the control flow velocity is 0.6ml/min; Discard washing lotion,, collect eluent in evaporating dish, put evaporate to dryness in the water-bath, use the dissolve with methanol residue with 90% ethanol 100ml wash-out; Move in the 5ml measuring bottle, add methyl alcohol, shake up, promptly get to scale;

Chromatographic condition and system suitability test: with carbowax-20M and methyl silicone rubber SE-30 is immobile liquid, and coating concentration is respectively 20% and 10%, 2: 0.5 mixes the dress post; Column temperature is 180 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and chlorination is copied into the solution that every 1ml contains 0.2mg, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder 1000mg and add injection water 20ml and make dissolving or get transfusion 200ml and be concentrated into 20ml or water intaking injection 20ml; Add 3mol/L sulfuric acid solution 10ml; Extract 1 time each 100ml, combined chloroform liquid with the chloroform jolting; Be settled to 8ml with the K-D concentrator is concentrated, promptly get;

The embodiment of the invention 2:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(1) getting freeze-dried powder 400mg adds injection water 5ml and makes dissolving or get that transfusion 200ml is concentrated into 5ml or water intaking injection 20ml is concentrated into 5ml; Get above-mentioned medicine respectively, add ethanol 100ml, stir; Filter, sediment dissolves sediment with washing with alcohol 4 times with distilled water 2ml; Get 3ml and put in the test tube, add 10 of 10% alpha-Naphthol ethanolic solutions after the low-grade fever, shake up; Slowly add concentrated sulphuric acid 0.2ml along tube wall again, at two liquid junction place displaing amaranth rings;

(2) getting freeze-dried powder 400mg adds injection water 20ml and makes dissolving or get transfusion 200ml and be concentrated into 20ml or water intaking injection 20ml; Put in the separating funnel, extract, discard methenyl choloride liquid with the jolting of 10ml methenyl choloride; Water layer extracts 4 times with water-saturated n-butanol, and each 40ml merges normal butyl alcohol liquid; With ammonia solution washing 1 time, each 50ml discards ammonia solution; Normal butyl alcohol liquid evaporate to dryness in water-bath, residue add methyl alcohol 0.5ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb ₁And the Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 0.5mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 20 μ l of above-mentioned two kinds of solution, put on same silica gel g thin-layer plate thick 1000 μ m respectively; With methenyl choloride: methyl alcohol: water=50: 1: 0.5 is developping agent, and controlled humidity launches where necessary, takes out, and dries, and spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating several minutes, puts under the ultraviolet lamp 200nm after placing more than 1 hour again and inspects; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Heavy metal: get freeze-dried powder 20mg and add injection water 2ml and make dissolving or get transfusion 8ml and be concentrated into 2ml or water intaking injection 2ml; Put in the crucible, evaporate to dryness in the water-bath according to the inspection of Chinese Pharmacopoeia heavy metal inspection technique, contains heavy metal and must not cross 5/1000000ths;

Pyrogen: get freeze-dried powder 600mg and add injection water 30ml and make dissolving or get transfusion 300ml and be concentrated into 30ml or water intaking injection 30ml; Be diluted to 100ml with sodium chloride injection or 5～10% glucose injections, by the dosage of rabbit body weight 1kg injection 10ml, according to the inspection of Chinese Pharmacopoeia pyrogen test, should be up to specification;

Assay:

(1) ginsenoside Re

The preparation of reference substance solution: precision takes by weighing ginsenoside Re's reference substance, puts in the measuring bottle, adds dissolve with methanol and makes and contain ginsenoside Re 2mg among every 1ml, shakes up, and promptly gets;

The preparation of typical curve: precision is measured reference substance solution 0,20,40,60,80,100 μ l, puts respectively in the 10ml tool plug test tube, puts the water-bath Back stroke and desolvates, and takes out immediately; Put coldly, add 2% vanillic aldehyde glacial acetic acid solution 1ml, perchloric acid 1.5ml, shake up; Put in 40 ℃ of water-baths and heated 25 minutes, take out, with water cooling 3 minutes; Add glacial acetic acid 10ml, shake up, according to the Chinese Pharmacopoeia AAS; Measuring absorbance log in the 400nm wavelength, is that ordinate, concentration are horizontal ordinate with the absorbance log, the drawing standard curve;

The preparation of need testing solution: get freeze-dried powder 200mg and add injection water 30ml and make dissolving or get transfusion 50ml and be concentrated into 30ml or water intaking injection 30ml; Precision is measured above-mentioned medicine respectively, put in the separating funnel, and with chloroform extraction 5 times, each 10ml, combined chloroform extract; With distilled water washing 4 times, each 10ml discards chloroform solution, and washing lotion and above-mentioned water layer merge, and put in the separating funnel; Extract 1 time with water saturated normal butyl alcohol, each 60ml merges n-butanol extracting liquid, adds anhydrous sodium sulfate 1g, stirs; Put to clarification, normal butyl alcohol liquid is moved in the evaporating dish, with a small amount of normal butyl alcohol washing anhydrous sodium sulfate, washing lotion is incorporated in the evaporating dish, evaporate to dryness; With a small amount of dissolved in distilled water residue, be added on the macroporous absorbent resin DA-201 post, treat that liquid level is near cotton layer after, with distilled water 100ml wash-out, the control flow velocity is 0.3ml/min; Discard washing lotion,, collect eluent in evaporating dish, put evaporate to dryness in the water-bath, use the dissolve with methanol residue with 40% ethanol 70ml wash-out; Move in the 5ml measuring bottle, add methyl alcohol, shake up, promptly get to scale;

Chromatographic condition and system suitability test: with carbowax-20M and methyl silicone rubber SE-30 is immobile liquid, and coating concentration is respectively 5% and 10%, 0.5: 0.5 mixes the dress post; Column temperature is 180 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and chlorination is copied into the solution that every 1ml contains 0.05mg, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder 1000mg and add injection water 100ml and make dissolving or get transfusion 500ml and be concentrated into 100ml or water intaking injection 100ml; Add 1mol/L sulfuric acid solution 2ml; Extract 5 times each 10ml, combined chloroform liquid with the chloroform jolting; Be settled to 2ml with the K-D concentrator is concentrated, promptly get;

The embodiment of the invention 3:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(1) getting freeze-dried powder 750mg adds injection water 10ml and makes dissolving or get that transfusion 400ml is concentrated into 10ml or water intaking injection 50ml is concentrated into 10ml; Get above-mentioned medicine respectively, add ethanol 50ml, stir; Filter, sediment dissolves sediment with washing with alcohol 2 times with distilled water 5ml; Get 1ml and put in the test tube, add 5 of 5% alpha-Naphthol ethanolic solutions after the low-grade fever, shake up; Slowly add concentrated sulphuric acid 0.5ml along tube wall again, at two liquid junction place displaing amaranth rings;

(2) getting freeze-dried powder 750mg adds injection water 5ml and makes dissolving or get transfusion 400ml and be concentrated into 500ml or water intaking injection 50ml; Put in the separating funnel, extract, discard methenyl choloride liquid with the jolting of 30ml methenyl choloride; Water layer extracts 2 times with water-saturated n-butanol, and each 50ml merges normal butyl alcohol liquid; With ammonia solution washing 2 times, each 30ml discards ammonia solution; Normal butyl alcohol liquid evaporate to dryness in water-bath, residue add methyl alcohol 1ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb ₁And the Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 1mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 5-10 μ l of above-mentioned two kinds of solution, put on same silica gel g thin-layer plate thick 500 μ m respectively; With methenyl choloride: methyl alcohol: water=15: 6: 1 is developping agent, and controlled humidity launches where necessary, takes out, and dries, and spray is with 10% ethanol solution of sulfuric acid, 105 ℃ of heating several minutes, puts under the ultraviolet lamp 365nm after placing more than 1 hour again and inspects; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Heavy metal: get freeze-dried powder 30mg and add injection water 2ml and make dissolving or get transfusion 10ml and be concentrated into 2ml or water intaking injection 2ml; Put in the crucible, evaporate to dryness in the water-bath according to the inspection of Chinese Pharmacopoeia heavy metal inspection technique, contains heavy metal and must not cross 5/1000000ths;

Pyrogen: get freeze-dried powder 500mg and add injection water 30ml and make dissolving or get transfusion 200ml and be concentrated into 30ml or water intaking injection 30ml; Be diluted to 100ml with sodium chloride injection or 5～10% glucose injections, by the dosage of rabbit body weight 1kg injection 10ml, according to the inspection of Chinese Pharmacopoeia pyrogen test, should be up to specification;

Assay:

(1) ginsenoside Re

The preparation of reference substance solution: precision takes by weighing ginsenoside Re's reference substance, puts in the measuring bottle, adds dissolve with methanol and makes and contain ginsenoside Re 1.6mg among every 1ml, shakes up, and promptly gets;

The preparation of typical curve: precision is measured reference substance solution 0,20,40,60,80,100 μ l, puts respectively in the 10ml tool plug test tube, puts the water-bath Back stroke and desolvates, and takes out immediately; Put coldly, add 5% vanillic aldehyde glacial acetic acid solution 0.2ml, perchloric acid 0.8ml, shake up; Put in 60 ℃ of water-baths and heated 15 minutes, take out, with water cooling 2 minutes; Add glacial acetic acid 5ml, shake up, according to the Chinese Pharmacopoeia AAS; Measuring absorbance log in the 544nm wavelength, is that ordinate, concentration are horizontal ordinate with the absorbance log, the drawing standard curve;

The preparation of need testing solution: get freeze-dried powder 300mg and add injection water 20ml and make dissolving or get transfusion 100ml and be concentrated into 20ml or water intaking injection 20ml; Precision is measured above-mentioned medicine respectively, put in the separating funnel, and with chloroform extraction 3 times, each 20ml, combined chloroform extract; With distilled water washing 2 times, each 5ml discards chloroform solution, and washing lotion and above-mentioned water layer merge, and put in the separating funnel; Extract 3 times with water saturated normal butyl alcohol, each 50ml merges n-butanol extracting liquid, adds anhydrous sodium sulfate 3g, stirs; Put to clarification, normal butyl alcohol liquid is moved in the evaporating dish, with a small amount of normal butyl alcohol washing anhydrous sodium sulfate, washing lotion is incorporated in the evaporating dish, evaporate to dryness; With a small amount of dissolved in distilled water residue, be added on the macroporous absorbent resin DA-201 post, treat that liquid level is near cotton layer after, with distilled water 100ml wash-out, the control flow velocity is 0.4ml/min; Discard washing lotion,, collect eluent in evaporating dish, put evaporate to dryness in the water-bath, use the dissolve with methanol residue with 70% ethanol 80ml wash-out; Move in the 5ml measuring bottle, add methyl alcohol, shake up, promptly get to scale;

Chromatographic condition and system suitability test: with carbowax-20M and methyl silicone rubber SE-30 is immobile liquid, and coating concentration is respectively 10% and 5%, 1: 1 mixes the dress post; Column temperature is 180 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and chlorination is copied into the solution that every 1ml contains 0.1mg, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder 800mg and add injection water 50ml and make dissolving or get transfusion 450ml and be concentrated into 50ml or water intaking injection 50ml; Add 1.8mol/L sulfuric acid solution 5ml; Extract 3 times with the chloroform jolting, be respectively 50ml, 30ml, 30ml, combined chloroform liquid; Be settled to 5ml with the K-D concentrator is concentrated, promptly get;

Determination method: precision is measured reference substance solution and each 5 μ l of need testing solution respectively, and inject gas chromatograph calculates, and promptly gets; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml.

The embodiment of the invention 4:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(2) getting freeze-dried powder 1000mg adds injection water 100ml and makes dissolving or get transfusion 500ml and be concentrated into 100ml or water intaking injection 100ml; Put in the separating funnel, extract 1 time, each 60ml with water saturated normal butyl alcohol; Merge extract, put evaporate to dryness in the water-bath, residue dissolves with distilled water 10ml; Add on the DA-201 resin column for preparing in advance internal diameter 0.5～2cm, long 10cm; After filling the DA-201 resin of 10cm, recharge the 5g neutral alumina; 80% methanol solution wash-out is used in water 50ml washing again, collects eluent, and evaporate to dryness, residue add methyl alcohol 5ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 0.5mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 3 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=1: 15: 10: 1,10 ℃ of lower floor's solution with held is developping agent, launches; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp 200nm and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Protein: get freeze-dried powder 20mg and add injection water 1ml and make dissolving or get transfusion 10ml and be concentrated into 1ml or water intaking injection 1ml; The 30% sulfosalicylic acid solution 1ml that adds new preparation, mixing was placed 5 minutes, muddiness must not occur;

Tannin: get freeze-dried powder 20mg and add injection water 1ml and make dissolving or get transfusion 10ml and be concentrated into 1ml or water intaking injection 1ml; The physiological sodium chloride solution 5ml that contains 1% Ovum Gallus domesticus album that adds new preparation in case of necessity, filters with the 0.45um miillpore filter, places 10 minutes, muddiness or deposition must not occur;

Resin: get freeze-dried powder 100mg and add injection water 5ml and make dissolving or get transfusion 50ml and be concentrated into 5ml or water intaking injection 5ml; Add 1 of hydrochloric acid, placed 30 minutes, deposition must not occur;

Oxalates: get freeze-dried powder 40mg and add injection water 2ml and make dissolving or get transfusion 20ml and be concentrated into 2ml or water intaking injection 2ml; Regulate pH value to 1-2 with watery hydrochloric acid, filter, filtrating is regulated pH value to 5-6, adds 3% calcium chloride solution 2-3 and drips, and places 10 minutes, muddiness or deposition must not occur;

Potassium ion: get freeze-dried powder 40mg and add injection water 2ml and make dissolving or get transfusion 20ml and be concentrated into 2ml or water intaking injection 2ml; Evaporate to dryness is measured according to Chinese Pharmacopoeia potassium ion determination method, should be up to specification;

Arsenic salt: get freeze-dried powder 20mg and add injection water 1ml and make dissolving or get transfusion 10ml and be concentrated into 1ml or water intaking injection 1ml; Precision is measured, and evaporate to dryness in crucible adds 2% magnesium nitrate ethanolic solution 3ml, lights; In the afterburnt, burning with little heated earlier makes charing, blazing to ashing fully at 500-600 ℃ again; Put coldly, add hydrochloric acid 5ml and water 21ml makes dissolving, according to the inspection of Chinese Pharmacopoeia arsenic salt inspection technique; Arsenic content must not cross 2/1000000ths in every 1ml finished product;

The preparation of need testing solution: get freeze-dried powder 200mg and add injection water 10ml and make dissolving or get transfusion 100ml and be concentrated into 10ml or water intaking injection 10ml; Add physiological sodium chloride solution and be diluted to 50ml, mixing promptly gets;

Assay:

(1) ginsenoside Re

The preparation of need testing solution: get freeze-dried powder 400mg and add injection water 10ml and make dissolving or get transfusion 200ml and be concentrated into 10ml or water intaking injection 10ml; Precision is measured above-mentioned medicine respectively, put in the separating funnel, and with chloroform extraction 1 time, each 30ml, combined chloroform extract; With distilled water washing 1 time, each 2ml discards chloroform solution, and washing lotion and above-mentioned water layer merge, and put in the separating funnel; Extract 5 times with water saturated normal butyl alcohol, each 40ml merges n-butanol extracting liquid, adds anhydrous sodium sulfate 5g, stirs; Put to clarification, normal butyl alcohol liquid is moved in the evaporating dish, with a small amount of normal butyl alcohol washing anhydrous sodium sulfate, washing lotion is incorporated in the evaporating dish, evaporate to dryness; With a small amount of dissolved in distilled water residue, be added on the macroporous absorbent resin DA-201 post, treat that liquid level is near cotton layer after, with distilled water 100ml wash-out, the control flow velocity is 0.6ml/min; Discard washing lotion,, collect in the eluent hand evaporating dish, put evaporate to dryness in the water-bath, use the dissolve with methanol residue with 90% ethanol 100ml wash-out; Move in the 5ml measuring bottle, add methyl alcohol, shake up, promptly get to scale;

The embodiment of the invention 5:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(2) getting freeze-dried powder 400mg adds injection water 20ml and makes dissolving or get transfusion 200ml and be concentrated into 20ml or water intaking injection 20ml; Put in the separating funnel, extract 4 times, each 10ml with water saturated normal butyl alcohol; Merge extract, put evaporate to dryness in the water-bath, residue dissolves with distilled water 1ml; Add on the DA-201 resin column for preparing in advance internal diameter 0.5～2cm, long 20cm; After filling the DA-201 resin of 15cm, recharge the 1g neutral alumina; 20% methanol solution wash-out is used in water 200ml washing again, collects eluent, and evaporate to dryness, residue add methyl alcohol 0.5ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 3mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 20 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=10: 2: 1: 10,10 ℃ of lower floor's solution with held are developping agent, launch; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp 500nm and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Protein: get freeze-dried powder 10mg and add injection water 1ml and make dissolving or get transfusion 4ml and be concentrated into 1ml or water intaking injection 1ml; The 30% sulfosalicylic acid solution 1ml that adds new preparation, mixing was placed 5 minutes, muddiness must not occur;

Tannin: get freeze-dried powder 10mg and add injection water 1ml and make dissolving or get transfusion 4ml and be concentrated into 1ml or water intaking injection 1ml; The physiological sodium chloride solution 5ml that contains 1% Ovum Gallus domesticus album that adds new preparation in case of necessity, filters with the 0.45um miillpore filter, places 10 minutes, muddiness or deposition must not occur;

Resin: get freeze-dried powder 50mg and add injection water 5ml and make dissolving or get transfusion 20ml and be concentrated into 5ml or water intaking injection 5ml; Add 1 of hydrochloric acid, placed 30 minutes, deposition must not occur;

Oxalates: get freeze-dried powder 20mg and add injection water 2ml and make dissolving or get transfusion 8ml and be concentrated into 2ml or water intaking injection 2ml; Regulate pH value to 1-2 with watery hydrochloric acid, filter, filtrating is regulated pH value to 5-6, adds 3% calcium chloride solution 2-3 and drips, and places 10 minutes, muddiness or deposition must not occur;

Potassium ion: get freeze-dried powder 20mg and add injection water 2ml and make dissolving or get transfusion 8ml and be concentrated into 2ml or water intaking injection 2ml; Evaporate to dryness is measured according to Chinese Pharmacopoeia potassium ion determination method, should be up to specification;

Arsenic salt: get freeze-dried powder 10mg and add injection water 1ml and make dissolving or get transfusion 4ml and be concentrated into 1ml or water intaking injection 1ml; Precision is measured, and evaporate to dryness in crucible adds 2% magnesium nitrate ethanolic solution 3ml, lights; In the afterburnt, burning with little heated earlier makes charing, blazing to ashing fully at 500-600 ℃ again; Put coldly, add hydrochloric acid 5ml and water 21ml makes dissolving, according to the inspection of Chinese Pharmacopoeia arsenic salt inspection technique; Arsenic content must not cross 2/1000000ths in every 1ml finished product;

The preparation of need testing solution: get freeze-dried powder 100mg and add injection water 10ml and make dissolving or get transfusion 40ml and be concentrated into 10ml or water intaking injection 10ml; Add physiological sodium chloride solution and be diluted to 50ml, mixing promptly gets;

Assay:

(1) ginsenoside Re

The embodiment of the invention 6:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(2) getting freeze-dried powder 700mg adds injection water 50ml and makes dissolving or get transfusion 400ml and be concentrated into 50ml or water intaking injection 50ml; Put in the separating funnel, extract 2 times, each 30ml with water saturated normal butyl alcohol; Merge extract, put evaporate to dryness in the water-bath, residue dissolves with distilled water 3ml; Add on the DA-201 resin column for preparing in advance internal diameter 1～1.5cm, long 15cm; After filling the DA-201 resin of 12cm, recharge the 2g neutral alumina; 40% methanol solution 50ml wash-out is used in water 100ml washing again, collects eluent, and evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 1mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=4: 8: 3: 4,10 ℃ of lower floor's solution with held are developping agent, launch; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp 365nm and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color;

Inspection:

PH value: should be 3.0-6.5;

Protein: get freeze-dried powder 15mg and add injection water 1ml and make dissolving or get transfusion 6ml and be concentrated into 1ml or water intaking injection 1ml; The 30% sulfosalicylic acid solution 1ml that adds new preparation, mixing was placed 5 minutes, muddiness must not occur;

Tannin: get freeze-dried powder 15mg and add injection water 1ml and make dissolving or get transfusion 8ml and be concentrated into 1ml or water intaking injection 1ml; The physiological sodium chloride solution 5ml that contains 1% Ovum Gallus domesticus album that adds new preparation in case of necessity, filters with the 0.45um miillpore filter, places 10 minutes, muddiness or deposition must not occur;

Resin: get freeze-dried powder 80mg and add injection water 5ml and make dissolving or get transfusion 40ml and be concentrated into 5ml or water intaking injection 5ml; Add 1 of hydrochloric acid, placed 30 minutes, deposition must not occur;

Oxalates: get freeze-dried powder 30mg and add injection water 2ml and make dissolving or get transfusion 10ml and be concentrated into 2ml or water intaking injection 2ml; Regulate pH value to 1-2 with watery hydrochloric acid, filter, filtrating is regulated pH value to 5-6, adds 3% calcium chloride solution 2-3 and drips, and places 10 minutes, muddiness or deposition must not occur;

Potassium ion: get freeze-dried powder 30mg and add injection water 2ml and make dissolving or get transfusion 15ml and be concentrated into 2ml or water intaking injection 2ml; Evaporate to dryness is measured according to Chinese Pharmacopoeia potassium ion determination method, should be up to specification;

Arsenic salt: get freeze-dried powder 15mg and add injection water 1ml and make dissolving or get transfusion 8ml and be concentrated into 1ml or water intaking injection 1ml; Precision is measured, and evaporate to dryness in crucible adds 2% magnesium nitrate ethanolic solution 3ml, lights; In the afterburnt, burning with little heated earlier makes charing, blazing to ashing fully at 500-600 ℃ again; Put coldly, add hydrochloric acid 5ml and water 21ml makes dissolving, according to the inspection of Chinese Pharmacopoeia arsenic salt inspection technique; Arsenic content must not cross 2/1000000ths in every 1ml finished product;

The preparation of need testing solution: get freeze-dried powder 150mg and add injection water 10ml and make dissolving or get transfusion 80ml and be concentrated into 10ml or water intaking injection 10ml; Add physiological sodium chloride solution and be diluted to 50ml, mixing promptly gets;

The material that boosts inspection: get freeze-dried powder and add the injection water and just dissolve or get transfusion or water intaking injection; With former concentration, press the dosage of rat body weight 1kg injection 10ml, according to the Chinese Pharmacopoeia material inspection technique inspection of boosting, should be up to specification;

Assay:

(1) ginsenoside Re

The embodiment of the invention 7:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

Chromatographic condition and system suitability test: glass column 1-2m * 3mm id is an immobile liquid with OV-17, and OV-17 is made up of 50% methyl, 50% phenyl polysiloxane, and being coated with the stain amount is 3%, carrier Shimalite W (AW-DMCS) 120 orders; Column temperature is 160 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of reference substance solution: it is an amount of to get the cantharidin reference substance, and accurate the title decides, and chlorination is copied into the solution that every 1ml contains 0.02mg, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder 100g and add injection water 20ml and make dissolving or get that transfusion 100ml is concentrated into about 20ml or the injection 20ml that fetches water; Precision is measured above-mentioned medicine respectively, adds 3mol/L sulfuric acid solution 2ml, the accurate again 2ml chloroform that adds, and jolting is extracted, and standing demix is centrifugal in case of necessity, gets subnatant and promptly gets;

Determination method: precision is measured reference substance solution and each 2 μ l of need testing solution respectively, and inject gas chromatograph calculates; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml;

The embodiment of the invention 8:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

Chromatographic condition and system suitability test: glass column 1-2m * 3mm id is an immobile liquid with OV-17, and OV-17 is made up of 50% methyl, 50% phenyl polysiloxane, and being coated with the stain amount is 0.5%, carrier Shimalite W (AW-DMCS) 60 orders; Column temperature is 160 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of reference substance solution: it is an amount of to get the cantharidin reference substance, and accurate the title decides, and chlorination is copied into the solution that every 1ml contains 0.005mg, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder 30g and add injection water 50ml and make dissolving or get that transfusion 30ml is concentrated into about 5ml or the injection 5ml that fetches water; Precision is measured above-mentioned medicine respectively, adds 1mol/L sulfuric acid solution 0.5ml, the accurate again 0.5ml chloroform that adds, and jolting is extracted, and standing demix is centrifugal in case of necessity, gets subnatant and promptly gets;

Determination method: precision is measured reference substance solution and each 12 μ l of need testing solution respectively, and inject gas chromatograph calculates; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml;

The embodiment of the invention 9:

For infusion solution, product is light yellow clear liquid to light brown;

Differentiate:

(2) getting freeze-dried powder 700mg adds injection water 50ml and makes dissolving or get transfusion 400ml and be concentrated into 50ml or water intaking injection 50ml; Put in the separating funnel, extract 2 times, each 30ml with water saturated normal butyl alcohol; Merge extract, put evaporate to dryness in the water-bath, residue dissolves with distilled water 3ml; Add on the DA-201 resin column for preparing in advance internal diameter 1～1.5cm, long 15cm; After filling the DA-201 resin of 12cm, recharge the 2g neutral alumina; 40% methanol solution 50ml wash-out is used in water 100ml washing again, collects eluent, and evaporate to dryness, residue add methyl alcohol 1ml makes dissolving, as need testing solution; Other gets ginsenoside Re, Rg ₁, Rb and Astragaloside IV reference substance, add methyl alcohol and process the mixed solution that every 1ml contains 1mg, as reference substance solution; According to appendix thin-layered chromatography test of Chinese Pharmacopoeia version in 2005, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate; With chloroform: ethyl acetate: methyl alcohol: water=4: 8: 3: 4,10 ℃ of lower floor's solution with held are developping agent, launch; Take out, dry, spray is with 10% ethanol solution of sulfuric acid; 105 ℃ of heating several minutes, put under the ultraviolet lamp 365nm and inspect; In the test sample chromatogram, with the corresponding position of reference substance chromatogram on, show the fluorescence spot of same color; Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

Chromatographic condition and system suitability test: glass column 1-2m * 3mm id is an immobile liquid with OV-17, and OV-17 is made up of 50% methyl, 50% phenyl polysiloxane, and being coated with the stain amount is 1.5%, carrier Shimalite W (AW-DMCS) 80-100 order; Column temperature is 160 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of reference substance solution: get the cantharidin reference substance, the accurate title, decide, and chlorination is copied into the solution that every 1ml contains 0.01mg, as reference substance solution;

The preparation of need testing solution: get freeze-dried powder 70g and add injection water 10ml and make dissolving or get that transfusion 80ml is concentrated into about 10ml or the injection 10ml that fetches water; Precision is measured above-mentioned medicine respectively, adds 1.8mol/L sulfuric acid solution 1ml, the accurate again 1ml chloroform that adds, and jolting is extracted, and standing demix is centrifugal in case of necessity, gets subnatant and promptly gets;

Determination method: precision is measured reference substance solution and each 4 μ l of need testing solution respectively, and inject gas chromatograph calculates; In this ejection preparation, containing cantharidin in the freeze drying powder injection is 0.0005%-0.05%; Containing cantharidin in the infusion solution is 0.00001-0.001mg/ml; Containing cantharidin in the liquid drugs injection is 0.0001-0.005mg/ml.

Claims

1. the detection method of an Aidi injection preparation is calculated according to composition by weight, and Aidi injection preparation mainly is prepared from Chinese blister beetle 0.5-5, genseng 20-80, Radix Astragali 200-600, wilsonii 100-200; Its preparation method is: above four flavors, and the genseng section is extracted 1-5 time with the 20-85% alcohol heating reflux; Merge extract; Filter, reclaim ethanol, soup is subsequent use; Three flavor boilings such as the dregs of a decoction and all the other Chinese blister beetles 1-6 time, collecting decoction filters; Filtrating merges with Ginseng extract, and with stone sulphur method precipitation process 1-3 time, the gained supernatant adds ethanol to be made and contain alcohol and measure and reach 50-99%; Hold over night is got the supernatant decompression recycling ethanol to there not being the alcohol flavor, processes freeze drying powder injection, infusion solution or liquid drugs injection respectively according to conventional method again; It is characterized in that: said detection method is: proterties: for freeze drying powder injection, product is the loose block of light brown;

For infusion solution, product is the clear liquid of light brown;

For liquid drugs injection, product is the clear liquid of light brown;

Differentiate:

Inspection:

PH value: should be 3.0-6.5;

Pyrogen: get freeze-dried powder 300-600mg and add injection water 30ml and make dissolving or get transfusion 100-300ml and be concentrated into 30ml or water intaking injection 30ml; Be diluted to 100ml with sodium chloride injection or 5～10% glucose injections, by the dosage of rabbit body weight 1kg injection 10ml, according to the inspection of Chinese Pharmacopoeia pyrogen test, should be up to specification:

Assay:

(1) ginsenoside Re

Chromatographic condition and system suitability test: glass column 1-2m * 3mm i d is an immobile liquid with OV-17, and OV-17 is made up of 50% methyl, 50% phenyl polysiloxane, and being coated with the stain amount is 0.5-3%, carrier Shimalite W AW-DMCS 60-120 order; Column temperature is 160 ± 10 ℃; Number of theoretical plate is pressed the cantharidin peak and is calculated, and should be not less than 1500;

The preparation of need testing solution: get freeze-dried powder respectively and add the injection water and make dissolving or get transfusion and concentrate or the injection of directly fetching water, add the 1-3mol/L sulfuric acid solution, precision adds people's chloroform again, and jolting is extracted, and standing demix is centrifugal, gets subnatant and promptly gets;

2. according to the detection method of the said Aidi injection preparation of claim 1, it is characterized in that: concrete detection method is: proterties: for freeze drying powder injection, product is the loose block of light brown;

For infusion solution, product is the clear liquid of light brown;

For liquid drugs injection, product is the clear liquid of light brown;

Differentiate:

Inspection:

PH value: should be 3.0-6.5;

Assay:

(1) ginsenoside Re

The preparation of reference substance solution: precision takes by weighing ginsenoside Re's reference substance, puts in the measuring bottle, adds dissolve with methanol and makes and contain ginsenoside Re 1-2mg among every 1ml, shakes up, and promptly gets:

The preparation of need testing solution: get freeze-dried powder 30-100g and add injection water 5-20ml and make dissolving or get that transfusion 30-100ml is concentrated into about 5-20ml or the injection 5-20ml that fetches water; Precision is measured above-mentioned medicine respectively, adds 1-3mol/L sulfuric acid solution 0.5-2ml, the accurate again 0.5-2ml chloroform that adds, and jolting is extracted, and standing demix is centrifugal, gets subnatant and promptly gets;