CN1899400A - Medicinal preparation containing ginseng and aconite root in raw material and its preparing method and quality control method - Google Patents

Medicinal preparation containing ginseng and aconite root in raw material and its preparing method and quality control method Download PDF

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CN1899400A
CN1899400A CN 200610103234 CN200610103234A CN1899400A CN 1899400 A CN1899400 A CN 1899400A CN 200610103234 CN200610103234 CN 200610103234 CN 200610103234 A CN200610103234 A CN 200610103234A CN 1899400 A CN1899400 A CN 1899400A
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preparation
solution
injection
water
liquid
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CN1899400B (en
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孙明珍
艾劼
刘路
瞿冰
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Shenzhen Zifu Pharmaceutical Co., Ltd.
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SHENZHEN ZIFU PHARMACEUTICAL CO Ltd
XUANHONG MEDICINE TECHNOLOGY Co Ltd TIANJIN
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Abstract

The present invention relates to a kind of medicine preparation with ginseng component and aconite root component and its preparation process and quality control method. Of the medicine preparation, each unit contains total ginsenoside in 1-90 mg, total ginseng polysaccharide in 2-282 mg, and total aconite root alkaloid in 0.02-22 mg. The medicine preparation may have other synergistic medicine component added. Its preparation process includes extracting the total ginsenoside, the total ginseng polysaccharide and the total aconite root alkaloid; adding supplementary material; and forming different preparation forms.

Description

The pharmaceutical preparation and preparation method thereof and the method for quality control that contain Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material
Technical field
The present invention belongs to field of pharmaceutical preparations, specifically, relates to the pharmaceutical preparation that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material and preparation method thereof and method of quality control.
Background technology
Ancient prescription " shenfu decoction ", its side is made up of Radix Ginseng, Radix Aconiti Lateralis Preparata two flavor medicines.According to Chinese medical theory, this side is the side of recuperating depleted YANG and rescuing the patient from collapse, supplementing QI to prevent collapse, clinically is mainly used in fainting of yang-energy sudden collapse and takes off disease.
Contain saponins, polysaccharide composition in the Radix Ginseng; Containing the alkaloids composition in the Radix Aconiti Lateralis Preparata, mainly is aconite alkaloids; Ester alkaloid is a kind of, poisonous in the aconite alkaloids.
There is SHENFU ZHUSHEYE to sell in the market, the disclosed patent that has multinomial relevant SHENFU ZHUSHEYE, ginseng aconite injection to apply in China simultaneously with freeze-dried powder.But it does not all have in the preparation finished product to keep polysaccharide composition in the Radix Ginseng as a kind of main effective ingredient, number is the publication of CN96117458.7, CN03135691.5, CN03135701.6, CN02149368.5, CN200510059785.0, CN200410013725.0, CN200510047701.1 as China's application (patent).
Polysaccharide composition in the Radix Ginseng has various active, can bring into play antineoplastic action from number of ways as the polysaccharide composition in the Radix Ginseng; Polysaccharide composition in the Radix Ginseng all has obvious facilitation to the specific immunity and the nonspecific immunity of body; And the polysaccharide composition in the Radix Ginseng has multiple physiologically actives such as hypoglycemic activity.Polysaccharide composition in the Radix Ginseng can improve the resistance of shock patient and its recovery is had facilitation.
Further development quality is stable, controlled, the effective ingredient kind keeps the attached pharmaceutical preparation of ginseng more complete, that active constituent content is higher, more complete reservation and develop the drug effect of traditional ancient prescription, can select for clinical use provides how better medication, also be the needs that the better development Chinese medicine is learned.
Summary of the invention
The inventor finds in experimentation unexpectedly: for the pharmaceutical preparation of Radix Ginseng and Radix Aconiti Lateralis Preparata, the pharmacological action that has kept the pharmaceutical preparation of the polysaccharide composition in the Radix Ginseng in the preparation finished product obviously is better than not keeping the pharmaceutical preparation of the polysaccharide composition in the Radix Ginseng.
The inventor explores through a large amount of research on the basis of aforementioned discovery, in conjunction with novel unique technique, has developed the pharmaceutical preparation that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.Contain the saponin component of Radix Ginseng, the polysaccharide composition of Radix Ginseng, the alkaloids composition of Radix Aconiti Lateralis Preparata in this pharmaceutical preparation.
An object of the present invention is to provide the pharmaceutical preparation that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
At traditional ancient prescription, on the basis of proved recipe, utilize traditional Chinese medicine theory, theoretical and the modern medicine theory of modern Chinese medicine, by at ancient prescription, increase and decrease flavour of a drug on the proved recipe, change the dose proportioning, the no incompatibility of increase, can play synergistic Chinese medicine, natural drug, Chinese medicine extract, the natural drug extract, effective ingredient in Chinese, natural drug effective site, Chinese medicine active compound monomer, natural drug active compound monomer, medicines such as chemicals, to ancient prescription, proved recipe is further furtherd investigate and secondary development, be the exploitation better efficacy, safety is better, a shortcut of the new drug that the research and development cost is lower.
Contain Radix Ginseng, Radix Aconiti Lateralis Preparata in the raw material in the pharmaceutical preparation of the present invention, simultaneously in this pharmaceutical preparation, can not add or add one or more no incompatibility, can play medicines such as synergistic Chinese medicine, natural drug, Chinese medicine extract, natural drug extract, effective ingredient in Chinese, natural drug effective site, Chinese medicine active compound monomer, natural drug active compound monomer, chemicals, as Radix Glycyrrhizae, the Radix Astragali, Radix Glycyrrhizae extract, Radix Astragali extract, naloxone hydrochloride, dopamine hydrochloride etc.
Radix Ginseng in this pharmaceutical preparation and Radix Aconiti Lateralis Preparata are wild product or artificial culture product, are genuine medicinal materials or non-genuine medicinal materials.Radix Ginseng as described can be Radix Ginseng or SHANSHEN or Park Ginseng or Panax Ginseng etc.
Described Radix Ginseng and Radix Aconiti Lateralis Preparata are concocted processed product that forms or the processed product that forms according to the modernism process of preparing Chinese medicine for its bright medical material or according to traditional method simultaneously.Radix Ginseng as described can be Radix Ginseng or sun-dried SHANSHEN or Radix Ginseng or Radix Ginseng Rubra etc., and described Radix Aconiti Lateralis Preparata can be Radix Aconiti Lateralis Preparata or Radix Aconiti Lateralis Preparata or Radix Aconiti Lateralis Preparata etc.
The pharmaceutical preparation that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material of the present invention can be any pharmaceutically useful dosage form, and these dosage forms comprise oral solid formulation, oral semi-solid preparation, oral liquid, injection.
Described oral solid formulation can be tablet, hard capsule, soft capsule, pill, drop pill, solid dispersion, powder, granule, fine granule, pellet, microcapsule, microspheres agent and other pharmaceutically acceptable oral solid formulation.Described oral liquid can be oral liquid and other pharmaceutically acceptable oral liquid.Described injection can be injection, freeze-drying preparation for injection, sterile packaged preparation for injection, glucose injection, sodium chloride injection and other pharmaceutically acceptable injection.
Pharmaceutical preparation of the present invention, preferably tablet, hard capsule, soft capsule, solid dispersion, oral liquid, injection, freeze-drying preparation for injection, sterile packaged preparation for injection, glucose injection, sodium chloride injection.More preferably tablet, hard capsule, solid dispersion, oral liquid, injection, freeze-drying preparation for injection.More preferably tablet, hard capsule, oral liquid, injection, freeze-drying preparation for injection.
Pharmaceutical preparation of the present invention exists with unit dosage form, and described unit dosage form is meant single-dose preparations, as every of injection, and every bottle, every of oral formulations, every etc.
Pharmaceutical preparation of the present invention, Radix Ginseng in the raw material: the ratio of Radix Aconiti Lateralis Preparata is 1-10: 1-10, preferred 1-4: 1-8, more preferably 1-2: 1-4, most preferably 1: 2.
Pharmaceutical preparation of the present invention in the preparation of per unit dosage, contains Radix Ginseng 0.1g-25g by raw material, Radix Aconiti Lateralis Preparata 0.2g-50g; Preferably contain Radix Ginseng 0.15g-20g, Radix Aconiti Lateralis Preparata 0.3g-40g; More preferably contain Radix Ginseng 0.2g-14g, Radix Aconiti Lateralis Preparata 0.4g-28g; More preferably contain Radix Ginseng 0.2g-11g, Radix Aconiti Lateralis Preparata 0.4g-22g.
Pharmaceutical preparation of the present invention in the preparation of per unit dosage, contains Radix Ginseng total saponins 1mg～90mg, Radix Ginseng total polysaccharides 2mg～282mg, Radix Aconiti Lateralis Preparata total alkaloids 0.02mg～22mg.Preferably contain Radix Ginseng total saponins 2mg～60mg, Radix Ginseng total polysaccharides 4mg～188mg, Radix Aconiti Lateralis Preparata total alkaloids 0.05mg～15mg.More preferably contain Radix Ginseng total saponins 2.5mg～45mg, Radix Ginseng total polysaccharides 6mg～141mg, Radix Aconiti Lateralis Preparata total alkaloids 0.06mg～11mg.
Concrete can be: in the preparation of per unit dosage, contain Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) meter, be 8mg～15mg; The Radix Ginseng total polysaccharides is 25mg～47mg with glucose meter; Radix Aconiti Lateralis Preparata total alkaloids 0.2mg～3.6mg.
Or in the preparation of per unit dosage, contain Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) meter, be 16mg～30mg; The Radix Ginseng total polysaccharides is 50mg～94mg with glucose meter; Radix Aconiti Lateralis Preparata total alkaloids 0.4mg～7.2mg.
Or in the preparation of per unit dosage, contain Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) meter, be 40mg～75mg; The Radix Ginseng total polysaccharides is 125mg～235mg with glucose meter; Radix Aconiti Lateralis Preparata total alkaloids 1mg～18mg.
Or in the preparation of per unit dosage, contain Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) meter, be 2.5mg～5mg; The Radix Ginseng total polysaccharides is 8mg～16mg with glucose meter; Radix Aconiti Lateralis Preparata total alkaloids 0.06mg～1.2mg.
Ester alkaloid in the Radix Aconiti Lateralis Preparata is a kind of, poisonous in the aconite alkaloids.Pharmaceutical preparation of the present invention in the preparation of per unit dosage, contains aconite alkaloids with aconitine (C 24H 47NO 11) meter, must not be higher than 10.0mg, preferably must not be higher than 5.0mg, more preferably must not be higher than 3.0mg, more preferably must not be higher than 2.0mg, more preferably must not be higher than 1.0mg.
The amount average molecular weight is 30～36 * 10 in the pharmaceutical preparation of the present invention 4, 8～12 * 10 4, 3.5～7.5 * 10 4The polysaccharide composition of Radix Ginseng in one or more, wherein measuring average molecular weight is 33.0445 * 10 4, or 9.8878 * 10 4, or 5.4087 * 10 4The content of polysaccharide composition of Radix Ginseng higher.
Pharmaceutical preparation of the present invention also contains where necessary or uses medicine acceptable auxiliary (or carrier), described adjuvant is pharmaceutically acceptable preparation adjuvant.
Preferably contain the saponin component of the Radix Ginseng of 0.05%～90% (percentage by weight), the polysaccharide composition of Radix Ginseng and the alkaloids composition of Radix Aconiti Lateralis Preparata in the pharmaceutical preparation of the present invention, and the pharmaceutically available adjuvant of 99.95%～0% (percentage by weight).The content of the alkaloids composition of the polysaccharide composition of the saponin component of Radix Ginseng, Radix Ginseng and Radix Aconiti Lateralis Preparata more preferably 0.08%～80% in described preparation, and more preferably 0.1%～75%, more preferably 0.15%～70%.
For oral solid formulation and oral semi-solid preparation, described adjuvant is selected from diluent, wetting agent, binding agent, disintegrating agent, fluidizer, antiplastering aid, lubricant, color and regulator thereof, solid dispersion carrier material, antioxidant, surfactant, stabilizing agent, PH regulator and other pharmaceutically acceptable oral solid formulation, oral semi-solid preparation with one or more the material in the adjuvant, and every kind of adjuvant can not select or select for use one or more the material in this kind adjuvant.
Described diluent is one or more the material that is selected from starch, amylum pregelatinisatum, Icing Sugar, dextrin, lactose, microcrystalline Cellulose, mannitol, sorbitol, calcium sulfate, calcium carbonate and other the pharmaceutically acceptable diluent;
Described wetting agent is one or more the material that is selected from ethanol, water and other the pharmaceutically acceptable wetting agent;
Described binding agent is one or more the material that is selected from hypromellose, ethyl cellulose, sodium carboxymethyl cellulose, methylcellulose, hyprolose, starch slurry, polyvidone, gelatin, Polyethylene Glycol, 50% to 70% sucrose solution, sodium alginate soln and other the pharmaceutically acceptable binding agent;
Described disintegrating agent is one or more the material that is selected from carboxymethyl starch sodium, low-substituted hydroxypropyl cellulose, cross-linking sodium carboxymethyl cellulose, dried starch, polyvinylpolypyrrolidone, gas-producing disintegrant and other the pharmaceutically acceptable disintegrating agent;
Described fluidizer, antiplastering aid, lubricant are one or more the materials that is selected from Pulvis Talci, micropowder silica gel, magnesium stearate, polyethylene glycols, sodium laurylsulfate, magnesium laurylsulfate, hydrogenated vegetable oil and other pharmaceutically acceptable fluidizer, antiplastering aid, the lubricant;
Described color and regulator thereof are one or more the materials that is selected from medicinal pigment, food coloring, essence and other pharmaceutically acceptable color and the regulator thereof;
Described solid dispersion carrier material is to be selected from polyethylene glycols, cellulose derivative, organic acid, surfactant-based, the polyvidone class, saccharide and alcohols, cellulose family, the polyacrylic resin class, cupreol, cholesterol, cholesterol ester stearic acid, tripalmitin, castor oil hydrogenated, Oleum Ricini wax, Cera Flava, the material of one or more in Brazil wax and other pharmaceutically acceptable solid dispersion carrier material, every class solid dispersion carrier material can not select or select for use one or more the material in such solid dispersion carrier material;
Described pH regulator agent can be at least a pharmaceutically acceptable material that is used to regulate pH value, and the material of described adjusting pH value is one or more the material that is selected from the pharmaceutically acceptable material that is used for regulating pH value of alkali compounds, buffer system, acid and other.
For oral liquid, described adjuvant is selected from one or more the material in solvent, solubilizing agent, cosolvent, cosolvent, antiseptic, correctives, coloring agent, pH regulator agent, antioxidant, complexing of metal ion agent and other the pharmaceutically acceptable liquid preparation additives, and every kind of adjuvant can not select or select for use one or more the material in this kind adjuvant.
Described solvent is to be selected from water, certain density ethanol, glycerol, propylene glycol and other pharmaceutically acceptable liquid preparation with one or more the material in the solvent;
Described antiseptic is to be selected from parabens, sorbic acid and salt thereof, benzoic acid and salt thereof such as methyl parahydroxybenzoate, ethyl ester, propyl ester, butyl ester, Oleum menthae and other pharmaceutically acceptable liquid preparation with one or more the material in the antiseptic;
Described correctives is to be selected from sucrose, simple syrup, rob, sorbitol, glycerol, mannitol, saccharin sodium, Herba Menthae Haplocalycis volatile oil and other pharmaceutically acceptable liquid preparation with one or more the material in the correctives.
For injection, described adjuvant is selected from pharmaceutically acceptable solvent for injection, solubilizing agent, antibacterial, antioxidant, stabilizing agent, chelating agent, analgesics, isoosmotic adjusting agent, buffer agent, pH regulator agent, and one or more the material in the pharmaceutically acceptable medicine with other miscellaneous function, every kind of adjuvant can not select or select for use one or more the material in this kind adjuvant.
Described solvent for injection is one or more the material that is selected from water for injection, certain density ethanol, glycerol, propylene glycol, benzyl alcohol and other the pharmaceutically acceptable solvent for injection;
Described solubilizing agent is to be selected from polysorbate 20, polyoxyethylene sorbitan monoleate, polysorbate 40, polysorbate 60 and other pharmaceutically acceptable injection with one or more the material in the solubilizing agent;
Described antibacterial is to be selected from hydroxypropyl butyl ester, hydroxypropyl methyl ester, benzyl alcohol, phenol, chlorobutanol and other pharmaceutically acceptable injection with one or more the material in the antibacterial;
Described antioxidant is to be selected from sodium pyrosulfite, sodium sulfite, sodium sulfite, sodium thiosulfate and other pharmaceutically acceptable injection with one or more the material in the antioxidant;
Described chelating agent is one or more the material that is selected from Calcium Disodium Versenate, cyclohexanediamine four sodium acetates, N-hydroxyl diethylamine three acetic acid, diethyl triamine six acetic acid and other the pharmaceutically acceptable chelating agent.
Described pH regulator agent is one or more the material that is selected from certain density sodium hydroxide solution, certain density hydrochloric acid solution, phosphoric acid, acetic acid-sodium acetate buffer solution, acetate buffer solution, phosphate buffered solution, citrate buffer solution, citric acid-disodium hydrogen phosphate buffer solution and other the pharmaceutically acceptable pH regulator agent.
Wherein said freeze-drying preparation for injection can also not add or add one or more the acceptable an amount of excipient of medicine in preparation.Described excipient is selected from one or more the material in mannitol, sorbitol, glycine, lactose, sodium chloride, glucose and other the pharmaceutically acceptable excipient.The preferred mannitol of described excipient, sorbitol.
Pharmaceutical preparation of the present invention is determined usage and dosage according to patient's concrete condition in use, but uses every day 1～4 time, uses the pharmaceutical preparation of 1～20 unit dose at every turn, as 1～20 of each use or.
Another purpose of the present invention provides the preparation method that contains the pharmaceutical preparation of Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
The preparation method of pharmaceutical preparation of the present invention is as follows:
Prescription: Radix Ginseng, Radix Aconiti Lateralis Preparata, a synergistic medicine (add or do not add), adjuvant (add or do not add)
Preparation method summary: extract the saponin component of the Radix Ginseng in Radix Ginseng and the Radix Aconiti Lateralis Preparata, the polysaccharide composition of Radix Ginseng, the alkaloids composition of Radix Aconiti Lateralis Preparata, add or do not added synergistic medicine, add or do not add adjuvant, make the oral solid formulation that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material, oral semi-solid preparation, oral liquid, injection.
Preparation method
One, the extraction of the alkaloids composition of the polysaccharide composition of the saponin component of the Radix Ginseng in Radix Ginseng and the Radix Aconiti Lateralis Preparata, Radix Ginseng, Radix Aconiti Lateralis Preparata:
(1) processing of Radix Ginseng: Radix Ginseng adopts certain density ethanol extraction (extracting method adopts reflux extraction or ultrasonic extraction or percolation or infusion process or continuous extraction or decompression extraction method), the gained extracting solution filters, filtrate recycling ethanol is to there not being the alcohol flavor, adjustment liquor strength extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.5～4.5g, use water saturated n-butanol extraction, merge n-butyl alcohol liquid, reclaim n-butyl alcohol to there not being the n-butyl alcohol flavor, change water-soluble, filter, get the saponin component intermediate of Radix Ginseng, standby;
(2) processing of medicine residues of Radix Ginseng: the medicinal residues of Radix Ginseng after by certain density ethanol extraction are earlier with water extraction (extracting method adopts decocting method or reflux extraction or ultrasonic extraction or percolation or infusion process or continuous extraction or decompression extraction method), the gained extracting solution filters, adjusting liquor strength is 0.2～2.4g crude drug/ml, temperature is 20～70 ℃, adds 85～95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 70～90%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, it is (or not centrifugal to merge supernatant, filter the back and directly adjust liquor strength), adjusting the supernatant liquor strength is 0.7～6g crude drug/ml, adds 85～95% ethanol and makes and contain alcohol and measure and reach 60～80%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, get supernatant (or not centrifugal, directly get filtrate after the filtration)
Previous step gained supernatant or filtrate are the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid;
Previous step gained supernatant adopts the ultrafilter membrane ultrafiltration of molecular cut off 100K～900K, and ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or with of the ultrafilter membrane ultrafiltration of previous step gained supernatant elder generation with molecular cut off 100K～900K, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 1K～10K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
(3) processing of Radix Aconiti Lateralis Preparata: the Radix Aconiti Lateralis Preparata medical material is earlier with water extraction (extracting method adopts decocting method or reflux extraction or ultrasonic extraction or percolation or infusion process or continuous extraction or decompression extraction method), the gained extracting solution filters, the relative density of adjusting medicinal liquid is 1.0～1.25,20～65 ℃ of temperature, ethanol precipitate with ethanol with 85～95% makes ethanol content reach 60～80%, standing over night, filter, filtrate recycling ethanol is to there not being the alcohol flavor, the relative density of adjusting medicinal liquid is that the ethanol of adding 85～95% in 1.05～1.45 o'clock makes pure content reach 75～90%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid;
Filtrate being heated to boiled, and keep little and boiled 10～60 minutes, cooling, cold preservation filters, and promptly gets the Radix Aconiti Lateralis Preparata intermediate, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor for the first time, adjustment medicinal liquid extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.7～2.5g, the dichloromethane extraction of 0.5～3 times of amount of each adding 2～9 times, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Two, play the processing of synergistic medicine:
(4) play the processing of synergistic medicine: playing a synergistic medicine is Chinese medicine or natural drug, extracts its effective site or effective ingredient monomer, and (make with extra care) is water-soluble, regulates pH value to 5.5～8.5, and filtration promptly gets intermediate, and is standby; Playing synergistic medicine is chemicals, and water-soluble, pH value transfers to 5.5～8.5, filters, and promptly gets intermediate, standby; (remarks: can be not water-soluble when being used to prepare oral solid formulation, oral semi-solid preparation)
Three, the preparation of each pharmaceutical preparation finished product:
(5) intermediate of merging " (1) ", " (2) ", " (3) " gained, mixing;
(6) intermediate with " (1) ", " (2) ", " (3) ", " (4) " gained merges, mixing, gained medicinal liquid filter, and are condensed into clear paste and [or " (5) " gained medicinal liquid are filtered, be condensed into clear paste, the intermediate not water-soluble with " (4) " mixes], add an amount of oral solid formulation, oral semi-solid preparation auxiliary materials and mixing, the system soft material, granulate, oven dry, tabletting or incapsulate is made the sheet or the capsule that contain Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(7) " (5) " gained medicinal liquid being adjusted volume with purified water or water for injection is 0.1～2.0g crude drug/ml to being equivalent to liquor strength, regulates pH value to 5.5～8.5, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter that [or filtrate adds " (the 4) " intermediate of gained and an amount of oral liquid with adjuvant or do not add adjuvant, or filtrate adds an amount of oral liquid with adjuvant or do not add adjuvant; Mixing, (filtration)], regulate pH value to 5.5～8.5, add purified water or water for injection and adjust volume to full dose, survey pH value, filter, according to dosage divide to be filled in the oral liquid packing container, seal, the conventional method sterilization promptly gets the oral liquid that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(8) " (5) " gained medicinal liquid being adjusted volume with water for injection is 0.1～2.0g crude drug/ml to being equivalent to liquor strength, regulates pH value to 5.5～8.5, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter that [or filtrate adds " (the 4) " intermediate of gained and an amount of injection with adjuvant or do not add adjuvant, or filtrate adds an amount of injection with adjuvant or do not add adjuvant; Mixing, (filtration)], (regulating pH value to 5.5～8.5), adding 0.02～0.5% active carbon is heated to and boils, and keeps little and boils 10～60 minutes, filters, regulate pH value to 5.5～8.5, add the injection water and adjust volume, survey pH value, fine straining (crossing the sintered glass filter in 0.22～0.50 μ m microporous filter membrane or similar aperture) to full dose, according to dosage divide and be filled to the infusion pump dress with in the container, seal, the conventional method sterilization promptly gets the injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(9) " (5) " gained medicinal liquid being adjusted volume with water for injection is 0.3～2.0g crude drug/ml to being equivalent to liquor strength, regulates pH value to 5.5～8.5, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter [or filtrate add " (the 4) " intermediate of gained and an amount of injection with adjuvant, freeze-drying preparation for injection with adjuvant or do not add adjuvant, or an amount of injection of filtrate adding with adjuvant, freeze-drying preparation for injection with adjuvant or do not add adjuvant; Mixing, (filtration)], regulate pH value to 5.5～8.5, adding 0.02～0.5% active carbon is heated to and boils, keeping little boiled 10～60 minutes, filter, regulate pH value to 5.5～8.5, add the injection water and adjust volume, survey pH value, fine straining (crossing the sintered glass filter in 0.22～0.50 μ m microporous filter membrane or similar aperture) to full dose, aseptic subpackagedly contain 1～8ml medicinal liquid to every control injection vial, lyophilization, is rolled lid at gland, promptly gets the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(10) freeze drying process is as follows:
The a pre-freeze stage: common pre-freeze, pre-freeze temperature-60 ℃～-40 ℃, about 2.5～6 hours of pre-freeze time; Or select pre-freeze repeatedly, pre-freeze temperature-60 ℃～-40 ℃, the temperature of rising again-15 ℃～-7 ℃, about 3～9 hours of pre-freeze time.
The b sublimation drying stage: when the temperature of condenser reduce to-60 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-30 ℃～-15 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water.
C is drying stage again: shelf temperature slowly rises to-5 ℃～30 ℃, judges exsiccant terminal point with the vacuum descent method again, again 3～12 hours drying times.
Preferred manufacturing procedure
Pharmaceutical preparation preferred manufacturing procedure of the present invention is:
One, the extraction of the alkaloids composition of the polysaccharide composition of the saponin component of the Radix Ginseng in Radix Ginseng and the Radix Aconiti Lateralis Preparata, Radix Ginseng, Radix Aconiti Lateralis Preparata:
(1) processing of Radix Ginseng: Radix Ginseng adds 3～9 times of amounts respectively (for the first time because be dried medical material, add 1～3 times of amount) 60～80% alcohol reflux or supersound extraction 1～4 time, each 0.5～3 hour, merge extractive liquid,, filter, filtrate recycling ethanol is to there not being the alcohol flavor, adjustment liquor strength extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.75～3.0g, the water saturated n-butanol extraction of 0.7～3 times of amount of each adding 2～9 times merges n-butyl alcohol liquid, reclaims n-butyl alcohol to there being the n-butyl alcohol flavor, change water-soluble, filter, get the saponin component intermediate of Radix Ginseng, standby;
(2) processing of medicine residues of Radix Ginseng: the medicine residues of Radix Ginseng after the alcohol extraction adds 6～13.5 times of water gagings respectively (for the first time because be dried medical material, add 1～5 times of amount) decoct to extract or reflux, extract, 2～6 times, each 0.5～4 hour, merge extractive liquid, filtered, adjusting liquor strength is 0.4～1.6g crude drug/ml, temperature is 20～60 ℃, adds 90～95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 75～85%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, it is (or not centrifugal to merge supernatant, filter the back and directly adjust liquor strength, be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid), adjusting the supernatant liquor strength is 1.0～4.0g crude drug/ml, adding 90～95% ethanol makes and contains the alcohol amount and reach 65～75%, standing over night filters, and precipitation is dry, after adding the suitable quantity of water dissolving, centrifugal, get supernatant (or not centrifugal, directly get filtrate after the filtration, be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid)
Previous step gained supernatant or filtrate are the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid;
Previous step gained supernatant adopts the ultrafilter membrane ultrafiltration of molecular cut off 100K～700K, and ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or with of the ultrafilter membrane ultrafiltration of previous step gained supernatant elder generation with molecular cut off 100K～700K, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 1K～8K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
(3) processing of Radix Aconiti Lateralis Preparata: the Radix Aconiti Lateralis Preparata medical material adds 4～12 times of water gagings (for the first time because be dried medical material, add 1～4 times of amount), decoct extraction or reflux, extract, 2～6 times, each 0.5～3 hour, merge extractive liquid,, filter, the relative density of adjusting medicinal liquid is 1.01～1.21,20～60 ℃ of temperature, ethanol precipitate with ethanol with 90～95% makes ethanol content reach 65～75%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, the relative density of adjusting medicinal liquid is that the ethanol of adding 90～95% in 1.15～1.40 o'clock makes pure content reach 80～90%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid;
Filtrate being heated to boiled, and keep little and boiled 15～55 minutes, cooling, cold preservation filters, and promptly gets the Radix Aconiti Lateralis Preparata intermediate, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor for the first time, adjustment medicinal liquid extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.8～2.0g, the dichloromethane extraction of 0.8～2 times of amount of each adding 3～7 times, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Two, play the processing of synergistic medicine:
(4) play the processing of synergistic medicine: playing a synergistic medicine is Chinese medicine or natural drug, extracts its effective site or effective ingredient monomer, and (make with extra care) is water-soluble, regulates pH value to 6～8, and filtration promptly gets intermediate, and is standby; Playing synergistic medicine is chemicals, and water-soluble, pH value transfers to 6～8, filters, and promptly gets intermediate, standby; (remarks: can be not water-soluble when being used to prepare oral solid formulation, oral semi-solid preparation)
Three, the preparation of each pharmaceutical preparation finished product:
(5) intermediate of merging " (1) ", " (2) ", " (3) " gained, mixing;
(6) intermediate with " (1) ", " (2) ", " (3) ", " (4) " gained merges, mixing, gained medicinal liquid filter, and are condensed into clear paste and [or " (5) " gained medicinal liquid are filtered, be condensed into clear paste, the intermediate not water-soluble with " (4) " mixes], add an amount of oral solid formulation, oral semi-solid preparation auxiliary materials and mixing, the system soft material, granulate, oven dry, tabletting or incapsulate is made the sheet or the capsule that contain Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(7) " (5) " gained medicinal liquid being adjusted volume with purified water or water for injection is 0.1～1.5g crude drug/ml to being equivalent to liquor strength, regulates pH value to 6.0～8.0, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter that [or filtrate adds " (the 4) " intermediate of gained and an amount of oral liquid with adjuvant or do not add adjuvant, or filtrate adds an amount of oral liquid with adjuvant or do not add adjuvant; Mixing, (filtration)], regulate pH value to 6.0～8.0, add purified water or water for injection and adjust volume to full dose, survey pH value, filter, according to dosage divide to be filled in the oral liquid packing container, seal, the conventional method sterilization promptly gets the oral liquid that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(8) " (5) " gained medicinal liquid being adjusted volume with water for injection is 0.1～1.5g crude drug/ml to being equivalent to liquor strength, regulates pH value to 6.0～8.0, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter that [or filtrate adds " (the 4) " intermediate of gained and an amount of injection with adjuvant or do not add adjuvant, or filtrate adds an amount of injection with adjuvant or do not add adjuvant; Mixing, (filtration)], (regulating pH value to 6.0～8.0), adding 0.02～0.3% active carbon is heated to and boils, and keeps little and boils 10～40 minutes, filters, regulate pH value to 6.0～8.0, add the injection water and adjust volume, survey pH value, fine straining (crossing the sintered glass filter in 0.22～0.45 μ m microporous filter membrane or similar aperture) to full dose, according to dosage divide and be filled to the infusion pump dress with in the container, seal, the conventional method sterilization promptly gets the injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.(9) " (5) " gained medicinal liquid being adjusted volume with water for injection is 0.5～2.0g crude drug/ml to being equivalent to liquor strength, regulates pH value to 6.0～8.0, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter [or filtrate add " (the 4) " intermediate of gained and an amount of injection with adjuvant, freeze-drying preparation for injection with adjuvant or do not add adjuvant, or an amount of injection of filtrate adding with adjuvant, freeze-drying preparation for injection with adjuvant or do not add adjuvant; Mixing, (filtration)], regulate pH value to 6.0～8.0, adding 0.02～0.3% active carbon is heated to and boils, keeping little boiled 10～40 minutes, filter, regulate pH value to 6.0～8.0, add the injection water and adjust volume, survey pH value, fine straining (crossing the sintered glass filter in 0.22～0.45 μ m microporous filter membrane or similar aperture) to full dose, aseptic subpackagedly contain 1～6ml medicinal liquid to every control injection vial, lyophilization, is rolled lid at gland, promptly gets the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(10) freeze drying process is as follows:
The a pre-freeze stage: common pre-freeze, pre-freeze temperature-60 ℃～-45 ℃, about 2.5～6 hours of pre-freeze time; Or select pre-freeze repeatedly, pre-freeze temperature-60 ℃～-45 ℃, the temperature of rising again-13 ℃～-8 ℃, about 3～9 hours of pre-freeze time.
The b sublimation drying stage: when the temperature of condenser reduce to-60 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-26 ℃～-15 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water.
C is drying stage again: shelf temperature slowly rises to 0 ℃～25 ℃, judges exsiccant terminal point with the vacuum descent method again, again 3～12 hours drying times.
The method of " extraction of the polysaccharide composition of the saponin component of the Radix Ginseng in Radix Ginseng and the Radix Aconiti Lateralis Preparata, Radix Ginseng, the alkaloids composition of Radix Aconiti Lateralis Preparata " in the preparation method of pharmaceutical preparation of the present invention can be more preferably:
(1) processing of Radix Ginseng: Radix Ginseng adds 5～7 times of amounts respectively (for the first time because be dried medical material, add 1～2 times of amount) 65～75% alcohol reflux 2～3 times, each 1.5～2.5 hours, merge extractive liquid,, filter, filtrate recycling ethanol is to there not being the alcohol flavor, adjustment liquor strength extremely every 1ml is equivalent to contain Radix Ginseng crude drug 1.0～2.0g, the water saturated n-butanol extraction of 0.8～1.2 times of amount of each adding 5～7 times merges n-butyl alcohol liquid, reclaims n-butyl alcohol to there being the n-butyl alcohol flavor, change water-soluble, filter, get the saponin component intermediate of Radix Ginseng, standby;
(2) processing of medicine residues of Radix Ginseng: the medicine residues of Radix Ginseng after the alcohol extraction adds 8～10 times of water gagings respectively (for the first time because be dried medical material, add 2～4 times of amounts) decoct to extract or reflux, extract, 2～4 times, each 1.5～2.5 hours, merge extractive liquid, filtered, adjusting liquor strength is 0.6～1.0g crude drug/ml, temperature is 20～45 ℃, adds 95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 75～85%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, it is (or not centrifugal to merge supernatant, filter the back and directly adjust liquor strength, be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid), adjusting the supernatant liquor strength is 1.5～2.5g crude drug/ml, adding 95% ethanol makes and contains the alcohol amount and reach 65～75%, standing over night filters, and precipitation is dry, after adding the suitable quantity of water dissolving, centrifugal, get supernatant (or not centrifugal, directly get filtrate after the filtration, be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid)
Previous step gained supernatant or filtrate are the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid;
Previous step gained supernatant adopts the ultrafilter membrane ultrafiltration of molecular cut off 100K～600K, and ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or with of the ultrafilter membrane ultrafiltration of previous step gained supernatant elder generation with molecular cut off 100K～600K, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 1K～8K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
(3) processing of Radix Aconiti Lateralis Preparata: the Radix Aconiti Lateralis Preparata medical material adds 7～9 times of water gagings (for the first time because be dried medical material, add 1～3 times of amount), decoct extraction or reflux, extract, 2～4 times, each 0.5～1.5 hour, merge extractive liquid,, filter, the relative density of adjusting medicinal liquid is 1.05～1.16,40～55 ℃ of temperature, ethanol precipitate with ethanol with 95% makes ethanol content reach 65～75%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, the relative density of adjusting medicinal liquid is that the ethanol of adding 95% in 1.21～1.35 o'clock makes pure content reach 80～90%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid;
Filtrate being heated to boiled, and keep little and boiled 30～45 minutes, cooling, cold preservation filters, and promptly gets the Radix Aconiti Lateralis Preparata intermediate, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor for the first time, adjustment medicinal liquid extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.8～1.2g, the dichloromethane extraction of 0.8～1.2 times of amount of each adding 4～6 times, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection.
A further object of the present invention provides the method for quality control that contains the pharmaceutical preparation of Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.Make the stable and controllable for quality of pharmaceutical preparation of the present invention.
The present invention preferably provides the method for quality control that contains the injection of Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material, comprises following discriminating, inspection, assay, finger printing.
Described discriminating, inspection, assay, finger printing and one concrete grammar wherein can be selected for use respectively or combination in any is used, and are used for the quality control of injection that a kind of raw material contains Radix Ginseng and Radix Aconiti Lateralis Preparata, freeze-drying preparation for injection, sterile packaged preparation for injection.
Described discriminating, inspection, assay, finger printing comprise Radix Ginseng, ginsenoside Rg 1, ginsenoside Re's discriminating, the discriminating of Radix Aconiti Lateralis Preparata, the limit examine of aconite alkaloids, ginsenoside Rg 1With the assay of polysaccharide composition in the assay of Re, Determination of Total Saponin Content in Panax Ginseng, the Radix Ginseng, the finger printing of the ginsenoside in ginseng crude drug's finger printing, Radix Aconiti Lateralis Preparata medicinal materials fingerprint, the preparation and alkaloidal finger printing.
The method of quality control of injection, freeze-drying preparation for injection, sterile packaged preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in described a kind of raw material is as follows:
[1] differentiates
[1.1] Radix Ginseng, ginsenoside Rg 1, the ginsenoside Re discriminating:
Get the injection content 1.0g for preparing, add water 30ml dissolving, add chloroform 30～50ml, jolting, water intaking liquid evaporate to dryness, residue adds water 2ml makes dissolving, added water saturated n-butyl alcohol 8～12ml supersound process 25～40 minutes, get n-butyl alcohol liquid and put in the separatory funnel, add the ammonia solution of 2～4 times of amounts, washing, washing liquid discards, get n-butyl alcohol liquid evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution.Other gets ginseng crude drug 1g (crossing 60 mesh sieves), adds ethanol 20～40ml, and reflux 20～40 minutes filters, and filtrate evaporate to dryness, residue add water 30ml makes dissolving, filters, and filtrate is made ginseng crude drug's solution with method.Get the ginsenoside Rg more respectively 1, ginsenoside Re's reference substance is an amount of, add methanol and make the solution that every 1ml contains 2mg respectively, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 2.5～5 μ l of above-mentioned four kinds of solution, put respectively on same silica gel g thin-layer plate, (14～16: 38～42: 21～23: 9～11) lower floor's solution of placing below 10 ℃ is developing solvent with chloroform-ethyl acetate-methanol-water, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃.In the test sample chromatograph, with the corresponding position of ginseng crude drug's chromatograph on, show the speckle of same color; With the corresponding position of reference substance chromatograph on, show the speckle of same color.
[1.2] discriminating of Radix Aconiti Lateralis Preparata:
Get the injection content 3.5g for preparing, add water 30ml dissolving, regulate pH value to 9～11 with ammonia solution, the jolting that adds diethyl ether is extracted 2～4 times, each 40～50ml, and ether liquid low temperature evaporate to dryness, residue adds dehydrated alcohol 2ml makes dissolving, as need testing solution.Other gets Radix Aconiti Lateralis Preparata medicinal material coarse powder 20g, puts in the tool plug conical flask the 130～170ml that adds diethyl ether, jolting 9～12 minutes, add ammonia solution 9～11ml, jolting 25～35 minutes was placed 1～2 hour, divided and got the ether layer, volatilize, residue adds dehydrated alcohol 2ml makes dissolving, as Radix Aconiti Lateralis Preparata medical material solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 12～30 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate (thick 500 μ m), chromatography cylinder is earlier saturated with ammonia, with petroleum ether-ether-acetone (4.8～5.2: 2.8～3.2: 2.8～3.2) be developing solvent, launch, take out, dry, spray is with bismuth potassium iodide test solution, and it is clear to be heated to speckle colour developing.In the test sample chromatograph, with the corresponding position of Radix Aconiti Lateralis Preparata medical material chromatograph on, show the speckle of same color.
[2] check
[2.1] limit examine of aconite alkaloids:
The preparation of [2.1.1] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the aconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 50ml measuring bottle, adds the 0.01mol/L hydrochloric acid solution and makes dissolving, and be diluted to scale, shakes up, promptly.
The preparation precision of [2.1.2] standard curve is measured above-mentioned reference substance solution 0.2～0.3ml, 0.4～0.6ml, 0.65～0.85ml, 0.9～1.1ml, 1.4～1.6ml, 1.65～1.85ml, 1.9～2.1ml, put in the separatory funnel respectively, add 0.01mol/L hydrochloric acid solution 1.85～1.65ml respectively, 1.6～1.4ml, 1.35～1.15ml, 1.1～0.9ml, 0.6～0.4ml, 0.35～0.15ml, 0.00ml, accurate respectively adding acetic acid-sodium-acetate buffer (is got 0.2mol/L acetum 250ml, with 0.2mol/L sodium acetate solution adjust pH to 3.1) 8～12ml, bromocresol green liquid (is got bromocresol green 50mg, add 0.05mol/L sodium hydroxide solution 1.6ml grinding and make dissolving, add water to 100ml, extract 3 times with the chloroform jolting, each 30ml, discard chloroform solution) 1.5～2.5ml, chloroform 8～12ml, jolting 2～6 minutes is left standstill, and divides and gets chloroform solution, retinue is blank, measures absorbance at 415nm wavelength place according to spectrophotometric degree method (appendix VA of Chinese Pharmacopoeia version in 2005).With concentration is abscissa, and absorbance is a vertical coordinate, the drawing standard curve.
The injection content 0.35g for preparing is got in the preparation of [2.1.3] need testing solution, and accurate the title decides, and adds water 10ml dissolving, be transferred in the separatory funnel, add ammonia solution and regulate pH value to 10～11, extract 3～5 times with the chloroform jolting of equivalent, combined chloroform liquid, evaporate to dryness.Residue adds 0.01mol/L hydrochloric acid solution gradation dissolving, and changes in the 10ml measuring bottle, is diluted to scale and shakes up, promptly.
[2.1.4] algoscopy precision is measured above-mentioned need testing solution 2ml, puts respectively in the separatory funnel, measures absorbance according to " preparation of standard curve " item down from " each precision adds acetic acid-sodium-acetate buffer 10ml " in accordance with the law, calculates, promptly.
Every of the injection that [2.1.5] prepares contains aconite alkaloids with aconitine (C 24H 47NO 11) meter, must not be higher than 1.0mg.
[3] assay
[3.1] ginsenoside Rg 1Assay with Re:
Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D).
[3.1.1] chromatographic condition is filler with the octadecylsilane chemically bonded silica, acetonitrile-0.1% phosphoric acid solution (17～23: 75～85) be mobile phase, detect wavelength 203nm.Number of theoretical plate calculates by the ginsenoside Re peak should be not less than 5000.
The preparation of [3.1.2] reference substance solution ginsenoside Rg of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1And ginsenoside Re's reference substance is an amount of, accurately claims surely, adds methanol and makes every 1ml respectively and contain the ginsenoside Rg 10.3mg, the solution of ginsenoside Re 0.2mg, promptly.
The injection content 0.35g for preparing under the content uniformity item is got in the preparation of [3.1.3] need testing solution, accurate claims surely, puts in the 5ml measuring bottle, is dissolved in water and is diluted to scale, shakes up, promptly.
Accurate respectively reference substance solution and each 10～30 μ l of need testing solution of drawing of [3.1.4] algoscopy inject chromatograph of liquid, measure, promptly.
Every of the injection that [3.1.5] prepares contains the ginsenoside Rg 1, the ginsenoside Re the content sum should be 0.2～2.0mg.
[3.2] Determination of Total Saponin Content in Panax Ginseng:
The preparation of [3.2.1] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance is an amount of, accurate claims surely, adds methanol and makes every 1ml and contain ginsenoside Rb 10.3mg solution, promptly.
The preparation precision of [3.2.2] standard curve is measured above-mentioned ginsenoside Rb 1Reference substance solution 0.05～0.15ml, 0.15～0.25ml, 0.25～0.35ml, 0.35～0.45ml, 0.45～0.55ml, 0.55～0.65ml, 0.65～0.75ml puts respectively in the tool plug test tube, fling to solvent, each adds 5% vanillin-glacial acetic acid solution (fresh preparation) 0.15～0.25ml and perchloric acid 0.6～1.0ml, heats 12～18min in 55～65 ℃ of water-baths, takes out, cooling rapidly, add glacial acetic acid 3～7ml, shake up, simultaneously with reagent corresponding as blank, measure absorbance in 556nm wavelength place according to spectrophotography (appendix VA of Chinese Pharmacopoeia version in 2005), with the absorbance is vertical coordinate, and sampling amount is an abscissa, the drawing standard curve.
The injection content 0.35g for preparing under the content uniformity item is got in the preparation of [3.2.3] sample solution, the accurate title, decide, adding distil water 10ml dissolving is put in the separatory funnel, extracts 3～5 times with the chloroform jolting, each 8～12ml, discard chloroform solution, water liquid extracts 3～5 times with water saturated n-butyl alcohol jolting again, each 8～12ml, merge n-butyl alcohol liquid, the saturated water washing of reuse n-butyl alcohol 2～3 times, each 8～12ml discards water liquid, n-butyl alcohol liquid is put evaporate to dryness in the water-bath, residue adds dissolve with methanol, is transferred in the 10ml measuring bottle, and is diluted to scale, shake up, promptly.
[3.2.4] algoscopy precision is measured sample solution 0.1ml and is put in the tool plug test tube, measures absorbance from " flinging to solvent " down according to " standard curve preparation " in accordance with the law, calculating, promptly.
Every of the injection that [3.2.5] prepares contains Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) meter, should be 2.0～20.0mg.
[3.3] assay of polysaccharide composition in the Radix Ginseng
The preparation of [3.3.1] reference substance solution phosphorus pentoxide drying under reduced pressure of learning from else's experience is an amount of to the glucose reference substance of constant weight, accurately claims surely, and adding distil water is made the solution that every 1ml contains glucose 0.2mg, promptly.
The preparation precision of [3.3.2] standard curve is measured glucose reference substance solution 0.05～0.15ml, 0.15～0.25ml, 0.25～0.35ml, 0.35～0.45ml, 0.45～0.55ml, 0.55～0.65ml, 0.65～0.75ml, 0.75～0.85ml puts respectively in the tool plug test tube, add water respectively and make into 1.0ml, each accurate freshly prepared 0.2% anthrone sulphuric acid (sulfuric acid concentration is 80%) solution 6～10ml that adds shakes up, and heats 8～12 minutes in 100 ℃ of water-baths, rapidly after the cooling, retinue is blank, measures absorbance at 620nm wavelength place according to spectrophotography (appendix VA of Chinese Pharmacopoeia version in 2005), is abscissa with concentration, absorbance is a vertical coordinate, the drawing standard curve.
The injection content 3.5g for preparing under the content uniformity item is got in the preparation of [3.3.3] sample solution, accurate claim fixed, add water 25～35ml dissolving after, adding ethanol makes and contains the alcohol amount and reach 75～85%, centrifugal, add dissolved in distilled water after precipitation volatilizes ethanol, and be settled in the 100ml measuring bottle.Get above-mentioned solution 1～3ml and put in the 25ml measuring bottle, thin up shakes up to scale, promptly.
[3.3.4] algoscopy precision is measured 0.2ml in tool plug test tube, measures absorbance from " adding distil water is to 1.0ml respectively " down by " preparation of standard curve " item in accordance with the law, calculates, promptly.
Every of the injection that [3.3.5] prepares contains polysaccharide composition in the Radix Ginseng with glucose meter, should be 4～60mg.
[4] finger printing
With reference to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D), measure in conjunction with the requirement of finger printing
[4.1] ginseng crude drug's finger printing
[4.1.1] medical material title and source
Radix Ginseng in this Radix Ginseng medicinal materials fingerprint is selected Radix Ginseng Rubra for use, and Radix Ginseng Rubra is the dry root and rhizome of cultivation product (practise and claim " Park Ginseng ") after steaming of Araliaceae Radix Ginseng Panaxginseng C.A.Mey..
The mensuration of [4.1.2] ginseng crude drug finger printing
[4.1.2.1] chromatographic condition chromatographic column: ODS HYPERSIL C 18(post of Φ 4.6mm * 250mm), packing material size 5 μ m; Mobile phase: acetonitrile and water are mobile phase, and gradient condition sees the following form
Gradient condition
Time (min Acetonitrile (%) Water (%)
0 20 55 60 10～20 25～35 45～55 10～20 90～80 75～65 55～45 90～80
Column temperature: 25～35 ℃; Analysis time: 60～120min; Flow velocity: 0.6～1.0ml/min; 100～120 ℃ of ELSD drift tubes, flow rate of carrier gas 2.0～4.0ml/min.Number of theoretical plate is with ginsenoside Rb 1The peak meter should be not less than 5000.
The preparation of [4.1.2.2] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance 2mg, the accurate title, decide, and puts in the 5ml measuring bottle, adds dissolve with methanol and be settled to scale, shakes up, promptly.
Radix Ginseng Rubra medicinal powder (crossing 20 mesh sieves) 2g is got in the preparation of [4.1.2.3] need testing solution, and accurate the title decides, and adds 70% alcohol reflux 2 times, and each 0.8～1.5 hour, add 6～8 times of amounts for the first time, add 5～7 times of amounts for the second time; The extracting solution filtration, filtrate recycling ethanol concentrates, and extracts 4～6 times with the water saturated n-butyl alcohol jolting of equivalent, merges n-butyl alcohol liquid, and washs with the ammonia solution of equivalent; Cleaning mixture discards, and n-butyl alcohol liquid evaporate to dryness, residue are dissolved in water and are transferred in the 10ml measuring bottle, are settled to scale, shake up, promptly.
Accurate respectively reference substance solution and each 5～20 μ l of need testing solution of drawing of [4.1.2.4] algoscopy inject chromatograph of liquid, measure, promptly.
[4.1.2.5] result is with ginsenoside Rb 1The peak is set at the object of reference peak, and with the logarithm of its peak area as 1.000, according to ginsenoside Rb 1The retention time at peak is calculated, calibrate 10 total peaks altogether, its relative retention time is respectively: 0.056 ± 10%, 0.102 ± 10%, 0.372 ± 10%, 0.632 ± 10%, 0.931 ± 10%, 1.000,1.040 ± 10%, 1.084 ± 10%, 1.181 ± 10%, 1.662 ± 10%; And the regulation relative retention time is that the relative peak area at 0.632 ± 10%, 1.040 ± 10%, 1.084 ± 10%, 1.181 ± 10% peak is 0.682～1.266,0.346～0.644,0.262～0.486,0.103～0.191 than (peak area logarithm ratio).
[4.2] Radix Aconiti Lateralis Preparata medicinal materials fingerprint
The title and the source of [4.2.1] Radix Aconiti Lateralis Preparata medical material
Radix Aconiti Lateralis Preparata is the processed goods of the daughter root of ranunculaceae plant Aconitum carmichjaelii Debx. Aconitum carmichaeli Debx..Radix Aconiti Lateralis Preparata in this Radix Aconiti Lateralis Preparata medicinal materials fingerprint is selected Radix Aconiti Lateralis Preparata for use.
The mensuration of [4.2.2] Radix Aconiti Lateralis Preparata medicinal materials fingerprint
[4.2.2.1] chromatographic condition: chromatographic column: ZORBAX SB C 18(Φ 4.6mm * 150mm), packing material size 5 μ m; Mobile phase: methanol and 0.1% phosphoric acid-0.04% triethylamine-aqueous solution are mobile phase, and gradient condition sees the following form
Gradient condition
Time (min) Methanol (%) 0.1% phosphoric acid-0.04% triethylamine-aqueous solution (%)
0 10 45 60 25～35 25～35 37～47 37～47 75～65 75～65 63～53 63～53
Column temperature: 25～35 ℃; Detect wavelength: 235nm; Analysis time: 60～120min; Flow velocity: 0.7～1.3ml/min; Number of theoretical plate calculates with the mesaconitine peak should be not less than 2000.
The preparation of [4.2.2.2] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the mesaconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, promptly.
The preparation Radix Aconiti Lateralis Preparata medicinal material coarse powder 10g of [4.2.2.3] need testing solution spends the night with ammonia solution 3～5ml, ether 40～60ml merceration, filters; Medicinal residues add diethyl ether: chloroform (2.8～3.2: 0.8～1.2) mixed solution 40～60ml, supersound process 25～40min, filter, medicinal residues wash 3～4 times with mixed solution, each 14～16ml, and washing liquid and filtrate merge, the low temperature evaporate to dryness, residue adds methanol 5ml dissolving, with the filter membrane filtration of 0.45 μ m, can measure.
Accurate respectively inner mark solution 15～25 μ l and need testing solution 30～50 μ l of drawing of [4.2.2.4] algoscopy inject chromatograph of liquid, measure, promptly.
[4.2.2.5] result is set at the object of reference peak with the mesaconitine peak, retention time according to the mesaconitine peak is calculated, calibrate 8 total peaks altogether, its relative retention time is respectively: 0.146 ± 10%, 0.254 ± 10%, 0.531 ± 10%, 0.630 ± 10%, 0.727 ± 10%, 0.833 ± 10%, 1.000,1.114 ± 10%; And with the stable and the biggest relative retention time of peak area be the peak area at 0.531 ± 10% peak as 1.000, and the regulation relative retention time is that the relative peak area ratio at 0.727 ± 10%, 1.114 ± 10% peak is 0.182～0.272,0.373～0.552.
[4.3] ginsenoside's finger printing
[4.3.1] chromatographic condition chromatographic column: ODS HYPERSIL C 18(post of Φ 4.6mm * 250mm), packing material size 5 μ m; Acetonitrile and water are mobile phase, and gradient condition sees the following form:
Gradient condition
Time (min) Acetonitrile (%) Water (%)
0 20 55 60 10～20 25～35 45～55 10～20 90～80 75～65 55～45 90～80
Column temperature: 25～35 ℃; Analysis time: 60～120min; Flow velocity: 0.6～1.0ml/min; ELSD parameter: 100～120 ℃ of drift tubes, flow rate of carrier gas 2.0～4.0ml/min.Number of theoretical plate is with ginsenoside Rb 1The peak calculates should be not less than 5000.
The preparation of [4.3.2] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance 2mg, the accurate title, decide, and puts in the 5ml measuring bottle, adds dissolve with methanol and be settled to scale, shakes up, promptly.
The injection content 0.35g for preparing is got in the preparation of [4.3.3] need testing solution, the accurate title, decide, be dissolved in water to 8～12ml, extract 4～6 times, merge n-butyl alcohol liquid with the water-saturated n-butanol jolting of equivalent, ammonia solution with equivalent washs 1～2 time, divide and to get n-butyl alcohol liquid, reclaim n-butyl alcohol, residue is with dissolved in distilled water and be settled in the 5ml measuring bottle, shake up, promptly.
Accurate respectively reference substance solution and each 5～15 μ l of need testing solution of drawing of [4.3.4] algoscopy inject chromatograph of liquid, measure, promptly.
[4.3.5] result is with ginsenoside Rb 1The peak is set at the object of reference peak, and with the logarithm of its peak area as 1.000, according to ginsenoside Rb 1The retention time at peak is calculated, calibrate 10 total peaks altogether, its relative retention time is respectively: 0.441 ± 10%, 0.632 ± 10%, 0.942 ± 10%, 1.000,1.042 ± 10%, 1.062 ± 10%, 1.089 ± 10%, 1.188 ± 10%, 1.701 ± 10%, 1.737 ± 10%; And the regulation relative retention time is that the relative peak area at 0.441 ± 10%, 0.632 ± 10%, 1.042 ± 10%, 1.089 ± 10% peak is respectively 0.688～1.148,0.673～1.121,0.692～1.154,0.688～1.146 than (peak area logarithm ratio).
[4.4] fingerprint of alkaloid
[4.4.1] chromatographic condition chromatographic column: ZORBAX SB C 18(post of Φ 4.6mm * 150mm), packing material size 5 μ m; Methanol and 0.1% phosphoric acid-0.04% triethylamine aqueous solution are mobile phase, and gradient condition sees the following form:
Gradient condition
Time (min) Methanol (%) 0.1% phosphoric acid-0.04% triethylamine aqueous solution (%)
0 10 45 60 25～35 25～35 37～47 37～47 75～65 75～65 63～53 63～53
25～35 ℃ of column temperatures; Detect wavelength: 235nm; Analysis time: 60～120min; Number of theoretical plate calculates with the mesaconitine peak should be not less than 2000.
The preparation of [4.4.2] inner mark solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the mesaconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 25ml measuring bottle, adds 1% phosphoric acid solution 3ml, is diluted to scale with methanol, shakes up, promptly.
The preparation precision of [4.4.3] need testing solution is measured the injection content 1.0g for preparing, be dissolved in water to 8～12ml, put in the separatory funnel, add ammonia solution adjust pH to 9～11, with the E-C (2.8～3.2: mixed liquor jolting extraction 0.8～1.2) 4～6 times of equivalent, combining extraction liquid, low temperature volatilizes, and residue adds methanol 1ml makes dissolving, adds inner mark solution 40～60 μ l, filter, as need testing solution.
Accurate respectively inner mark solution 15～25 μ l and need testing solution 30～50 μ l of drawing of [4.4.4] algoscopy inject chromatograph of liquid, measure, promptly.
[4.4.5] result is set at the mesaconitine peak at the object of reference peak of relative retention time, retention time according to the mesaconitine peak is calculated, calibrate 5 total peaks altogether, its relative retention time is respectively: 0.259 ± 10%, 0.342 ± 10%, 0.532 ± 10%, 0.724 ± 10%, 1.000 ± 10%; And with the stable and the biggest relative retention time of peak area be the peak area at 0.532 ± 10% peak as 1.000, and the regulation relative retention time is that the relative peak area ratio at 0.724 ± 10% peak is 0.455～0.683.
Contain injection, the freeze-drying preparation for injection of Radix Ginseng and Radix Aconiti Lateralis Preparata, the method for quality control of sterile packaged preparation for injection in described a kind of raw material, its each concrete steps are as follows:
[1] differentiates
[1.1] Radix Ginseng, ginsenoside Rg 1, the ginsenoside Re discriminating:
Get the injection content 1.0g for preparing, add water 30ml dissolving, add chloroform 40ml, jolting, water intaking liquid evaporate to dryness, residue adds water 2ml makes dissolving, added water saturated n-butyl alcohol 10ml supersound process 30 minutes, get n-butyl alcohol liquid and put in the separatory funnel, add the ammonia solution of 3 times of amounts, washing, washing liquid discards, get n-butyl alcohol liquid evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution.Other gets ginseng crude drug 1g (crossing 60 mesh sieves), adds ethanol 30ml, and reflux 30 minutes filters, and filtrate evaporate to dryness, residue add water 30ml makes dissolving, filters, and filtrate is made ginseng crude drug's solution with method.Get the ginsenoside Rg more respectively 1, ginsenoside Re's reference substance is an amount of, add methanol and make the solution that every 1ml contains 2mg respectively, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 2.5～5 μ l of above-mentioned four kinds of solution, put respectively on same silica gel g thin-layer plate, (15: 40: 22: 10) lower floor's solution of placing below 10 ℃ was developing solvent with chloroform-ethyl acetate-methanol-water, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃.In the test sample chromatograph, with the corresponding position of ginseng crude drug's chromatograph on, show the speckle of same color; With the corresponding position of reference substance chromatograph on, show the speckle of same color.
[1.2] discriminating of Radix Aconiti Lateralis Preparata:
Get the injection content 3.5g for preparing, add water 30ml dissolving, regulate pH value to 10 with ammonia solution, the jolting that adds diethyl ether is extracted 3 times, each 45ml, and ether liquid low temperature evaporate to dryness, residue adds dehydrated alcohol 2ml makes dissolving, as need testing solution.Other gets Radix Aconiti Lateralis Preparata medicinal material coarse powder 20g, puts in the tool plug conical flask, and the 150ml that adds diethyl ether, jolting 10 minutes adds ammonia solution 10ml, and jolting 30 minutes was placed 1～2 hour, divided and got the ether layer, volatilized, and residue adds dehydrated alcohol 2ml makes dissolving, as Radix Aconiti Lateralis Preparata medical material solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 12～30 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate (thick 500 μ m), chromatography cylinder is earlier saturated with ammonia, is developing solvent with petroleum ether-ether-acetone (5: 3: 3), launches, take out, dry, spray is with bismuth potassium iodide test solution, and it is clear to be heated to speckle colour developing.In the test sample chromatograph, with the corresponding position of Radix Aconiti Lateralis Preparata medical material chromatograph on, show the speckle of same color.
[2] check
[2.1] limit examine of aconite alkaloids:
The preparation of [2.1.1] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the aconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 50ml measuring bottle, adds the 0.01mol/L hydrochloric acid solution and makes dissolving, and be diluted to scale, shakes up, promptly.
The preparation precision of [2.1.2] standard curve is measured above-mentioned reference substance solution 0.25ml, 0.50ml, 0.75ml, 1.00ml, 1.50ml, 1.75ml, 2.00ml, put in the separatory funnel respectively, add 0.01mol/L hydrochloric acid solution 1.75ml respectively, 1.50ml, 1.25ml, 1.00ml, 0.50ml, 0.25ml, 0.00ml, accurate respectively adding acetic acid-sodium-acetate buffer (is got 0.2mol/L acetum 250ml, with 0.2mol/L sodium acetate solution adjust pH to 3.1) 10ml, bromocresol green liquid (is got bromocresol green 50mg, add 0.05mol/L sodium hydroxide solution 1.6ml grinding and make dissolving, add water to 100ml, extract 3 times with the chloroform jolting, each 30ml, discard chloroform solution) 2ml, chloroform 10ml, jolting 3 minutes is left standstill, and divides and gets chloroform solution, retinue is blank, measures absorbance at 415nm wavelength place according to spectrophotometric degree method (appendix VA of Chinese Pharmacopoeia version in 2005).With concentration is abscissa, and absorbance is a vertical coordinate, the drawing standard curve.
The injection content 0.35g for preparing is got in the preparation of [2.1.3] need testing solution, and accurate the title decides, and adds water 10ml dissolving, is transferred in the separatory funnel, adds ammonia solution and regulates pH value to 10～11, extracts combined chloroform liquid, evaporate to dryness 4 times with the chloroform jolting of equivalent.Residue adds 0.01mol/L hydrochloric acid solution gradation dissolving, and changes in the 10ml measuring bottle, is diluted to scale and shakes up, promptly.
[2.1.4] algoscopy precision is measured above-mentioned need testing solution 2ml, puts respectively in the separatory funnel, measures absorbance according to " preparation of standard curve " item down from " each precision adds acetic acid-sodium-acetate buffer 10ml " in accordance with the law, calculates, promptly.
Every of the injection that [2.1.5] prepares contains aconite alkaloids with aconitine (C 24H 47NO 11) meter, must not be higher than 1.0mg.
[3] assay
[3.1] ginsenoside Rg 1Assay with Re:
Measure according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D).
[3.1.1] chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica, and acetonitrile-0.1% phosphoric acid solution (20: 80) is a mobile phase, detects wavelength 203nm.Number of theoretical plate calculates by the ginsenoside Re peak should be not less than 5000.
The preparation of [3.1.2] reference substance solution ginsenoside Rg of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1And ginsenoside Re's reference substance is an amount of, accurately claims surely, adds methanol and makes every 1ml respectively and contain the ginsenoside Rg 10.3mg, the solution of ginsenoside Re 0.2mg, promptly.
The injection content 0.35g for preparing under the content uniformity item is got in the preparation of [3.1.3] need testing solution, accurate claims surely, puts in the 5ml measuring bottle, is dissolved in water and is diluted to scale, shakes up, promptly.
Accurate respectively reference substance solution and each the 20 μ l of need testing solution of drawing of [3.1.4] algoscopy inject chromatograph of liquid, measure, promptly.
Every of the injection that [3.1.5] prepares contains the ginsenoside Rg 1, the ginsenoside Re the content sum should be 0.8～1.4mg.
[3.2] Determination of Total Saponin Content in Panax Ginseng:
The preparation of [3.2.1] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance is an amount of, accurate claims surely, adds methanol and makes every 1ml and contain ginsenoside Rb 10.3mg solution, promptly.
The preparation precision of [3.2.2] standard curve is measured above-mentioned ginsenoside Rb 1Reference substance solution 0.1ml, 0.2ml, 0.3ml, 0.4ml, 0.5ml, 0.6ml 0.7ml puts respectively in the tool plug test tube, fling to solvent, each adds 5% vanillin-glacial acetic acid solution (fresh preparation) 0.2ml and perchloric acid 0.8ml, heats 15min in 60 ℃ of water-baths, takes out, cooling rapidly, add glacial acetic acid 5ml, shake up, simultaneously with reagent corresponding as blank, measure absorbance in 556nm wavelength place according to spectrophotography (appendix VA of Chinese Pharmacopoeia version in 2005), with the absorbance is vertical coordinate, and sampling amount is an abscissa, the drawing standard curve.
The injection content 0.35g for preparing under the content uniformity item is got in the preparation of [3.2.3] sample solution, the accurate title, decide, adding distil water 10ml dissolving is put in the separatory funnel, extracts 4 times with the chloroform jolting, each 10ml, discard chloroform solution, water liquid extracts 4 times with water saturated n-butyl alcohol jolting again, each 10ml, merge n-butyl alcohol liquid, the saturated water washing of reuse n-butyl alcohol 2 times, each 10ml discards water liquid, n-butyl alcohol liquid is put evaporate to dryness in the water-bath, residue adds dissolve with methanol, is transferred in the 10ml measuring bottle, and is diluted to scale, shake up, promptly.
[3.2.4] algoscopy precision is measured sample solution 0.1ml and is put in the tool plug test tube, measures absorbance from " flinging to solvent " down according to " standard curve preparation " in accordance with the law, calculating, promptly.
Every of the injection that [3.2.5] prepares contains Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) meter, should be 8.0～15.0mg.
[3.3] assay of polysaccharide composition in the Radix Ginseng
The preparation of [3.3.1] reference substance solution phosphorus pentoxide drying under reduced pressure of learning from else's experience is an amount of to the glucose reference substance of constant weight, accurately claims surely, and adding distil water is made the solution that every 1ml contains glucose 0.2mg, promptly.
The preparation precision of [3.3.2] standard curve is measured glucose reference substance solution 0.1ml, 0.2ml, 0.3ml, 0.4ml, 0.5ml, 0.6ml, 0.7ml 0.8ml puts respectively in the tool plug test tube, add water respectively and make into 1.0ml, each accurate freshly prepared 0.2% anthrone sulphuric acid (sulfuric acid concentration is 80%) solution 8ml that adds shakes up, and heating is 10 minutes in 100 ℃ of water-baths, rapidly after the cooling, retinue is blank, measures absorbance at 620nm wavelength place according to spectrophotography (an appendix V of Chinese Pharmacopoeia version in 2005 A), is abscissa with concentration, absorbance is a vertical coordinate, the drawing standard curve.
The injection content 3.5g for preparing under the content uniformity item is got in the preparation of [3.3.3] sample solution, accurate claim fixed, add water 30ml dissolving after, adding ethanol makes and contains the alcohol amount and reach 80%, centrifugal, add dissolved in distilled water after precipitation volatilizes ethanol, and be settled in the 100ml measuring bottle.Get above-mentioned solution 2ml and put in the 25ml measuring bottle, thin up shakes up to scale, promptly.
[3.3.4] algoscopy precision is measured 0.2ml in tool plug test tube, measures absorbance from " adding distil water is to 1.0ml respectively " down by " preparation of standard curve " item in accordance with the law, calculates, promptly.
Every of the injection that [3.3.5] prepares contains polysaccharide composition in the Radix Ginseng with glucose meter, should be 27～47mg.
[4] finger printing
With reference to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D), measure in conjunction with the requirement of finger printing
[4.1] ginseng crude drug's finger printing
[4.1.1] medical material title and source
Radix Ginseng in this Radix Ginseng medicinal materials fingerprint is selected Radix Ginseng Rubra for use, and Radix Ginseng Rubra is the dry root and rhizome of cultivation product (practise and claim " Park Ginseng ") after steaming of Araliaceae Radix Ginseng Panaxginseng C.A.Mey..
The mensuration of [4.1.2] ginseng crude drug finger printing
[4.1.2.1] chromatographic condition chromatographic column: ODS HYPERSIL C 18(post of Φ 4.6mm * 250mm), packing material size 5 μ m; Mobile phase: acetonitrile and water are mobile phase, and gradient condition sees the following form
Gradient condition
Time (min Acetonitrile (%) Water (%)
0 20 55 60 15 30 50 15 85 70 50 85
Column temperature: 30 ℃; Analysis time: 60min; Flow velocity: 0.8ml/min; 110 ℃ of ELSD drift tubes, flow rate of carrier gas 3.0ml/min.Number of theoretical plate is with ginsenoside Rb 1The peak meter should be not less than 5000.
The preparation of [4.1.2.2] reference substance solution: the phosphorus pentoxide drying under reduced pressure of learning from else's experience is to ginsenoside Rb1's reference substance 2mg of constant weight, accurately claims surely, puts in the 5ml measuring bottle, adds dissolve with methanol and is settled to scale, shakes up, promptly.
Radix Ginseng Rubra medicinal powder (crossing 20 mesh sieves) 2g is got in the preparation of [4.1.2.3] need testing solution, and accurate the title decides, and adds 70% alcohol reflux twice, and each 1 hour, add 7 times of amounts for the first time, add 6 times of amounts for the second time; The extracting solution filtration, filtrate recycling ethanol concentrates, and extracts 5 times with the water saturated n-butyl alcohol jolting of equivalent, merges n-butyl alcohol liquid, and washs with the ammonia solution of equivalent; Cleaning mixture discards, and n-butyl alcohol liquid evaporate to dryness, residue are dissolved in water and are transferred in the 10ml measuring bottle, are settled to scale, shake up, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of [4.1.2.4] algoscopy inject chromatograph of liquid, measure, promptly.
[4.1.2.5] result is with ginsenoside Rb 1The peak is set at the object of reference peak, and with the logarithm of its peak area as 1.000, according to ginsenoside Rb 1The retention time at peak is calculated, calibrate 10 total peaks altogether, its relative retention time is respectively: 0.056 ± 10%, 0.102 ± 10%, 0.372 ± 10%, 0.632 ± 10%, 0.931 ± 10%, 1.000,1.040 ± 10%, 1.084 ± 10%, 1.181 ± 10%, 1.662 ± 10%; And the regulation relative retention time is that the relative peak area at 0.632 ± 10%, 1.040 ± 10%, 1.084 ± 10%, 1.181 ± 10% peak is 0.682～1.266,0.346～0.644,0.262～0.486,0.103～0.191 than (peak area logarithm ratio).
[4.2] Radix Aconiti Lateralis Preparata medicinal materials fingerprint
The title and the source of [4.2.1] Radix Aconiti Lateralis Preparata medical material
Radix Aconiti Lateralis Preparata is the processed goods of the daughter root of ranunculaceae plant Aconitum carmichjaelii Debx. Aconitum carmichaeli Debx..Radix Aconiti Lateralis Preparata in this Radix Aconiti Lateralis Preparata medicinal materials fingerprint is selected Radix Aconiti Lateralis Preparata for use.
The mensuration of [4.2.2] Radix Aconiti Lateralis Preparata medicinal materials fingerprint
[4.2.2.1] chromatographic condition: chromatographic column: ZORBAX SBC 18(post of Φ 4.6mm * 150mm), packing material size 5 μ m; Mobile phase: methanol and 0.1% phosphoric acid-0.04% triethylamine-aqueous solution are mobile phase, and gradient condition sees the following form
Gradient condition
Time (min) Methanol (%) 0.1% phosphoric acid-0.04% triethylamine-aqueous solution (%)
0 10 45 60 30 30 42 42 70 70 58 58
Column temperature: 30 ℃; Detect wavelength: 235nm; Analysis time: 60min; Flow velocity: 1.0ml/min; Number of theoretical plate calculates with the mesaconitine peak should be not less than 2000.
The preparation of [4.2.2.2] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the mesaconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, promptly.
The preparation Radix Aconiti Lateralis Preparata medicinal material coarse powder 10g of [4.2.2.3] need testing solution spends the night with ammonia solution 4ml, ether 50ml merceration, filters; Medicinal residues add diethyl ether: chloroform (3: 1) mixed solution 50ml, and supersound process 30min filters, and medicinal residues wash 3～4 times with mixed solution, each 15ml, washing liquid and filtrate merge, the low temperature evaporate to dryness, residue adds methanol 5ml dissolving, with the filter membrane filtration of 0.45 μ m, can measure.
Accurate respectively inner mark solution 20 μ l and the need testing solution 40 μ l of drawing of [4.2.2.4] algoscopy inject chromatograph of liquid, measure, promptly.
[4.2.2.5] result is set at the object of reference peak with the mesaconitine peak, retention time according to the mesaconitine peak is calculated, calibrate 8 total peaks altogether, its relative retention time is respectively: 0.146 ± 10%, 0.254 ± 10%, 0.531 ± 10%, 0.630 ± 10%, 0.727 ± 10%, 0.833 ± 10%, 1.000,1.114 ± 10%; And with the stable and the biggest relative retention time of peak area be the peak area at 0.531 ± 10% peak as 1.000, and the regulation relative retention time is that the relative peak area ratio at 0.727 ± 10%, 1.114 ± 10% peak is 0.182～0.272,0.373～0.552.
[4.3] ginsenoside's finger printing
[4.3.1] chromatographic condition chromatographic column: ODS HYPERSIL C 18(post of Φ 4.6mm * 250mm), packing material size 5 μ m; Acetonitrile and water are mobile phase, and gradient condition sees the following form:
Gradient condition
Time (min) Acetonitrile (%) Water (%)
0 20 55 60 15 30 50 15 85 70 50 85
Column temperature: 30 ℃; Analysis time: 60min; Flow velocity: 0.8ml/min; ELSD parameter: 110 ℃ of drift tubes, flow rate of carrier gas 3.0ml/min.Number of theoretical plate is with ginsenoside Rb 1The peak calculates should be not less than 5000.
The preparation of [4.3.2] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance 2mg, the accurate title, decide, and puts in the 5ml measuring bottle, adds dissolve with methanol and be settled to scale, shakes up, promptly.
The injection content 0.35g for preparing is got in the preparation of [4.3.3] need testing solution, the accurate title, decide, be dissolved in water to 10ml, extract 5 times, merge n-butyl alcohol liquid with the water-saturated n-butanol jolting of equivalent, ammonia solution with equivalent washs 1 time, divide and to get n-butyl alcohol liquid, reclaim n-butyl alcohol, residue is with dissolved in distilled water and be settled in the 5ml measuring bottle, shake up, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of [4.3.4] algoscopy inject chromatograph of liquid, measure, promptly.
[4.3.5] result is with ginsenoside Rb 1The peak is set at the object of reference peak, and with the logarithm of its peak area as 1.000, according to ginsenoside Rb 1The retention time at peak is calculated, calibrate 10 total peaks altogether, its relative retention time is respectively: 0.441 ± 10%, 0.632 ± 10%, 0.942 ± 10%, 1.000,1.042 ± 10%, 1.062 ± 10%, 1.089 ± 10%, 1.188 ± 10%, 1.701 ± 10%, 1.737 ± 10%; And the regulation relative retention time is that the relative peak area at 0.441 ± 10%, 0.632 ± 10%, 1.042 ± 10%, 1.089 ± 10% peak is respectively 0.688～1.148,0.673～1.121,0.692～1.154,0.688～1.146 than (peak area logarithm ratio).
[4.4] fingerprint of alkaloid
[4.4.1] chromatographic condition chromatographic column: ZORBAX SB C 18(post of Φ 4.6mm * 150mm), packing material size 5 μ m; Methanol and 0.1% phosphoric acid-0.04% triethylamine aqueous solution are mobile phase, and gradient condition sees the following form:
Gradient condition
Time (min) Methanol (%) 0.1% phosphoric acid-0.04% triethylamine aqueous solution (%)
0 10 45 60 30 30 42 42 70 70 58 58
30 ℃ of column temperatures; Detect wavelength: 235nm; Analysis time: 60min; Number of theoretical plate calculates with the mesaconitine peak should be not less than 2000.
The preparation of [4.4.2] inner mark solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the mesaconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 25ml measuring bottle, adds 1% phosphoric acid solution 3ml, is diluted to scale with methanol, shakes up, promptly.
The preparation precision of [4.4.3] need testing solution is measured the injection content 1.0g for preparing, be dissolved in water to 10ml, put in the separatory funnel, add ammonia solution adjust pH to 10, use the mixed liquor jolting of the E-C (3: 1) of equivalent to extract 5 times, combining extraction liquid, low temperature volatilizes, and residue adds methanol 1ml makes dissolving, adds inner mark solution 50 μ l, filter, as need testing solution.
Accurate respectively inner mark solution 20 μ l and the need testing solution 40 μ l of drawing of [4.4.4] algoscopy inject chromatograph of liquid, measure, promptly.
[4.4.5] result is set at the mesaconitine peak at the object of reference peak of relative retention time, retention time according to the mesaconitine peak is calculated, calibrate 5 total peaks altogether, its relative retention time is respectively: 0.259 ± 10%, 0.342 ± 10%, 0.532 ± 10%, 0.724 ± 10%, 1.000 ± 10%; And with the stable and the biggest relative retention time of peak area be the peak area at 0.532 ± 10% peak as 1.000, and the regulation relative retention time is that the relative peak area ratio at 0.724 ± 10% peak is 0.455～0.683.
Pharmaceutical preparation of the present invention, kept the polysaccharide composition in the Radix Ginseng, can add synergistic medicine, more comprehensively keep and developed the good drug effect of traditional ancient prescription " shenfu decoction ", adopted advanced method of quality control simultaneously, as fingerprint pattern technology, make the stable and controllable for quality of pharmaceutical preparation of the present invention, change the relatively poor shortcoming of quality controllability of Chinese medicine preparation.Can bring into play better clinical effect, select for clinical use provides how better medication.
Oral solid formulation of the present invention carries taking convenience, significantly reduced injection volume, changed the Chinese medicine amount big, carry the shortcoming of taking inconvenience; The existing commercially available SHENFU ZHUSHEYE of the kind analogy of the effective ingredient of injection of the present invention is many, can more comprehensively keep and develop the curative effect of ancient prescription, and onset is rapid; Freeze-drying preparation for injection of the present invention is on the basis of all advantages of injection, and the storage time is longer, and character is more stable, and onset is rapid.The inventor is by a large amount of quadrature screening experiment simultaneously, and preferred suitable preparation method can adopt hyperfiltration technique in the present invention, removes macromole impurity, also can remove small molecular weight impurity again, better guarantees stability of formulation.
The preparation method of pharmaceutical preparation of the present invention simultaneously is not loaded down with trivial details, can control cost preferably.The preparation nature of making is stable, and is quality controllable, Orally-administrable or drug administration by injection.Can be used for treating fainting of yang-energy sudden collapse and take off disease (infectivity, losing blood property, hypovolemic shock etc.); Also can be used for due to the yang deficiency (deficiency of vital energy) palpitation with fear, palpitation with a distress feeling, breath with cough, have a stomach-ache, have loose bowels, arthromyodynia etc., and other former " shenfu decoction " disease of controlling.Can select for clinical use provides how better medication.
The main pharmacodynamics experiment of pharmaceutical preparation of the present invention shows that pharmaceutical preparation of the present invention has good pharmacological action.
From the experiment of the main pharmacodynamics of pharmaceutical preparation of the present invention as can be seen, " the injection ginseng attached (lyophilizing) " that the present invention makes, the main pharmacodynamics experiment shows under Isodose, (Sanjiu Pharmaceutical Industry Co., Ltd., Ya'an City produces with the SHENFU ZHUSHEYE of selling in the market for it, lot number: 041103) compare, the time-to-live of injection ginseng attached (lyophilizing) high dose group prolongation mice normal pressure anoxia enduring is longer significantly, and the effect that increases coronary flow, inhibition CK, LDH release is stronger.Proved beneficial effect of the present invention.
The specific embodiment
Below will the invention will be further described by embodiment, these descriptions are not that content of the present invention is done further qualification.One skilled in the art will understand that the equal replacement that technical characterictic of the present invention is done, or corresponding the improvement, still belong within protection scope of the present invention.
Embodiment 1
Prescription:
Prescription 1 Prescription 2 Prescription 3
Radix Ginseng (Radix Ginseng Rubra) 1000g Radix Ginseng (Radix Ginseng Rubra) 900g Radix Ginseng (Radix Ginseng Rubra) 800g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2100g Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2200g
Starch 200.0g Starch 230.0g Starch 250.0g
Icing Sugar 100.0g Dextrin 70.0g Lactose 50.0g
Microcrystalline Cellulose 80.0g Microcrystalline Cellulose 80.0g Microcrystalline Cellulose 80.0g
Carboxymethyl starch sodium 20.0g Carboxymethyl starch sodium 20.0g Carboxymethyl starch sodium 20.0g
95% ethanol In right amount 95% ethanol In right amount 95% ethanol In right amount
Magnesium stearate 4g Magnesium stearate 4g Magnesium stearate 4g
Make 3000/or 3000 capsules
Preparation method:
(1) Radix Ginseng adds 6 times of amounts (because be dried medical material, adding 1 times of amount for the first time), 70% alcohol reflux 2 times, each 2 hours respectively, filter, filtrate recycling ethanol is not to there being the alcohol flavor, and medicinal liquid is concentrated into every 1ml and is equivalent to contain Radix Ginseng crude drug 1.45～1.55g, and the water-saturated n-butanol that at every turn adds equivalent extracts 6 times, merge n-butyl alcohol liquid, reclaim n-butyl alcohol to there not being the n-butyl alcohol flavor, change water-soluble, filter, get ginsenoside's intermediate, standby;
(2) the Radix Ginseng Rubra medicinal residues after the alcohol extraction add 9 times of water gagings respectively (for the first time because be dried medical material, add 3 times of amounts) decoct to extract or reflux, extract, 3 times, each 2 hours, filter, being concentrated into liquor strength is 0.75～0.85g crude drug/ml, temperature is 38～42 ℃, adds 95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 80%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, merge supernatant, adjusting the supernatant liquor strength is 1.95～2.05g crude drug/ml, adds 95% ethanol and makes and contain the alcohol amount and reach 70%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, it is (or not centrifugal to get supernatant, directly get filtrate after the filtration), previous step gained supernatant or filtrate are the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 100K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 200K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 500K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 200K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 5K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 500K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 2K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 500K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 5K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
(3) the Radix Aconiti Lateralis Preparata medical material adds 8 times of water gagings (for the first time because be dried medical material, add 2 times of amounts), decoct extraction or reflux, extract, 3 times, each 1 hour, filter, be concentrated into relative density of medicine liquid 1.09～1.13,47～53 ℃ of temperature, ethanol precipitate with ethanol with 95% makes ethanol content reach 70%, and standing over night filters, filtrate recycling ethanol is to there not being the alcohol flavor, the relative density of adjusting medicinal liquid is that the ethanol of adding 95% in 1.27～1.31 o'clock makes pure content reach 85%, and standing over night filters, filtrate recycling ethanol is to there not being the alcohol flavor, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
Filtrate being heated to boiled, and keep little and boiled 38～45 minutes, cooling, cold preservation filters, and promptly gets the Radix Aconiti Lateralis Preparata intermediate, and is standby;
Or for the first time alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor, adjusts medicinal liquid and is equivalent to contain Radix Ginseng crude drug 0.95～1.05g to every 1ml, adds the dichloromethane extraction 5 times of 1.0～1.1 times of amounts at every turn, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
(4) intermediate with " (1) ", " (2) ", " (3) " gained merges, and mixing filters, and is condensed into clear paste;
(5) will cross 60～80 mesh sieves except that the adjuvant 95% ethanol, the magnesium stearate, mixing with " (4) " gains mixing, adds 95% an amount of ethanol, make soft material, pushed 20～24 mesh sieves and granulate, 40 ℃ to 65 ℃ dry 1h to 4h cross 20 mesh sieve granulate, add magnesium stearate, mixing
(6) with above-mentioned granule tabletting, promptly get the sheet that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material;
(7), promptly get the tablet that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in the raw material with " (6) " gained sheet bag film-coat or sugar coating;
(8) " (5) " gained is particles filled to capsule, with the polishing of gained capsule, promptly get the capsule that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material then.
Embodiment 2
Prescription:
Prescription 4 Prescription 5 Prescription 6 Prescription 7 Prescription 8
Radix Ginseng (Radix Ginseng Rubra) 1000g 1000g 1000g 1000g 1000g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g 2000g 2000g 2000g 2000g
Add purified water or water for injection extremely 5000ml 8000ml 10000ml 12000ml 15000ml
Make 1000 attached oral liquids of ginseng
Prescription 9 Prescription 10 Prescription 11 Prescription 12 Prescription 13
Radix Ginseng (Radix Ginseng Rubra) 1000g 1000g 1000g 1000g 1000g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g 2000g 2000g 2000g 2000g
Add the injection water extremely 3000ml 5000ml 8000ml 10000ml 12000ml
Make 1000 SHENFU ZHUSHEYE
Prescription 14 Prescription 15 Prescription 16 Prescription 17 Prescription 18
Radix Ginseng (Radix Ginseng Rubra) 1000g 1000g 1000g 1000g 1000g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g 2000g 2000g 2000g 2000g
Mannitol 100g 150g 200g 250g 300g
Add the injection water extremely 3000ml 3000ml 3000ml 3000ml 3000ml
Make 1000 injection ginsengs attached (lyophilizing)
Prescription 19 Prescription 20 Prescription 21
Radix Ginseng (Radix Ginseng Rubra) 1000g 1000g 1000g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 1000g 2000g 2000g
Sorbitol 150g 200g 225g
Add the injection water extremely 3000ml 3000ml 3000ml
Make 1000 injection ginsengs attached (lyophilizing)
Prescription 22 Prescription 23 Prescription 24 Prescription 25 Prescription 26
Radix Ginseng (Radix Ginseng Rubra) 1100g 1200g 1500g 900g 800g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 1900g 1800g 1500g 2100g 2200g
Mannitol 150g 150g 200g 250g 300g
Add the injection water extremely 3000ml 3000ml 3000ml 3000ml 3000ml
Make 1000 injection ginsengs attached (lyophilizing)
Preparation method:
Remarks: the preparation method of similar preparation prescription is identical in the present embodiment.
(1) Radix Ginseng adds 6 times of amounts (because be dried medical material, adding 1 times of amount for the first time), 70% alcohol reflux 2 times, each 2 hours respectively, filter, filtrate recycling ethanol is not to there being the alcohol flavor, and medicinal liquid is concentrated into every 1ml and is equivalent to contain Radix Ginseng crude drug 1.45～1.55g, and the water-saturated n-butanol that at every turn adds equivalent extracts 6 times, merge n-butyl alcohol liquid, reclaim n-butyl alcohol to there not being the n-butyl alcohol flavor, change water-soluble, filter, get ginsenoside's intermediate, standby;
(2) the Radix Ginseng Rubra medicinal residues after the alcohol extraction add 9 times of water gagings respectively (for the first time because be dried medical material, add 3 times of amounts) decoct to extract or reflux, extract, 3 times, each 2 hours, filter, being concentrated into liquor strength is 0.75～0.85g crude drug/ml, temperature is 38～42 ℃, adds 95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 80%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, merge supernatant, adjusting the supernatant liquor strength is 1.95～2.05g crude drug/ml, adds 95% ethanol and makes and contain the alcohol amount and reach 70%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, it is (or not centrifugal to get supernatant, directly get filtrate after the filtration, be used for the preparation of oral liquid)
Previous step gained supernatant or filtrate are the polysaccharide composition intermediate of Radix Ginseng, and is standby, is used for the preparation of oral liquid;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 100K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, is used for the preparation of oral liquid, injection;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 200K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, is used for the preparation of oral liquid, injection;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 500K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, is used for the preparation of oral liquid, injection;
Or previous step gained supernatant is used the ultrafilter membrane ultrafiltration of molecular cut off 200K earlier, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 5K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, be used for the preparation of oral liquid, injection;
Or previous step gained supernatant is used the ultrafilter membrane ultrafiltration of molecular cut off 500K earlier, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 2K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, be used for the preparation of oral liquid, injection;
Or previous step gained supernatant is used the ultrafilter membrane ultrafiltration of molecular cut off 500K earlier, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 5K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, be used for the preparation of oral liquid, injection;
(3) the Radix Aconiti Lateralis Preparata medical material adds 8 times of water gagings (because be dried medical material, adding 2 times of amounts for the first time), decocts extraction or reflux, extract, 3 times, each 1 hour, filter, be concentrated into relative density of medicine liquid 1.09～1.13,47～53 ℃ of temperature, the ethanol precipitate with ethanol with 95% makes ethanol content reach 70%, standing over night, filter, filtrate recycling ethanol does not extremely have the alcohol flavor, and the relative density of adjusting medicinal liquid is that the ethanol of adding 95% in 1.27～1.31 o'clock makes pure content reach 85%, standing over night, filter, filtrate recycling ethanol filters to there not being the alcohol flavor, and filtrate being heated to boiled, keeping little boiled 38～45 minutes, cooling, cold preservation filters, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
Or for the first time alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor, adjusts medicinal liquid and is equivalent to contain Radix Ginseng crude drug 0.95～1.05g to every 1ml, adds the dichloromethane extraction 5 times of 1.0～1.1 times of amounts at every turn, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
(4) intermediate with " (1) ", " (2) ", " (3) " gained merges, and mixing adds purified water or water for injection to full dose, regulates pH value to 7.0, is heated to and boils, and keep little and boiled 38～42 minutes, cooling, cold preservation is spent the night; Filter, filtrate is regulated pH value to 7.0, adds purified water or water for injection and adjusts volume to full dose, surveys pH value, filters, and according to dosage divides to be filled in the oral liquid packing container, seals, and the conventional method sterilization promptly gets the oral liquid that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(5) intermediate with " (1) ", " (2) ", " (3) " gained merges, and mixing adds to the full amount of water for injection, and regulates pH value to 7.0, is heated to and boils, and keep little and boiled 40～45 minutes, cooling, cold preservation is spent the night; Filter, (regulating pH value to 7.0) adds 0.1% active carbon and is heated to and boils, keeping little boiled 20～25 minutes, filter, regulate pH value to 7.0, add the injection water and adjust volume to full dose, survey pH value, cross 0.45 μ m microporous filter membrane, according to dosage divide to be filled to the infusion pump dress, seal with in the container, the conventional method sterilization promptly gets the injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(6) intermediate with " (1) ", " (2) ", " (3) " gained merges, and mixing adds to the full amount of water for injection, and regulates pH value to 7.0, is heated to and boils, and keep little and boiled 40～45 minutes, cooling, cold preservation is spent the night; Filter, add mannitol/or sorbitol in the filtrate, stir evenly, regulate pH value to 7.0, add 0.1% active carbon and be heated to and boil, keep little and boiled 20～25 minutes, filter, regulate pH value to 7.0, add the injection water and adjust volume to full dose, survey pH value, cross 0.45 μ m microporous filter membrane, aseptic subpackaged to control injection vial, lyophilization, gland, roll lid, promptly get the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(7) freeze drying process is as follows:
The a pre-freeze stage: pre-freeze temperature-56 ℃～-45 ℃, about 2.5～6 hours of pre-freeze time;
The b sublimation drying stage: when the temperature of condenser reduce to-56 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-21 ℃～-19 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water.
C is drying stage again: shelf temperature slowly rises to 0 ℃～20 ℃, judges exsiccant terminal point with the vacuum descent method again, again 4～11 hours drying times.
Embodiment 3
Prescription:
Prescription 27
Radix Ginseng (Radix Ginseng Rubra) 1000g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g
Mannitol 150g
Add the injection water extremely 3000ml
Make 1000 injection ginsengs attached (lyophilizing)
Preparation method:
(1) Radix Ginseng adds 6 times of amounts (because be dried medical material, adding 1 times of amount for the first time), 70% alcohol reflux 2 times, each 2 hours respectively, filter, filtrate recycling ethanol is not to there being the alcohol flavor, and medicinal liquid is concentrated into every 1ml and is equivalent to contain Radix Ginseng crude drug 1.45～1.55g, and the water-saturated n-butanol that at every turn adds equivalent extracts 6 times, merge n-butyl alcohol liquid, reclaim n-butyl alcohol to there not being the n-butyl alcohol flavor, change water-soluble, filter, get ginsenoside's intermediate, standby;
(2) the Radix Ginseng Rubra medicinal residues after the alcohol extraction add 9 times of water gagings (for the first time because be dried medical material, add 3 times of amounts) respectively and decoct and extract or reflux, extract, 3 times each 2 hours, filter, being concentrated into liquor strength is 0.75～0.85g crude drug/ml, and temperature is 37～43 ℃, adds 95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 80%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, merge supernatant, adjusting the supernatant liquor strength is 1.95～2.05g crude drug/ml, adds 95% ethanol and makes and contain the alcohol amount and reach 70%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, get supernatant, supernatant earlier with the ultrafilter membrane ultrafiltration of molecular cut off 500K, is removed small-molecular weight impurity with the gained ultrafiltrate with the ultrafilter membrane of molecular cut off 5K then, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
(3) the Radix Aconiti Lateralis Preparata medical material adds 8 times of water gagings (because be dried medical material, adding 2 times of amounts for the first time), decocts extraction or reflux, extract, 3 times, each 1 hour, filter, be concentrated into relative density of medicine liquid 1.09～1.13,47～53 ℃ of temperature, the ethanol precipitate with ethanol with 95% makes ethanol content reach 70%, standing over night, filter, filtrate recycling ethanol does not extremely have the alcohol flavor, and the relative density of adjusting medicinal liquid is that the ethanol of adding 95% in 1.27～1.31 o'clock makes pure content reach 85%, standing over night, filter, filtrate recycling ethanol filters to there not being the alcohol flavor, and filtrate being heated to boiled, keeping little boiled 38～45 minutes, cooling, cold preservation filters, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
(4) intermediate with " (1) ", " (2) ", " (3) " gained merges, and mixing adds to the full amount of water for injection, and regulates pH value to 7.0, is heated to and boils, and keep little and boiled 40～45 minutes, cooling, cold preservation is spent the night; Filter, add mannitol in the filtrate, stir evenly, regulate pH value to 7.0, add 0.1% active carbon and be heated to and boil, keep little and boiled 20～25 minutes, filter, regulate pH value to 7.0, add the injection water and adjust volume to full dose, survey pH value, cross 0.45 μ m microporous filter membrane, aseptic subpackaged to control injection vial, lyophilization, gland, roll lid, promptly get the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.(remarks: regulate pH value and use the 1mol/L sodium hydroxide solution, or use the 1mol/L hydrochloric acid solution.)
(5) freeze drying process is as follows:
The a pre-freeze stage: select pre-freeze repeatedly, pre-freeze temperature-56 ℃～-45 ℃, the temperature of rising again-11 ℃～-10 ℃, about 4～7 hours of pre-freeze time.
The b sublimation drying stage: when the temperature of condenser reduce to-56 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-21 ℃～-19 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water.
C is drying stage again: shelf temperature slowly rises to 0 ℃～20 ℃, judges exsiccant terminal point with the vacuum descent method again, again 4～11 hours drying times.
Embodiment 4
Prescription:
Prescription 28
Radix Ginseng (Radix Ginseng Rubra) 1000g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g
Mannitol 150g
Add the injection water extremely 3000ml
Make 1000 injection ginsengs attached (lyophilizing)
Preparation method:
(1) Radix Ginseng adds 6 times of amounts (because be dried medical material, adding 1 times of amount for the first time), 70% alcohol reflux 2 times, each 2 hours respectively, filter, filtrate recycling ethanol is not to there being the alcohol flavor, and medicinal liquid is concentrated into every 1ml and is equivalent to contain Radix Ginseng crude drug 1.45～1.55g, and the water-saturated n-butanol that at every turn adds equivalent extracts 6 times, merge n-butyl alcohol liquid, reclaim n-butyl alcohol to there not being the n-butyl alcohol flavor, change water-soluble, filter, get ginsenoside's intermediate, standby;
(2) the Radix Ginseng Rubra medicinal residues after the alcohol extraction add 9 times of water gagings (for the first time because be dried medical material, add 3 times of amounts) respectively and decoct and extract or reflux, extract, 3 times each 2 hours, filter, being concentrated into liquor strength is 0.75～0.85g crude drug/ml, and temperature is 37～43 ℃, adds 95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 80%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, merge supernatant, adjusting the supernatant liquor strength is 1.95～2.05g crude drug/ml, adds 95% ethanol and makes and contain the alcohol amount and reach 70%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, get supernatant, supernatant earlier with the ultrafilter membrane ultrafiltration of molecular cut off 500K, is removed small-molecular weight impurity with the gained ultrafiltrate with the ultrafilter membrane of molecular cut off 5K then, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
(3) the Radix Aconiti Lateralis Preparata medical material adds 8 times of water gagings (for the first time because be dried medical material, add 2 times of amounts), decoct extraction or reflux, extract, 3 times, each 1 hour, filter, be concentrated into relative density of medicine liquid 1.09～1.13,47～53 ℃ of temperature, the ethanol precipitate with ethanol with 95% makes ethanol content reach 70%, standing over night, filter, filtrate recycling ethanol is not to there being the alcohol flavor, adjusts medicinal liquid and is equivalent to contain Radix Ginseng crude drug 0.95～1.05g to every 1ml, adds the dichloromethane extraction 5 times of 1.0～1.1 times of amounts at every turn, combined dichloromethane liquid, reclaim dichloromethane to the greatest extent, change water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
(4) intermediate with " (1) ", " (2) ", " (3) " gained merges, and mixing adds to the full amount of water for injection, and regulates pH value to 7.0, is heated to and boils, and keep little and boiled 40～45 minutes, cooling, cold preservation is spent the night; Filter, add mannitol/or sorbitol in the filtrate, stir evenly, regulate pH value to 7.0, add 0.1% active carbon and be heated to and boil, keep little and boiled 20～25 minutes, filter, regulate pH value to 7.0, add the injection water and adjust volume to full dose, survey pH value, cross 0.45 μ m microporous filter membrane, aseptic subpackaged to control injection vial, lyophilization, gland, roll lid, promptly get the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.(remarks: regulate pH value and use the 1mol/L sodium hydroxide solution, or use the 1mol/L hydrochloric acid solution.)
(5) freeze drying process is as follows:
The a pre-freeze stage: select pre-freeze repeatedly, pre-freeze temperature-56 ℃～-45 ℃, the temperature of rising again-11 ℃～-10 ℃, about 4～7 hours of pre-freeze time.
The b sublimation drying stage: when the temperature of condenser reduce to-56 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-21 ℃～-19 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water.
C is drying stage again: shelf temperature slowly rises to 0 ℃～20 ℃, judges exsiccant terminal point with the vacuum descent method again, again 4～11 hours drying times.
Embodiment 5
Prescription:
Prescription 29 Prescription 30 Prescription 31 Prescription 32 Prescription 33
Radix Ginseng (Radix Ginseng Rubra) 1000g 1200g 1500g 900g 800g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g 1800g 1500g 2100g 2200g
Mannitol 150g 150g 200g 250g 300g
Add the injection water extremely 3000ml 3000ml 3000ml 3000ml 3000ml
Make 1000 injection ginsengs attached (lyophilizing)
Preparation method:
(1) Radix Ginseng adds 6 times of amounts (because be dried medical material, adding 1 times of amount for the first time), 70% alcohol reflux 2 times, each 2 hours respectively, filter, filtrate recycling ethanol is not to there being the alcohol flavor, and medicinal liquid is concentrated into every 1ml and is equivalent to contain Radix Ginseng crude drug 1.45～1.55g, and the water-saturated n-butanol that at every turn adds equivalent extracts 6 times, merge n-butyl alcohol liquid, reclaim n-butyl alcohol to there not being the n-butyl alcohol flavor, change water-soluble, filter, get ginsenoside's intermediate, standby;
(2) the Radix Ginseng Rubra medicinal residues after the alcohol extraction add 9 times of water gagings (for the first time because be dried medical material, add 3 times of amounts) respectively and decoct and extract or reflux, extract, 3 times each 2 hours, filter, being concentrated into liquor strength is 0.75～0.85g crude drug/ml, and temperature is 37～43 ℃, adding 95% ethanol precipitate with ethanol makes and contains the alcohol amount and reach 80%, standing over night filters, and precipitation is dry, after adding the suitable quantity of water dissolving, centrifugal, merge supernatant, adjusting the supernatant liquor strength is 1.95～2.05g crude drug/ml, adding 95% ethanol makes and contains the alcohol amount and reach 70%, standing over night filters, and precipitation is dry, after adding the suitable quantity of water dissolving, centrifugal, get supernatant
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 100K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 200K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 500K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 200K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 5K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 200K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 2K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
Or the molecular cut off 500K ultrafilter membrane ultrafiltration of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 2K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
(3) the Radix Aconiti Lateralis Preparata medical material adds 8 times of water gagings (because be dried medical material, adding 2 times of amounts for the first time), decocts extraction or reflux, extract, 3 times, each 1 hour, filter, be concentrated into relative density of medicine liquid 1.09～1.13,47～53 ℃ of temperature, the ethanol precipitate with ethanol with 95% makes ethanol content reach 70%, standing over night, filter, filtrate recycling ethanol does not extremely have the alcohol flavor, and the relative density of adjusting medicinal liquid is that the ethanol of adding 95% in 1.27～1.31 o'clock makes pure content reach 85%, standing over night, filter, filtrate recycling ethanol filters to there not being the alcohol flavor, and filtrate being heated to boiled, keeping little boiled 35～45 minutes, cooling, cold preservation filters, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
Or for the first time alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor, adjusts medicinal liquid and is equivalent to contain Radix Ginseng crude drug 0.95～1.05g to every 1ml, adds the dichloromethane extraction 5 times of 1.0～1.1 times of amounts at every turn, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
(4) intermediate with " (1) ", " (2) ", " (3) " gained merges, and mixing adds to the full amount of water for injection, and regulates pH value to 7.0, is heated to and boils, and keep little and boiled 40～45 minutes, cooling, cold preservation is spent the night; Filter, add mannitol in the filtrate, stir evenly, regulate pH value to 7.0, add 0.1% active carbon and be heated to and boil, keep little and boiled 20～25 minutes, filter, regulate pH value to 7.0, add the injection water and adjust volume to full dose, survey pH value, cross 0.45 μ m microporous filter membrane, aseptic subpackaged to control injection vial, lyophilization, gland, roll lid, promptly get the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(5) freeze drying process is as follows:
The a pre-freeze stage: select pre-freeze repeatedly, pre-freeze temperature-56 ℃～-45 ℃, the temperature of rising again-11 ℃～-10 ℃, about 4～7 hours of pre-freeze time.
The b sublimation drying stage: when the temperature of condenser reduce to-56 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-21 ℃～-19 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water.
C is drying stage again: shelf temperature slowly rises to 0 ℃～20 ℃, judges exsiccant terminal point with the vacuum descent method again, again 4～11 hours drying times.
Embodiment 6
Prescription:
Prescription 34 Prescription 35 Prescription 36 Prescription 37 Prescription 38
Radix Ginseng (Radix Ginseng Rubra) 1000g 1000g 1000g 1000g 700g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g 2000g 2000g 2000g 1400g
Naloxone hydrochloride 0.08g 0.10g 0.12g 0.15g 0.15g
Mannitol 150g 150g 175g 175g 120g
Add the injection water extremely 3000ml 3000ml 3000ml 3000ml 3000ml
Make 1000 freeze-drying preparation for injection
Prescription 39 Prescription 40 Prescription 41 Prescription 42 Prescription 43
Radix Ginseng (Radix Ginseng Rubra) 1000g 1000g 1000g 1000g 700g
Radix Aconiti Lateralis Preparata (Radix Aconiti Lateralis Preparata) 2000g 2000g 2000g 2000g 1400g
Dopamine hydrochloride 4g 6g 8g 10g 10g
Mannitol 150g 150g 175g 175g 120g
Add the injection water extremely 3000ml 3000ml 3000ml 3000ml 3000ml
Make 1000 freeze-drying preparation for injection
Preparation method:
(1) Radix Ginseng adds 6 times of amounts (because be dried medical material, adding 1 times of amount for the first time), 70% alcohol reflux 2 times, each 2 hours respectively, filter, filtrate recycling ethanol is not to there being the alcohol flavor, and medicinal liquid is concentrated into every 1ml and is equivalent to contain Radix Ginseng crude drug 1.45～1.55g, and the water-saturated n-butanol that at every turn adds equivalent extracts 6 times, merge n-butyl alcohol liquid, reclaim n-butyl alcohol to there not being the n-butyl alcohol flavor, change water-soluble, filter, get ginsenoside's intermediate, standby;
(2) the Radix Ginseng Rubra medicinal residues after the alcohol extraction add 9 times of water gagings (for the first time because be dried medical material, add 3 times of amounts) respectively and decoct and extract or reflux, extract, 3 times each 2 hours, filter, being concentrated into liquor strength is 0.75～0.85g crude drug/ml, and temperature is 37～43 ℃, adding 95% ethanol precipitate with ethanol makes and contains the alcohol amount and reach 80%, standing over night filters, and precipitation is dry, after adding the suitable quantity of water dissolving, centrifugal, merge supernatant, adjusting the supernatant liquor strength is 1.95～2.05g crude drug/ml, adding 95% ethanol makes and contains the alcohol amount and reach 70%, standing over night filters, and precipitation is dry, after adding the suitable quantity of water dissolving, centrifugal, get supernatant
Previous step gained supernatant adopts the ultrafilter membrane ultrafiltration of molecular cut off 100K, and ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 200K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of previous step gained supernatant employing molecular cut off 500K, ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 200K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 5K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 200K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 2K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 500K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 2K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
Or the ultrafilter membrane ultrafiltration of molecular cut off 500K of previous step gained supernatant elder generation, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 5K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby;
(3) the Radix Aconiti Lateralis Preparata medical material adds 8 times of water gagings (because be dried medical material, adding 2 times of amounts for the first time), decocts extraction or reflux, extract, 3 times, each 1 hour, filter, be concentrated into relative density of medicine liquid 1.09～1.13,47～53 ℃ of temperature, the ethanol precipitate with ethanol with 95% makes ethanol content reach 70%, standing over night, filter, filtrate recycling ethanol does not extremely have the alcohol flavor, and the relative density of adjusting medicinal liquid is that the ethanol of adding 95% in 1.27～1.31 o'clock makes pure content reach 85%, standing over night, filter, filtrate recycling ethanol filters to there not being the alcohol flavor, and filtrate being heated to boiled, keeping little boiled 38～45 minutes, cooling, cold preservation filters, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
Or for the first time alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor, adjusts medicinal liquid and is equivalent to contain Radix Ginseng crude drug 0.95～1.05g to every 1ml, adds the dichloromethane extraction 5 times of 1.0～1.1 times of amounts at every turn, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby;
(4) intermediate with " (1) ", " (2) ", " (3) " gained merges, and mixing adds to the full amount of water for injection, and regulates pH value to 7.0, is heated to and boils, and keep little and boiled 40～45 minutes, cooling, cold preservation is spent the night; Filter, add in the filtrate naloxone hydrochloride/or dopamine hydrochloride (naloxone hydrochloride/or dopamine hydrochloride be dissolved in the low amounts of water, adjust about pH value of water solution to 7, and filter), mannitol, stir evenly, regulate about pH value to 7.0, add 0.1% active carbon and be heated to and boil, keeping little boiled 20～25 minutes, filter, regulate about pH value to 7.0, add the injection water and adjust volume to full dose, survey pH value, cross 0.45 μ m microporous filter membrane, aseptic subpackaged to control injection vial, lyophilization, gland, roll lid, promptly get the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material.
(5) freeze drying process is as follows:
The a pre-freeze stage: pre-freeze temperature-56 ℃～-45 ℃, about 2.5～6 hours of pre-freeze time; Or select pre-freeze repeatedly, pre-freeze temperature-56 ℃～-45 ℃, the temperature of rising again-11 ℃～-10 ℃, about 4～7 hours of pre-freeze time.
The b sublimation drying stage: when the temperature of condenser reduce to-56 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-21 ℃～-19 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water.
C is drying stage again: shelf temperature slowly rises to 0 ℃～20 ℃, judges exsiccant terminal point with the vacuum descent method again, again 4～11 hours drying times.
Embodiment 7
It below is the Pharmacodynamic test of active extract of pharmaceutical preparation of the present invention.
1 experiment purpose and content
Clinical application according to injection ginseng attached (lyophilizing), on animal, carry out the main pharmacodynamics checking, mainly carry out following experiment: injection ginseng attached (lyophilizing) high, medium and low dosage group is to the influence of anoxia enduring time-to-live of mice normal pressure, hypophysis is held the influence of Acute Myocardial Ischemia in Rats due to the foline (Pit), influence to acute rat blood stasis model, influence to the isolated rat heart ischemical reperfusion injury, to the influence of myocardial infarction due to the anesthetized dog coronary ligation, to the hemodynamic influence of anesthetized dog.
2 experiment materials
2.1 be subjected to the reagent thing: injection ginseng attached (lyophilizing) is provided lot number by Xuanhong Medicine Technology Co., Ltd., Tianjin: 040710, and adopt the embodiment of the invention 3 used prescriptions and preparation method to make.Face with preceding 0.9% sodium chloride injection and be mixed with needed concentration with certain volume.
2.2 medicine and reagent:
SHENFU ZHUSHEYE, Sanjiu Pharmaceutical Industry Co., Ltd., Ya'an City, lot number: 041103 (being the used medicine of the attached former medicine group of the ginseng of indication in this experiment);
Radix Salviae Miltiorrhizae Injection, Zhengda Qingchunbao Pharmaceutical Co., Ltd, lot number: 0405202;
Nitroglycerin injection, Beijing Yimin Pharmaceutical Co., Ltd., lot number: 040217;
Heparin sodium injection, the biological thousand red pharmaceutical Co. Ltds in Changzhou, lot number: 040615;
Sodium citrate, the sincere chemical reagent company limited in Shanghai, lot number: 040128;
Adrenalin hydrochloride, Tianjin gold credit aminoacid company limited, lot number: 0403031;
Pentobarbital sodium, Beijing chemical reagents corporation, lot number: 020402;
Pituitrin, Shanghai No.1 Bio-Chemical Pharmacetical Industry Co., Ltd, lot number: 040501;
Urethane, Shanghai chemical reagents corporation of China Drug Co., lot number 20021104;
Lactic acid dehydrogenase (LDH-L) test kit, the medical company limited of sysmex Jinan Sysmex, lot number: ZG4001;
Creatine kinase (CK) test kit, the medical company limited of sysmex Jinan Sysmex, lot number: ZG4004;
Sodium chloride, AR level, the sincere chemical reagent company limited in Shanghai, lot number: 050705;
Potassium chloride, AR level, Shanghai Ling Feng chemical reagent company limited, lot number: 021031;
Calcium chloride, AR level, Shanghai Ling Feng chemical reagent company limited, lot number: 030619;
Sodium bicarbonate, AR level, last marine rainbow photoinitiator chemical factory, lot number: 030604;
Glucose, the AR level, last Nereid analyses Chemical Industry Science Co., Ltd, lot number: 030823;
Activated partial prothrombin time (APTT) is measured test kit, Beijing Steellex Scientific Instrument Company's lot number: ST20201-20;
Fibrinogen (FIB) is measured test kit, Beijing Steellex Scientific Instrument Company's lot number: ST20401-16;
Prothrombin time (PT) is measured test kit, Beijing Steellex Scientific Instrument Company's lot number: ST10101-22;
Rockwell liquid preparation: NaCl 9.0g/L, KCl 0.42g/L, CaCl 20.24g/L, NaHCO 30.2g/L, Glu 1.0g/L;
Formaldehyde: Nanjing chemical reagent factory, lot number: 20040721.
2.3 experimental apparatus
RM-6000 eight road physiology monitor and adnexaes, Japanese NIHON KOHDEN company;
The DH140 artificial ventilator, Zhejiang Medical university medical instrument trial (demonstration) plant; DHG-9053A type Constant Temp. Oven, the medical thermostatic equipment in Shanghai factory; The last ware electronic balance of FA2104, Shanghai balance equipment factory; JNA type precision torsion balance, Shanghai Second Balance Factory; Electrocardiograph, Japanese NIHON KOHDEN company; Vitalab 200 type automatic clinical chemistry analyzers, Dutch vital scientific company; BT01-100 type constant flow pump, Baoding LanGe constant flow pump Co., Ltd; Perfusion device: self-designed L angendorff perfusion device;
LMS-2B type two road physiology monitors, Chengdu Instruement Factory; The WC/09-05 temperature chamber, Chinese Chongqing Yinhe Experimental Equipment Co., Ltd..
2.4 animal
Kunming mouse, body weight 18～22g; The SD rat, body weight 180-220g; Rabbit, body weight 2.0～2.4kg, male and female half and half are supplied with by Shanghai Si Laike laboratory animal responsibility company limited.The quality certification: SCXK (Shanghai) 2003-0003.
The hybrid dog, body weight 7-10kg, the male and female dual-purpose by the hybrid dog of Nanjing University of Traditional Chinese Medicine's animal center purchase from suburbs, is raised and train at this center before the experiment, after quarantine and anthelmintic, is used for this experiment.
3 experimental techniques and result
Data are represented (X ± S) with mean ± standard deviation.All measurement datas adopt the Student-t check, and enumeration data adopts rank test.
Before rate of change (incidence rate) %=(after the X administration-X administration before)/X administration * 100%
Be observation after the administration after the X administration, be observation before the administration before the X administration.
3.1 influence to mice normal pressure anoxia enduring
3.1.1 experimental technique: get 120 of Kunming mouses, male and female half and half, be divided into 6 groups at random, promptly matched group (giving isometric normal saline), positive drug group (Radix Salviae Miltiorrhizae Injection 20.0g crude drug/kg), the attached former medicine group of ginseng (and 3.2g crude drug/kg), injection ginseng attached (lyophilizing) high, medium and low (3.2,1.6,0.8g crude drug/kg).Each group is all by the tail intravenously administrable, and the administration volume is 20ml/kg.Behind the intravenously administrable 15min, mice is placed the airtight wide mouthed bottle of 125ml (built-in sodica calx 20g).Stopping with the mice mouth breathing is the dead mouse sign, the time-to-live of record mice, surpass 60min in 60min.
3.1.2 experimental result:
Experimental result sees Table 1.
Table 1 injection ginseng attached (lyophilizing) is to the protective effect of mice normal pressure anoxia enduring (X ± SD)
Group Dosage (the g crude drug/kg) Number of animals (only) Breathing hold time (min)
Matched group Radix Salviae Miltiorrhizae Injection group is joined attached low dose group ginseng attached middle school dosage group and is joined attached high dose group and join attached former medicine group - 20.0 0.8 1.6 3.2 3.2 20 20 20 20 20 20 26.97±8.44 35.28±7.85 ** 30.86±5.80 32.02±6.65 * 35.63±7.36 ** 31.48±5.19 *▲
Compare with matched group: *P＜0.05, *P＜0.01.
Compare with injection ginseng attached (lyophilizing) high dose group: ▲: P＜0.05.
The result shows, injection is joined attached (lyophilizing) height, middle dosage group and is joined the time-to-live that attached former medicine group all can prolong mice normal pressure anoxia enduring, with remarkable (P＜0.01 of matched group comparing difference, P＜0.05, and join attached (lyophilizing) high dose group mice time-to-live and significantly be longer than former medicine group (P＜0.05) P＜0.05).
3.2 influence to rat heart muscle ischemia due to the pituitrin (Pit)
3.2.1 experimental technique: get 56 of healthy SD rats, male and female half and half, body weight 180～220g, be divided into 7 groups at random, that is: normal group, model group (above two groups give isometric normal saline), positive drug group (nitroglycerin 0.5mg/kg), injection ginseng attached (lyophilizing) basic, normal, high (0.4,0.8,3 dosage groups of 1.6g crude drug/kg), the attached former medicine group of ginseng (1.6g crude drug/kg).Water 12h is can't help in the rat fasting before the experiment, and dorsal position is got in lumbar injection urethane 1g/kg anesthesia, and it is subcutaneous that needle electrode carefully thrusts extremity, measures the II lead electrocardiogram, and 15min is stablized in the operation back, and the record normal ECG.Each treated animal is respectively through the tail vein injection administration, and behind the 15min, except that the normal control group, each group is no more than 5s inject time respectively through sublingual vein injection of pituitrin 1.5u/kg.The electrocardiogram of 15s, 30s, 1min, 2min, 3min, 5min, 7min, 10min, 15min behind the immediate record injection of pituitrin.
Observation index mainly is decided to be the variation of T wave height, changes in heart rate.It is baseline that the T wave height is measured with the PR section, and every time point is surveyed 3 successive wave modes, gets its meansigma methods.Record also calculates the changing value (no matter raise or reduce, get the absolute value of variation) of T wave height.For the index that dynamic observes of sequential relationships such as variation of T wave height and heart rate, on each time point, with t check between the changing value work group after the medication.
20.3.2.2 experimental result:
Experimental result sees Table 2, table 3.
(1) the attached influence to rats with myocardial ischemia ECG T wave displacement due to the pituitrin of injection ginseng sees Table 2.
(X ± S) (n=8) of the influence of rats with myocardial ischemia ECG T wave due to the table 2 pair pituitrin
Group Dosage T(mv)
Before the modeling After the modeling
15s 30s 1min 2min 3min 5min 7min 10min 15min
The former medicine group of dosage group rate of change % Shenfu high dose group rate of change % Shenfu rate of change % in the normal group rate of change % model group rate of change % monobel group rate of change % Shenfu low dose group rate of change % Shenfu -- -- 0.5 mg /kg 0.4 g/k g 0.8 g/k g 1.6 g/k g 1.6 g/k g 0.22 ±0.04 0.21 ±0.08 0.28 ±0.08 0.26 ±0.07 0.23 ±0.04 0.27 ±0.05 0.28 ±0.09 0.21 ±0.05 * 6.72 ±7.40 ** 0.45 ±0.31 216.89 ±133.84 0.34 ±0.08 39.51 ± 31.91 ** 0.41 ±0.13 63.75 ± 32.82 ** 0.29 ±0.09 42.62 ± 30.15 ** 0.27 ±0.10 19.67 ± 16.87 ** 0.36 ± 0.15 36.45 ±10.84 *** 0.21 ±0.05 3.32 ±4.61 * 0.09 ±0.30 137.40 ±129.83 0.25 ±0.09 19.34 ±19.27 * 0.28 ±0.23 76.19 ±54.93 0.29 ±0.07 26.64 ±27.43 * 0.28 ±0.03 15.79 ±11.15 * 0.30 ±0.10 29.59 ±13.32 ** 0.21 ±0.05 4.95 ±5.49 * 0.23 ±0.11 51.22 ±50.54 0.18 ±0.16 46.96 ±48.93 0.25 ±0.10 28.08 ±16.37 0.24 ±0.08 27.92 ±20.08 0.24 ±0.06 20.17 ±15.81 0.23 ±0.15 42.00 ±23.15* 0.21 ±0.04 2.65 ±4.59 0.24 ±0.05 31.79 ±45.61 0.18 ±0.16 43.52 ±60.21 0.26 ±0.07 17.29 ±15.82 0.26 ±0.06 30.59 ±26.67 0.23 ±0.07 18.93 ±17.37 0.22 ±0.12 35.29 ±21.84 0.21 ±0.05 * 6.61 ±6.06 * 0.27 ±0.04 46.51 ±39.70 0.19 ±0.19 51.30 ±59.32 0.26 ±0.05 14.56 ±14.35 0.27 ±0.05 27.40 ±21.53 0.27 ±0.08 12.46 ±15.04 * 0.25 ±0.12 31.77 ±16.26 * 0.22 ±0.04 4.60 ±4.86 * 0.24 ±0.05 31.60 ±26.75 0.24 ±0.12 20.33 ±9.68 0.29 ±0.04 26.91 ±15.75 0.29 ±0.08 40.13 ±26.76 0.29 ±0.08 19.73 ±9.96 0.27 ±0.11 19.13 ±14.24 0.22 ±0.04 3.99 ±3.79 * 0.24 ±0.06 24.88 ±22.79 0.24 ±0.09 22.49 ±13.07 0.27 ±0.04 20.98 ±17.97 0.30 ±0.18 52.10 ±54.52 0.25 ±0.08 20.34 ±16.91 0.27 ±0.12 24.94 ±18.75 0.22 ±0.05 4.64 ±6.66 * 0.26 ±0.07 ±36.24 ±39.83 0.26 ±0.10 12.17 ±13.51 0.29 ±0.05 22.59 ±22.66 0.25 ±0.09 51.80 ±25.35 0.26 ±0.07 17.63 ±10.15 0.26 ±0.10 20.62 ±14.77 0.21 ±0.04 * 2.65 ±4.59 * 0.27 ±0.07 42.04 ±41.43 0.27 ±0.12 14.80 ±13.35 0.28 ±0.06 18.15 ±16.07 0.27 ±0.07 36.13 ±37.19 0.27 ±0.06 13.35 ±4.57 0.29 ±0.11 17.54 ±20.76
Compare with model group *P＜0.05, *P＜0.01; Compare * P＜0.05 with the attached high dose group of ginseng
Behind the rats in normal control group intravenous injection pituitrin, electrocardio takes place obviously to change, and the first phase, (1～30s) tangible T wave height occurs alarmmed; The second phase (after the 30s), present the low flat or inversion of tangible T ripple.
Injection ginseng attached (lyophilizing) high, medium and low dosage and former medicine group can significantly be resisted the electrocardio T ripple displacement (P＜0.01) that pituitrin causes at the 15s time point, join attached (lyophilizing) height, middle dosage and former medicine group and can significantly resist the electrocardio T ripple displacement (P＜0.05) that pituitrin causes at the 30s time point, and there is certain dose-effect relationship between the high, medium and low dosage, joins attached high dose group and compare in 15s, 30s, 1min, 5min also significant difference (P＜0.05) with former medicine.
(2) the attached influence to rats with myocardial ischemia heart rate due to the pituitrin of injection ginseng the results are shown in Table 3.
(X ± S) (n=8) of the influence of rats with myocardial ischemia heart rate due to the table 3 pair pituitrin
Group Dosage Heart rate (beats/min)
Before the modeling After the modeling
15s 30s 1min 2min 3min 5mim 7min 10min 15min
The former medicine group of dosage group rate of change % Shenfu high dose group rate of change % Shenfu rate of change % in the normal group rate of change % model group rate of change % monobel group rate of change % Shenfu low dose group rate of change % Shenfu -- -- 0.5 mg/kg 0.4 g/kg 0.8 g/kg 1.6 g/kg 1.6 g/kg 402.4 ±69.1 422.9 ±64.2 338.4 ±73.6 377.4 ±61.8 355.1 ±67.1 405.0 ±33.5 345.0 ±59.5 399.4 ±69.6 0.011 ±0.010 * 388.0 ±80.6 0.088 ±0.091 294.4 ±66.5 0.202 ±0.132 332.8 ±58.8 0.183 ±0.151 299.3 ±93.3 0.175 ±0.166 351.5 ±46.1 0.131 ±0.101 275.6 ±76.1 0.205 ±0.138 402.3 ±71.7 0.008 ±0.013 ** 344.6 ±76.3 0.180 ±0.155 252.4 ±70.5 0.259 ±0.182 293.5 ±87.9 0.253 ±0.215 248.0 ±69.2 0.276 ±0.232 278.5 ±67.9 0.306 ±0.182 280.1 ±71.1 0.189 ±0.149 406.0 ±62.4 ** 0.023 ±0.041 ** 233.6 ±58.8 0.443 ±0.125 240.0 ±69.2 0.300 ±0.187 246.1 ±85.4 0.358 ±0.237 233.4 ±57.8 0.316 ±0.214 280.0 ±66.4 0.305 ±0.169 253.9 ±84.1 0.254 ±0.220 408.4 ±61.3 ** 0.026 ±0.054 ** 254.4 ±53.6 0.398 ±0.096 238.5 ±58.5 0.286 ±0.181 252.5 ±57.2 0.312 ±0.211 258.3 ±44.8 0.250 ±0.173 250.8 ±62.3 0.379 ±0.147 248.8 ±58.4 0.274 ±0.131 * 408.4 ±61.3 ** 0.026 ±0.054 ** 276.9 ±30.0 0.336 ±0.096 245.1 ±44.7 0.257 ±0.160 263.4 ±47.0 0.286 ±0.175 262.1 ±38.9 0.247 ±0.127 278.1 ±43.8 0.312 ±0.105 248.0 ±58.2 0.277 ±0.127 407.1 ±59.6 ** 0.023 ±0.055 ** 268.1 ±45.4 0.363 ±0.078 255.1 ±49.5 0.235 ±0.135 * 271.0 ±48.7 0.274 ±0.165 236.4 ±53.2 0.308 ±0.202 279.1 ±42.3 0.310 ±0.100 255.8 ±56.8 0.252 ±0.137 411.3 ±48.5 ** 0.044 ±0.111 ** 279.3 ±27.6 0.332 ±0.071 262.1 ±38.3 0.213 ±0.136 * 276.3 ±44.9 0.250 ±0.174 259.3 ±28.0 0.253 ±0.121 294.5 ±31.8 0.272 ±0.072 265.6 ±51.4 0.222 ±0.130 410.1 ±45.6 ** 0.046 ±0.111 ** 284.4 ±30.7 0.320 ±0.071 259.9 ±30.3 0.235 ±0.135 277.9 ±46.1 0.249 ±0.159 269.1 ±34.5 0.305 ±0.299 305.5 ±33.1 0.245 ±0.069 * 269.6 ±54.3 0.209 ±0.144 409.3 ±46.9 ** 0.048 ±0.110 ** 306.1 ±47.1 0.267 ±0.114 275.6 ±18.7 0.196 ±0.124 306.8 ±51.8 0.244 ±0.109 261.4 ±74.1 0.342 ±0.354 342.4 ±45.5 0.179 ±0.068 303.3 ±62.5 0.151 ±0.133
Compare with model group *P＜0.05, *P＜0.01;
Behind the rats in normal control group intravenous injection pituitrin, the rat heart rate is obviously slowed down, not recover yet to 15min.Injection ginseng attached (lyophilizing) high dose can be alleviated the decreased heart rate due to the pituitrin when 10min, join attached former medicine and when 2min, can alleviate decreased heart rate due to the pituitrin, with model group than difference remarkable (P＜0.05), compare with the attached former medicine of ginseng the using of alleviating due to the pituitrin of decreased heart rate and there is no significant difference (P＞0.05) but join attached (lyophilizing) high dose.
3.2.3 conclusion:
Injection ginseng attached (lyophilizing) high, medium and low dosage and former medicine can obviously resist between the variation of the electrocardiogram T section that pituitrin causes and the high, medium and low dosage and have certain dose-effect relationship, join attached (lyophilizing) high dose and former medicine and can alleviate decreased heart rate due to the pituitrin at some time point, prompting injection ginseng attached (lyophilizing) has the effect of myocardial ischemia due to certain antagonism pituitrin.
3.3 to the hemorheological influence of stasis syndrome rat model
3.3.1 experimental implementation method:
Get 56 of SD male rats, body weight 350～450g, be divided into 7 groups at random, it is the normal control group, model group (above two groups give isometric normal saline), positive drug group (10g crude drug/kg Radix Salviae Miltiorrhizae Injection), injection ginseng attached (lyophilizing) is low, in, high (0.4,0.8,1.6g 3 dosage groups of crude drug/kg), join attached former medicine group (1.6g crude drug/kg), the administration volume is 5ml/kg, vein successive administration 3 days, after the 2nd administration, except that the normal control group, each treated animal subcutaneous injection adrenalin hydrochloride injection (Adr) 0.8ml/kg totally 2 times, interval 4h, before injecting Adr for the second time, place frozen water to soak 5min rat, cause the Blood stasis model.Fasting, the 15min administration is 1 time before operation, with 10% chloral hydrate anesthesia (300mg/kg, ip), face upward the position and fix the blood-letting of carotid artery intubate, sodium citrate with 3.8% or heparin sodium normal saline solution (500ul/ml) detect hemorheological every index by anticoagulant in 1: 9.
3.3.2 experiment detects index
(1) to the influence of rat platelet aggregation rate
Sodium citrate anticoagulant blood with 3.8% is with the centrifugal 10min of 800r/min, get platelet rich plasma (PRP), remainder is centrifugal with 3000r/min, get platelet poor plasma (PPP), utilize LG-PABER type platelet aggregation and thrombin analyser record maximum agglutination rate and calculate suppression ratio by following formula.
Assemble suppression ratio (%)
=[(model group maximum agglutination rate-administration group maximum agglutination rate)/model group maximum agglutination rate] * 100%
(2) blood viscosity detects:
1. whole blood viscosity: adopt LG-R-80 series blood viscosity instrument to measure the blood of anticoagulant heparin.
2. plasma viscosity: after the whole blood test finished, the residue blood sample was centrifugal with 3000r/min, carries out the test of plasma viscosity.
3. erythrocyte deformability, erythrocyte aggregation ability detect: the blood of anticoagulant heparin is adopted LG-R-190 type red blood cell deformation/ability of aggregation analyzer.
(3) detection of blood coagulation system:
Sodium citrate anticoagulant blood with 3.8% is with the centrifugal 10min of 3000r/min, separate platelet poor plasma, adopt LG-PABER type platelet aggregation and thrombin analysis-e/or determining activated partial prothrombin time (APTT), prothrombin time (PT), plasma fibrinogen (FIB).
3.3.3 experimental result
Experimental result sees Table 4～table 7.
(1) injection ginseng attached (lyophilizing) the results are shown in Table 4 to the influence of Blood stasis rat model platelet aggregation rate.
Table 4 injection ginseng attached (lyophilizing) is to the influence of Blood stasis rat model platelet aggregation rate (X ± s)
Group Dosage (the g crude drug/kg) Number of animals (only) Platelet maximum agglutination rate (%) Assemble suppression ratio (%)
The former medicine group of dosage group Shenfu high dose group Shenfu in the Normal group model group danshen injections group Shenfu low dose group Shenfu - - 10.0 0.4 0.8 1.6 1.6 8 8 8 8 8 8 8 22.69±6.57 ** 35.45±6.91 26.49±4.99 * 32.86±5.69 27.59±6.56 * 27.04±4.64 * 27.71±6.28 * - - 26.68 14.13 14.99 26.98 14.56
Compare with model group: *P＜0.01, *P＜0.05.
(2) injection ginseng attached (lyophilizing) the results are shown in Table 5 to the influence of Blood stasis rat model red blood cell deformation and ability of aggregation.
Table 5 injection ginseng attached (lyophilizing) is to the influence of Blood stasis rat model red blood cell deformation and ability of aggregation (X ± s)
Group Dosage g/kg The animal number of elements The erythrocyte aggregation ability Erythrocyte deformability
MAXD SS MAXDI SSS
The former medicine group of dosage group Shenfu high dose group Shenfu in the Normal group model group danshen injections group Shenfu low dose group Shenfu - - 10.0 0.4 0.8 1.6 1.6 8 8 8 8 8 8 8 1.29±0.74 * 1.89±0.58 1.23±0.31 * 1.67±0.36 1.37±0.43 1.41±0.20 * 1.43±0.19 264.9±33.7 * 387.0±114.2 257.4±61.8 * 341.5±73.3 285.3±89.0 292.9±39.0 * 298.8±37.4 0.479±0.028 0.444±0.067 0.469±0.023 0.450±0.053 0.450±0.043 0.455±0.060 0.466±0.044 236.8±19.0 219.6±37.2 235.8±13.3 217.6±39.2 217.5±30.6 221.4±34.8 233.5±23.2
Compare with model group: *P＜0.05; *P＜0.01.
(3) injection ginseng attached (lyophilizing) the results are shown in Table 6 to the influence of Blood stasis rat model APTT, PT, FIB.
Table 6 injection ginseng attached (lyophilizing) is to the influence of Blood stasis rat model APTT, PT, FIB (n=8, X ± s)
Group Dosage (g/kg) APTT (s) PT(s) FIB(g/l)
Normal control group model group Radix Salviae Miltiorrhizae Injection group is joined attached low dose group - - 10.0 0.4 27.56±3.86 * 21.74±6.59 27.13±4.91 25.16±4.77 21.49±3.04 * 17.39±3.16 21.23±2.08 * 18.50±3.04 1.25±0.19 * 1.63±0.36 1.27±0.24 1.58±0.47
Ginseng attached middle school dosage group is joined attached high dose group and is joined attached former medicine group 0.8 1.6 1.6 26.70±3.51 27.61±5.69 25.40±3.85 18.94±3.05 20.90±2.31 * 21.16±2.14 * 1.49±0.26 1.25±0.21 * 1.47±0.40 *
Compare with model group: *P＜0.05.
(4) injection ginseng attached (lyophilizing) the results are shown in Table 7 to the influence of Blood stasis rat model blood viscosity.
Table 7 injection ginseng attached (lyophilizing) is to the influence of Blood stasis rat model blood viscosity (n=8, X ± s)
Group Dosage g/kg Whole blood viscosity (CP) Plasma viscosity (100s -1)
Height is cut (200s -1) In cut (30s -1) In cut (5s -1) Low (the 1s that cuts -1)
The former medicine group of dosage group Shenfu high dose group Shenfu in the Normal group model group danshen injections group Shenfu low dose group Shenfu - - 10 0.4 0.8 1.6 1.6 4.83±0.47 5.27±0.61 5.02±0.40 5.03±0.34 5.10±0.22 4.94±0.77 5.09±0.22 6.09±0.43 6.43±0.82 5.80±0.52 6.17±0.46 6.18±0.73 6.05±1.14 6.13±0.75 9.90±2.20 10.54±0.71 9.58±1.46 12.57±4.37 10.65±1.35 10.28±1.50 10.83±1.29 20.24±3.93 ** 26.20±3.16 21.36±3.17 ** 25.31±7.56 22.45±3.17 * 21.64±2.67 ** 21.73±3.66 * 1.44±0.21 * 1.80±0.36 1.33±0.26 ** 1.62±0.24 1.46±0.24 * 1.35±0.25 * 1.45±0.25 *
Compare with model group: *P＜0.05; *P＜0.01
Result: joining attached (lyophilizing) high dose can anticoagulant, reduce the erythrocyte aggregation ability, reduce whole blood viscosity (low cutting) and plasma viscosity, prolong blood plasma PT, reduce plasma fibrin (FIB) content, with model group comparing difference remarkable (P＜0.05, P＜0.01).But middle dosage is anticoagulant, reduction whole blood viscosity (low cutting) and plasma viscosity also, with model group comparing difference remarkable (P＜0.05).Joining attached former medicine can anticoagulant, reduce the erythrocyte aggregation ability, reduce whole blood viscosity (low cutting) and plasma viscosity prolongs blood plasma PT, reduces plasma fibrin (FIB) content, with model group comparing difference significantly (P＜0.05); But joining attached former medicine compares and there is no significant difference (P＞0.05) with ginseng attached (lyophilizing) high dose.
3.4 influence to the isolated rat heart ischemical reperfusion injury
3.4.1 experimental implementation:
(1) grouping and dosage
Get 56 of SD rats, body weight 200g～300g is divided into 7 groups at random, and 8 every group, male and female half and half.I.e. (1) normal control group: contain oxygen Rockwell liquid; (2) model group: nitrification Rockwell liquid; (3) positive drug group: nitrification Radix Salviae Miltiorrhizae Injection, 1.3mg/ml; (4) injection ginseng attached (lyophilizing) high dose group: the nitrification ginseng is attached, 0.31mg/ml; (5) dosage group in the injection ginseng attached (lyophilizing): nitrification is joined attached 0.16mg/ml; (6) injection ginseng attached (lyophilizing) low dose group: the nitrification ginseng is attached, 0.08mg/ml; (7) the attached former medicine group of ginseng: the former SHENFU ZHUSHEYE of nitrification, 0.31mg/ml.Used medicine is all prepared with Rockwell liquid.
Rat tail vein injecting heparin sodium (1000U/kg) hits dizzyly behind the 15min, open breast, heart is taken out rapidly, move in the culture dish of 4 ℃ of Rockwell liquid of contain, wash out heart blood as far as possible, under liquid level, rapidly aorta is connected on the sleeve pipe of perfusion, and pricks fixing with toe-in.Carry out perfusion to contain oxygen Rockwell liquid immediately.Perfusion pressure 70cm water column, 38 ± 0.5 ℃ of perfusion temperature, flow velocity 11.5 ± 0.5ml/min.Tangle apex and link to each other with frog heart clip, be connected in two road physiology monitors and trace changes in heart rate with transducer.Recovering beat of heart and heart rate are stablized 30min after showing normally, experimentize again, arrhythmia person occurs during this period and discard.
(2) method:
Adopt the modeling of anoxia reperfusion injury method:
Model group: with after containing oxygen Rockwell liquid perfusion 30min and making heart working stable, regulate three-way cock, use in advance that nitrification Rockwell liquid pours into 40min as anoxia instead, recover to contain oxygen Rockwell liquid perfusion 40min again to perfusion pipe one side of nitrification Rockwell liquid in advance.Cause anoxia reperfusion injury model.
Positive drug group and each administration group: method is the same substantially, only pours into 40min adding different pharmaceutical when nitrification Rockwell liquid is done the anoxia perfusion in advance.
Normal control group: to contain oxygen Rockwell liquid perfusion 110min.
Coronary flow and heart rate when record normally and again pours into back the 20th, 40min, the spill-out of lactic acid dehydrogenase (LDH), creatine kinase (CK) is calculated rate of change in the mensuration each point perfusate.Experiment with 10% formaldehyde fixed heart, is done the pathology histological examination after finishing.
Rate of descent %=(before stopping irritating-after pouring into again)/before stopping irritating * 100%
Climbing %=(perfusion back-before stopping irritating) again/before stopping irritating * 100%
3.4.2 statistical procedures adopts the Student-t check.
3.4.3 the results are shown in Table 8～table 11.
(1) injection ginseng attached (lyophilizing) the results are shown in Table 8 to the influence of anoxia reperfusion injury isolated rat heart heart rate.
Table 8 injection ginseng attached (lyophilizing) is to influence (X ± S) (n=8) of anoxia reperfusion injury isolated rat heart heart rate
Group Concentration mg/ml Heart rate (beatmin -1)
Before stopping irritating The multiple back 20min that irritates The multiple back 40min that irritates
Heart rate Rate of descent (%) Heart rate Rate of descent (%)
The former medicine group of dosage group Shenfu low dose group Shenfu in the positive group of the Normal group model group Shenfu high dose group Shenfu -- -- 1.33 0.31 0.16 0.08 0.31 243.5±29.43 255.0±33.18 252.0±22.22 258.0±15.71 265.5±13.51 245.8±42.83 274.5±30.38 213.5± 60.47 ** 168.0±40.06 217.5±47.33 * 223.5± 15.63 ** 210±27.21 * 196.5±44.41 223.5± 28.64 ** 13.2±19.88 ** 34.4±10.56 14.5±14.92 ** 13.2±6.87 ** 20.8±10.56 * 19.41±14.2 * 17.7±12.78 * 212±43.08 ** 129.0±36.14 202.5± 47.33 ** 198±19.24 ** 184.5± 23.95 ** 179.8±55.06 * 213.0± 30.59 ** 13.4±11.20 ** 49.1±12.96 20.4±15.32 ** 23.3±5.46 ** 30.4±9.58 ** 26.0±21.93 * 21.83± 12.06 **
Compare with model group, *P＜0.05; *P＜0.01.
Experimental result: each group equal zero difference of heart rate before stopping irritating, when pouring into back 20min and 40min again, injection ginseng attached (lyophilizing) high, medium and low dosage group heart rate all is lower than model group, and rate of descent and model group are relatively, statistical significance (P＜0.01, P＜0.05) is arranged.Injection ginseng attached (lyophilizing) high dose group heart rate rate of descent is compared no difference of science of statistics (P＞0.05) with join attached former medicine with concentration.
(2) injection ginseng attached (lyophilizing) the results are shown in Table 9 to the influence of anoxia reperfusion injury isolated rat heart arteria coronaria perfusion flow.
Table 9 injection ginseng attached (lyophilizing) is to influence (X ± S) (n=8) of anoxia reperfusion injury isolated rat heart arteria coronaria perfusion flow
Group Concentration mg/ml Perfusion flow (mlmin -1)
Before stopping irritating The multiple back 20min that irritates The multiple back 40min that irritates
Perfusion flow Rate of descent (%) Perfusion flow Rate of descent (%)
The former medicine group of dosage group Shenfu low dose group Shenfu in the positive group of the Normal group model group Shenfu high dose group Shenfu -- -- 1.33 0.31 0.16 0.08 0.31 6.6±0.56 6.6±0.60 6.1±0.9 6.5±0.44 6.3±0.47 5.9±0.85 6.5±0.61 6.1±0.71 4.4±0.35 5.0±1.05 5.5±0.56 ** 5.1±0.40 ** 4.1±0.75 5.4±0.57 **/ 7.9±5.74 ** 33.4±8.66 17.4± 10.55 ** 14.9±5.31 ** 18.7±5.46 ** 29.4±15.15 17.4±3.10 ** 5.8±1.08 3.5±0.40 4.8±1.07 ** 5.0±0.58 ** 4.4±0.61 ** 3.4±0.92 4.9±0.64 ** 13.4± 10.59 ** 47.0±3.07 21.6± 11.89 ** 22.9±7.39 ** 30.2±6.35 ** 39.81±20.95 24.4±5.54 **
Compare with model group, *P＜0.05; *P＜0.01.
Experimental result: each group is perfusion flow and the relatively more equal zero difference of normal group before stopping irritating, when pouring into back 20min and 40min again, injection ginseng attached (lyophilizing) height, middle dosage group perfusion flow are apparently higher than model group, and the low and model group of its rate of descent has significant difference (P＜0.01).Join attached (lyophilizing) low dose group perfusion flow rate of descent and all be lower than model group, but not statistically significant (P＞0.05).Join attached (lyophilizing) three dosage groups and in influence, present certain dose-effect relationship the arteria coronaria perfusion flow.Join attached (lyophilizing) high dose group arteria coronaria perfusion flow rate of descent and be lower than the attached former medicine group of ginseng, but no difference of science of statistics (P＞0.05).
(3) injection ginseng attached (lyophilizing) the results are shown in Table 10 to the influence of anoxia reperfusion injury isolated rat heart CK.
Experimental result: each group is CK value and the relatively more equal zero difference of normal group before stopping irritating, when pouring into back 20min and 40min again, injection ginseng attached (lyophilizing) high, medium and low dosage group CK value significantly is lower than model group, its climbing and model group are relatively, be lower than model group, significant difference (P＜0.01) is arranged.Join attached (lyophilizing) three dosage groups and can suppress all that CK overflows due to the myocardial cell injury, present certain dose-effect relationship.Join attached (lyophilizing) high dose group CK value climbing and be lower than with concentration and join attached former medicine group, but no difference of science of statistics (P＞0.05).
Table 10 injection ginseng attached (lyophilizing) is to influence (X ± S) (n=8) of anoxia reperfusion injury isolated rat heart CK
Group Concentration mg/ml CK(U·L -1)
Before stopping irritating The multiple back 20min that irritates The multiple back 40min that irritates
CK Climbing (%) CK Climbing (%)
The former medicine group of dosage group Shenfu low dose group Shenfu in the positive group of the Normal group model group Shenfu high dose group Shenfu -- -- 1.33 0.31 0.16 0.08 0.31 13.3± 6.25 13.9± 5.64 12.1± 5.44 13.5± 4.11 14.1± 2.59 18.3± 5.80 13.1±4.7 8.5±4.60 ** 51.9±20.75 15.3±9.91 ** 13.5±5.68 ** 16.5±5.21 ** 20.6±7.37 ** 14.4±3.70 ** -36.4± 24.47 ** 307.8±201.34 56.3± 113.07 ** 2.2±34.06 ** 18.6±37.86 ** 25.7±60.97 ** 18.5±40.62 ** 7.9±5.03 ** 58.63±17.62 16±11.58 ** 15.3±5.39 ** 21.6±6.82 ** 31.4±18.49 ** 18.3±8.9 **6 -41.49±19.95 ** 359.2±161.89 53.4±91.25 ** 18.0±41.26 ** 56.2±50.65 ** 89.5±118.61 ** 44.3±57.83 **
Compare with model group *P＜0.05; *P＜0.01.
(4) injection ginseng attached (lyophilizing) the results are shown in Table 11 to the influence of anoxia reperfusion injury isolated rat heart LDH.
Table 11 injection ginseng attached (lyophilizing) is to influence (X ± S) (n=8) of anoxia reperfusion injury isolated rat heart LDH
Group Concentration (mg/m l) LDH/(U·L -1)
Before stopping irritating The multiple back 20min that irritates The multiple back 40min that irritates
LDH Climbing (%) LDH Climbing (%)
The former medicine group of dosage group Shenfu low dose group Shenfu in the positive group of the Normal group model group Shenfu high dose group Shenfu -- -- 1.33 0.31 0.16 0.08 0.31 11.3±2.87 11.1±5.87 11.8±3.41 14.3±2.82 14.4±3.70 13.8±3.58 14.3±2.49 8.9±1.96 ** 26.1±8.27 11.6±3.70 ** 12.8±1.75 ** 15.4±4.53 ** 15.8±6.04 * 15.6±3.20 **▲ -17.56±23.76 ** 185.3±170.67 15.8±74.89 * -5.8±29.81 ** 7.9±22.42 * 20.1±50.40 * 11.1±23.5 * 8.5±2.20 ** 23.4±9.20 12.4±4.14 ** 12.4±2.07 ** 17.5±5.50 21.5±8.59 17.9±6.18 *▲ -20.0± 26.67 ** 158.1±148.79 20.8±73.86 * -8.6±29.86 ** 25.4±40.90 * 66.9±78.06 25.6±37.4 *
Compare with model group, *P＜0.05; *P＜0.01.High dose group and matched group comparison ▲ P＜0.05.
Experimental result: each group is LDH value and the relatively more equal zero difference of normal group before stopping irritating, when pouring into back 20min and 40min again, injection ginseng attached (lyophilizing) is high, middle dosage group LDH value all is lower than model group, and its climbing and model group are relatively, statistical significance (P＜0.01, P＜0.05) is arranged.Join attached (lyophilizing) low dose group when pouring into back 20min again, the LDH climbing is lower than model group (P＜0.05), and when pouring into back 40min again, climbing also is lower than model group, but not statistically significant (P＞0.05).Join attached (lyophilizing) three dosage groups and can suppress all that LDH overflows due to the myocardial cell injury, present certain dose-effect relationship.Join attached (lyophilizing) high dose group LDH value and all be lower than with concentration and join attached former medicine group, and have certain difference (P＜0.05), high dose group LDH climbing to be lower than to join attached former medicine group with concentration, but no difference of science of statistics (P＞0.05).
3.4.4 conclusion:
Injection ginseng attached (lyophilizing) height, middle dosage group are when pouring into back 20min and 40min again, and heart rate, perfusion flow rate of descent and CK, LDH climbing all are lower than model group (P＜0.01, P＜0.05).Prompting ginseng attached (lyophilizing) is high, middle dosage group can be improved the heart rate decline that the anoxia reperfusion injury is caused, and increases coronary flow simultaneously, suppresses CK, LDH and discharges.The effect that wherein joining attached (lyophilizing) high dose group increases coronary flow, inhibition CK, LDH release demonstrates the trend of joining attached former medicine group with concentration that is better than.
Injection ginseng attached (lyophilizing) low dose group is when pouring into back 20min and 40min again, and heart rate rate of descent, CK climbing all have certain difference (P＜0.05, P＜0.01) with model group, and when pouring into back 20min again, the LDH climbing is lower than model group (P＜0.05).Prompting injection ginseng attached (lyophilizing) low dose group can be improved heart rate decline, inhibition CK, the LDH release that ischemia-reperfusion causes.
Anoxia reperfusion injury isolated rat heart is the result show; injection ginseng attached (lyophilizing) is mainly by improving the protective effect of bringing into play heart of overflowing of heart rate decline that the anoxia reperfusion injury caused, coronary artery dilator, inhibition myocardium enzyme; have certain coronary artery dilator simultaneously, increase coronary flow; improve the effect that heart rate that the anoxia reperfusion injury caused descends; three dosage groups are to arteria coronaria perfusion flow and CK; in the influence of LDH, all present certain dose-effect relationship.
Isolated rat heart ischemical reperfusion injury tissue pathology checking report shows: injection ginseng attached (lyophilizing) has the certain protection effect to damage due to the isolated rat myocardial ischemia-reperfusion.
3.5 influence to myocardial infarction due to the anesthetized dog coronary ligation
3.5.1 experimental technique
Get 35 of domesticated dogs, body weight is 7-10kg, is divided into 7 groups at random, every group of 5 animals, male and female dual-purpose: (1) sham operated rats: give the isometric(al) normal saline.(2) model group: give the isometric(al) normal saline.(3) positive drug Radix Salviae Miltiorrhizae Injection matched group: 2.0g crude drug/kg.(4) injection ginseng attached (lyophilizing) low dose group: 0.2g crude drug/kg.(5) dosage group in the injection ginseng attached (lyophilizing): 0.4g crude drug/kg.(6) injection ginseng attached (lyophilizing) high dose group: 0.8g crude drug/kg.(7) the attached former medicine matched group of ginseng: 0.4g crude drug/kg.The used medicine of each treated animal is dissolved in 0.9% sodium chloride injection, and the administration volume is 1ml/kg.
After domesticated dog was anaesthetized with 3% pentobarbital sodium (30mg/kg) by the forelimb small saphenous vein, back of the body position was fixed on the operating-table, cuts off the hair of cervical region, chest and left hind inboard.It is subcutaneous that needle electrode is inserted the dog extremity, recording ecg (ECG).Separate trachea and insert tracheal intubation.Separate left side femoral artery, femoral vein, separate femoral vein and insert venous cannulation, slowly constant speed input normal saline (about 1ml/min); Separate femoral artery insertion arterial cannulation (being full of the heparin-saline of 500u/ml in the pipe) and connect pressure transducer (TP-400T), measure systolic arterial pressure (SAP), auterial diastole pressure (DAP), mean arterial pressure (MAP) through amplifying (AP-641G), calibration sensitivity is 13.33kPa (100mmHg)/cm.The line connection board that connects monitor directly triggers heart rate numeration instrument (AT-601G) recorded heart rate (HR) by arterial pressure.Fourth, fifth intercostal is opened breast in the left side, exposes heart, and connects artificial respirator.Open pericardium, be sewn in thoracic wall, make the pocket cradle.Expose the right auricle, quiet notes heparin 5-10mg/kg makes heparinization, clamp the right auricle with auricle clamp, make a pocket sealing, suture is through in a bit of rubber tube, line is colluded into rubber tube with little steel wire, cut an osculum in pocket mouth center, (internal diameter 4-5mm, the mouth of pipe are the oblique angle, and nearly mouth of pipe place is that circle is expanded to insert coronary sinus cannula rapidly, the oblique angle face avoids the mouth of pipe and Dou Bi to be close to, the portion of expanding is for making coronary sinus blood unlikely excessive), with pocket suture tension in the small rubber hose, withstand intubate by small rubber hose, to avoid hemorrhage, rapidly intubate is accurately inserted coronary sinus, place, ligation right auricle pocket suture, fixedly intubate.The far-end of this intubate connects a tee T to be on the waiting list blood.With descending 1/3 place, a sham operated rats not ligation of threading in No. 0 silk thread ligation ramus descendens anterior arteriae coronariae sinistrae.12 the mapping point seams in position are put the epicardial electrogram electrode near selecting infarct, and selecting a control point away from infarct, carry out mapping with hand-held insulated metal point-like electrode by the mapping dot sequency, through pick off (JB-642G), amplifier (AB-621G), with eight road physiology monitor record epicardial electrogram (EECG) (calibrations: 1mm=1mv).Ligation 15min is after systolic arterial pressure (SAP), diastolic pressure (DAP), mean arterial pressure (MAP), heart rate (HR) are directly write down in femoral vein constant flow pump constant speed administration (1ml/min), and traces EECG and arterial pressure (BP) curve.Before the record ligation, after ligation 15min and the administration 5,15,30,60,90, EECG, the SAP of 120min, DAP, MAP, HR.Add up ST field offset ∑-ST of each mapping point EECG, and the ST section is raised 〉=the above N-ST of 2mv.Each group is respectively at before the administration and behind the administration 60min, and left ventricle is got blood and surveyed arterial blood oxygen, and coronary sinus is got blood and surveyed coronary sinus blood oxygen, both oxygen difference reflecting myocardium oxygen consumptions; Each group is respectively at before the ligation, femoral vein is got blood behind ligation 15min, the administration 60min, with kit measurement serum CK, LDH value.After experiment finishes, take out heart immediately, behind normal saline flush away blood, take by weighing heavy whole-heartedly and ventricular weight, and the ventricle crosscut become 5, and place 37 ℃ of 1%TTC solution 15min that dyes, cut off the non-infarct that each myocardium sheet is colored, undyed infarct cardiac muscle is weighed, obtain infarction size heavily respectively divided by heavy or ventricle whole-heartedly and account for heavy whole-heartedly or account for the heavy percentage rate of ventricle.
Myocardial oxygen consumption (kpa/min)=MAP*HR
3.5.2 experimental result
(1) injection ginseng attached (lyophilizing) is to the myocardial infarction BP (influence of SAP, DAP, MAP, HR and myocardial oxygen consumption due to the anesthetized dog coronary ligation
Experimental result shows: injection ginseng attached (lyophilizing) height, in the dosage group can reduce DAP, MAP, HR and the myocardial oxygen consumption of myocardial infarction anesthetized dog due to the coronary ligation, with model group notable difference (P＜0.05 is arranged relatively, P＜0.01), the dosage group relatively has significant difference (P＜0.05) with the attached former medicine group of ginseng and wherein.
(2) injection ginseng attached (lyophilizing) is to anesthetized dog arteriovenous blood oxygen partial pressure (Po 2), arteriovenous blood partial pressure of carbon dioxide (Pco 2) influence
The result shows: injection ginseng attached (lyophilizing) is high, middle dosage group and model group relatively, the arteriovenous oxygen difference that can significantly suppress due to the coronary ligation increases (P＜0.05), and wherein the dosage group with join some point of attached former medicine group comparison significant difference (P＜0.01) arranged.
(3) injection ginseng attached (lyophilizing) is to the influence of myocardial infarction ∑-ST due to the anesthetized dog coronary ligation
The result shows: injection ginseng attached (lyophilizing) high, medium and low dosage group can reduce myocardial infarction anesthetized dog ∑-ST due to the coronary ligation, part-time point and remarkable (P＜0.05 of model group comparing difference, P＜0.01), show that injection ginseng attached (lyophilizing) can obviously alleviate degree of myocardial ischemia, and wherein the dosage group there is significant difference (P＜0.01) with some point of the attached former medicine group of ginseng comparison.
(4) injection ginseng attached (lyophilizing) is to the influence of myocardial infarction N-ST due to the anesthetized dog coronary ligation
The result shows: injection ginseng attached (lyophilizing) high, medium and low dosage group can reduce myocardial infarction anesthetized dog N-ST due to the coronary ligation, part-time point and remarkable (P＜0.05 of model group comparing difference, P＜0.01), and wherein the dosage group relatively has significant difference (P＜0.01) with some point of the attached former medicine group of ginseng.
(5) injection ginseng attached (lyophilizing) is to the serum CK of myocardial infarction due to the anesthetized dog coronary ligation, the influence of LDH.
Experimental result shows: injection ginseng attached (lyophilizing) high, medium and low dosage group, can reduce the release of CK, LDH in the serum after the myocardial infarction due to the anesthetized dog coronary ligation, with model group comparing difference significantly (P＜0.01).
(6) injection ginseng attached (lyophilizing) is to the influence of myocardial infarction infarction size due to the anesthetized dog coronary ligation
Experimental result shows: injection ginseng attached (lyophilizing) is high, middle dosage group can reduce myocardial infarction infarction size due to the anesthetized dog coronary ligation, with remarkable (P＜0.05 of model group comparing difference, P＜0.01), and wherein the dosage group relatively has significant difference (P＜0.01) with the attached former medicine group of ginseng.
3.5.3 conclusion:
The myocardial infarction experimental result shows due to the anesthetized dog coronary ligation: injection ginseng attached (lyophilizing) height, in the dosage group can reduce DAP, MAP, HR and the myocardial oxygen consumption of myocardial infarction anesthetized dog due to the coronary ligation, arteriovenous oxygen difference due to the inhibition coronary ligation increases, part-time point and model group comparing difference remarkable (P＜0.05, P＜0.01); High, medium and low dosage group can reduce myocardial infarction anesthetized dog ∑-ST due to the coronary ligation, and N-ST, part-time point and model group comparing difference be (P＜0.05, P＜0.01) significantly; High, medium and low dosage group can reduce due to the anesthetized dog coronary ligation after the myocardial infarction release of LDH and CK in the serum, and is with model group comparing difference significantly (P＜0.05), remarkable with former medicine group of last index and middle dosage group comparing difference.Show that injection ginseng attached (lyophilizing) can obviously alleviate degree of myocardial ischemia.
3.6 to the hemodynamic influence of anesthetized dog [5-6]
3.6.1 experimental technique:
Get 30 of domesticated dogs, the male and female dual-purpose is divided into 6 groups at random, every group of 5 animals, i.e. (1) normal control group: give the isometric(al) normal saline; (2) positive drug injection of danshen group: 2.0g crude drug/kg.(3) injection ginseng attached (lyophilizing) low dose group: 0.2g crude drug/kg.(4) dosage group in the injection ginseng attached (lyophilizing): 0.4g crude drug/kg.(5) injection ginseng attached (lyophilizing) high dose group: 0.8g crude drug/kg.(6) the attached former medicine group of ginseng: 0.4g crude drug/kg, the used medicine of each treated animal is dissolved in 0.9% sodium chloride injection, the administration volume is 1ml/kg.
With the subcutaneous cephalic vein anesthesia of 3% pentobarbital sodium 30mg/kg forelimb, back of the body position is fixed on the operating-table, and the inboard unhairing of cervical region, chest and left hind is standby.Separate trachea and insert tracheal intubation; Separate femoral vein and insert venous cannulation, slowly constant speed input normal saline (about 1ml/min); Separate femoral artery and insert arterial cannulation (being full of the heparin-saline of 500u/ml in the pipe), to measure arteriotony.Under the artificial respiration, open breast, cut off pericardium in the 4th intercostal, be sewn in thoracic wall, separate root of ascending aorta and ramus descendens anterior arteriae coronariae sinistrae, place the electromagnetic blood flowmeter probe (12mm of suitable internal diameter, 2mm), be connected in and measure cardiac output (CO) and coronary flow (CBF) on the electromagnetic blood flowmeter.Left ventricular cannulation (being full of heparin-saline in the pipe) in left ventricle apex wound inserts left ventricle, is measured left indoor pressure (LVSP), left chamber diastasis pressure (LVEDP), the maximum climbing speed (LVdp/dtmax) of intraventricular pressure; It is subcutaneous that needle electrode is inserted the dog extremity, recording ecg (ECG).After waiting to stablize, with constant flow pump through femoral vein administration (1ml/min), directly write down systolic arterial pressure (SAP), diastolic pressure (DAP), mean arterial pressure (MAP), heart rate (HR), cardiac output (CO), coronary flow (CBF), and on eight road physiology monitors, trace ECG, left indoor pressure (LVSP), LVdp/dtmax, LVEDP and arterial pressure (BP) curve.Measure and calculate the BP (SAP of each time period anesthetized dog of administration front and back, DAP, MAP), HR, CO, CBF, LVSP, LVEDP, + dp/dtmax (myocardial contraction parameter),-dp/dtmax (myocardial relaxation parameter), t～dp/dtmax (left chamber begins to be contracted to left indoor pressure climbing speed time to peak), ejection time, SV (stroke volume), CI (cardiac index), SI (SI), LVWI (stroke work index), TTI (total oxygen consumption index), TPVR (total peripheral vascular resistance).
Computing formula:
CI (cardiac index)=CO/ body surface area; SV (stroke volume)=CO*1000/HR;
SI (SI)=CI*1000/HR; TTI (total oxygen consumption index)=MAP*HR* ejection time;
TPVR (total peripheral vascular resistance)=MAP/CO;
LVWI (stroke work index)=CI*1.025* (SAP-0.667) * 13.6*0.001
3.6.2 experimental result
(1) injection ginseng attached (lyophilizing) is to the influence of anesthetized dog SAP, DAP, MAP, HR
The result shows: the DAP that injection ginseng attached (lyophilizing) is high, middle dosage group can reduce anesthetized dog, decreased heart rate is with normal control group comparing difference remarkable (P＜0.05, P＜0.01).
(2) injection ginseng attached (lyophilizing) is to the influence of anesthetized dog CO, CBF, SV, CI, SI
The result shows: injection ginseng attached (lyophilizing) is high, middle dosage group can increase anesthetized dog coronary flow (CBF), SI (SI), stroke volume (SV), with normal control group comparing difference remarkable (P＜0.05, P＜0.01); High, middle dosage group can obviously increase anesthetized dog cardiac output (CO), cardiac index (CI), (with normal control group P＜0.01 relatively), and wherein dosage group and the attached former medicine group comparing difference of ginseng significantly (P＜0.05).
(3) to the influence of anesthetized dog ejection time, LVSP, LVEDP
The result shows: injection ginseng attached (lyophilizing) three each dosage groups all do not make significant difference to anesthetized dog ejection time, LVSP, LVEDP.
(4) to anesthetized dog+dp/dtmax ,-influence of dp/dtmax, t-dp/dtmax
The result shows: injection ginseng attached (lyophilizing) three dosage groups to anesthetized dog+dp/dtmax ,-dp/dtmax and t-dp/dtmax do not make significant difference (with the normal control group relatively, P＞0.05).
(5) to the influence of anesthetized dog LVWI, TTI, TPVR
The result shows: injection ginseng attached (lyophilizing) is high, in two dosage groups can reduce anesthetized dog total peripheral resistance (TPVR), total oxygen consumption (TTI), compare with the normal control group, significant difference (P＜0.05, P＜0.01), and wherein dosage group and the attached former medicine group comparing difference of ginseng significantly (P＜0.05).
3.6.3 experiment conclusion:
The experiment of anesthetized dog hemodynamics shows: the DAP that injection ginseng attached (lyophilizing) is high, middle dosage group can reduce anesthetized dog, decreased heart rate, reduce anesthetized dog total peripheral resistance (TPVR), total oxygen consumption (TTI), increase anesthetized dog cardiac output (CO), cardiac index (CI), with normal control group comparing difference remarkable (P＜0.05, P＜0.01).High, middle dosage group can increase anesthetized dog coronary flow (CBF), SI (SI), stroke volume (SV), with normal control group comparing difference remarkable (P＜0.05, P＜0.01) and remarkable with former medicine group of last index and middle dosage group comparing difference.
More than two experimental results show: injection ginseng attached (lyophilizing) has the improvement effect to the myocardial ischemia and the cardiac function of anesthetized dog anterior descending coronary caused by ligature, on the basis that does not reduce the heart blood supply function, the alleviate myocardial ischemia degree.
Learn experimental result as can be known by this main pharmacodynamics, " injection ginseng attached (lyophilizing) " stable in properties that the present invention makes, the main pharmacodynamics experiment shows under Isodose, it is compared with the SHENFU ZHUSHEYE of selling in the market, the time-to-live of high dose group prolongation mice normal pressure anoxia enduring is longer significantly, and it is stronger that high dose group increases the effect of coronary flow, inhibition CK, LDH release.Proved beneficial effect of the present invention.
Embodiment 8
Get and write out a prescription 12 among the embodiment 2, prescription 13 employed raw materials and the injection made by corresponding preparation method (having kept the polysaccharide composition in the Radix Ginseng of 5K～500K molecular weight), detect (sample volume is converted accordingly by the weight of the corresponding crude drug of weight of preparation finished product content among embodiment 3, the embodiment 4) by the method in " containing injection, the freeze-drying preparation for injection of Radix Ginseng and Radix Aconiti Lateralis Preparata, the method for quality control of sterile packaged preparation for injection in a kind of raw material " in " summary of the invention ", meet the regulation in this method of quality control.
Embodiment 9
Get embodiment 3, embodiment 4 employed raw materials and gained preparation, detect by the method in " containing injection, the freeze-drying preparation for injection of Radix Ginseng and Radix Aconiti Lateralis Preparata, the method for quality control of sterile packaged preparation for injection in a kind of raw material " in " summary of the invention ", meet the regulation in this method of quality control.

Claims (17)

1, contains the pharmaceutical preparation of Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material, it is characterized in that containing in the described preparation saponin component in the Radix Ginseng, the polysaccharide composition in the Radix Ginseng, the alkaloids composition in the Radix Aconiti Lateralis Preparata.
2, pharmaceutical preparation according to claim 1, it is characterized in that the raw material of described pharmaceutical preparation is mainly Radix Ginseng, Radix Aconiti Lateralis Preparata, Radix Ginseng: the ratio of Radix Aconiti Lateralis Preparata is 1-10: 1-10, or 1-4: 1-8, or 1-2: 1-4, or 1: 2.
3, pharmaceutical preparation according to claim 1 is characterized in that described Radix Ginseng and Radix Aconiti Lateralis Preparata are wild product or artificial culture product, are genuine medicinal materials or non-genuine medicinal materials.
4, pharmaceutical preparation according to claim 1 is characterized in that described Radix Ginseng and Radix Aconiti Lateralis Preparata concoct the processed product form or concoct the processed product that forms according to modernism for its bright medical material or according to traditional method.
5, pharmaceutical preparation according to claim 1 is characterized in that containing Radix Ginseng total saponins 1mg～90mg in the preparation of per unit dosage Radix Ginseng total polysaccharides 2mg～282mg, Radix Aconiti Lateralis Preparata total alkaloids 0.02mg～22mg; Perhaps contain Radix Ginseng total saponins 2mg～60mg, Radix Ginseng total polysaccharides 4mg～188mg, Radix Aconiti Lateralis Preparata total alkaloids 0.05mg～15mg; Perhaps contain Radix Ginseng total saponins 2.5mg～45mg, Radix Ginseng total polysaccharides 6mg～141mg, Radix Aconiti Lateralis Preparata total alkaloids 0.06mg～11mg.
6, pharmaceutical preparation according to claim 1 is characterized in that in the preparation of per unit dosage, contains Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) count 8mg～15mg, the Radix Ginseng total polysaccharides is 25mg～47mg with the glucose meter, Radix Aconiti Lateralis Preparata total alkaloids 0.2mg～3.6mg;
Or in the preparation of per unit dosage, contain Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) count 16mg～30mg, the Radix Ginseng total polysaccharides is 50mg～94mg with the glucose meter, Radix Aconiti Lateralis Preparata total alkaloids 0.4mg～7.2mg;
Or in the preparation of per unit dosage, contain Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) count 40mg～75mg, the Radix Ginseng total polysaccharides is 125mg～235mg with the glucose meter, Radix Aconiti Lateralis Preparata total alkaloids 1mg～18mg;
Or in the preparation of per unit dosage, contain Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) count 2.5mg～5mg, the Radix Ginseng total polysaccharides is 8mg～16mg with the glucose meter, Radix Aconiti Lateralis Preparata total alkaloids 0.06mg～1.2mg.
7, pharmaceutical preparation according to claim 1 is characterized in that in the preparation of per unit dosage, contains aconite alkaloids with aconitine (C 24H 47NO 11) meter, must not be higher than 10.0mg, maybe must not be higher than 5.0mg, maybe must not be higher than 3.0mg, maybe must not be higher than 2.0mg, maybe must not be higher than 1.0mg.
8, pharmaceutical preparation according to claim 1 is characterized in that described preparation is oral solid formulation, oral semi-solid preparation, oral liquid or injection;
Described oral solid formulation can be tablet, hard capsule, soft capsule, pill, drop pill, solid dispersion, powder, granule, fine granule, pellet, microcapsule, microspheres agent or other pharmaceutically acceptable oral solid formulation;
Described oral liquid can be oral liquid or other pharmaceutically acceptable oral liquid;
Described injection can be injection, freeze-drying preparation for injection, sterile packaged preparation for injection, glucose injection, sodium chloride injection or other pharmaceutically acceptable injection.
9, pharmaceutical preparation according to claim 1 is characterized in that described preparation is tablet, hard capsule, soft capsule, solid dispersion, oral liquid, injection, freeze-drying preparation for injection, sterile packaged preparation for injection, glucose injection or sodium chloride injection.
10, pharmaceutical preparation according to claim 1 is characterized in that pharmaceutical preparation of the present invention also contains where necessary or uses medicine acceptable auxiliary (or carrier), described adjuvant is pharmaceutically acceptable preparation adjuvant;
For oral solid formulation and oral semi-solid preparation, described adjuvant is selected from diluent, wetting agent, binding agent, disintegrating agent, fluidizer, antiplastering aid, lubricant, color and regulator thereof, solid dispersion carrier material, antioxidant, surfactant, stabilizing agent, PH regulator and other pharmaceutically acceptable oral solid formulation, oral semi-solid preparation with one or more the material in the adjuvant, and every kind of adjuvant can not select or select for use one or more the material in this kind adjuvant;
Described diluent is one or more the material that is selected from starch, amylum pregelatinisatum, Icing Sugar, dextrin, lactose, microcrystalline Cellulose, mannitol, sorbitol, calcium sulfate, calcium carbonate and other the pharmaceutically acceptable diluent;
Described wetting agent is one or more the material that is selected from ethanol, water and other the pharmaceutically acceptable wetting agent;
Described binding agent is one or more the material that is selected from hypromellose, ethyl cellulose, sodium carboxymethyl cellulose, methylcellulose, hyprolose, starch slurry, polyvidone, gelatin, Polyethylene Glycol, 50% to 70% sucrose solution, sodium alginate soln and other the pharmaceutically acceptable binding agent;
Described disintegrating agent is one or more the material that is selected from carboxymethyl starch sodium, low-substituted hydroxypropyl cellulose, cross-linking sodium carboxymethyl cellulose, dried starch, polyvinylpolypyrrolidone, gas-producing disintegrant and other the pharmaceutically acceptable disintegrating agent;
Described fluidizer, antiplastering aid, lubricant are one or more the materials that is selected from Pulvis Talci, micropowder silica gel, magnesium stearate, polyethylene glycols, sodium laurylsulfate, magnesium laurylsulfate, hydrogenated vegetable oil and other pharmaceutically acceptable fluidizer, antiplastering aid, the lubricant;
Described color and regulator thereof are one or more the materials that is selected from medicinal pigment, food coloring, essence and other pharmaceutically acceptable color and the regulator thereof;
Described solid dispersion carrier material is to be selected from polyethylene glycols, cellulose derivative, organic acid, surfactant-based, the polyvidone class, saccharide and alcohols, cellulose family, the polyacrylic resin class, cupreol, cholesterol, cholesterol ester stearic acid, tripalmitin, castor oil hydrogenated, Oleum Ricini wax, Cera Flava, the material of one or more in Brazil wax and other pharmaceutically acceptable solid dispersion carrier material, every class solid dispersion carrier material can not select or select for use one or more the material in such solid dispersion carrier material;
Described pH regulator agent can be at least a pharmaceutically acceptable material that is used to regulate pH value, and the material of described adjusting pH value is one or more the material that is selected from the pharmaceutically acceptable material that is used for regulating pH value of alkali compounds, buffer system, acid and other;
For oral liquid, described adjuvant is selected from one or more the material in solvent, solubilizing agent, cosolvent, cosolvent, antiseptic, correctives, coloring agent, pH regulator agent, antioxidant, complexing of metal ion agent and other the pharmaceutically acceptable liquid preparation additives, and every kind of adjuvant can not select or select for use one or more the material in this kind adjuvant;
Described solvent is to be selected from water, certain density ethanol, glycerol, propylene glycol and other pharmaceutically acceptable liquid preparation with one or more the material in the solvent;
Described antiseptic is to be selected from parabens, sorbic acid and salt thereof, benzoic acid and salt thereof such as methyl parahydroxybenzoate, ethyl ester, propyl ester, butyl ester, Oleum menthae and other pharmaceutically acceptable liquid preparation with one or more the material in the antiseptic;
Described correctives is to be selected from sucrose, simple syrup, rob, sorbitol, glycerol, mannitol, saccharin sodium, Herba Menthae Haplocalycis volatile oil and other pharmaceutically acceptable liquid preparation with one or more the material in the correctives;
For injection, described adjuvant is selected from pharmaceutically acceptable solvent for injection, solubilizing agent, antibacterial, antioxidant, stabilizing agent, chelating agent, analgesics, isoosmotic adjusting agent, buffer agent, pH regulator agent, and one or more the material in the pharmaceutically acceptable medicine with other miscellaneous function, every kind of adjuvant can not select or select for use one or more the material in this kind adjuvant;
Described solvent for injection is one or more the material that is selected from water for injection, certain density ethanol, glycerol, propylene glycol, benzyl alcohol and other the pharmaceutically acceptable solvent for injection;
Described solubilizing agent is to be selected from polysorbate 20, polyoxyethylene sorbitan monoleate, polysorbate 40, polysorbate 60 and other pharmaceutically acceptable injection with one or more the material in the solubilizing agent;
Described antibacterial is to be selected from hydroxypropyl butyl ester, hydroxypropyl methyl ester, benzyl alcohol, phenol, chlorobutanol and other pharmaceutically acceptable injection with one or more the material in the antibacterial;
Described antioxidant is to be selected from sodium pyrosulfite, sodium sulfite, sodium sulfite, sodium thiosulfate and other pharmaceutically acceptable injection with one or more the material in the antioxidant;
Described chelating agent is one or more the material that is selected from Calcium Disodium Versenate, cyclohexanediamine four sodium acetates, N-hydroxyl diethylamine three acetic acid, diethyl triamine six acetic acid and other the pharmaceutically acceptable chelating agent;
Described pH regulator agent is one or more the material that is selected from certain density sodium hydroxide solution, certain density hydrochloric acid solution, phosphoric acid, acetic acid-sodium acetate buffer solution, acetate buffer solution, phosphate buffered solution, citrate buffer solution, citric acid-disodium hydrogen phosphate buffer solution and other the pharmaceutically acceptable pH regulator agent;
Wherein said freeze-drying preparation for injection can also not add or add one or more the acceptable an amount of excipient of medicine in preparation.Described excipient is selected from one or more the material in mannitol, sorbitol, glycine, lactose, sodium chloride, glucose and other the pharmaceutically acceptable excipient.The preferred mannitol of described excipient, sorbitol.
11, pharmaceutical preparation according to claim 1 is characterized in that the amount average molecular weight is 30～36 * 10 in the described pharmaceutical preparation 4, 8～12 * 10 4, 3.5～7.5 * 10 4The polysaccharide composition of Radix Ginseng in one or more, wherein measuring average molecular weight is 33.0445 * 10 4, or 9.8878 * 10 4, or 5.4087 * 10 4The content of polysaccharide composition of Radix Ginseng higher.
12, the preparation method of the described pharmaceutical preparation of a kind of claim 1 is characterized in that the preparation method of described pharmaceutical preparation is:
Extract the saponin component of the Radix Ginseng in Radix Ginseng and the Radix Aconiti Lateralis Preparata, the polysaccharide composition of Radix Ginseng, the alkaloids composition of Radix Aconiti Lateralis Preparata, add or do not added synergistic medicine, add or do not add adjuvant, make the oral solid formulation that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material, oral semi-solid preparation, oral liquid, injection.
13, the preparation method of the described pharmaceutical preparation of a kind of claim 1 is characterized in that the preparation method of described pharmaceutical preparation may further comprise the steps:
One, the extraction of the alkaloids composition of the polysaccharide composition of the saponin component of the Radix Ginseng in Radix Ginseng and the Radix Aconiti Lateralis Preparata, Radix Ginseng, Radix Aconiti Lateralis Preparata:
(1) processing of Radix Ginseng: Radix Ginseng adopts certain density ethanol extraction (extracting method adopts reflux extraction or ultrasonic extraction or percolation or infusion process or continuous extraction or decompression extraction method), the gained extracting solution filters, filtrate recycling ethanol is to there not being the alcohol flavor, adjustment liquor strength extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.5～4.5g, use water saturated n-butanol extraction, merge n-butyl alcohol liquid, reclaim n-butyl alcohol to there not being the n-butyl alcohol flavor, change water-soluble, filter, get the saponin component intermediate of Radix Ginseng, standby;
(2) processing of medicine residues of Radix Ginseng: the medicinal residues of Radix Ginseng after by certain density ethanol extraction are earlier with water extraction (extracting method adopts decocting method or reflux extraction or ultrasonic extraction or percolation or infusion process or continuous extraction or decompression extraction method), the gained extracting solution filters, adjusting liquor strength is 0.2～2.4g crude drug/ml, temperature is 20～70 ℃, adds 85～95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 70～90%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, it is (or not centrifugal to merge supernatant, filter the back and directly adjust liquor strength), adjusting the supernatant liquor strength is 0.7～6g crude drug/ml, adds 85～95% ethanol and makes and contain alcohol and measure and reach 60～80%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, get supernatant (or not centrifugal, directly get filtrate after the filtration)
Previous step gained supernatant or filtrate are the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid;
Previous step gained supernatant adopts the ultrafilter membrane ultrafiltration of molecular cut off 100K～900K, and ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or with of the ultrafilter membrane ultrafiltration of previous step gained supernatant elder generation with molecular cut off 100K～900K, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 1K～10K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
(3) processing of Radix Aconiti Lateralis Preparata: the Radix Aconiti Lateralis Preparata medical material is earlier with water extraction (extracting method adopts decocting method or reflux extraction or ultrasonic extraction or percolation or infusion process or continuous extraction or decompression extraction method), the gained extracting solution filters, the relative density of adjusting medicinal liquid is 1.0～1.25,20～65 ℃ of temperature, ethanol precipitate with ethanol with 85～95% makes ethanol content reach 60～80%, standing over night, filter, filtrate recycling ethanol is to there not being the alcohol flavor, the relative density of adjusting medicinal liquid is that the ethanol of adding 85～95% in 1.05～1.45 o'clock makes pure content reach 75～90%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid;
Filtrate being heated to boiled, and keep little and boiled 10～60 minutes, cooling, cold preservation filters, and promptly gets the Radix Aconiti Lateralis Preparata intermediate, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor for the first time, adjustment medicinal liquid extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.7～2.5g, the dichloromethane extraction of 0.5～3 times of amount of each adding 2～9 times, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Two, play the processing of synergistic medicine:
(4) play the processing of synergistic medicine: playing a synergistic medicine is Chinese medicine or natural drug, extracts its effective site or effective ingredient monomer, and (make with extra care) is water-soluble, regulates pH value to 5.5～8.5, and filtration promptly gets intermediate, and is standby; Playing synergistic medicine is chemicals, and water-soluble, pH value transfers to 5.5～8.5, filters, and promptly gets intermediate, standby; Can be not water-soluble when wherein being used to prepare oral solid formulation, oral semi-solid preparation;
Three, the preparation of each pharmaceutical preparation finished product:
(5) intermediate of merging " (1) ", " (2) ", " (3) " gained, mixing;
(6) intermediate with " (1) ", " (2) ", " (3) ", " (4) " gained merges, mixing, gained medicinal liquid filter, and are condensed into clear paste and [or " (5) " gained medicinal liquid are filtered, be condensed into clear paste, the intermediate not water-soluble with " (4) " mixes], add an amount of oral solid formulation, oral semi-solid preparation auxiliary materials and mixing, the system soft material, granulate, oven dry, tabletting or incapsulate is made the sheet or the capsule that contain Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material; (7) " (5) " gained medicinal liquid being adjusted volume with purified water or water for injection is 0.1～2.0g crude drug/ml to being equivalent to liquor strength, regulates pH value to 5.5～8.5, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter that [or filtrate adds " (the 4) " intermediate of gained and an amount of oral liquid with adjuvant or do not add adjuvant, or filtrate adds an amount of oral liquid with adjuvant or do not add adjuvant; Mixing, (filtration)], regulate pH value to 5.5～8.5, add purified water or water for injection and adjust volume to full dose, survey pH value, filter, according to dosage divide to be filled in the oral liquid packing container, seal, the conventional method sterilization promptly gets the oral liquid that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material;
(8) " (5) " gained medicinal liquid being adjusted volume with water for injection is 0.1～2.0g crude drug/ml to being equivalent to liquor strength, regulates pH value to 5.5～8.5, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter that [or filtrate adds " (the 4) " intermediate of gained and an amount of injection with adjuvant or do not add adjuvant, or filtrate adds an amount of injection with adjuvant or do not add adjuvant; Mixing, (filtration)], (regulating pH value to 5.5～8.5), adding 0.02～0.5% active carbon is heated to and boils, and keeps little and boils 10～60 minutes, filters, regulate pH value to 5.5～8.5, add the injection water and adjust volume, survey pH value, fine straining (crossing the sintered glass filter in 0.22～0.50 μ m microporous filter membrane or similar aperture) to full dose, according to dosage divide and be filled to the infusion pump dress with in the container, seal, the conventional method sterilization promptly gets the injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material; (9) " (5) " gained medicinal liquid being adjusted volume with water for injection is 0.3～2.0g crude drug/ml to being equivalent to liquor strength, regulates pH value to 5.5～8.5, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter [or filtrate add " (the 4) " intermediate of gained and an amount of injection with adjuvant, freeze-drying preparation for injection with adjuvant or do not add adjuvant, or an amount of injection of filtrate adding with adjuvant, freeze-drying preparation for injection with adjuvant or do not add adjuvant; Mixing, (filtration)], regulate pH value to 5.5～8.5, adding 0.02～0.5% active carbon is heated to and boils, keeping little boiled 10～60 minutes, filter, regulate pH value to 5.5～8.5, add the injection water and adjust volume, survey pH value, fine straining (crossing the sintered glass filter in 0.22～0.50 μ m microporous filter membrane or similar aperture) to full dose, aseptic subpackagedly contain 1～8ml medicinal liquid to every control injection vial, lyophilization, is rolled lid at gland, promptly gets the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material;
(10) freeze drying process is as follows:
The a pre-freeze stage: common pre-freeze, pre-freeze temperature-60 ℃～-40 ℃, about 2.5～6 hours of pre-freeze time; Or select pre-freeze repeatedly, pre-freeze temperature-60 ℃～-40 ℃, the temperature of rising again-15 ℃～-7 ℃, about 3～9 hours of pre-freeze time;
The b sublimation drying stage: when the temperature of condenser reduce to-60 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-30 ℃～-15 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water;
C is drying stage again: shelf temperature slowly rises to-5 ℃～30 ℃, judges exsiccant terminal point with the vacuum descent method again, again 3～12 hours drying times.
14, the preparation method of pharmaceutical preparation according to claim 13 is characterized in that the preparation method of described pharmaceutical preparation may further comprise the steps:
One, the extraction of the alkaloids composition of the polysaccharide composition of the saponin component of the Radix Ginseng in Radix Ginseng and the Radix Aconiti Lateralis Preparata, Radix Ginseng, Radix Aconiti Lateralis Preparata:
(1) processing of Radix Ginseng: Radix Ginseng adds 3～9 times of amounts respectively (for the first time because be dried medical material, add 1～3 times of amount) 60～80% alcohol reflux or supersound extraction 1～4 time, each 0.5～3 hour, merge extractive liquid,, filter, filtrate recycling ethanol is to there not being the alcohol flavor, adjustment liquor strength extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.75～3.0g, the water saturated n-butanol extraction of 0.7～3 times of amount of each adding 2～9 times merges n-butyl alcohol liquid, reclaims n-butyl alcohol to there being the n-butyl alcohol flavor, change water-soluble, filter, get the saponin component intermediate of Radix Ginseng, standby;
(2) processing of medicine residues of Radix Ginseng: the medicine residues of Radix Ginseng after the alcohol extraction adds 6～13.5 times of water gagings respectively (for the first time because be dried medical material, add 1～5 times of amount) decoct to extract or reflux, extract, 2～6 times, each 0.5～4 hour, merge extractive liquid,, filter, adjusting liquor strength is 0.4～1.6g crude drug/ml, and temperature is 20～60 ℃, adds 90～95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 75～85%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, the merging supernatant (or not centrifugal, filter the back and directly adjust liquor strength, be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid), adjusting the supernatant liquor strength is 1.0～4.0g crude drug/ml, add 90～95% ethanol and make and contain the alcohol amount and reach 65～75%, standing over night filters, precipitation is dry, after adding the suitable quantity of water dissolving, centrifugal, get supernatant; Or not centrifugal, directly get filtrate after the filtration, be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid;
Previous step gained supernatant or filtrate are the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid;
Previous step gained supernatant adopts the ultrafilter membrane ultrafiltration of molecular cut off 100K～700K, and ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or with of the ultrafilter membrane ultrafiltration of previous step gained supernatant elder generation with molecular cut off 100K～700K, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 1K～8K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
(3) processing of Radix Aconiti Lateralis Preparata: the Radix Aconiti Lateralis Preparata medical material adds 4～12 times of water gagings (for the first time because be dried medical material, add 1～4 times of amount), decoct extraction or reflux, extract, 2～6 times, each 0.5～3 hour, merge extractive liquid,, filter, the relative density of adjusting medicinal liquid is 1.01～1.21,20～60 ℃ of temperature, ethanol precipitate with ethanol with 90～95% makes ethanol content reach 65～75%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, the relative density of adjusting medicinal liquid is that the ethanol of adding 90～95% in 1.15～1.40 o'clock makes pure content reach 80～90%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid;
Filtrate being heated to boiled, and keep little and boiled 15～55 minutes, cooling, cold preservation filters, and promptly gets the Radix Aconiti Lateralis Preparata intermediate, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor for the first time, adjustment medicinal liquid extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.8～2.0g, the dichloromethane extraction of 0.8～2 times of amount of each adding 3～7 times, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Two, play the processing of synergistic medicine:
(4) play the processing of synergistic medicine: playing a synergistic medicine is Chinese medicine or natural drug, extracts its effective site or effective ingredient monomer, and (make with extra care) is water-soluble, regulates pH value to 6～8, and filtration promptly gets intermediate, and is standby; Playing synergistic medicine is chemicals, and water-soluble, pH value transfers to 6～8, filters, and promptly gets intermediate, standby; Can be not water-soluble when being used to prepare oral solid formulation, oral semi-solid preparation;
Three, the preparation of each pharmaceutical preparation finished product:
(5) intermediate of merging " (1) ", " (2) ", " (3) " gained, mixing;
(6) intermediate with " (1) ", " (2) ", " (3) ", " (4) " gained merges, mixing, gained medicinal liquid filter, and are condensed into clear paste and [or " (5) " gained medicinal liquid are filtered, be condensed into clear paste, the intermediate not water-soluble with " (4) " mixes], add an amount of oral solid formulation, oral semi-solid preparation auxiliary materials and mixing, the system soft material, granulate, oven dry, tabletting or incapsulate is made the sheet or the capsule that contain Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material;
(7) " (5) " gained medicinal liquid being adjusted volume with purified water or water for injection is 0.1～1.5g crude drug/ml to being equivalent to liquor strength, regulates pH value to 6.0～8.0, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter that [or filtrate adds " (the 4) " intermediate of gained and an amount of oral liquid with adjuvant or do not add adjuvant, or filtrate adds an amount of oral liquid with adjuvant or do not add adjuvant; Mixing, (filtration)], regulate pH value to 6.0～8.0, add purified water or water for injection and adjust volume to full dose, survey pH value, filter, according to dosage divide to be filled in the oral liquid packing container, seal, the conventional method sterilization promptly gets the oral liquid that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material;
(8) " (5) " gained medicinal liquid being adjusted volume with water for injection is 0.1～1.5g crude drug/ml to being equivalent to liquor strength, regulates pH value to 6.0～8.0, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter that [or filtrate adds " (the 4) " intermediate of gained and an amount of injection with adjuvant or do not add adjuvant, or filtrate adds an amount of injection with adjuvant or do not add adjuvant; Mixing, (filtration)], (regulating pH value to 6.0～8.0), adding 0.02～0.3% active carbon is heated to and boils, and keeps little and boils 10～40 minutes, filters, regulate pH value to 6.0～8.0, add the injection water and adjust volume, survey pH value, fine straining (crossing the sintered glass filter in 0.22～0.45 μ m microporous filter membrane or similar aperture) to full dose, according to dosage divide and be filled to the infusion pump dress with in the container, seal, the conventional method sterilization promptly gets the injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material;
(9) " (5) " gained medicinal liquid being adjusted volume with water for injection is 0.5～2.0g crude drug/ml to being equivalent to liquor strength, regulates pH value to 6.0～8.0, is heated to and boils, keep little and boiled 20～60 minutes, and cooling, cold preservation is spent the night; Filter [or filtrate add " (the 4) " intermediate of gained and an amount of injection with adjuvant, freeze-drying preparation for injection with adjuvant or do not add adjuvant, or an amount of injection of filtrate adding with adjuvant, freeze-drying preparation for injection with adjuvant or do not add adjuvant; Mixing, (filtration)], regulate pH value to 6.0～8.0, adding 0.02～0.3% active carbon is heated to and boils, keeping little boiled 10～40 minutes, filter, regulate pH value to 6.0～8.0, add the injection water and adjust volume, survey pH value, fine straining (crossing the sintered glass filter in 0.22～0.45 μ m microporous filter membrane or similar aperture) to full dose, aseptic subpackagedly contain 1～6ml medicinal liquid to every control injection vial, lyophilization, is rolled lid at gland, promptly gets the freeze-drying preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in a kind of raw material;
(10) freeze drying process is as follows:
The a pre-freeze stage: common pre-freeze, pre-freeze temperature-60 ℃～-45 ℃, about 2.5～6 hours of pre-freeze time; Or select pre-freeze repeatedly, pre-freeze temperature-60 ℃～-45 ℃, the temperature of rising again-13 ℃～-8 ℃, about 3～9 hours of pre-freeze time;
The b sublimation drying stage: when the temperature of condenser reduce to-60 ℃～below-45 ℃, open vacuum pump, the temperature of shelf is arranged on-26 ℃～-15 ℃, goods slowly heat up, and observe the distillation interface, all walk only the end of sublimation drying stage to free water;
C is drying stage again: shelf temperature slowly rises to 0 ℃～25 ℃, judges exsiccant terminal point with the vacuum descent method again, again 3～12 hours drying times.
15, the preparation method of pharmaceutical preparation according to claim 14 is characterized in that the method for " extraction of the polysaccharide composition of the saponin component of the Radix Ginseng in Radix Ginseng and the Radix Aconiti Lateralis Preparata, Radix Ginseng, the alkaloids composition of Radix Aconiti Lateralis Preparata " in the preparation method of described pharmaceutical preparation can be more preferably:
(1) processing of Radix Ginseng: Radix Ginseng adds 5～7 times of amounts respectively (for the first time because be dried medical material, add 1～2 times of amount) 65～75% alcohol reflux 2～3 times, each 1.5～2.5 hours, merge extractive liquid,, filter, filtrate recycling ethanol is to there not being the alcohol flavor, adjustment liquor strength extremely every 1ml is equivalent to contain Radix Ginseng crude drug 1.0～2.0g, the water saturated n-butanol extraction of 0.8～1.2 times of amount of each adding 5～7 times merges n-butyl alcohol liquid, reclaims n-butyl alcohol to there being the n-butyl alcohol flavor, change water-soluble, filter, get the saponin component intermediate of Radix Ginseng, standby;
(2) processing of medicine residues of Radix Ginseng: the medicine residues of Radix Ginseng after the alcohol extraction adds 8～10 times of water gagings respectively (for the first time because be dried medical material, add 2～4 times of amounts) decoct to extract or reflux, extract, 2～4 times, each 1.5～2.5 hours, merge extractive liquid,, filter, adjusting liquor strength is 0.6～1.0g crude drug/ml, and temperature is 20～45 ℃, adds 95% ethanol precipitate with ethanol and makes and contain the alcohol amount and reach 75～85%, standing over night, filter, precipitation is dry, add the suitable quantity of water dissolving after, centrifugal, the merging supernatant (or not centrifugal, filter the back and directly adjust liquor strength, be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid), adjusting the supernatant liquor strength is 1.5～2.5g crude drug/ml, add 95% ethanol and make and contain the alcohol amount and reach 65～75%, standing over night filters, precipitation is dry, after adding the suitable quantity of water dissolving, centrifugal, get supernatant; Or not centrifugal, directly get filtrate after the filtration, be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid;
Previous step gained supernatant or filtrate are the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid;
Previous step gained supernatant adopts the ultrafilter membrane ultrafiltration of molecular cut off 100K～600K, and ultrafiltrate is the polysaccharide composition intermediate of Radix Ginseng, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or with of the ultrafilter membrane ultrafiltration of previous step gained supernatant elder generation with molecular cut off 100K～600K, then the gained ultrafiltrate is removed small-molecular weight impurity with the ultrafilter membrane of molecular cut off 1K～8K, promptly get the polysaccharide composition intermediate of Radix Ginseng, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
(3) processing of Radix Aconiti Lateralis Preparata: the Radix Aconiti Lateralis Preparata medical material adds 7～9 times of water gagings (for the first time because be dried medical material, add 1～3 times of amount), decoct extraction or reflux, extract, 2～4 times, each 0.5～1.5 hour, merge extractive liquid,, filter, the relative density of adjusting medicinal liquid is 1.05～1.16,40～55 ℃ of temperature, ethanol precipitate with ethanol with 95% makes ethanol content reach 65～75%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, the relative density of adjusting medicinal liquid is that the ethanol of adding 95% in 1.21～1.35 o'clock makes pure content reach 80～90%, standing over night filters, and filtrate recycling ethanol is to there not being the alcohol flavor, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for oral solid formulation, oral semi-solid preparation, the preparation of oral liquid;
Filtrate being heated to boiled, and keep little and boiled 30～45 minutes, cooling, cold preservation filters, and promptly gets the Radix Aconiti Lateralis Preparata intermediate, and is standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection;
Or alcohol deposit fluid reclaims ethanol to there not being the alcohol flavor for the first time, adjustment medicinal liquid extremely every 1ml is equivalent to contain Radix Ginseng crude drug 0.8～1.2g, the dichloromethane extraction of 0.8～1.2 times of amount of each adding 4～6 times, combined dichloromethane liquid reclaims dichloromethane to the greatest extent, changes water-soluble, filter, promptly get the Radix Aconiti Lateralis Preparata intermediate, standby, can be used for the preparation of oral solid formulation, oral semi-solid preparation, oral liquid, injection.
16, the method for quality control of the described pharmaceutical preparation of a kind of claim 1, it is characterized in that containing in the described a kind of raw material that preferably provides the method for quality control of the injection of Radix Ginseng and Radix Aconiti Lateralis Preparata, comprise following discriminating, inspection, assay, finger printing;
Described discriminating, inspection, assay, finger printing and one concrete grammar wherein can be selected for use respectively or combination in any is used, and are used for the quality control of injection that a kind of raw material contains Radix Ginseng and Radix Aconiti Lateralis Preparata, freeze-drying preparation for injection, sterile packaged preparation for injection;
The method of quality control of injection, freeze-drying preparation for injection, sterile packaged preparation for injection that contains Radix Ginseng and Radix Aconiti Lateralis Preparata in described a kind of raw material is as follows:
[1] differentiates
[1.1] Radix Ginseng, ginsenoside Rg 1, the ginsenoside Re discriminating:
Get the injection content 1.0g for preparing, add water 30ml dissolving, add chloroform 30～50ml, jolting, water intaking liquid evaporate to dryness, residue adds water 2ml makes dissolving, added water saturated n-butyl alcohol 8～12ml supersound process 25～40 minutes, get n-butyl alcohol liquid and put in the separatory funnel, add the ammonia solution of 2～4 times of amounts, washing, washing liquid discards, get n-butyl alcohol liquid evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution.Other gets ginseng crude drug 1g (crossing 60 mesh sieves), adds ethanol 20～40ml, and reflux 20～40 minutes filters, and filtrate evaporate to dryness, residue add water 30ml makes dissolving, filters, and filtrate is made ginseng crude drug's solution with method.Get the ginsenoside Rg more respectively 1, ginsenoside Re's reference substance is an amount of, add methanol and make the solution that every 1ml contains 2mg respectively, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 2.5～5 μ l of above-mentioned four kinds of solution, put respectively on same silica gel g thin-layer plate, (14～16: 38～42: 21～23: 9～11) lower floor's solution of placing below 10 ℃ is developing solvent with chloroform-ethyl acetate-methanol-water, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃.In the test sample chromatograph, with the corresponding position of ginseng crude drug's chromatograph on, show the speckle of same color; With the corresponding position of reference substance chromatograph on, show the speckle of same color;
[1.2] discriminating of Radix Aconiti Lateralis Preparata:
Get the injection content 3.5g for preparing, add water 30ml dissolving, regulate pH value to 9～11 with ammonia solution, the jolting that adds diethyl ether is extracted 2～4 times, each 40～50ml, and ether liquid low temperature evaporate to dryness, residue adds dehydrated alcohol 2ml makes dissolving, as need testing solution.Other gets Radix Aconiti Lateralis Preparata medicinal material coarse powder 20g, puts in the tool plug conical flask the 130～170ml that adds diethyl ether, jolting 9～12 minutes, add ammonia solution 9～11ml, jolting 25～35 minutes was placed 1～2 hour, divided and got the ether layer, volatilize, residue adds dehydrated alcohol 2ml makes dissolving, as Radix Aconiti Lateralis Preparata medical material solution.Test according to thin layer chromatography (appendix VIB of Chinese Pharmacopoeia version in 2005), draw each 12～30 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate (thick 500 μ m), chromatography cylinder is earlier saturated with ammonia, with petroleum ether-ether-acetone (4.8～5.2: 2.8～3.2: 2.8～3.2) be developing solvent, launch, take out, dry, spray is with bismuth potassium iodide test solution, and it is clear to be heated to speckle colour developing.In the test sample chromatograph, with the corresponding position of Radix Aconiti Lateralis Preparata medical material chromatograph on, show the speckle of same color;
[2] check
[2.1] limit examine of aconite alkaloids:
The preparation of [2.1.1] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the aconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 50ml measuring bottle, adds the 0.01mol/L hydrochloric acid solution and makes dissolving, and be diluted to scale, shakes up, promptly;
The preparation precision of [2.1.2] standard curve is measured above-mentioned reference substance solution 0.2～0.3ml, 0.4～0.6ml, 0.65～0.85ml, 0.9～1.1ml, 1.4～1.6ml, 1.65～1.85ml, 1.9～2.1ml, put in the separatory funnel respectively, add 0.01mol/L hydrochloric acid solution 1.85～1.65ml respectively, 1.6～1.4ml, 1.35～1.15ml, 1.1～0.9ml, 0.6～0.4ml, 0.35～0.15ml, 0.00ml, accurate respectively adding acetic acid-sodium-acetate buffer (is got 0.2mol/L acetum 250ml, with 0.2mol/L sodium acetate solution adjust pH to 3.1) 8～12ml, bromocresol green liquid (is got bromocresol green 50mg, add 0.05mol/L sodium hydroxide solution 1.6ml grinding and make dissolving, add water to 100ml, extract 3 times with the chloroform jolting, each 30ml, discard chloroform solution) 1.5～2.5ml, chloroform 8～12ml, jolting 2～6 minutes, leave standstill, divide and get chloroform solution, retinue is blank, measures absorbance at 415nm wavelength place according to spectrophotometric degree method (an appendix V of Chinese Pharmacopoeia version in 2005 A).With concentration is abscissa, and absorbance is a vertical coordinate, the drawing standard curve;
The injection content 0.35g for preparing is got in the preparation of [2.1.3] need testing solution, and accurate the title decides, and adds water 10ml dissolving, be transferred in the separatory funnel, add ammonia solution and regulate pH value to 10～11, extract 3～5 times with the chloroform jolting of equivalent, combined chloroform liquid, evaporate to dryness.Residue adds 0.01mol/L hydrochloric acid solution gradation dissolving, and changes in the 10ml measuring bottle, is diluted to scale and shakes up, promptly;
[2.1.4] algoscopy precision is measured above-mentioned need testing solution 2ml, puts respectively in the separatory funnel, measures absorbance according to " preparation of standard curve " item down from " each precision adds acetic acid-sodium-acetate buffer 10ml " in accordance with the law, calculates, promptly;
Every of the injection that [2.1.5] prepares contains aconite alkaloids with aconitine (C 24H 47NO 11) meter, must not be higher than 1.0mg;
[3] assay
[3.1] ginsenoside Rg 1Assay with Re:
Measure according to high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2005).
[3.1.1] chromatographic condition is filler with the octadecylsilane chemically bonded silica, acetonitrile-0.1% phosphoric acid solution (17～23: 75～85) be mobile phase, detect wavelength 203nm.Number of theoretical plate calculates by the ginsenoside Re peak should be not less than 5000;
The preparation of [3.1.2] reference substance solution ginsenoside Rg of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1And ginsenoside Re's reference substance is an amount of, accurately claims surely, adds methanol and makes every 1ml respectively and contain the ginsenoside Rg 10.3mg, the solution of ginsenoside Re 0.2mg, promptly;
The injection content 0.35g for preparing under the content uniformity item is got in the preparation of [3.1.3] need testing solution, accurate claims surely, puts in the 5ml measuring bottle, is dissolved in water and is diluted to scale, shakes up, promptly.
Accurate respectively reference substance solution and each 10～30 μ l of need testing solution of drawing of [3.1.4] algoscopy inject chromatograph of liquid, measure, promptly;
Every of the injection that [3.1.5] prepares contains the ginsenoside Rg 1, the ginsenoside Re the content sum should be 0.2～2.0mg;
[3.2] Determination of Total Saponin Content in Panax Ginseng:
The preparation of [3.2.1] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance is an amount of, accurate claims surely, adds methanol and makes every 1ml and contain ginsenoside Rb 10.3mg solution, promptly;
The preparation precision of [3.2.2] standard curve is measured above-mentioned ginsenoside Rb 1Reference substance solution 0.05～0.15ml, 0.15～0.25ml, 0.25～0.35ml, 0.35～0.45ml, 0.45～0.55ml, 0.55～0.65ml, 0.65～0.75ml puts respectively in the tool plug test tube, fling to solvent, each adds 5% vanillin-glacial acetic acid solution (fresh preparation) 0.15～0.25ml and perchloric acid 0.6～1.0ml, heats 12～18min in 55～65 ℃ of water-baths, takes out, cooling rapidly, add glacial acetic acid 3～7ml, shake up, simultaneously with reagent corresponding as blank, measure absorbance in 556nm wavelength place according to spectrophotography (an appendix V of Chinese Pharmacopoeia version in 2005 A), with the absorbance is vertical coordinate, and sampling amount is an abscissa, the drawing standard curve;
The injection content 0.35g for preparing under the content uniformity item is got in the preparation of [3.2.3] sample solution, the accurate title, decide, adding distil water 10ml dissolving is put in the separatory funnel, extracts 3～5 times with the chloroform jolting, each 8～12ml, discard chloroform solution, water liquid extracts 3～5 times with water saturated n-butyl alcohol jolting again, each 8～12ml, merge n-butyl alcohol liquid, the saturated water washing of reuse n-butyl alcohol 2～3 times, each 8～12ml discards water liquid, n-butyl alcohol liquid is put evaporate to dryness in the water-bath, residue adds dissolve with methanol, is transferred in the 10ml measuring bottle, and is diluted to scale, shake up, promptly;
[3.2.4] algoscopy precision is measured sample solution 0.1ml and is put in the tool plug test tube, measures absorbance from " flinging to solvent " down according to " standard curve preparation " in accordance with the law, calculating, promptly;
Every of the injection that [3.2.5] prepares contains Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) meter, should be 2.0～20.0mg;
[3.3] assay of polysaccharide composition in the Radix Ginseng
The preparation of [3.3.1] reference substance solution phosphorus pentoxide drying under reduced pressure of learning from else's experience is an amount of to the glucose reference substance of constant weight, accurately claims surely, and adding distil water is made the solution that every 1ml contains glucose 0.2mg, promptly;
The preparation precision of [3.3.2] standard curve is measured glucose reference substance solution 0.05～0.15ml, 0.15～0.25ml, 0.25～0.35ml, 0.35～0.45ml, 0.45～0.55ml, 0.55～0.65ml, 0.65～0.75ml, 0.75～0.85ml puts respectively in the tool plug test tube, add water respectively and make into 1.0ml, each accurate freshly prepared 0.2% anthrone sulphuric acid (sulfuric acid concentration is 80%) solution 6～10ml that adds shakes up, and heats 8～12 minutes in 100 ℃ of water-baths, rapidly after the cooling, retinue is blank, measures absorbance at 620nm wavelength place according to spectrophotography, is abscissa with concentration, absorbance is a vertical coordinate, the drawing standard curve;
The injection content 3.5g for preparing under the content uniformity item is got in the preparation of [3.3.3] sample solution, accurate claim fixed, add water 25～35ml dissolving after, adding ethanol makes and contains the alcohol amount and reach 75～85%, centrifugal, add dissolved in distilled water after precipitation volatilizes ethanol, and be settled in the 100ml measuring bottle.Get above-mentioned solution 1～3ml and put in the 25ml measuring bottle, thin up shakes up to scale, promptly;
[3.3.4] algoscopy precision is measured 0.2ml in tool plug test tube, measures absorbance from " adding distil water is to 1.0ml respectively " down by " preparation of standard curve " item in accordance with the law, calculates, promptly;
Every of the injection that [3.3.5] prepares contains polysaccharide composition in the Radix Ginseng with glucose meter, should be 4～60mg;
[4] finger printing
With reference to high performance liquid chromatography, measure in conjunction with the requirement of finger printing
[4.1] ginseng crude drug's finger printing
[4.1.1] medical material title and source
Radix Ginseng in this Radix Ginseng medicinal materials fingerprint is selected Radix Ginseng Rubra for use, and Radix Ginseng Rubra is the dry root and rhizome of cultivation product (practise and claim " Park Ginseng ") after steaming of Araliaceae Radix Ginseng Panaxginseng C.A.Mey.;
The mensuration of [4.1.2] ginseng crude drug finger printing
[4.1.2.1] chromatographic condition chromatographic column: ODS HYPERSIL C 18(post of Φ 4.6mm * 250mm), packing material size 5 μ m; Mobile phase: acetonitrile and water are mobile phase, and gradient condition sees the following form
Gradient condition Time (min Acetonitrile (%) Water (%) 0 20 55 60 10～20 25～35 45～55 10～20 90～80 75～65 55～45 90～80
Column temperature: 25～35 ℃; Analysis time: 60～120min; Flow velocity: 0.6～1.0ml/min; 100～120 ℃ of ELSD drift tubes, flow rate of carrier gas 2.0～4.0ml/min.Number of theoretical plate is with ginsenoside Rb 1The peak meter should be not less than 5000;
The preparation of [4.1.2.2] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance 2mg, the accurate title, decide, and puts in the 5ml measuring bottle, adds dissolve with methanol and be settled to scale, shakes up, promptly;
Radix Ginseng Rubra medicinal powder (crossing 20 mesh sieves) 2g is got in the preparation of [4.1.2.3] need testing solution, and accurate the title decides, and adds 70% alcohol reflux 2 times, and each 0.8～1.5 hour, add 6～8 times of amounts for the first time, add 5～7 times of amounts for the second time; The extracting solution filtration, filtrate recycling ethanol concentrates, and extracts 4～6 times with the water saturated n-butyl alcohol jolting of equivalent, merges n-butyl alcohol liquid, and washs with the ammonia solution of equivalent; Cleaning mixture discards, and n-butyl alcohol liquid evaporate to dryness, residue are dissolved in water and are transferred in the 10ml measuring bottle, are settled to scale, shake up, promptly;
Accurate respectively reference substance solution and each 5～20 μ l of need testing solution of drawing of [4.1.2.4] algoscopy inject chromatograph of liquid, measure, promptly;
[4.1.2.5] result is with ginsenoside Rb 1The peak is set at the object of reference peak, and with the logarithm of its peak area as 1.000, according to ginsenoside Rb 1The retention time at peak is calculated, calibrate 10 total peaks altogether, its relative retention time is respectively: 0.056 ± 10%, 0.102 ± 10%, 0.372 ± 10%, 0.632 ± 10%, 0.931 ± 10%, 1.000,1.040 ± 10%, 1.084 ± 10%, 1.181 ± 10%, 1.662 ± 10%; And the regulation relative retention time is that the relative peak area at 0.632 ± 10%, 1.040 ± 10%, 1.084 ± 10%, 1.181 ± 10% peak is 0.682～1.266,0.346～0.644,0.262～0.486,0.103～0.191 than (peak area logarithm ratio);
[4.2] Radix Aconiti Lateralis Preparata medicinal materials fingerprint
The title and the source of [4.2.1] Radix Aconiti Lateralis Preparata medical material
Radix Aconiti Lateralis Preparata is the processed goods of the daughter root of ranunculaceae plant Aconitum carmichjaelii Debx. Aconitum carmichaeli Debx..Radix Aconiti Lateralis Preparata in this Radix Aconiti Lateralis Preparata medicinal materials fingerprint is selected Radix Aconiti Lateralis Preparata for use;
The mensuration of [4.2.2] Radix Aconiti Lateralis Preparata medicinal materials fingerprint
[4.2.2.1] chromatographic condition chromatographic column: ZORBAX SB C 18(post of Φ 4.6mm * 150mm), packing material size 5 μ m; Mobile phase: methanol and 0.1% phosphoric acid-0.04% triethylamine-aqueous solution are mobile phase, and gradient condition sees the following form
Gradient condition Time (min) Methanol (%) 0.1% phosphoric acid-0.04% triethylamine-aqueous solution (%) 0 10 45 60 25～35 25～35 37～47 37～47 75～65 75～65 63～53 63～53
Column temperature: 25～35 ℃; Detect wavelength: 235nm; Analysis time: 60～120min; Flow velocity: 0.7～1.3ml/min; Number of theoretical plate calculates with the mesaconitine peak should be not less than 2000;
The preparation of [4.2.2.2] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the mesaconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, promptly;
The preparation Radix Aconiti Lateralis Preparata medicinal material coarse powder 10g of [4.2.2.3] need testing solution spends the night with ammonia solution 3～5ml, ether 40～60ml merceration, filters; Medicinal residues add diethyl ether: chloroform (2.8～3.2: 0.8～1.2) mixed solution 40～60ml, supersound process 25～40min, filter, medicinal residues wash 3～4 times with mixed solution, each 14～16ml, and washing liquid and filtrate merge, the low temperature evaporate to dryness, residue adds methanol 5ml dissolving, with the filter membrane filtration of 0.45 μ m, can measure;
Accurate respectively inner mark solution 15～25 μ l and need testing solution 30～50 μ l of drawing of [4.2.2.4] algoscopy inject chromatograph of liquid, measure, promptly;
[4.2.2.5] result is set at the object of reference peak with the mesaconitine peak, retention time according to the mesaconitine peak is calculated, calibrate 8 total peaks altogether, its relative retention time is respectively: 0.146 ± 10%, 0.254 ± 10%, 0.531 ± 10%, 0.630 ± 10%, 0.727 ± 10%, 0.833 ± 10%, 1.000,1.114 ± 10%; And with the stable and the biggest relative retention time of peak area be the peak area at 0.531 ± 10% peak as 1.000, and the regulation relative retention time is that the relative peak area ratio at 0.727 ± 10%, 1.114 ± 10% peak is 0.182～0.272,0.373～0.552;
[4.3] ginsenoside's finger printing
[4.3.1] chromatographic condition chromatographic column: ODS HYPERSIL C 18(post of Φ 4.6mm * 250mm), packing material size 5 μ m; Acetonitrile and water are mobile phase, and gradient condition sees the following form:
Gradient condition Time (min) Acetonitrile (%) Water (%) 0 20 55 60 10～20 25～35 45～55 10～20 90～80 75～65 55～45 90～80
Column temperature: 25～35 ℃; Analysis time: 60～120min; Flow velocity: 0.6～1.0ml/min; ELSD parameter: 100～120 ℃ of drift tubes, flow rate of carrier gas 2.0～4.0ml/min.Number of theoretical plate is with ginsenoside Rb 1The peak calculates should be not less than 5000;
The preparation of [4.3.2] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance 2mg, the accurate title, decide, and puts in the 5ml measuring bottle, adds dissolve with methanol and be settled to scale, shakes up, promptly;
The injection content 0.35g for preparing is got in the preparation of [4.3.3] need testing solution, the accurate title, decide, be dissolved in water to 8～12ml, extract 4～6 times, merge n-butyl alcohol liquid with the water-saturated n-butanol jolting of equivalent, ammonia solution with equivalent washs 1～2 time, divide and to get n-butyl alcohol liquid, reclaim n-butyl alcohol, residue is with dissolved in distilled water and be settled in the 5ml measuring bottle, shake up, promptly;
Accurate respectively reference substance solution and each 5～15 μ l of need testing solution of drawing of [4.3.4] algoscopy inject chromatograph of liquid, measure, promptly;
[4.3.5] result is with ginsenoside Rb 1The peak is set at the object of reference peak, and with the logarithm of its peak area as 1.000, according to ginsenoside Rb 1The retention time at peak is calculated, calibrate 10 total peaks altogether, its relative retention time is respectively: 0.441 ± 10%, 0.632 ± 10%, 0.942 ± 10%, 1.000,1.042 ± 10%, 1.062 ± 10%, 1.089 ± 10%, 1.188 ± 10%, 1.701 ± 10%, 1.737 ± 10%; And the regulation relative retention time is that the relative peak area at 0.441 ± 10%, 0.632 ± 10%, 1.042 ± 10%, 1.089 ± 10% peak is respectively 0.688～1.148,0.673～1.121,0.692～1.154,0.688～1.146 than (peak area logarithm ratio);
[4.4] fingerprint of alkaloid
[4.4.1] chromatographic condition chromatographic column: ZORBAX SB C 18(post of Φ 4.6mm * 150mm), packing material size 5 μ m; Methanol and 0.1% phosphoric acid-0.04% triethylamine aqueous solution are mobile phase, and gradient condition sees the following form:
Gradient condition Time (min) Methanol (%) 0.1% phosphoric acid-0.04% triethylamine aqueous solution (%) 0 10 45 60 25～35 25～35 37～47 37～47 75～65 75～65 63～53 63～53
25～35 ℃ of column temperatures; Detect wavelength: 235nm; Analysis time: 60～120min; Number of theoretical plate calculates with the mesaconitine peak should be not less than 2000;
The preparation of [4.4.2] inner mark solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the mesaconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 25ml measuring bottle, adds 1% phosphoric acid solution 3ml, is diluted to scale with methanol, shakes up, promptly;
The preparation precision of [4.4.3] need testing solution is measured the injection content 1.0g for preparing, be dissolved in water to 8～12ml, put in the separatory funnel, add ammonia solution adjust pH to 9～11, with the E-C (2.8～3.2: mixed liquor jolting extraction 0.8～1.2) 4～6 times of equivalent, combining extraction liquid, low temperature volatilizes, and residue adds methanol 1ml makes dissolving, adds inner mark solution 40～60 μ l, filter, as need testing solution;
Accurate respectively inner mark solution 15～25 μ l and need testing solution 30～50 μ l of drawing of [4.4.4] algoscopy inject chromatograph of liquid, measure, promptly;
[4.4.5] result is set at the mesaconitine peak at the object of reference peak of relative retention time, retention time according to the mesaconitine peak is calculated, calibrate 5 total peaks altogether, its relative retention time is respectively: 0.259 ± 10%, 0.342 ± 10%, 0.532 ± 10%, 0.724 ± 10%, 1.000 ± 10%; And with the stable and the biggest relative retention time of peak area be the peak area at 0.532 ± 10% peak as 1.000, and the regulation relative retention time is that the relative peak area ratio at 0.724 ± 10% peak is 0.455～0.683.
17, the method for quality control of pharmaceutical preparation according to claim 16, it is characterized in that containing in described a kind of raw material injection, the freeze-drying preparation for injection of Radix Ginseng and Radix Aconiti Lateralis Preparata, the method for quality control of sterile packaged preparation for injection, its each concrete steps are as follows:
[1] differentiates
[1.1] Radix Ginseng, ginsenoside Rg 1, the ginsenoside Re discriminating:
Get the injection content 1.0g for preparing, add water 30ml dissolving, add chloroform 40ml, jolting, water intaking liquid evaporate to dryness, residue adds water 2ml makes dissolving, added water saturated n-butyl alcohol 10ml supersound process 30 minutes, get n-butyl alcohol liquid and put in the separatory funnel, add the ammonia solution of 3 times of amounts, washing, washing liquid discards, get n-butyl alcohol liquid evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution.Other gets ginseng crude drug 1g (crossing 60 mesh sieves), adds ethanol 30ml, and reflux 30 minutes filters, and filtrate evaporate to dryness, residue add water 30ml makes dissolving, filters, and filtrate is made ginseng crude drug's solution with method.Get the ginsenoside Rg more respectively 1, ginsenoside Re's reference substance is an amount of, add methanol and make the solution that every 1ml contains 2mg respectively, in contrast product solution.Test according to thin layer chromatography (appendix VIB of Chinese Pharmacopoeia version in 2005), draw each 2.5～5 μ l of above-mentioned four kinds of solution, put respectively on same silica gel g thin-layer plate, (15: 40: 22: 10) lower floor's solution of placing below 10 ℃ was developing solvent with chloroform-ethyl acetate-methanol-water, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃.In the test sample chromatograph, with the corresponding position of ginseng crude drug's chromatograph on, show the speckle of same color; With the corresponding position of reference substance chromatograph on, show the speckle of same color;
[1.2] discriminating of Radix Aconiti Lateralis Preparata:
Get the injection content 3.5g for preparing, add water 30ml dissolving, regulate pH value to 10 with ammonia solution, the jolting that adds diethyl ether is extracted 3 times, each 45ml, and ether liquid low temperature evaporate to dryness, residue adds dehydrated alcohol 2ml makes dissolving, as need testing solution.Other gets Radix Aconiti Lateralis Preparata medicinal material coarse powder 20g, puts in the tool plug conical flask, and the 150ml that adds diethyl ether, jolting 10 minutes adds ammonia solution 10ml, and jolting 30 minutes was placed 1～2 hour, divided and got the ether layer, volatilized, and residue adds dehydrated alcohol 2ml makes dissolving, as Radix Aconiti Lateralis Preparata medical material solution.Test according to thin layer chromatography (appendix VIB of Chinese Pharmacopoeia version in 2005), draw each 12～30 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate (thick 500 μ m), chromatography cylinder is earlier saturated with ammonia, is developing solvent with petroleum ether-ether-acetone (5: 3: 3), launches, take out, dry, spray is with bismuth potassium iodide test solution, and it is clear to be heated to speckle colour developing.In the test sample chromatograph, with the corresponding position of Radix Aconiti Lateralis Preparata medical material chromatograph on, show the speckle of same color;
[2] check
[2.1] limit examine of aconite alkaloids:
The preparation of [2.1.1] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the aconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 50ml measuring bottle, adds the 0.01mol/L hydrochloric acid solution and makes dissolving, and be diluted to scale, shakes up, promptly;
The preparation precision of [2.1.2] standard curve is measured above-mentioned reference substance solution 0.25ml, 0.50ml, 0.75ml, 1.00ml, 1.50ml, 1.75ml, 2.00ml, put in the separatory funnel respectively, add 0.01mol/L hydrochloric acid solution 1.75ml respectively, 1.50ml, 1.25ml, 1.00ml, 0.50ml, 0.25ml, 0.00ml, accurate respectively adding acetic acid-sodium-acetate buffer (is got 0.2mol/L acetum 250ml, with 0.2mol/L sodium acetate solution adjust pH to 3.1) 10ml, bromocresol green liquid (is got bromocresol green 50mg, add 0.05mol/L sodium hydroxide solution 1.6ml grinding and make dissolving, add water to 100ml, extract 3 times with the chloroform jolting, each 30ml, discard chloroform solution) 2ml, chloroform 10ml, jolting 3 minutes is left standstill, and divides and gets chloroform solution, retinue is blank, measures absorbance at 415nm wavelength place according to spectrophotometric degree method (appendix VA of Chinese Pharmacopoeia version in 2005).With concentration is abscissa, and absorbance is a vertical coordinate, the drawing standard curve;
The injection content 0.35g for preparing is got in the preparation of [2.1.3] need testing solution, and accurate the title decides, and adds water 10ml dissolving, is transferred in the separatory funnel, adds ammonia solution and regulates pH value to 10～11, extracts combined chloroform liquid, evaporate to dryness 4 times with the chloroform jolting of equivalent.Residue adds 0.01mol/L hydrochloric acid solution gradation dissolving, and changes in the 10ml measuring bottle, is diluted to scale and shakes up, promptly;
[2.1.4] algoscopy precision is measured above-mentioned need testing solution 2ml, puts respectively in the separatory funnel, measures absorbance according to " preparation of standard curve " item down from " each precision adds acetic acid-sodium-acetate buffer 10ml " in accordance with the law, calculates, promptly;
Every of the injection that [2.1.5] prepares contains aconite alkaloids with aconitine (C 24H 47NO 11) meter, must not be higher than 1.0mg;
[3] assay
[3.1] ginsenoside Rg 1Assay with Re:
According to high effective liquid chromatography for measuring;
[3.1.1] chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica, and acetonitrile-0.1% phosphoric acid solution (20: 80) is a mobile phase, detects wavelength 203nm.Number of theoretical plate calculates by the ginsenoside Re peak should be not less than 5000;
The preparation of [3.1.2] reference substance solution ginsenoside Rg of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1And ginsenoside Re's reference substance is an amount of, accurately claims surely, adds methanol and makes every 1ml respectively and contain the ginsenoside Rg 10.3mg, the solution of ginsenoside Re 0.2mg, promptly;
The injection content 0.35g for preparing under the content uniformity item is got in the preparation of [3.1.3] need testing solution, accurate claims surely, puts in the 5ml measuring bottle, is dissolved in water and is diluted to scale, shakes up, promptly.
Accurate respectively reference substance solution and each the 20 μ l of need testing solution of drawing of [3.1.4] algoscopy inject chromatograph of liquid, measure, promptly;
Every of the injection that [3.1.5] prepares contains the ginsenoside Rg 1, the ginsenoside Re the content sum should be 0.8～1.4mg;
[3.2] Determination of Total Saponin Content in Panax Ginseng:
The preparation of [3.2.1] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance is an amount of, accurate claims surely, adds methanol and makes every 1ml and contain ginsenoside Rb 10.3mg solution, promptly;
The preparation precision of [3.2.2] standard curve is measured above-mentioned ginsenoside Rb 1Reference substance solution 0.1ml, 0.2ml, 0.3ml, 0.4ml, 0.5ml, 0.6ml, 0.7ml puts respectively in the tool plug test tube, fling to solvent, each adds 5% vanillin-glacial acetic acid solution (fresh preparation) 0.2ml and perchloric acid 0.8ml, heats 15min in 60 ℃ of water-baths, take out, cooling adds glacial acetic acid 5ml rapidly, shake up, the while as blank, in 556nm wavelength place is measured absorbance according to spectrophotography with reagent corresponding, with the absorbance is vertical coordinate, and sampling amount is an abscissa, the drawing standard curve;
The injection content 0.35g for preparing under the content uniformity item is got in the preparation of [3.2.3] sample solution, the accurate title, decide, adding distil water 10ml dissolving is put in the separatory funnel, extracts 4 times with the chloroform jolting, each 10ml, discard chloroform solution, water liquid extracts 4 times with water saturated n-butyl alcohol jolting again, each 10ml, merge n-butyl alcohol liquid, the saturated water washing of reuse n-butyl alcohol 2 times, each 10ml discards water liquid, n-butyl alcohol liquid is put evaporate to dryness in the water-bath, residue adds dissolve with methanol, is transferred in the 10ml measuring bottle, and is diluted to scale, shake up, promptly;
[3.2.4] algoscopy precision is measured sample solution 0.1ml and is put in the tool plug test tube, measures absorbance from " flinging to solvent " down according to " standard curve preparation " in accordance with the law, calculating, promptly;
Every of the injection that [3.2.5] prepares contains Radix Ginseng total saponins with ginsenoside Rb 1(C 54H 92O 23) meter, should be 8.0～15.0mg;
[3.3] assay of polysaccharide composition in the Radix Ginseng
The preparation of [3.3.1] reference substance solution phosphorus pentoxide drying under reduced pressure of learning from else's experience is an amount of to the glucose reference substance of constant weight, accurately claims surely, and adding distil water is made the solution that every 1ml contains glucose 0.2mg, promptly;
The preparation precision of [3.3.2] standard curve is measured glucose reference substance solution 0.1ml, 0.2ml, 0.3ml, 0.4ml, 0.5ml, 0.6ml, 0.7ml 0.8ml puts respectively in the tool plug test tube, add water respectively and make into 1.0ml, each accurate freshly prepared 0.2% anthrone sulphuric acid (sulfuric acid concentration is 80%) solution 8ml that adds shakes up, and heating is 10 minutes in 100 ℃ of water-baths, rapidly after the cooling, retinue is blank, measures absorbance at 620nm wavelength place according to spectrophotography (an appendix V of Chinese Pharmacopoeia version in 2005 A), is abscissa with concentration, absorbance is a vertical coordinate, the drawing standard curve;
The injection content 3.5g for preparing under the content uniformity item is got in the preparation of [3.3.3] sample solution, accurate claim fixed, add water 30ml dissolving after, adding ethanol makes and contains the alcohol amount and reach 80%, centrifugal, add dissolved in distilled water after precipitation volatilizes ethanol, and be settled in the 100ml measuring bottle.Get above-mentioned solution 2ml and put in the 25ml measuring bottle, thin up shakes up to scale, promptly.
[3.3.4] algoscopy precision is measured 0.2ml in tool plug test tube, measures absorbance from " adding distil water is to 1.0ml respectively " down by " preparation of standard curve " item in accordance with the law, calculates, promptly;
Every of the injection that [3.3.5] prepares contains polysaccharide composition in the Radix Ginseng with glucose meter, should be 27～47mg;
[4] finger printing
With reference to high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2005), measure in conjunction with the requirement of finger printing
[4.1] ginseng crude drug's finger printing
[4.1.1] medical material title and source
Radix Ginseng in this Radix Ginseng medicinal materials fingerprint is selected Radix Ginseng Rubra for use, and Radix Ginseng Rubra is the dry root and rhizome of cultivation product (practise and claim " Park Ginseng ") after steaming of Araliaceae Radix Ginseng Panaxginseng C.A.Mey.;
The mensuration of [4.1.2] ginseng crude drug finger printing
[4.1.2.1] chromatographic condition chromatographic column: ODS HYPERSIL C 18(post of Φ 4.6mm * 250mm), packing material size 5 μ m; Mobile phase: acetonitrile and water are mobile phase, and gradient condition sees the following form
Gradient condition Time (min Acetonitrile (%) Water (%) 0 20 55 60 15 30 50 15 85 70 50 85
Column temperature: 30 ℃; Analysis time: 60min; Flow velocity: 0.8ml/min; 110 ℃ of ELSD drift tubes, flow rate of carrier gas 3.0ml/min.Number of theoretical plate is with ginsenoside Rb 1The peak meter should be not less than 5000;
The preparation of [4.1.2.2] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to ginsenoside Rb1's reference substance 2mg of constant weight, accurately claims surely, puts in the 5ml measuring bottle, adds dissolve with methanol and is settled to scale, shakes up, promptly;
Radix Ginseng Rubra medicinal powder (crossing 20 mesh sieves) 2g is got in the preparation of [4.1.2.3] need testing solution, and accurate the title decides, and adds 70% alcohol reflux twice, and each 1 hour, add 7 times of amounts for the first time, add 6 times of amounts for the second time; The extracting solution filtration, filtrate recycling ethanol concentrates, and extracts 5 times with the water saturated n-butyl alcohol jolting of equivalent, merges n-butyl alcohol liquid, and washs with the ammonia solution of equivalent; Cleaning mixture discards, and n-butyl alcohol liquid evaporate to dryness, residue are dissolved in water and are transferred in the 10ml measuring bottle, are settled to scale, shake up, promptly;
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of [4.1.2.4] algoscopy inject chromatograph of liquid, measure, promptly;
[4.1.2.5] result is with ginsenoside Rb 1The peak is set at the object of reference peak, and with the logarithm of its peak area as 1.000, according to ginsenoside Rb 1The retention time at peak is calculated, calibrate 10 total peaks altogether, its relative retention time is respectively: 0.056 ± 10%, 0.102 ± 10%, 0.372 ± 10%, 0.632 ± 10%, 0.931 ± 10%, 1.000,1.040 ± 10%, 1.084 ± 10%, 1.181 ± 10%, 1.662 ± 10%; And the regulation relative retention time is that the relative peak area at 0.632 ± 10%, 1.040 ± 10%, 1.084 ± 10%, 1.181 ± 10% peak is 0.682～1.266,0.346～0.644,0.262～0.486,0.103～0.191 than (peak area logarithm ratio);
[4.2] Radix Aconiti Lateralis Preparata medicinal materials fingerprint
The title and the source of [4.2.1] Radix Aconiti Lateralis Preparata medical material
Radix Aconiti Lateralis Preparata is the processed goods of the daughter root of ranunculaceae plant Aconitum carmichjaelii Debx. Aconitum carmichaeli Debx.; Radix Aconiti Lateralis Preparata in this Radix Aconiti Lateralis Preparata medicinal materials fingerprint is selected Radix Aconiti Lateralis Preparata for use;
The mensuration of [4.2.2] Radix Aconiti Lateralis Preparata medicinal materials fingerprint
[4.2.2.1] chromatographic condition chromatographic column: ZORBAX SB C 18(post of Φ 4.6mm * 150mm), packing material size 5 μ m; Mobile phase: methanol and 0.1% phosphoric acid-0.04% triethylamine-aqueous solution are mobile phase, and gradient condition sees the following form
Gradient condition Time (min) Methanol (%) 0.1% phosphoric acid-0.04% triethylamine-aqueous solution (%) 0 10 45 60 30 30 42 42 70 70 58 58
Column temperature: 30 ℃; Detect wavelength: 235nm; Analysis time: 60min; Flow velocity: 1.0ml/min; Number of theoretical plate calculates with the mesaconitine peak should be not less than 2000;
The preparation of [4.2.2.2] reference substance solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the mesaconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, promptly;
The preparation Radix Aconiti Lateralis Preparata medicinal material coarse powder 10g of [4.2.2.3] need testing solution spends the night with ammonia solution 4ml, ether 50ml merceration, filters; Medicinal residues add diethyl ether: chloroform (3: 1) mixed solution 50ml, and supersound process 30min filters, and medicinal residues wash 3～4 times with mixed solution, each 15ml, washing liquid and filtrate merge, the low temperature evaporate to dryness, residue adds methanol 5ml dissolving, with the filter membrane filtration of 0.45 μ m, can measure;
Accurate respectively inner mark solution 20 μ l and the need testing solution 40 μ l of drawing of [4.2.2.4] algoscopy inject chromatograph of liquid, measure, promptly;
[4.2.2.5] result is set at the object of reference peak with the mesaconitine peak, retention time according to the mesaconitine peak is calculated, calibrate 8 total peaks altogether, its relative retention time is respectively: 0.146 ± 10%, 0.254 ± 10%, 0.531 ± 10%, 0.630 ± 10%, 0.727 ± 10%, 0.833 ± 10%, 1.000,1.114 ± 10%; And with the stable and the biggest relative retention time of peak area be the peak area at 0.531 ± 10% peak as 1.000, and the regulation relative retention time is that the relative peak area ratio at 0.727 ± 10%, 1.114 ± 10% peak is 0.182～0.272,0.373～0.552;
[4.3] ginsenoside's finger printing
[4.3.1] chromatographic condition chromatographic column: ODS HYPERSIL C 18(post of Φ 4.6mm * 250mm), packing material size 5 μ m; Acetonitrile and water are mobile phase, and gradient condition sees the following form:
Gradient condition Time (min) Acetonitrile (%) Water (%) 0 20 55 60 15 30 50 15 85 70 50 85
Column temperature: 30 ℃; Analysis time: 60min; Flow velocity: 0.8ml/min; ELSD parameter: 110 ℃ of drift tubes, flow rate of carrier gas 3.0ml/min.Number of theoretical plate is with ginsenoside Rb 1The peak calculates should be not less than 5000;
The preparation of [4.3.2] reference substance solution ginsenoside Rb of phosphorus pentoxide drying under reduced pressure that learn from else's experience to constant weight 1Reference substance 2mg, the accurate title, decide, and puts in the 5ml measuring bottle, adds dissolve with methanol and be settled to scale, shakes up, promptly;
The injection content 0.35g for preparing is got in the preparation of [4.3.3] need testing solution, the accurate title, decide, be dissolved in water to 10ml, extract 5 times, merge n-butyl alcohol liquid with the water-saturated n-butanol jolting of equivalent, ammonia solution with equivalent washs 1 time, divide and to get n-butyl alcohol liquid, reclaim n-butyl alcohol, residue is with dissolved in distilled water and be settled in the 5ml measuring bottle, shake up, promptly;
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of [4.3.4] algoscopy inject chromatograph of liquid, measure, promptly;
[4.3.5] result is with ginsenoside Rb 1The peak is set at the object of reference peak, and with the logarithm of its peak area as 1.000, according to ginsenoside Rb 1The retention time at peak is calculated, calibrate 10 total peaks altogether, its relative retention time is respectively: 0.441 ± 10%, 0.632 ± 10%, 0.942 ± 10%, 1.000,1.042 ± 10%, 1.062 ± 10%, 1.089 ± 10%, 1.188 ± 10%, 1.701 ± 10%, 1.737 ± 10%; And the regulation relative retention time is that the relative peak area at 0.441 ± 10%, 0.632 ± 10%, 1.042 ± 10%, 1.089 ± 10% peak is respectively 0.688～1.148,0.673～1.121,0.692～1.154,0.688～1.146 than (peak area logarithm ratio);
[4.4] fingerprint of alkaloid
[4.4.1] chromatographic condition chromatographic column: ZORBAX SB C 18(post of Φ 4.6mm * 150mm), packing material size 5 μ m; Methanol and 0.1% phosphoric acid-0.04% triethylamine aqueous solution are mobile phase, and gradient condition sees the following form:
Gradient condition Time (min) Methanol (%) 0.1% phosphoric acid-0.04% triethylamine aqueous solution (%) 0 10 45 60 30 30 42 42 70 70 58 58
30 ℃ of column temperatures; Detect wavelength: 235nm; Analysis time: 60min; Number of theoretical plate calculates with the mesaconitine peak should be not less than 2000;
The preparation of [4.4.2] inner mark solution is learnt from else's experience the phosphorus pentoxide drying under reduced pressure to the mesaconitine reference substance 5mg of constant weight, accurately claims surely, puts in the 25ml measuring bottle, adds 1% phosphoric acid solution 3ml, is diluted to scale with methanol, shakes up, promptly;
The preparation precision of [4.4.3] need testing solution is measured the injection content 1.0g for preparing, be dissolved in water to 10ml, put in the separatory funnel, add ammonia solution adjust pH to 10, use the mixed liquor jolting of the E-C (3: 1) of equivalent to extract 5 times, combining extraction liquid, low temperature volatilizes, and residue adds methanol 1ml makes dissolving, adds inner mark solution 50 μ l, filter, as need testing solution;
Accurate respectively inner mark solution 20 μ l and the need testing solution 40 μ l of drawing of [4.4.4] algoscopy inject chromatograph of liquid, measure, promptly;
[4.4.5] result is set at the mesaconitine peak at the object of reference peak of relative retention time, retention time according to the mesaconitine peak is calculated, calibrate 5 total peaks altogether, its relative retention time is respectively: 0.259 ± 10%, 0.342 ± 10%, 0.532 ± 10%, 0.724 ± 10%, 1.000 ± 10%; And with the stable and the biggest relative retention time of peak area be the peak area at 0.532 ± 10% peak as 1.000, and the regulation relative retention time is that the relative peak area ratio at 0.724 ± 10% peak is 0.455～0.683.
CN200610103234A 2006-07-14 2006-07-14 Detection method for injection containing ginseng and aconite root in raw material Active CN1899400B (en)

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CN102058643A (en) * 2009-11-17 2011-05-18 天津天士力制药股份有限公司 Ginseng saponin H extract and preparation method thereof
CN102495012A (en) * 2011-12-05 2012-06-13 中国水产科学研究院黄海水产研究所 Method for identifying sugar-doped dried sea cucumber
CN102707008A (en) * 2012-06-15 2012-10-03 湖南麓山天然植物制药有限公司 Detection method of gynostemma total saponin granules
CN104083462A (en) * 2014-07-28 2014-10-08 成都中医药大学附属医院 Pharmaceutical composition for treating hepatic failure as well as preparation method and use thereof
CN104083463A (en) * 2014-07-28 2014-10-08 成都中医药大学附属医院 New application of medicine composition
CN104849369A (en) * 2015-05-13 2015-08-19 济南康众医药科技开发有限公司 Quality detection method of ephedra sinica-aconitum napellus-liquorice medicine
CN106705574A (en) * 2015-11-17 2017-05-24 上海东富龙科技股份有限公司 Solid preparation fabrication equipment and method
CN107677528A (en) * 2017-09-20 2018-02-09 上海贞元诊断用品科技有限公司 Calibration object and quality-control product of a kind of anticoagulant heparin detection reagent and preparation method thereof
US10220064B2 (en) * 2016-11-08 2019-03-05 Macau University Of Science And Technology Carbohydrate composition extracted from Panax ginseng and its use in the treatment of ischemic conditions
CN110907580A (en) * 2019-12-20 2020-03-24 东阿阿胶股份有限公司 Detection method of HPLC (high performance liquid chromatography) characteristic spectrum of Baoyuan decoction
CN111257254A (en) * 2020-03-19 2020-06-09 中国水产科学研究院黄海水产研究所 Method for measuring glycogen content in oyster tissue
CN111366672A (en) * 2020-04-24 2020-07-03 劲牌有限公司 Detection method of health wine fingerprint
CN113030359A (en) * 2021-01-28 2021-06-25 成都第一制药有限公司 Detection method for various index components in motherwort injection and quality control method of motherwort injection
CN115710318A (en) * 2022-11-22 2023-02-24 安徽山河药用辅料股份有限公司 Preparation method of high-viscosity sodium carboxymethylcellulose for injection

Cited By (18)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102058643A (en) * 2009-11-17 2011-05-18 天津天士力制药股份有限公司 Ginseng saponin H extract and preparation method thereof
CN102058643B (en) * 2009-11-17 2013-09-11 天士力制药集团股份有限公司 Ginseng saponin H extract and preparation method thereof
CN102495012A (en) * 2011-12-05 2012-06-13 中国水产科学研究院黄海水产研究所 Method for identifying sugar-doped dried sea cucumber
CN102707008A (en) * 2012-06-15 2012-10-03 湖南麓山天然植物制药有限公司 Detection method of gynostemma total saponin granules
CN104083463B (en) * 2014-07-28 2017-02-15 成都中医药大学附属医院 Application of medicine composition for preparation of alcohol addiction or/and dependence drugs
CN104083463A (en) * 2014-07-28 2014-10-08 成都中医药大学附属医院 New application of medicine composition
CN104083462B (en) * 2014-07-28 2017-02-01 成都中医药大学附属医院 Pharmaceutical composition for treating hepatic failure as well as preparation method and use thereof
CN104083462A (en) * 2014-07-28 2014-10-08 成都中医药大学附属医院 Pharmaceutical composition for treating hepatic failure as well as preparation method and use thereof
CN104849369A (en) * 2015-05-13 2015-08-19 济南康众医药科技开发有限公司 Quality detection method of ephedra sinica-aconitum napellus-liquorice medicine
CN106705574A (en) * 2015-11-17 2017-05-24 上海东富龙科技股份有限公司 Solid preparation fabrication equipment and method
US10220064B2 (en) * 2016-11-08 2019-03-05 Macau University Of Science And Technology Carbohydrate composition extracted from Panax ginseng and its use in the treatment of ischemic conditions
CN107677528A (en) * 2017-09-20 2018-02-09 上海贞元诊断用品科技有限公司 Calibration object and quality-control product of a kind of anticoagulant heparin detection reagent and preparation method thereof
CN110907580A (en) * 2019-12-20 2020-03-24 东阿阿胶股份有限公司 Detection method of HPLC (high performance liquid chromatography) characteristic spectrum of Baoyuan decoction
CN111257254A (en) * 2020-03-19 2020-06-09 中国水产科学研究院黄海水产研究所 Method for measuring glycogen content in oyster tissue
CN111366672A (en) * 2020-04-24 2020-07-03 劲牌有限公司 Detection method of health wine fingerprint
CN113030359A (en) * 2021-01-28 2021-06-25 成都第一制药有限公司 Detection method for various index components in motherwort injection and quality control method of motherwort injection
CN115710318A (en) * 2022-11-22 2023-02-24 安徽山河药用辅料股份有限公司 Preparation method of high-viscosity sodium carboxymethylcellulose for injection
CN115710318B (en) * 2022-11-22 2024-02-02 安徽山河药用辅料股份有限公司 Preparation method of high-viscosity sodium carboxymethylcellulose for injection

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