CN100512849C - Use of extracted total timosaponin for producing myocardial ischemia treating products - Google Patents

Abstract

The present invention relates to the new use of total timosaponin as one Chinese medicine extract. The extract has timosaponin B-II compound as main component in the content of 30-70 % and is obtained through conventional extraction, separation and purification process. The extract has powerful pharmaceutical effect of diagnosing and treating myocardial ischemia and relevant diseases. It has simple preparation process, high yield, stable chemical property and safety and use wide application range.

Description

Extraced total timosaponin is used to prepare the purposes of myocardial ischemia treating products

Technical field

The present invention relates to medicine, Food ﹠ Drink technical field, specifically relate to the purposes of Chinese medicine extract, more particularly relate to the purposes that extraced total timosaponin is used to prepare myocardial ischemia treating products.

Background technology

(1) research overview of the Rhizoma Anemarrhenae

The Rhizoma Anemarrhenae (Rhizoma Anemarrhenae) is a liliaceous plant Rhizoma Anemarrhenae Anemarrhena asphodeloides Bunge rhizome, and another name garlic clove grass, even female is born in mountain region, dry hills or area, grassland, main product Hebei.Annual spring, autumn excavate, and remove fibrous root and earth, dry, and practise claiming " RHIZOMA ANEMARRHENAE with peet "; Remove crust, dry title " unpeeled RHIZOMA ANEMARRHENAE ".

The Rhizoma Anemarrhenae is a perennial herb.Phyllopodium gives birth to, and linear, matter is hard slightly, long 20～70cm, wide 0.3～0.6cm.High 40～the 60cm of scape, scattered flakey squamella; Spend on 2～6 cluster roominess rachises tapel 6, yellow or violet; Stamen 3, with interior wheel tapel to life.Capsule is long avette, tool 6 vertical ribs.5～August of florescence, fruit phase 8～JIUYUE.

The Rhizoma Anemarrhenae is cold in nature, bitter in the mouth, sweet, and the effect that has nourishing YIN to lower pathogenic fire, moisturizes laxation, clearing away heat-fire promotes the production of body fluid and moisturizes; Be used for traditionally that fever caused by exogenous pathogenic factors, lung-heat type cough, osteopyrexia and fever, interior-heat are quenched one's thirst, dryness of the intestine, constipation.Consumption 6～12g.

The chemical constituent of the Rhizoma Anemarrhenae is mainly 1-timosaponin A-1-I, 1-timosaponin A-1-II, 1-timosaponin A-1-III, 1-timosaponin A-1-IV, timosaponin B-I, timosaponin B-II, timosaponin H ₁, timosaponin H ₂, timosaponin I ₁, timosaponin I ₂Deng, and contain chimonin, Isomangiferin, choline, nicotinic acid, pantothenic acid, polysaccharide and become to grade.Through the research of Chinese scholars, timosaponin is the main active in the Rhizoma Anemarrhenae.

The pharmacological action document of timosaponin constituents has following report:

1, timosaponin unit (is called for short: ZMS), timosaponin B learning and memory function (Pharmacology and Clinics of Chinese Materia Medica, 1995 that can significantly improve old rats; 11 (3): 18-21; Modern combination of Chinese and Western medicine magazine, 2000; 9 (18): 1755-6; China's gerontology magazine, 21 (5): 379-80; Chinese invention patent application " the steroid saponins and the derivant thereof that are used for the treatment of Alzheimer ", enters national November 14 2000 date, application number 99806158.1, publication number CN 1301166 at on 03 26th, 1999 applying date).

Chinese invention patent application " the steroid saponins and the derivant thereof that are used for the treatment of Alzheimer " discloses many saponin and sapogenin, particularly the saponin of those steroid class formations and the sapogenin purposes in the not normal and similar disease of treatment cognitive function also discloses Therapeutic Method and pharmaceutical composition.

Steroidal saponin has treatment alzheimer disease effect (Chinese invention patent application " steroidal saponin is prevented and treated the purposes of alzheimer disease and new steroidal saponin ", on 09 26th, 1997 applying date, application number 97119680.X, publication number CN1212966) in the Rhizoma Anemarrhenae.

Chinese invention patent application " steroidal saponin is prevented and treated the purposes of alzheimer disease and new steroidal saponin " discloses the purposes that formula I steroid saponin compound is prevented and treated alzheimer disease, new formula II steroidal saponin and contain the pharmaceutical composition of described formula I steroidal saponin.

2, timosaponin D has the effect of the myocardium I/R of protection damage, this effect may with mechanism relevant (Capital Medical College's journal, 1994 such as its anti-PAF, antiplatelet aggregation and removing free radical; 15 (2): 138).

Timosaponin I ₁, timosaponin B-I, timosaponin B-II, timosaponin B-III, 1-timosaponin A-1-III have significant anti human platelet aggregation and prolong activity (Clin Chim Acta, 1999 of clotting time; 289 (1-2): 79-88).

3, Rhizoma Anemarrhenae total steroidal saponin has effect (the Chinese invention patent application " new medical use of total steroidal saponin and preparation method thereof in the Chinese medicine Rhizoma Anemarrhenae " of estrogen-like function of resisting osteoporosis, antiinflammatory, the generation of inhibition neutrophilic granulocyte peroxide, 02 month 26 days calendar year 2001 of the applying date, application number 01106195.2, publication number CN1370537).

Chinese invention patent application " new medical use of total steroidal saponin and preparation method thereof in the Chinese medicine Rhizoma Anemarrhenae " provides new medical use of total steroidal saponin in the Chinese medicine Rhizoma Anemarrhenae and preparation method thereof, proposed Rhizoma Anemarrhenae total steroidal saponin and had estrogen sample function of resisting osteoporosis, anti-inflammatory activity and have the effect that suppresses the generation of neutrophilic granulocyte peroxide, the utilization that can be Rhizoma Anemarrhenae resource provides new approach.Rhizoma ane marrhenae is through soaking and decoct, and again through liquid concentration, is to end at 1～3: 1 until the ratio of its weight and described rhizoma ane marrhenae weight behind the residue obtained liquid, and then absorb-elute, decolouring, decontamination, after reclaiming drying finished product.Process route cost of the present invention is low, and content of beary metal and persticide residue are little, the production process environmentally safe.

4, extraced total timosaponin is used to prepare purposes (the Chinese invention patent application " Rhizoma Anemarrhenae total saponins is used to prepare the purposes of control apoplexy medicine " of treatment apoplexy medicine, at on 05 09th, 2003 applying date, application number 03116824.8, publication number CN1451384A).

Chinese invention patent application " Rhizoma Anemarrhenae total saponins is used to prepare the purposes of control apoplexy medicine " discloses the purposes that extraced total timosaponin is used for preparation control apoplexy (that is: apoplexy) medicine or food.The disclosure according to the document, through animal experiment, extraced total timosaponin can obviously improve behavior symptom that cerebral ischemia causes, dwindles cerebral infarct volume, reduce the rats with cerebral ischemia cerebral edema.And think the purposes of having expanded extraced total timosaponin, for the control apoplexy provides a kind of new medicament sources.

(2) extraction separation method of Chinese herbal medicine effective ingredients commonly used

1, solvent extraction method

(1) principle: solvent extraction method is according to the dissolution properties of various compositions in solvent in the Chinese herbal medicine, select for use the active component dissolubility big, to not needing the little solvent of stripping composition dissolubility, and the method that effective ingredient is dissolved out in the medical material tissue.When solvent is added in the herbal raw material (needing suitably to pulverize), solvent is owing to diffusion, osmosis penetrate in the cell by cell wall gradually, dissolved solable matter, and cause concentration difference inside and outside the cell, so intracellular concentrated solution is constantly to external diffusion, solvent constantly enters in the medical material histiocyte again, so repeatedly come and go, when solution concentration reaches dynamic equilibrium inside and outside cell, this saturated solution is leached, continue repeatedly to add novel solvent, just can be bordering on complete stripping or the stripping of big portion to desirable ingredients.The dissolubility of medicinal herb components in solvent is directly relevant with solvent property.Solvent can be divided into hydrophilic organic solvent and lipotropy organic solvent, and dissolved material also has hydrophilic and lipophilic difference.Hydrophilic radical is many in the organic compound molecule structure, and its polarity is negligent of oil greatly; The hydrophilic radical that has is few, and its polarity is little and be negligent of water.The character of each kind solvent, equally also relevant with its molecular structure.Like this, the inventor just can remove to estimate their this type of character and the solvent of selecting for use by to the medicinal herb components structural analysis.Generally speaking,, bigger dissolubility will be arranged therein, i.e. the rule of so-called " similar mixing " as long as this character of the hydrophilic of medicinal herb components and lipotropy and solvent is suitable.This is to select appropriate solvent to extract one of foundation of required composition in Chinese herbal medicine.

(2) choice of Solvent: the key of utilization solvent extraction method is to select appropriate solvent.Solvent is selected suitably, just can be more successfully the composition of needs be extracted.Selective solvent will be noted following 3 points: 1. solvent is big to the effective ingredient dissolubility, and is little to the impurity dissolubility; 2. solvent can not play chemical change with the composition of Chinese medicine; 3. solvent want economical, be easy to get, safe in utilization etc.Common extraction solvent can be divided into following three classes:

1. water: water is a kind of strong polar solvent.Hydrophilic composition in the Chinese herbal medicine can both be gone out by water-soluble as the not too big polysaccharide of inorganic salt, saccharide, molecule, tannin, aminoacid, protein, acylate, alkaloid salt and glycoside etc.In order to increase the dissolubility of some composition, also often adopt sour water and aqueous alkali as extracting solvent.

Ethanol), methanol (but also claims: another name for), acetone etc., the most frequently used with ethanol 2. hydrophilic organic solvent: just general said and the miscible organic solvent of water (claim not only: as ethanol.Alcoholic acid solubility property is relatively good, and is stronger to the penetration capacity of Chinese herbal medicine cell.Outside hydrophilic composition isolating protein, phlegmatic temperament, pectin, starch and the part polysaccharide etc., big multipotency dissolves in ethanol.Be insoluble in the low-polarity component of water, the dissolubility in ethanol is also bigger.Can also adopt Different concentrations of alcohol to extract according to the character that is extracted material.More less than water consumption with ethanol extraction, extraction time is short, and it is also few to dissolve the water-solubility impurity that.Ethanol is organic solvent, though inflammable, toxicity is little, low price, and convenient sources has a locking equipment can reclaim repeatedly and use, and alcoholic acid extracting solution is difficult for moldy metamorphism.Owing to these reasons, be one of always the most frequently used method with the method for ethanol extraction.The character of methanol is similar with ethanol, boiling point lower (64 ℃), but toxic, should note during use.

3. lipophilic organic solvent: the organic solvent that just general said and water can not be miscible, as petroleum ether, benzene, chloroform, ether, ethyl acetate, dichloroethanes etc.These choice of Solvent performances are strong, can not or be not easy to propose hydrophilic impurities.But this kind solvent volatility is big, how inflammable (except the chloroform), generally poisonous, price is more expensive, equipment requirements is higher, and they penetrate plant tissue ability a little less than, often need to extract repeatedly for a long time and could extract fully.If contain more water in the medical material, just be difficult to leach its effective ingredient with this kind solvent, therefore, when extracting herbal raw material in a large number, directly using this kind solvent has certain limitation.

(3) extracting method: use the solvent extraction medicinal herb components, infusion process commonly used, percolation, decocting method, reflux extraction and continuous backflow extraction method etc.Simultaneously, factors such as the degree of grinding of raw material, extraction time, extraction temperature, appointed condition also can both influence extraction efficiency, must take in.

1. (be called for short: infusion process): the dipping genealogy of law is packed herbal powder or fragment in the proper container, adds The suitable solvent (as ethanol, rare alcohol or water), and the dipping medical material is with the stripping method of composition wherein to flood extraction method.This law is relatively simple, but leaching rate is relatively poor, and is solvent as water, and the easy moldy metamorphism of its extracting solution must note adding suitable preservatives.

2. (be called for short: percolation): percolation is that herbal powder is contained in the percolator to the seepage flow extraction method, constantly adds novel solvent, makes it penetrate medical material, flows out a kind of leaching method of leachate from top to bottom from the percolator bottom.When moving down when solvent infilters medicated powder, stripping composition proportion strengthens, its position is just replaced in the solution on upper strata or rare immersion, causes good concentration difference, and diffusion energy is carried out preferably, so leaching effect is better than infusion process.But should control flow velocity, in oozing transient, on powder, replenish novel solvent at any time, make till effective ingredient fully leaches in the medical material.Maybe extremely shallow or when oozing the volume that gushes liquid and being equivalent to heavy 10 times of crude drug when oozing the dropping liquid color, just can think and extract basically fully.The rare leachate that often will collect in mass production is as the usefulness of the solvent of another batch new raw material.

3. (be called for short: decocting method): decocting method is traditional leaching method that China uses the earliest to decoct extraction method.Used container is generally pottery, sand jar or copper, enamel ware, should not use iron pan, in order to avoid the medicinal liquid variable color.Preferably stir often during straight fire heating, in order to avoid that local medical material is heated is too high, burnt easily the paste.Big reaction pot, big copper pot, barrel are adopted in the pharmaceutical factory that steam-heating apparatus is arranged more, or feed Steam Heating in the pond of cement block.It is interconnection by pipeline also several can be decocted device, fries in shallow oil continuously and soaks.

4. heating and refluxing extraction method: use the organic solvent heating extraction, need to adopt the reflux device, in order to avoid the solvent evaporates loss.When operating in a small amount, can on round-bottomed flask, connect reflux condenser.The powder charge material is about 20%～60% of capacity in the bottle, and solvent soaked the about 1～2cm in medical material surface.Reflux in water-bath, the general maintenance, seethed with excitement 3～6 hours, puts cold filtration, and solubilizer in medicinal residues is made second and third time reflux and is made an appointment with half an hour respectively again, or to carrying till the most effective ingredient substantially.This method extraction efficiency adopts continuous extractions than the cold-maceration height more in the mass production.

5. continuous backflow extraction method: use volatile organic solvent and extract Chinese herbal medicine effective ingredients, no matter small test or large-scale production, all with continuous extraction for well, and need with quantity of solvent lessly, the extraction composition is also more complete.Laboratory fat-extraction device commonly used or title apparatus,Soxhlet's.Continuous extraction generally needs a few hours could extract fully.It is longer to extract the composition heated time, and the labile composition of case of thermal instability should not adopt this method.

2, separation and purification process:

Resulting extracts of Chinese herbal medicine of said extracted method or extract remain mixture, need further remove impurity, separate and make with extra care.

(1) solvent segregation: generally be with above-mentioned total extract, select three for use, the solvent of four kind of opposed polarity, by low polarity to high polarity proceed step by step extraction separation.Aqueous extract or ethanol extract often are jelly, be difficult to be dispersed in the low polar solvent, so can not extract fully, can admix an amount of inert filler, as kieselguhr or fiber powder etc., low temperature or natural drying then, after the pulverizing, to select for use solvent to extract successively, make each constituent in the total extract again, obtain according to the difference of its dissolubility in the opposed polarity solvent separating.Utilize the Chinese herbal medicine chemical constituent, the dissolubility in the opposed polarity solvent carries out separation and purification, is the most frequently used method.

(2) solvent extraction:

1. extraction: the solvent extraction extraction is called for short extraction again, is to utilize the difference of each composition partition coefficient in two kinds of immiscible solvents in the mixture and reach isolating method.If each composition partition coefficient in solvent differs big more during extraction, then separation efficiency is high more; If the effective ingredient in aqueous extract is lipophilic material, the general lipotropy organic solvent of using more, extract as benzene, chloroform or ether, if effective ingredient is to be partial to hydrophilic material, indissoluble is separated in lipophilic solvent, just need use weak lipophilic solvent, for example ethyl acetate, butanols etc. instead.Can also in chloroform, ether, add an amount of ethanol or methanol to increase its hydrophilic.When extracting flavones ingredient, how to extract with ethyl acetate and water.Then multiselect n-butyl alcohol, isoamyl alcohol and water extract to extract the strong saponin of hydrophilic.But, the common organic solvents hydrophilic is big more, and the effect of doing extraction with water is just bad more, and is because more hydrophilic impurities is followed, very big to the further refining influence of effective ingredient.

2. counter current continuous extraction method: be a kind of successive solvent extraction.Its device can have one, several or more extracting tube.Fill the contact surface during with increase solvent extraction in the pipe with little porcelain circle or little rustless steel wire ring.If a kind of infusion of Chinese herbal medicine need extract with the benzene lighter than water, ethyl acetate etc., then need water extracting liquid is contained in the extracting tube, and benzene, ethyl acetate are stored in the high-level container.Extract whether complete, but sample thief is analysed with thin layer chromatography, ply of paper and chromogenic reaction or precipitation are checked.

3. counter-current distribution method: counter-current distribution method claims CCD method, counter-current distribution or countercurrent distribution again.Counter-current distribution method is consistent with solvent counter-current extraction principle, but the application of sample amount is certain, and continuous in the solvent of a constant volume, reaches the separation of mixture through repeatedly being shifted the extraction distribution.

4. drop counter-current distribution method: the drop counter-current distribution method claims the droplet countercurrent chromatography method again.Be improved solvent extraction on the counter-current distribution method basis in recent years.To the same substantially counter-current distribution method of the selection of solvent system, but requirement can be separated at short notice, and can generate effective drop.Because mobile phase forms drop, contacting effectively with immobile phase in thin distribution extracting tube, rubbing constantly forms new surface, promotes the distribution of solute in solvent, so its separating effect is often good than counter-current distribution method.

(3) macroporous adsorbent resin method: macroporous adsorbent resin is a class organic polymer adsorbent that grows up the sixties in 20th century, have the good adsorption performance, be applied to the development of the extraction separation and the new Chinese medicine of Chinese herbal medicine chemical constituent surplus in the of nearly ten over year gradually.

Macroporous adsorbent resin is for absorption and screen the parting material that principle combines.Its adsorptivity is because the result of Van der Waals force or generation hydrogen bond.The screening principle is because itself cellular structure determines.Because absorption and screening principle, organic compound separates through certain solvent elution on macroporous adsorbent resin according to the difference of absorption affinity and the size of molecular weight.This make organic compound especially the purification of water soluble compound simplified greatly.The skeleton of macroporous adsorbent resin is generated by styrene and divinylbenzene polycondensation, because the adding of modifier, the polarity of macroporous adsorbent resin changes, and according to the surface nature of resin, that adsorbent resin generally is divided into is nonpolar, Semi-polarity and polarity three classes.

Nonpolar adsorption resin be by the very little monomer-polymer of dipole moment make not with the adsorbent resin of any functional group.Typical example is the adsorbent resin of styrene-divinylbenzene system, as D101, XAD-1, DiaionHP-10 macroporous adsorbent resin.

The Semi-polarity adsorbent resin refers to contain the adsorbent resin of ester group, as an acrylate or a crosslinked analog copolymer such as methacrylate and double methyl methacrylate.It is on the basis of nonpolar macroporous adsorption resin, adds acrylic acid methyl ester. or acrylonitrile polycondensation and forms, as the AB-8 macroporous adsorbent resin of the domestic frequent use of China.

Polar Adsorbent Resin is meant that amide-containing, itrile group, phenolic hydroxyl group etc. are nitrogenous, the adsorbent resin of oxygen, sulfur polar functionalities base.In addition, sometimes the ion exchange resin of ligand groups such as nitrogenous, oxygen, sulfur is called strong Polar Adsorbent Resin, the boundary of strong Polar Adsorbent Resin and ion exchange resin is difficult to difference.Polar macroporous adsorption resin can be formed by methyl methacrylate, acrylamide or the polycondensation of sulfoxide class, as the Diaion HP 2MG of Mitsubishi chemical industry, the XAD-10 of U.S. Rohm-hass company, XAD-9 macroporous adsorbent resin.

Compare with other adsorbent with active carbon, macroporous adsorbent resin has a lot of advantages, and is higher as the adsorptive selectivity to certain material; Physical and chemical stability and mechanical strength are better; Description is more, can change resin physics or chemical constitution as required; Adsorbent resin is generally spherical particle, and fluid resistance is less or the like.Thereby be widely used in chemical industry, medicine and other fields, more and more about the applied research report of macroporous adsorbent resin in natural product extraction is separated in recent years.Macroporous adsorbent resin centering herbal chemistry composition such as alkaloid, flavone, saponin, coumarin and some other glycoside compositions all have certain adsorption.Absorbability to sugar is very poor, and is stronger to the absorbability of pigment.

(4) sedimentation method: be in extracts of Chinese herbal medicine, to add some reagent to make the generation precipitation, with the method that obtains effective ingredient or remove impurity.As lead salt precipitation: lead salt precipitation is one of classical way of separating some medicinal herb components.Because lead acetate and Lead monosubacetate in water and alcoholic solution, can generate the lead salt or the complex salt precipitation of indissoluble with multiple medicinal herb components, so can utilize this character that effective ingredient is separated with impurity.Then lead salt precipitation is suspended in the novel solvent, passes to hydrogen sulfide gas, make and decompose and transfer insoluble vulcanized lead to and precipitate.

(5) salting out method: salting out method is in the water extract of Chinese herbal medicine, adds inorganic salt to finite concentration, or the state that reaches capacity, and can make the dissolubility of some composition in water reduce precipitation and separate out, and separate with the big impurity of water solublity.Be commonly used for the inorganic salt of saltouing sodium chloride, sodium sulfate, magnesium sulfate, ammonium sulfate etc. are arranged.

(6) dialysis: dialysis is to utilize small-molecule substance can pass through semipermeable membrane in solution, and macromolecular substances can not reach isolating method by the character of semipermeable membrane.Otherwise also macromolecular impurity can be stayed in the semipermeable membrane, and micromolecular material is entered in the outer solution of film by semipermeable membrane, and separation and purification in addition.

(7) crystallization, recrystallization and Steppecd crystallization: identify the Chinese herbal medicine chemical constituent, study its chemical constitution, must at first medicinal herb components be prepared into the pure product of monomer.At normal temperatures, the character of material own is the chemical compound of liquid, can carry out separation and purification with fractionating process or chromatography respectively.In general, the Chinese herbal medicine chemical constituent is solid material at normal temperatures mostly, all has the general character of crystalline solid, can reach the purpose of separation and purification according to the difference of dissolubility with crystallization process.

3, conventional drying method

(1) vacuum drying: be based on such ultimate principle: water saturation vapour pressure and temperature are closely related, under vacuum state, the boiling point lowering of water, i.e. operation operation at low temperatures just under vacuum, can avoid the destruction of nutritional labeling such as vitamin etc. at high temperature, improve rate of drying simultaneously.Vacuum drying is widely used in industries such as food, pharmacy, chemical industry, and China also develops and introduced various vacuum dryers, and its version is varied.Form commonly used mainly contains box type vacuum exsiccator, bipyramid formula vacuum desiccator, belt vacuum desiccator etc.These traditional Minton dryers mainly adopt heating such as hot blast, steam or electricity, utilize conduction of heat, convection current or radiation theory that heat is passed to material inside from the outside.It is low that vacuum drying has a baking temperature, and anoxia relatively in the hothouse can be avoided fat oxidation, and series of advantages such as pigment brown stain are suitable for the drying of heat sensitivity food material, and equipment cost, dry expense are also relatively low in addition.

(2) spray drying: be that fluidization technique is used for the exsiccant a kind of method of liquid material.Because of being wink-dry, be specially adapted to heat sensitive material, so the products obtained therefrom quality is good, keep original color, smell and taste, and easily dissolving.The research that utilizes spray drying to prepare microcapsule is carried out, it is that heart material is suspended in the solution of dress material, through centrifugal atomizer it is sprayed in the thermal current, the product of gained is the microcapsule that dress material bag heart material forms, this microcapsule powder can be used in direct compression, also can prepare capsule, syrup or suspensoid.

(3) lyophilization: be that the dry liquid material is frozen into solid, under the low-temperature reduced-pressure condition, utilize the distillation performance of icing, make the low-temperature material dehydration and reach exsiccant a kind of method.Because material is dry under high vacuum and cryogenic conditions, so the drying of some extremely thermo-labile article is well suited for.Wang Dalin has reported a kind of spraying ventilation lyophilizing new technique, be to utilize cold air or nitrogen as medium, the scars of flowing through rapidly make water sublimate, the product microgranule that makes of spraying lyophilizing is little, fast drying, time are short, evenly, good fluidity, and the good instant capacity of tool.In recent years, plaster material and the exsiccant research of sticky material have been obtained bigger progress, fluidization technology, spraying technique, inert carrier technology then grow up on this research basis.Rotatingandflashstreamingdrier, thermojet pneumatic drier, inert carrier drying machine all are fit to the drying of heat sensitive material and plaster material.These new achievements in research are used for Chinese medicine preparation production, with improving the technical merit of Chinese medicine processing greatly, enhance productivity.

(4) far infrared heating drying method: be a new dry technology, its drying principles is to change electric energy into far infrared radiation, thereby by the molecule absorption of medical material, produce resonance, cause the vibration and the rotation of molecule and atom, cause the object heating,, finally reach exsiccant purpose through thermal diffusion, evaporation and chemical change.Far-infrared ray drying can be saved electric energy 20%～50%, and effect is better.

(5) micro-wave drying method: be the new technique that develops rapidly a sixties in 20th century, microwave drying is actually by eddy-current heating and medium heating, make moisture and fat in the thing that is dried absorb microwave energy to some extent, thereby and it is changed into heat reach exsiccant purpose.But microwave drying killing microorganisms and mycete, and has disinfective action.The microwave heating installation of China's production at present has 915mhz and two frequencies of 2450mhz.

(3) research overview of myocardial ischemia

Myocardial ischemia is defined as relative oxygen and needs (or claim: oxygen consumes), and myocardial oxygen descends.That is to say that myocardial ischemia is meant that the hemoperfusion of heart reduces, and causes the oxygen supply of heart to reduce, energy metabolism of myocardial is undesired, can not support a kind of pathological state of heart operate as normal.The blood supply of heart is not unalterable, but exists fluctuation all the time, but this fluctuation impels the blood supply and demand constant relatively through body self adjusting, guarantees the heart operate as normal.If any reason causes myocardial ischemia, can not satisfy the heart working needs through the body adjusting, this has just constituted myocardial ischemia truly.

Lack enough blood flows and also be unfavorable for removing deleterious metabolite.In addition, the restoration of blood flow degree that may further increase the weight of to damage (is called for short: perfusion again).

The reason of heart ischemia: blood pressure reduction, aorta blood supply minimizing, coronary occlusion can directly cause heart blood supply to reduce; Valvulopathy, blood viscosity change, myocardium pathological changes own also can make heart blood supply reduce.Also have a kind of situation, heart blood supply does not reduce, but heart oxygen demand increased, and this is a kind of relative myocardial ischemia.

Clinical demonstration causes main, the modal cause of disease of myocardial ischemia, is coronary stricture.And the main cause of coronary stricture is an atherosclerosis.The heart disease that causes because of coronary atherosclerosis is exactly the coronary heart disease that the big daily life of a family is said, i.e. angina pectoris and acute myocardial infarction.So coronary heart disease is myocardial ischemia " arch-criminal ".

1, the metabolism of myocardial ischemia and physiological mechanism

Increasing laboratory and clinical research show that myocardial ischemia mainly is a kind of metabolic disease.Under the normal condition, fatty acid is the main energy-provision way of myocardium oxidative phosphorylation, and other substrate comprises glucose, aminoacid, acetone acid and lactic acid.Heart can absorb lactic acid (tremulous pulse lactic acid 20%) as its stand-by power source.Therefore, under the weary oxygen metabolism situation of ischemia, the lactic acid that is produced also is an important metabolic index of myocardial ischemia.Along with ischemia increases the weight of, intracellular ph value reduces, and the osmosis after the endocellular metabolism product is accumulated can cause the myocardial cell edema.

(1) acute myocardial ischemia: the contraction and the diastolic function that can influence heart.Diastolic dysfunction is often early than the variation of contractile function.Myocardial ischemia is relevant with the etiology of ischemia to the influence at once of ventricular compliance.Increase with ventricular compliance when oxygen begins for descending, and oxygen need increase and ventricular compliance significantly descends at once relevant (that is: ventricle becomes stiff).Ventricle need higher filling pressure (be called for short: LVEDP), to keep certain amount of fighting whenever.This moment patient may to show ventricular wall motion unusual, arrhythmia and conduction block.If coronary blood flow descends 80%, can cause that then ventricular systole is unable; Coronary blood flow descends 95%, the ventricle dyskinesis then occurs.When myocardial ischemia was serious, LVEDP raises can cause pulmonary edema.Thought in the past that ischemic myocardium can be irreversible damage and (be called for short: infarction) or immediately recover.

Think that at present ischemic myocardium also has other physiological pathway.Behind the transience severe cardiac myocardial ischemia, myocardium shrinkage function can recover (to be called for short: myocardial stunning) gradually.

(2) chronic myocardial ischemia: along with the degree of ischemia difference, myocardial cell shows a series of procedure of adaptation to ischemia during chronic myocardial ischemia, as myocardial stunning, hibernation, pre-ischemia and apoptosis.When acute ischemia increased the weight of, myocardial infarction appearred.These processes are overlapped, influence each other, and are influencing cardiac function in varying degrees.

Chronic severe ischemic can cause that the heart contraction work done descends as chronic ventricular wall motion unusual (be called for short: myocardial hibernation), hibernating myocardium is prevalent in reversible ischemic myocardial patient, also is found in acute coronary syndrome and serious left heart failure.Discover, 20%～60% ischemic myocardial patient cardiac function behind myocardial revascularization improves, the myocardial cell that ischemic area is described is not downright bad fully, but because hypoxic-ischemic is in adaptability low activity state, can recover for the function of these cells of back at recovery blood.Therefore, the detection of survival myocardium has crucial meaning to the assessment of patient's danger and the selection of Therapeutic Method, is the focus of Recent study.

(3) silent myocardial ischemia: though silent ischemia is meant the patient heart disease is arranged, asymptomatic and sign, and clinical examination has the evidence of myocardial ischemia, as the ST section change, heart muscle perfusion is damaged and ventricular wall motion is unusual etc.Usually after coronary occlusion takes place, following change promptly occurs: coronary artery blood flow reduces, local amount of blood supply and need the blood volume ratio that change take place, and CS-O2 reduces; Myocardium shrinkage function and diastolic function weaken; Left ventricular diastolic pressure raises; ECG changes, and for example the ST section is raised or forced down; Chest discomfort etc.But these unusually often take place within a short period of time, and the chest discomfort might appear in patient never.

That is to say that the silent myocardial ischemia is a kind of specific type of coronary heart disease, may reside among the various coronary heart disease types, but also individualism, and promptly there is the objective evidence of myocardial ischemia outbreak in the coronary disease patient, and is equal to symptom without angina pectoris or angina pectoris.

Relevant silent ischemia hypothesis: ischemic areas is less; Side Zhi Xunhuan is arranged; Ischemia Time is shorter; Angina pectoris warning system defectiveness; The patient denies; Pain threshold is more high.

The prognosis of silent ischemia is similar to angina pectoris, can cause acute myocardial infarction, so that sudden death, and because its asymptomatic Chang Buwei doctor and patient's attention.

2, the consequence of myocardial ischemia

Myocardial ischemia all may bring many adverse effects to heart and whole body.Myocardial ischemia can cause the significant change of cardiac function, and brings out a series of matters of aggravation, as myocardium infarction, arrhythmia, pulmonary edema, in addition dead.

Oxygen is the movable requisite material of myocardial cell, and oxygen is given cell by blood transport.Heart does not have " oxygen warehouse ", relies on myocardial blood flow fully, so in case ischemia can cause anoxia at once.Anoxybiotic direct result is that the myocardial cell aerobic metabolism weakens, and production capacity reduces, and energy essential when making cardiomotility is under-supply, causes that angina pectoris, arrhythmia, cardiac function descend.Simultaneously, metabolic refuse can not be removed effectively in time, easily has a negative impact.

Ischemia, anoxia, lack energy, finally can influence the contractile function of heart.Stop to shrink as if the cardiac muscle that has 20%～25%, left chamber function depletion can occur usually; If there is the cardiac muscle more than 40% not shrink, just have the depletion of severe cardiac pump function.If this situation takes place suddenly, breakneck cardiogenic shock will appear.Acute myocardial infarction is just normal relevant with this situation.

The myocardial ischemia chest pain is a kind of clinically very dangerous symptom, handles untimelyly to be in peril of one's life possibly.The crowd that easily sends out of myocardial ischemia chest pain: 1. elderly male or the women after climacteric; 2. hypertension, hyperlipemia (especially hypercholesterolemia), diabetes, smoking and overweight people are arranged; 3. family history person is arranged, as before the father 55 years old or mother coronary heart disease took place in 65 years old in the past, its children promptly belong to the people that familial history of coronary artery disease is arranged.Have the many more easy more morbidities of above risk factor.

Myocardial ischemia also can damage diastolic function.Shrink bad and diastole is bad combines, easily cause ventricular filling pressure to raise, cause pulmonary congestion, can cause that also complicated substance metabolism is disorderly and myocardial electrical activity is not normal.

Therefore,, cause of disease symptomatic treatment should be found accurately, just potential serious consequence can be avoided in case myocardial ischemia occurs.

People more and more recognize the serious consequence of myocardial ischemia at present.

Peri-operation period is handled and is concentrated on the application beta-blocker.The cause of disease that it is generally acknowledged many acute myocardial infarctions is broken relevant with coronary artery unstability atheromatous plaque.During beginning, plaque rupture causes ischemic event, causes coronary vasodilator thrombosis then.It is generally acknowledged that the calcium speckle that presents on the high electron beam ct is the high-risk object of plaque rupture.

3, the monitoring of myocardial ischemia

Myocardial ischemia needs the understanding of early diagnosis and treatment in time to be rooted in the hearts of the people.The method of clinical examination myocardial ischemia has: diastolic function, the myocardial metabolism of medical history, ECG (tranquillization, motion or dynamically ECG monitoring), Syst regional wall motion, cardiac muscle.

Research and propose, the rising of PCWP and characteristic wave mode thereof are the indexs of ischemia.But most researchs think that PAC is not a sensitivity index, should be as main monitoring method.

TEE is the high susceptibility index of monitoring myocardial ischemia.Myocardial ischemia shows as that new regional ventricular wall motion is unusual (to be called for short: RWMAs), systolic wall thickening reduces and ventricular dilatation during TEE.After extracorporeal circulation was shut down in the CABG operation, if RWMAs, then clinical consequences was bad.On the contrary, RWMAs before detected ischemia of ECG or the extracorporeal circulation and peri-operation period cardiac event incidence rate are irrelevant.Shortcoming or the problem of TEE comprise: the expense height; Can not obtain and insert the preceding variation of TEE; Real-time analysis TEE image can reduce precision of analysis in the art; TEE detects may disperse the attention of Anesthetist to more important vital sign; Whether the Anesthetist the qualified ultrasonic experiments of showing.

Also there are some scholars to use all multi index options and estimate myocardium keto consumption (abbreviation: MVO ₂) and myocardial ischemia, but its meaning of estimating the anesthesia patient still remains to be furtherd investigate.

4, the treatment of myocardial ischemia

(be called for short: ATP) supply with 60%～70% from the free fatty beta-oxidation, 20%～25% is glucose oxidase to the energy of normal myocardium, and 5%～10% separates for sugar alcohol.The free-fat acid oxidase produces the oxygen consumption height of the oxygen consumption of equivalent ATP than glucose oxidase, and high-level free-fat acid oxidase can obviously suppress the speed of glucose oxidase.Sympathetic activation during myocardial ischemia, catecholamine levels raises, and causes free fatty acid levels rising in the blood then; Free fatty acid oxidation increases to 80%～90% in the cardiac muscle energy resource supply; The coupling connection imbalance of glucose alcoholysis and glucose oxidase causes H in the myocardial cell ⁺, Ca ²⁺And Na ⁺Overload causes cell acidosis and infringement, and myocardium work efficiency is reduced, and power consumption simultaneously increases.Therefore, optimize energy metabolism of myocardial, especially suppress the free-fat acid oxidase, strengthen myocardial glucose metabolism and help alleviating the tissue injury that myocardial ischemia causes, improve myocardial function.

The traditional treatment of ischemic heart desease is based on anti symptom treatment, nitrate esters, epinephrine beta-blocker, calcium-channel antagonists etc. are by improving the hemodynamics of heart and whole body, playing the purpose that increases myocardium energy supply, reduces the cardiac muscle power consumption, is the main medicine of treatment myocardial ischemia.Along with further research and understanding, proposed by intervening metabolism link that cardiac energy produces, improving the new ideas that ischemic heart desease is treated in the mitochondrion energy metabolism to the myocardial ischemia metabolic process.

Trimetazidine (trimetazidine) is a kind of new drug that resists myocardial ischemia, and main by suppressing fatty acids metabolism, the increase glucose metabolism increases the cardiac energy supply.Trimetazidine can alleviate angina pectoris symptom, improves exercise tolerance, plays a role in the treatment of angina pectoris, ischemic cardiomyopathy etc.; To stable angina pectoris, trimetazidine can be used for the minor angina patient separately, reduces anginal attack times of patients with coronary heart disease and nitroglycerin consumption, improves exercise tolerance; Centering, severe angina pectoris, trimetazidine can be used as ancillary drug, merges with other traditional medicaments for resisting myocardial ischemia and uses.Singly use beta-blocker, calcium-channel antagonists, when nitrate control angina pectoris is not good, share trimetazidine and can obviously improve curative effect.To acute myocardial infarction, trimetazidine may alleviate the reperfusion injury behind the acute anterior myocardial infarction patient thrombolytic, reduces infarct size, influences remodeling ventricle behind the acute myocardial infarction.

At present mainly contain three aspects for the research that improves hibernating myocardium: the one, actively carry out myocardial revascularization, its curative effect has obtained certainly.The 2nd, intervene myocardial metabolism.This type of medicine mainly contains: the steatolysis inhibitor, as: nicotinic acid and derivant, insulin etc.; Carnitine acyl transferase-1 (be called for short: inhibitor CPT-1), as: L-carnitine, piperazine Ke Xilin, etomoxir etc.; Fatty acid beta-oxidation inhibitor, as: trimetazidine, thunder Lip river piperazine etc.The 3rd, cell transplantation currently also rests on the preclinical experimental stage.

At present, more existing small-scale clinical trials are observed the curative effect of medicine under different clinical settings of intervening myocardial metabolism.DIGAMI research is divided into two groups at random with the patient that 620 routine diabetes merge acute myocardial infarction, on the conventional therapy basis, one group gives insulin and glucose treatment, another group gives placebo, observe after 12 months and find, compare with matched group, the relative risk of insulin group death reduces by 29% (P=0.027).Diaz etc. also find to utilize the GIK treatment can significantly reduce the case fatality rate of patient behind the AMI in 3 months, but scale that should research is also smaller.

Some medicines can reach the purpose of myocardial preservation by the function of improving hibernating myocardium, as carvedilol.CHRIST-MAS finds that carvedilol was treated stable heart failure patient 4 months, can obviously improve the survival myocardium function, improves ejection fraction.In addition; studies show that in a large number; ACEI, that ARB class medicine is used for acute myocardial infarction is early stage, also can significantly improve the incidence rate of patient's be in hospital case fatality rate and cardiovascular event, and one of mechanism of action is to bring into play myocardium protecting action by the blocking-up renin-angiotensin system.

Statins may be by stabilize plaque, protect endothelium, improve fibrinolytic-blood coagulation system function performance myocardium protecting action.Clinical research is same finds that using Statins in early days at acute coronary syndrome can significantly improve patient's prognosis.About the statins myocardial preservation really cutter system, whether have the stable and organelle protective effect of film and be still waiting further research.

By literature search etc., up to the present, the extraced total timosaponin that still finds no timosaponin B-II and be main component is at the report that is used to prevent, diagnose, detect, protect, treat and study aspect myocardial ischemia and the relevant disease thereof.

Summary of the invention

The technical problem that will solve required for the present invention is the new purposes that discloses a kind of Chinese medicine extraced total timosaponin, to overcome the above-mentioned defective that prior art exists.

That is to say, the invention is intended to the activity of clear and definite extraced total timosaponin, and then extraced total timosaponin is used to prepare myocardial ischemia treating products in the myocardial ischemia application facet; Described myocardial ischemia treating products is meant the product that is used to prevent, diagnose, detect, protect, treat and study myocardial ischemia and relevant disease thereof, is to comprise in medicine, reagent, the food etc. one or more, preferred agents.

(1) technical conceive

The independent development original new drug is a present urgent task of China, China's Chinese medicine and pharmacy has a long history, also accumulated rich experience with Chinese herbal medicine prevention and treatment disease aspect, therefore seeking effective active component from Chinese medicine is a valid approach, also is the place of the advantage of Chinese original new drug development.

The inventor screens and proves the activity and the purposes of this total saponin extracts by the total saponin extracts of single rhizoma ane marrhenae being carried out the chemical constitution study of system.

According to literature search, the inventor found through experiments the active site extraced total timosaponin and has the number of significant pharmacologically active.And, prove that the main component in the Rhizoma Anemarrhenae active site extraced total timosaponin is a timosaponin B-II chemical compound through chemical constitution study.Thereby the inventor infer the Rhizoma Anemarrhenae in active clinical drug effects in aspect such as prevention, diagnosis, protection and treatment myocardial ischemia; also should mainly be by the active site extraced total timosaponin particularly the drug effect of timosaponin B-II bring into play, result of study also prove and confirmed extraced total timosaponin particularly timosaponin B-II have significant pharmacologically active.

Cardiovascular disease has become the important death cause of Chinese population, development treatment cardiovascular disease medicine, especially prevent, diagnose, protect and treat aspect product such as myocardial ischemia particularly medicine have significant social, economic benefit.

(2) preparation of extraced total timosaponin and authentication method thereof

The preparation method of the said extraced total timosaponin of the present invention comprises the steps:

(1) extract: it is some to get Rhizoma Anemarrhenae raw material, extracts, and gets extracting solution, i.e. Rhizoma Anemarrhenae total extract;

(2) separation and purification:, promptly get extraced total timosaponin with the extracting solution separation and purification.

Described extracting method comprises all operable methods well known in the art such as solvent extraction method.

The extracting method of solvent extraction method mentioned above is the conventional extracting method of this area, that is to say to comprise ultrasonic extraction commonly used, dipping extraction method, percolation extraction method, decoct in extraction method, heating and refluxing extraction method or the continuous backflow extraction method etc. one or more; Extraction time can be once or repeatedly.Simultaneously, factors such as the degree of grinding of raw material, extraction time, extraction temperature, appointed condition also can both influence extraction efficiency, must take in.Various extracting method comprise all technical data of ins and outs, all can be referring to relevant teaching material and relevant technical literature etc.

The routine that the employed extraction solvent of solvent extraction method mentioned above is this area is extracted solvent, that is to say one or more of extraction such as three classes that comprise common water, hydrophilic organic solvent or lipophilic organic solvent solvent;

Described water is to comprise a kind of in water, sour water or the aqueous alkali etc.;

Described hydrophilic organic solvent is general said and the miscible organic solvent of water, comprise in ethanol, ethanol water, methanol or the acetone etc. one or more, in preferred alcohol or the Different concentrations of alcohol aqueous solution etc. one or more, further preferred Different concentrations of alcohol aqueous solution;

Described lipophilic organic solvent is the organic solvent that generally said and water can not be miscible, comprise in petroleum ether, benzene, chloroform, ether, ethyl acetate, dichloromethane or the dichloroethanes etc. one or more, in preferred petroleum ether, chloroform, ether, ethyl acetate, dichloromethane or the dichloroethanes etc. one or more, one or more in further preferred chloroform, ether, ethyl acetate or the dichloromethane etc.

The resulting Rhizoma Anemarrhenae total extract of said extracted method needs further separation and purification.

Described isolation and purification method is all operable methods well known in the art, comprise solvent segregation, solvent extraction (comprise in extraction, counter current continuous extraction method, counter-current distribution method or the drop counter-current distribution method etc. one or more), the macroporous adsorbent resin method, the sedimentation method, salting out method, one or more in column chromatography or crystallization and recrystallization and the Steppecd crystallization etc.; These methods all are known technologies of this area, at needs further in the conclusive evidence, are easy to find all technical data that comprise ins and outs from relevant teaching material and relevant technical literature etc.

In order to make product property stable, easy to use and preserve, also can increase the step of an optimization, i.e. step 3:

(3) drying.

Described drying means is an operable method well known in the art, comprises that atmosphere pressure desiccation is as in oven for drying, hypobaric drying method, boulton process, spray drying method, freeze-drying, far infrared heating drying method or micro-wave drying method etc. one or more.

In the specific implementation, need adopt wherein suitable method, and choose measures necessary such as appropriate condition to reach the set goal according to existing fund, technology and relevant requirement.And these methods also all are known technologies of this area, at needs further in the conclusive evidence, all are to be easy to find all technical data that comprise ins and outs from relevant teaching material and relevant technical literature etc.

For example, the preparation method of the said extraced total timosaponin of the present invention can for:

(1) extracts: routinely rhizoma ane marrhenae is ground into coarse powder,, collects 5～30 times of medical material volume percolates, be extracting solution with 10～95% ethanol percolate extraction.

(2) divide high purification: it is 0.5～10:1 (medical material weight: liquor capacity), be sample liquid that extracting solution is concentrated to concentration.(nonpolar or low pole macroporous resin is for being the polystyrene type porous adsorbent resin of bridging materials with styrene by nonpolar or low pole macroporous resin adsorption with sample liquid, as D101, D201, ZTC-1, AB-8,1300-1 type macroporous resin etc.), remove impurity with water wash; The moisture lower alcohol eluting of reuse, lower alcohol is C such as methanol, ethanol or propanol ₁～C ₅Alcohols, its concentration are 10～40%, and volumetric usage is 5～24 times of resin bed volume, collect the lower alcohol eluent, are concentrated into driedly, and pulverize; Powder is with 1.5～2.5 times of amounts (acetone volume/powder weight) acetone solution, add 2～12 times of amounts (acetone volume/powder weight) acetone again, filter, filtrate is reclaimed solvent to 2～8 times amount (acetone volume/powder weight), leave standstill, separate out crystallization, filter, with the washing with acetone crystallization, promptly get extraced total timosaponin of the present invention.

Integrated application mass spectrum of the present invention (be called for short: MS), proton nmr spectra (be called for short: ¹H-NMR), carbon-13 nmr spectra (be called for short: ¹³C-NMR) and the nuclear magnetic resonance, NMR two-dimensional spectrum (be called for short: 2D-NMR) etc. the analysis technology has been carried out the structure evaluation to the timosaponin B-II in the extraced total timosaponin, and its content range is 30%～70%.

(3) pharmacologically active of extraced total timosaponin

The present invention has carried out many-sided test to extraced total timosaponin in the activity of the aspects such as prevention, diagnosis, protection, treatment and research of myocardial ischemia.

For test agent all is to take above preparation method to obtain, specific as follows:

(1) containing total saponins in the extraced total timosaponin is 40%～80%;

(2) extraced total timosaponin main component timosaponin B-II content is 30%～70%;

The in vitro tests of the prevention of extraced total timosaponin myocardial ischemia, diagnosis, protection and therapeutic activity

1, the dog coronary artery ligation is caused the therapeutical effect of myocardial infarction model

Adopt the ligation of anesthetized dog left anterior descending coronary artery to cause myocardial infarction model, observe the therapeutical effect of extraced total timosaponin.Experimental result shows, extraced total timosaponin 20mg/kg, 60mg/kg, 120mg/kg gastric infusion can significantly reduce the degree of myocardial ischemia behind the dog coronary ligation, the scope of dwindling myocardial ischemia, and action intensity and dosage have certain relation; It is consistent with the epicardial electrogram measurement result that quantitative tissue is learned testing result, compares with the normal saline matched group, and extraced total timosaponin can significantly reduce infarction size.Extraced total timosaponin can also significantly reduce the degree that serum lactic acid and creatine kinase raise behind the dog coronary ligation; Primary cellular defect when pointing out it to alleviate myocardial ischemia has protective effect to myocardial cell.Experimental result shows that degree of myocardial ischemia and scope when extraced total timosaponin can significantly alleviate the dog myocardial infarction reduce infarction size.

2, to the influence of anesthetized dog myocardial oxygen consumption

Experiment is intended to observe the influence of extraced total timosaponin gastric infusion to the anesthetized dog myocardial oxygen consumption.The result shows: myocardial oxygen consumption is compared no significant difference with physiology saline group behind the extraced total timosaponin low dose group gastric infusion, and middle and high dosage group can significantly reduce the anesthetized dog myocardial oxygen consumption; Three dosage groups all can obviously reduce myocardium oxygen uptake rate, increase coronary flow, reduce coronary resistance, and action intensity and dosage have certain relation.Extraced total timosaponin high dose group and positive control drug DIAOXINXUE KANG JIAONANG can significantly reduce total peripheral resistance, and each dosage group does not have obvious influence to the dog cardiac output.

3, the protective effect that rat myocardial ischemia and reperfusion is damaged

Laboratory observation the influence of extraced total timosaponin gastric infusion to myocardial damage due to the SD rat heart ischemia-reperfusion.Behind 3% pentobarbital sodium intraperitoneal anesthesia SD rat, left side the 4-5 intercostal is opened breast, and the ligation left anterior descending coronary artery is made the acute cardiac ischemia-reperfusion injury model.Irritate stomach and give extraced total timosaponin, measure serum creatine kinase behind the acute myocardial ischemia 60min reperfusion injury 120min and (be called for short: CK), lactic acid dehydrogenase (is called for short: LDH) activity and myocardial infarct size.The result shows: extraced total timosaponin low dose group (60mg/kg) Serum LDH, CK activity, myocardial infarction area/cardiac weight percentage ratio are starkly lower than model group (P＜0.05); (120,360mg/kg) Serum LDH, CK activity, myocardial infarction area/cardiac weight percentage ratio are starkly lower than model group (P＜0.01) to middle and high dosage group very much; DIAOXINXUE KANG JIAONANG group Serum LDH, CK activity, myocardial infarction area/cardiac weight percentage ratio highly significant are lower than model group (P＜0.01).The result shows that damage has protective effect to the extraced total timosaponin gastric infusion to rat myocardial ischemia and reperfusion.

(4) purposes of extraced total timosaponin

1, general introduction

The purpose of this invention is to provide a kind of product that is used to prevent, diagnose, detect, protect, treat and study myocardial ischemia and relevant disease thereof, comprise in medicine, reagent, the food etc. one or more, preferred agents.

Prove that by the pharmacologically active screening extraced total timosaponin that employing macroporous adsorbent resin 5～75% ethanol elution positions obtain prevents, diagnoses, detects, protects, treats and study the active site of myocardial ischemia and relevant disease thereof for it.

Show that through experimentation the extraced total timosaponin gastric infusion can significantly reduce the degree of myocardial ischemia behind the dog coronary ligation, reduces infarction size; Can significantly reduce the anesthetized dog myocardial oxygen consumption, increase coronary flow, reduce coronary resistance; Damage has protective effect to gastric infusion to rat myocardial ischemia and reperfusion.Completed acute toxicity testing proves, the mouse stomach administration surpasses 2.0g/kg to the maximum tolerated dose of this active site, be equivalent to 440 times of clinical recommended drug dosage, show that this effective site is safe and reliable, solved complicated component in the Chinese medicine compound, active constituent content is low and has contained the problem of toxic component.

In sum; the inventor has carried out theory study to extraced total timosaponin; through a large amount of particularly secular pharmacology tests of experimentation, find that the extraced total timosaponin of being addressed has the activity of significant prevention, diagnosis, detection, protection, treatment and research myocardial ischemia and relevant disease thereof.Therefore, extraced total timosaponin and compositions thereof can be used for preparing myocardial ischemia treating products, are the medicine that feedstock production forms with extraced total timosaponin of the present invention preferably.

2, the using method of extraced total timosaponin and compositions thereof and requirement

Extraced total timosaponin of the present invention can be united use separately or with other active component; comprise and be used to prepare the product that is used to prevent, diagnose, detect, protect, treat and study myocardial ischemia and relevant disease thereof; comprise medicine, reagent, food etc., especially medicine.

Aspect concrete use, extraced total timosaponin of the present invention can use separately, can also use with other many chemical substances.These chemical substances biologically active or have the function of treatment disease whether no matter, comprise miscellaneous function as collaborative amplification, antagonism or alleviate the side effect etc. of extraced total timosaponin, these chemical substances are to comprise in pharmaceutically acceptable carrier, food, natural product, chemical synthetic drug or the human medication etc. one or more; Preferably include in pharmaceutically acceptable carrier or the food etc. one or more; Further preferred pharmaceutically acceptable carrier.

" pharmaceutically acceptable carrier " used herein comprises one or more in any He all physiology suitable solvent, disperse medium, afterbirth, antibacterial and antifungal, isotonic agent or the absorption delay agent etc.The example of pharmaceutically acceptable carrier comprises one or more water, saline, phosphate-buffered saline, glucose, glycerol or ethanol etc. and in the compositions one or more thereof.In many cases, in said composition, preferably include isotonic agent, for example, sugar, such as in the polyhydric alcohol of mannitol, sorbitol, sorbitol or the sodium chloride etc. one or more.Pharmaceutically acceptable carrier can also comprise a spot of auxiliary substance, one or more in wetting agent or emulsifying agent, antiseptic or the buffer etc. for example, and they have strengthened the effect duration or the effectiveness of this extraced total timosaponin.

From concrete classification, said pharmaceutically acceptable carrier is meant the pharmaceutical carrier of medicine and pharmacology field routine, comprises excipient, as in starch or the water etc. one or more; Lubricant is as in glycerol or the magnesium stearate etc. one or more; Disintegrating agent is as microcrystalline Cellulose etc.; Filler is as in starch or the lactose etc. one or more; Bonding agent is as in pregelatinized Starch, dextrin, cellulose derivative, alginate, gelatin or the polyvinylpyrrolidone etc. one or more; Osmotic pressure regulator is as in glucose, sucrose, sorbitol or the mannitol etc. one or more; Diluent is as water etc.; Disintegrating agent is as in agar, calcium carbonate or the sodium bicarbonate etc. one or more; Absorption enhancer is as quaternary ammonium compound etc.; Surfactant is as hexadecanol etc.; Absorption carrier is as in Kaolin or the soap clay etc. one or more; Lubricant is as in Pulvis Talci, calcium stearate, magnesium stearate or the Polyethylene Glycol etc. one or more; In addition, can also in compositions, add other adjuvant, as in flavouring agent or the sweeting agent etc. one or more.

For example, the active component extraced total timosaponin is dissolved, suspendible or (for example be emulsifiable in the suitable aqueous solvent, distilled water, in normal saline or the Green's solution etc. one or more) or in the oil-based solvent (for example, vegetable oil is olive oil for example, Oleum sesami, Oleum Gossypii semen, in Semen Maydis oil or the propylene glycol etc. one or more) in, can make ejection preparation, wherein (for example can contain dispersant in the solvent, polyoxyethylene sorbitan monoleate, polyoxyethylene hardened castor oil 60, Polyethylene Glycol, benzyl alcohol, in chlorobutanol or the phenol etc. one or more), osmotic pressure regulator (for example, sodium chloride, glycerol, D9-mannose, in D-sorbitol or the glucose etc. one or more).In this case, if necessary, can add additive, for example solubilizing agent (for example, one or more in sodium salicylate or the sodium acetate etc.), stabilizing agent (for example, human serum albumin etc.), analgesic (for example, benzyl alcohol etc.) etc.

Of the present invention and extraced total timosaponin can also unite use with the form of compositions, particularly with other chemical substance such as medicine animal especially mammal is comprised that people or other animals treat compositions for use or similar compositions.Described mammal, comprise in people, mice, rat, sheep, monkey, cattle, pig, horse, rabbit, dog, chimpanzee, baboon, Adeps seu carnis Rhiopithecus roxellanae, macaque or the Rhesus Macacus etc. one or more, in preferred people, mice, rat, monkey, pig, rabbit or the dog etc. one or more, one or more in further preferred people, rat or the monkey etc.For example, extraced total timosaponin of the present invention can be added be suitable for to curee's Pharmaceutical composition in.Usually, this Pharmaceutical composition comprises extraced total timosaponin of the present invention and pharmaceutically acceptable carrier.

The compositions of extraced total timosaponin particularly pharmaceutical composition can have various forms, comprises in the dosage forms such as liquid, semisolid and solid for example one or more; Wherein said pharmaceutical composition comprises that the extraced total timosaponin for the treatment of effective dose is an active component, and one or more pharmaceutically acceptable carriers.

The pharmaceutical composition of extraced total timosaponin can adopt conventional production method well known in the art to make various dosage forms, and active component is mixed with one or more carriers, is made into required dosage form then.Described dosage form comprises one or more in tablet, capsule, granule, suspensoid, Emulsion, solution, syrup or the injection etc., takes one or more route of administration in oral or injection (comprise in intravenous injection, intravenous drip, intramuscular injection or the subcutaneous injection etc. one or more), the mucosa dialysis etc. to carry out prevention, diagnosis, detection, protection, treatment or the scientific research of myocardial ischemia and relevant disease thereof.

It is 0.5%～99% active component extraced total timosaponin that pharmaceutical composition preferably contains weight ratio, further preferably contain weight ratio and be 1%～95% active component extraced total timosaponin, most preferably contain weight ratio and be 5%～90% active component extraced total timosaponin.

The pharmaceutical composition of extraced total timosaponin generally must be aseptic and stable under the production condition of storage.Said composition can be mixed with solution, microemulsion, dispersion liquid, liposome or other is suitable for the ordered structure of high drug level.By with a kind of of this extraced total timosaponin of aequum and required mentioned component or combine to add in the appropriate solvent and then carry out aseptic filtration and prepare aseptic parenteral solution.Generally speaking, prepare dispersion liquid by this extraced total timosaponin being added in the aseptic solvent that contains basic disperse medium and required above-mentioned other composition.Under the situation of the sterile powder that is used to prepare aseptic parenteral solution, the preparation method of recommendation is vacuum drying and lyophilized preparation.For example, by passing through to keep required granular size such as the coating of lecithin, under the situation of dispersion liquid and, can keeping the adequate liquidity of solution by using surfactant.By comprising that in said composition the medicament (for example Monostearate or gelatin) that postpones to absorb can reach the prolongation absorption of injectable composition.

When being used for the patient, extraced total timosaponin dosage of the present invention is 5～20mg/kgd, and this dosage or consumption decide according to the age of patient or user and the situation of body weight and health or patient's symptom usually.

Extraced total timosaponin of the present invention and Pharmaceutical composition thereof can comprise the extraced total timosaponin of the present invention of " treatment effective dose " or " prevention effective dose "." treatment effective dose " is meant at the dosage of necessity and effectively reaches the amount of required therapeutic effect under the time.The treatment effective dose of extraced total timosaponin can cause that at this individuality the factors such as ability of required reaction change according to the patient's condition, age, sex and body weight and this extraced total timosaponin such as individuality.The treatment effective dose also refers to that the useful therapeutic effect of this extraced total timosaponin surpasses the amount of its any toxicity or harmful effect." prevention effective dose " is meant the amount that effectively reaches required preventive effect under necessary dosage and time.Because preventive dose is used for the ill preceding or early stage curee of disease, the prevention effective dose is usually less than the treatment effective dose.The typical non-limiting scope of the treatment of extraced total timosaponin of the present invention or prevention effective dose is 5～20mg/kg, more preferably 5～10mg/kg.Should note, dose value will change according to disease type of desiring to alleviate and seriousness, that is to say when being used for the patient that extraced total timosaponin dosage of the present invention or consumption decide according to the age of patient or user and the situation of body weight and health or patient's symptom usually.In addition; should understand; for any specific curee; should along with the time according to individual need and give with or supervision give with the people's of described compositions professional judgement and adjust the given dose system; and the dosage range that this paper sets only be illustrative, the scope or the practice of the compositions of can't requirement for restriction protecting.

That is to say, need be according to object, route of administration, institute's disease of treat and the situation etc. of treatment, variation extraced total timosaponin of the present invention at every turn and/or dosage or the consumption of every day.For example, give mammal through vein, adult (as body weight 60kg) especially, the single dose of described extraced total timosaponin is about 50～200mg, preferably about 100mg, preferred administration every day 1～3 time.Can adjust dosage unit, to propose the best required reaction of arch (for example, treatment or prevention are replied).For example, can single heavy dose of administration can give several divided doses or reduce or increase dosage in proportion according to the urgency of treatment situation in a period of time.The non-intestinal compositions that preparation is easy to the unified dosage unit form of administration and dosage is especially favourable.Dosage unit form used herein refers to be suitable for the physical separation unit of dosage unit of the mammalian subject of desire treatment; The calculating that each unit contains scheduled volume is used for together producing with required pharmaceutical carrier the active matter extraced total timosaponin of required therapeutic effect.The specification of dosage unit form of the present invention, determine and directly depend on the specific characteristic of following (a) this extraced total timosaponin and the particular treatment of desiring to reach or preventive effect and (b) interior in mixing this technology that is used for the treatment of the individual sensitivity extraced total timosaponin by following in restriction.

3, the pharmaceutical dosage form of extraced total timosaponin and compositions thereof and route of administration

The product that is used to prevent, diagnose, detect, protect, treat and study myocardial ischemia and relevant disease thereof of extraced total timosaponin of the present invention and preparation of compositions thereof, wherein the product according to the requirement of beverage, food technology field preparation can be used in prevention, protection and treats myocardial ischemia and relevant disease thereof; Can be used in patient's treatment or health care according to the product of the requirement of medical technical field preparation, can either be directly used in the medicine of preparation treatment or health care separately, also can mix with many chemical substances or make up, directly or indirectly be used to prepare the medicine of treatment or health care.Chemical substance described here is above described identical with this section.

In the present invention, required material comprises raw material of the present invention, above-mentioned matching used chemical substance etc., all should adopt the material of food stage or pharmaceutical grade according to practical situation and needs.

Extraced total timosaponin of the present invention and compositions thereof can be with the whole bag of tricks administration known in the art, although route of administration/administering mode of recommending in many therapeutic use is spray or oral administration.But the technical staff will appreciate that route of administration/administering mode changes with required result.In some concrete enforcement, the carrier that this reactive compound can avoid rapid release with this chemical compound of protection is preparation example such as empty release formulation together, comprises that graft transmission system, transdermal paste one or more in transmission system or the microcapsule transmission system etc.In addition, can also use biodegradable, biocompatible polymer, for example one or more in ethylene-ethyl acetate, polyanhydride, polyglycolic acid, collagen protein, polyorthoesters or the polylactic acid etc.Prepare the equal patent applied for of many methods of this preparation or generally known to those skilled in the art (referring to for example Sustained and Controlled Release Drug Delivery Systems, J.R.Robinson edits, Marcel Dekker, Inc., New York, 1978).

Extraced total timosaponin of the present invention and compositions thereof, usually by oral, snuffing is gone into, one or more modes in rectum or the parenteral etc., is applied to the patient who needs this treatment.

Be used for when oral, can be made into conventional solid preparation such as in tablet, powder, granule or the capsule etc. one or more.When implementing, extraced total timosaponin of the present invention can be together oral with for example inert diluent or assimilable edible carrier.This extraced total timosaponin (with its composition altogether, if desired) can also be wrapped in hard or soft shell gelatin capsules, is pressed into tablet or directly adds in curee's the meals.About oral therapeutic administration, described extraced total timosaponin can be added with excipient and use with one or more forms in edible tablet, buccal tablet agent, lozenge, capsule, suspension, syrup or wafer or the like.

For to give extraced total timosaponin of the present invention outside the parenterai administration, may need with preventing that the material of its inactivation from together giving to this extraced total timosaponin coating or with this extraced total timosaponin.The reactive compound that replenishes can also be added in the said composition.In the specific implementation, extraced total timosaponin of the present invention and one or more other medicines that can be used for the treatment of disease are prepared altogether and/or given altogether.Thisly unite use, can utilize this medicine that gives primely, therefore avoid possible toxicity or the complication relevant with various monotherapies than low dosage.

Make in liquid preparation such as water preparation, oil-suspending agent or other liquid preparation one or more, as in syrup, tincture or the elixir etc. one or more; When being used for parenteral, can be made in solution, water preparation or the oiliness suspending agent etc. of injection one or more.

In the above-described type of service, preferred form is one or more in tablet, coated tablet, capsule, suppository or the injection etc., further one or more in preferred tablet, capsule or the injection etc., optimizing injection especially.

In addition; the employed medicinal raw material of extraced total timosaponin also can be directly used in the product that preparation is used to prevent, diagnose, detect, protect, treat and study myocardial ischemia and relevant disease thereof in some cases separately; also can mix with many chemical substances or make up, directly or indirectly be used to prepare the product that is used to prevent, diagnose, detect, protect, treat and study myocardial ischemia and relevant disease thereof with the form of compositions.Chemical substance described here is above described identical with this section.

For example; the powder of the employed medicinal raw material of extraced total timosaponin is used for preparation and is used for the myocardial ischemia prevention; diagnosis; protection and treatment product be the various dosage forms of medicine particularly; or the powder of the employed medicinal raw material of extraced total timosaponin is used for preparation with relevant adjuvant and is used for prevention; diagnosis; detect; protection; the product of treatment and research myocardial ischemia and relevant disease thereof is the various dosage forms of medicine especially; or the powder of the employed medicinal raw material of extraced total timosaponin is used for prevention with relevant preparation; diagnosis; detect; protection; the product of treatment and research myocardial ischemia and relevant disease thereof such as medicine one are used from preparation and are used for prevention; diagnosis; detect; protection; treatment and research myocardial ischemia and the product of relevant disease such as the various dosage forms of medicine; or the powder of the employed medicinal raw material of extraced total timosaponin is used from preparation with relevant ancillary drug one and is used for prevention; diagnosis; detect; protection; treatment and research myocardial ischemia and the product of relevant disease such as the various dosage forms of medicine; as tablet; in capsule or the suspensoid etc. one or more, preferred capsule.

One of described method is that the powder fill with the employed medicinal raw material of extraced total timosaponin is a capsule, two of method be with the powder of the employed medicinal raw material of extraced total timosaponin and relevant preparation be used to prevent, diagnose, detect, protect, treat and study the product of myocardial ischemia and relevant disease thereof such as medicine together fill be capsule, three of method is to be capsule with the powder of the employed medicinal raw material of extraced total timosaponin with relevant ancillary drug fill together; Four of method is the powder of the employed medicinal raw material of extraced total timosaponin directly to be pressed together according to a conventional method with relevant adjuvant be tablet; five of method is the powder with the employed medicinal raw material of extraced total timosaponin; relevant preparation is used for prevention; diagnosis; detect; protection; the product of treatment and research myocardial ischemia and relevant disease thereof such as medicine are directly pressed according to a conventional method with relevant adjuvant together and are tablet, and six of method is the powder with the employed medicinal raw material of extraced total timosaponin; it is tablet etc. that relevant ancillary drug is directly pressed according to a conventional method with relevant adjuvant together.

Except that six kinds of above-mentioned basic skills, can also select other forms of the employed medicinal raw material of extraced total timosaponin or the employed medicinal raw material of extraced total timosaponin is carried out after method well known in the art handles, prepare the product that contains the employed medicinal raw material of extraced total timosaponin such as the medicine of various dosage forms.But, it should be noted that, in the employed medicinal raw material of above-mentioned direct use extraced total timosaponin, should be earlier according to the dosage requirement of employed extraced total timosaponin, conversion obtains the consumption of the employed medicinal raw material of extraced total timosaponin of required use.

In sum, extraced total timosaponin of the present invention and compositions thereof can be used for preventing, diagnose, detect, protect, treating and study the product of myocardial ischemia and relevant disease thereof, preferred agents and food, further preferred agents.

(4) technology speciality

The present invention is to being that the extraced total timosaponin of main component has been expanded new medical usage with the saponin, also for prevention, diagnosis, detect, protection, treatment and research myocardial ischemia and relevant disease thereof provide a kind of new medicament sources.Extraced total timosaponin safety and low toxicity of the present invention, pharmacological action is stronger, and its raw material sources are abundant, inexpensive, and preparation technology is simple, can be used for preparing the product of prevention, diagnosis, detection, protection, treatment and research myocardial ischemia and relevant disease thereof.

The present invention studies extraced total timosaponin targetedly, and the extraced total timosaponin pharmacological action is stronger, and its raw material sources are abundant, and preparation technology is simple, the yield height; And safe in utilization, one-object-many-purposes has been brought into play effect to greatest extent, and the scope of application is wide especially, therefore applies easily, can have a tremendous social and economic benefits in the short period of time.

The extraced total timosaponin stable in properties; use the quality of the pharmaceutical preparations of preparation stable; the effect of prevention, diagnosis, detection, protection, treatment and research myocardial ischemia and relevant disease thereof is obvious, so it is more suitable for preventing, diagnose, detect, protect, treating and study the suitability for industrialized production of myocardial ischemia and relevant disease product thereof.

In a word, active adaption of the present invention modern medical service and the job demand of scientific research field and the needs of human nature service, be the safe raw material that is used to prevent, diagnose, detect, protect, treat and study aspects such as myocardial ischemia and relevant disease thereof.

Description of drawings

Fig. 1 represents the influence of extraced total timosaponin to dog coronary ligation epicardial electrogram ST section summation (∑ ST);

Fig. 2 represents that extraced total timosaponin is to (the influence of N-ST) of dog coronary ligation myocardial infarct size;

Fig. 3 represents the influence of extraced total timosaponin to dog myocardial oxygen consumption (mL/min.100g-1);

Fig. 4 represents the influence of extraced total timosaponin to dog cardiac muscle oxygen uptake rate (%);

Fig. 5 represents the influence of extraced total timosaponin to dog coronary flow (ml/min);

Fig. 6 represents the influence of extraced total timosaponin to dog coronary resistance (Kpa/ml/min).

Wherein:

Expression solvent control group

The expression positive controls The expression small dose group

Dosage group in the expression

The heavy dose of group of expression

The specific embodiment

The present invention has studied the new pharmacological action of existing extraced total timosaponin and new purposes; a kind of raw material that can be used in products such as the prevention of preparation myocardial ischemia, diagnosis, protection and treatment is provided, has been convenient to the safe handling in medical industry and fields such as relevant industries such as food, beverage.

(1) preparation method

The preparation method of extraced total timosaponin of the present invention is:

(1) extract: routinely the rhizoma ane marrhenae of pulverizing is carried with 20～95% ethanol heat, the solvent volumetric usage is about 4～8 times of crude drug first; Maybe with the rhizoma ane marrhenae pulverized with 20～95% ethanol percolate extraction, the solvent volumetric usage is about 4～12 times of crude drug; Get extracting solution; With extracting liquid filtering, concentrated, the centrifugal or filtration of concentrated solution;

(2) divide high purification: (nonpolar or low pole macroporous resin is for being the polystyrene type porous adsorbent resin of bridging materials with styrene by nonpolar or low pole macroporous resin adsorption for the centrifuged supernatant of concentrated solution or filtrate, as D101, D201, ZTC-1, AB-8,1300-1 type macroporous resin etc.), impurity is removed in water and 20% following ethanol drip washing; The moisture lower alcohol eluting of reuse, lower alcohol is C1～C5 alcohols such as methanol, ethanol or propanol, and its concentration is 30～95%, and volumetric usage is 5～15 times of crude drug, collects the lower alcohol eluent, and being concentrated into does not have alcohol;

(3) drying: drying and crushing or spray drying are extraced total timosaponin of the present invention, product yield more than 1.5% of dose of making a living.

Extraced total timosaponin component analysis of the present invention: through silica gel, sephadexLH-20, reversed phase chromatography carrier column chromatographies such as ODS have obtained 6 chemical compounds, use chemistry, spectral method has been measured structure, be respectively: Sarsasapogenin-3-O-β-D-glucopyranosyl (1 → 2)-β-D-galactopyranoside (be called for short: 1-timosaponin A-1-III), Markogenin-3-O-β-D-glucopyranosyl (1 → 2)-β-D-galactopyranoside, 26-O-β-D-glucopyranosyl furan steroid-20 (22)-alkene-3 β, 26-glycol-3-O-β-D-glucopyranosyl (1 → 2)-β-D-galactopyranoside (is called for short: timosaponin B), Sarsasapogenin-3-O-β-D-glucopyranosyl (1 → 3)-O-β-D-glucopyranosyl (1 → 2)-β-D-galactopyranoside (is called for short: Xilin saponin B), 26-O-β-D-glucopyranosyl furan steroid-22,3 β, 26-triol-3-O-β-D-glucopyranosyl (1 → 2)-β-D-galactopyranoside (is called for short: timosaponin B-II), Sarsasapogenin.Detect through HPLC, content sum 〉=50% of timosaponin B-II, timosaponin B in the extraced total timosaponin, timosaponin B-II content 〉=30%, the best proportion of this extract is: timosaponin B-II, timosaponin B percentage composition example are 75～95:15～5.

(2) Chang Yong practical dosage form

The present invention prepares injectable powder and generally adopts conventional freeze-drying, as solvent, the steps include: to get extraced total timosaponin with water, adds excipient, is dissolved in water, and adds active carbon, filtration sterilization, and plug is partly rolled in fill, and lyophilization, tamponade are rolled lid and are got final product.Used excipient is selected from one or more in mannitol, gelatin hydrolysate, glucose, lactose, the dextran etc.Every bottle contains extraced total timosaponin 10～100mg.

The present invention prepares injectable powder also can adopt spray drying method, as solvent, the steps include: to get extraced total timosaponin with water, adds or do not add excipient (excipient is the same), be dissolved in water, add active carbon, filtration sterilization, spray drying, aseptic subpackaged, tamponade is rolled lid and is got final product.Every bottle contains extraced total timosaponin 10～100mg.

When the present invention prepared small-volume injection, preparation got final product as solvent with water for injection, also can add appropriate amount of auxiliary materials, and adjuvant is selected from one or more in ethanol, propylene glycol, glycerol, Polyethylene Glycol, benzyl benzoate, the dimethyl acetylamide.Every contains extraced total timosaponin 10～100mg.

The present invention prepares glucose infusion liquid or sodium chloride transfusion, with water for injection as solvent, add the preparation of an amount of glucose or sodium chloride and get final product, also can add appropriate amount of auxiliary materials, adjuvant is selected from one or more in ethanol, propylene glycol, glycerol, Polyethylene Glycol, benzyl benzoate, the dimethyl acetylamide.Every bottle contains extraced total timosaponin 10～100mg.

The present invention prepares oral formulations such as tablet, capsule, granule, oral liquid, and adjuvant can be lactose, starch, dextrin, stearate etc., technology preparation routinely.

In the present invention, the example of the above-described specific embodiment and the following stated all is in order to set forth the present invention better, is not to be used for limiting scope of invention.

Below by embodiment the present invention is described in detail.

The preparation method of embodiment 1, extraced total timosaponin

Rhizoma ane marrhenae after 100 kilograms of pulverizing drops into extraction pot, carries 3 times with 400 liters of heat of 70% ethanol successively at 90～100 ℃, and each extraction time is 90 minutes, merges 3 times extracting solution, is concentrated into 80 liters, and filtration is adjusted to 100 liters with filtrate with distilled water.Again slowly by the adsorption column of 350 liters of ZTC-1 type macroporous resins (Tianjin became the production of clarification technique company limited in positive day) is housed, remove impurity with 500 liters of distilled water, 1000 liter of 20% ethanol elution, 1000 liter of 45% ethanol elution of reuse, collect eluent, be concentrated into dried, after the drying and crushing yellow powder, be extraced total timosaponin of the present invention, weighing 2.7 kilograms altogether, is 2.7% by crude drug amount yield.

The another kind of preparation method of embodiment 2, extraced total timosaponin

Rhizoma ane marrhenae is thinly sliced, got 100kg,, collect the 2400L percolate with 30% ethanol 300L infiltration percolation extraction after 12 hours.Percolate is concentrated to 150L; Concentrated solution filters, filtrate feeds in the chromatographic column of the D101 type resin that 200L is housed, use distilled water 3000L, 20% ethanol 3000L, 60% ethanol 1200L eluting successively, collect 60% ethanol elution part, be concentrated into dried, after the drying and crushing yellow powder, be extraced total timosaponin of the present invention, weighing 3.1 kilograms altogether, is 3.1% by crude drug amount yield.

Another preparation method of embodiment 3, extraced total timosaponin

The dry medical material 2.5kg of the Rhizoma Anemarrhenae, with 50 liters of reflux, extract, of 75% alcoholic solution 2 times, each 2 hours, merge extracted twice liquid, reclaim under reduced pressure filters to get filtrate to there not being the alcohol flavor, filtrate transfers to relative density about 1.05 with distilled water, last AB-8 macroporous adsorptive resins washes the solubility impurity that anhydrates with water, continues and uses the different concentration ethanol eluting, collect 50% ethanol elution position, promptly get the eluent that is rich in phenolic acid glycoside and diterpene glycoside composition, reclaim under reduced pressure gets extraced total timosaponin 45g to doing.

Embodiment 4, extraced total timosaponin cause the therapeutical effect of myocardial infarction model to the dog coronary artery ligation

The present invention causes the therapeutical effect of myocardial infarction model to test by gastric infusion to the dog coronary artery ligation to extraced total timosaponin.

(1) test method

Hybrid dog (the The 2nd Army Medical College animal center provides) is divided into 5 groups at random: dosage group and extraced total timosaponin high dose group in negative control group, positive controls, extraced total timosaponin low dose group, the extraced total timosaponin, 5 every group.Negative control group is that the dog coronary artery ligation causes myocardial infarction model, gives the normal saline of equivalent; Positive controls DIAOXINXUE KANG JIAONANG (Chengdu buchu pharmaceutical Co. Ltd, lot number 0402061) as the contrast of extraced total timosaponin curative effect, clinical recommended dose is 0.6g/ days/people (being equivalent to total steroidal saponin 210mg), the people calculates with 60 kg body weight, be 3.5mg/kg/ people (total steroidal saponin amount), conversion is 7mg/kg for the dosage of dog, and design positive control pharmaceutical quantities is equivalent to be subjected to the middle dosage of reagent, so dosage is 21mg/kg.Extraced total timosaponin dog low dosage is 20mg/kg, and middle dosage is 60mg/kg, and high dose is 120mg/kg.

According to the new drug reviewing requirement, preparation dog coronary artery ligation causes myocardial infarction model, and (preparation method sees pharmacology's part of " Western medicine new drug preclinical study guideline compilation ", bureau of drug policy ﹠ administration of Ministry of Health of the People's Republic of China, 1993:61 for details; " pharmacological experimental methodology " second edition, People's Health Publisher, 1991:830,1113), each group is all with this model dog test.Each group is the back gastric infusion after coronary artery ligation, and negative control group is given with the volume normal saline.Observe following index then:

1. to the (influence of epicardial electrogram N-ST): the variation of 15min, 30min, 60min, 90min, 120min, 180min record epicardial electrogram after administration respectively of degree of myocardial ischemia (epicardial electrogram ∑ ST) and myocardial ischemia scope.The epicardial electrogram observation index: with the summation (∑ ST) that the ST section raises or leading more than the reduction 2mV counted (NST) and ST section lift-off value is the variation of index observing administration front and back, and calculating myocardium degree of ischemia (∑ ST) and scope (NST).To compare before different time measured value and the medication after the medication, between different time variation percentage rate (being 100% before the medication) is organized after the medication, compare.

2. the quantitative tissue of myocardial infarct size is learned the influence that detects: ligation is cored dirty after 3 hours, take by weighing weight whole-heartedly; Cut atrium and right ventricle, weigh up left ventricular mass, and under the coronary ligation line, be parallel to coronary sulcus left ventricle is cut into 4～6 of uniform thickness, clean, put 37 ℃ of dyeing of 0.05% chlorination nitro blue tetrazolium (N-BT) liquid 30 minutes with normal saline.Shaking dyeing liquor in the dyeing course frequently makes it fully to contact with cardiac muscle.After the dyeing immediately water wash away excess dyestuff.Infarcted region is not painted, and non-infarcted region skipper, cuts off coloured part, and uncoloured infarcted region is weighed, and calculates infarct weight and accounts for the percentage ratio that reaches left ventricular mass whole-heartedly.

3. to the influence of serum biochemical indicator behind the coronary ligation: respectively got right ventricle blood 3ml in 3 hours before the coronary ligation and after the administration, centrifugal 15 minutes of 3000rpm gets serum and surveys lactic acid dehydrogenase and (be called for short: LDH), serum creatine kinase (is called for short: CK).The mensuration employing LDH testing cassete method of LDH (lot number: 20040719, bio-engineering research institute is built up in Nanjing), CK (lot number: 20040802, bio-engineering research institute is built up in Nanjing).On UV751GD type ultraviolet spectrophotometer, carry out (Shanghai analytical tool factory).

(2) result of the test

1. to the influence of degree of myocardial ischemia (epicardial electrogram ∑ ST)

The result sees Fig. 1 for details.As can be seen from Figure 1, can alleviate degree of myocardial ischemia in dose dependent ground behind extraced total timosaponin group 20～120mg/kg intravenously administrable, ∑ ST significantly descends (P＜0.01), small dose group 60～90min and normal saline group difference highly significant (P＜0.01); Middle dosage group promptly has significant difference (P＜0.05) from 15min, and each time point and normal saline group relatively have significant difference during 30min, and 60～90min has highly significant difference (P＜0.01); High dose group 60～120min difference highly significant (P＜0.01).Extraced total timosaponin 20～120mg/kg irritates stomach the acute myocardial ischemia degree that the Canis familiaris L. coronary ligation causes is had significant therapeutic effect.

2. to the (influence of epicardial electrogram N-ST) of myocardial ischemia scope

Experimental result is seen Fig. 2.As can be seen from Figure 2, extraced total timosaponin can dwindle the myocardial ischemia scope in dose dependent ground, reduces N-ST value.Each time point N-ST value reduction rate of low dose group 20mg/kg and normal saline group compare with time point, significant difference during 30min (P＜0.05); In dosage group 60mg/kg N-ST value reduction rate and normal saline group are with the time point comparison during from 15～90min, there were significant differences (P＜0.05); High dose group (120mg/kg) compares from 30～180min reduction myocardial ischemia scope and normal saline group, and highly significant difference (P＜0.01) is arranged.

3. extraced total timosaponin is learned the influence that detects to the quantitative tissue of myocardial infarct size

Experimental result sees Table 1.As seen from Table 1, extraced total timosaponin 20～120mg/kg irritates stomach energy dose dependent ground minimizing myocardial infarct size.The basic, normal, high dosage group of extraced total timosaponin (20,60, the 120mg/kg) infarct/ratio whole-heartedly and the ratio of infarct/left ventricle and normal saline group relatively descend significantly (P＜0.05, P＜0.01).Wherein high dose group (120mg/kg) myocardial infarction area is with the ratio of weight and the ratio of infarct and left ventricular mass drop to 11.20 ± 1.82%, 19.32 ± 1.34% by 15.13 ± 1.51%, 26.30 ± 2.94% of normal saline group respectively whole-heartedly.

Table 1, extraced total timosaponin are measured the influence (x ± s, n=5/group) of myocardial infarct size to N-BT dyeing

^*P＜0.05, ^*Compare with the normal saline group P＜0.01

4. to the influence of serum biochemical indicator behind the coronary ligation

Experimental result sees Table 2,3.

Table 2, extraced total timosaponin are to the influence of coronary ligation tremulous pulse serum biochemical indicator (x ± s, n=5 only/group)

^*P＜0.05, ^*Compare with the normal saline group P＜0.01

Table 3, extraced total timosaponin are to the influence of coronary ligation tremulous pulse serum biochemical indicator (x ± s, n=5 only/group)

^*P＜0.05, ^*Compare with the normal saline group P＜0.01

Basic, normal, high dosage group of extraced total timosaponin and positive control drug DIAOXINXUE KANG JIAONANG group all can reduce the rising degree (P＜0.05) of Serum LDH and CK significantly.Wherein high dose group (120mg/kg) Serum LDH, CK can rise to 487.2 ± 145.3,612.0 ± 94.7 by 239.4 ± 72.0,297.7 ± 63.8 respectively, have only risen 2.05 ± 0.27 and 2.09 ± 0.29 times respectively.

Above result of the test explanation, extraced total timosaponin can obviously improve the rising degree that alleviates degree of myocardial ischemia and scope, minimizing myocardial infarct size, reduction Serum LDH and CK that causes myocardial infarction to cause after the dog coronary artery ligation.In addition, extraced total timosaponin also can significantly reduce the anesthetized dog myocardial oxygen consumption, damage has protective effect to rat myocardial ischemia and reperfusion.These effects all help prevention and treatment myocardial ischemia, and therefore, extraced total timosaponin can be used for preparing the medicine or the food for the treatment of myocardial ischemia.

Embodiment 5, extraced total timosaponin are to the influence of anesthetized dog myocardial oxygen consumption

The present invention irritates stomach to extraced total timosaponin the influence of anesthetized dog myocardial oxygen consumption is tested.

(1) test method

Hybrid dog (the The 2nd Army Medical College animal center provides) is divided into 5 groups at random: dosage group and extraced total timosaponin high dose group in negative control group, positive controls, extraced total timosaponin low dose group, the extraced total timosaponin, 5 every group.Negative control group is that the dog coronary artery ligation causes myocardial infarction model, gives the normal saline of equivalent; Positive controls DIAOXINXUE KANG JIAONANG (Chengdu buchu pharmaceutical Co. Ltd, lot number 0402061) as the contrast of extraced total timosaponin curative effect, clinical recommended dose is 0.6g/ days/people (being equivalent to total steroidal saponin 210mg), the people calculates with 60 kg body weight, be 3.5mg/kg/ people (total steroidal saponin amount), conversion is 7mg/kg for the dosage of dog, and design positive control pharmaceutical quantities is equivalent to be subjected to the middle dosage of reagent, so dosage is 21mg/kg.Extraced total timosaponin dog low dosage is 20mg/kg, and middle dosage is 60mg/kg, and high dose is 120mg/kg.

Experimental technique sees pharmacology's part of " Western medicine new drug preclinical study guideline compilation ", bureau of drug policy ﹠ administration of Ministry of Health of the People's Republic of China, 1993 for details; " pharmacological experimental methodology " second edition, People's Health Publisher, 1991).

(2) result of the test

1. to the influence of anesthetized dog myocardial oxygen consumption:

The extraced total timosaponin low dose group is irritated behind the stomach myocardial oxygen consumption and is compared there was no significant difference (P〉0.05) with physiology saline group; But middle dosage group 60～90min highly significant reduces dog myocardial oxygen consumption (P＜0.01), and difference is still remarkable during 120min; Promptly there were significant differences (P＜0.05) for high dose group 15min, 30min-180min difference highly significant (P＜0.01); DIAOXINXUE KANG group 15～120min compares difference highly significant (P＜0.01) before playing oxygen consumption and administration, still can significantly reduce myocardial oxygen consumption (P＜0.05) (the results are shown in Figure 3) during 180min.

2. to the influence of anesthetized dog cardiac muscle oxygen uptake rate:

Extraced total timosaponin can significantly reduce myocardium oxygen uptake rate after irritating stomach.Reduction rate is compared with time point with the normal saline group, and it is that there were significant differences (P＜0.05) that small dose group rises at 30min, difference highly significant during 60～120min (P＜0.01); Middle dosage group 15min promptly reduces anesthetized dog cardiac muscle oxygen uptake rate, the equal highly significant of each time point difference of 30min to 180min (P＜0.01); High dose group reduces (P＜0.01) with the normal saline group than the equal highly significant of myocardium oxygen uptake rate from 30min to 180min; It is that there were significant differences (P＜0.05) that positive control drug DIAOXINXUE KANG 15min rises, and 30min to 180min highly significant reduces myocardium oxygen uptake rate (P＜0.01).(the results are shown in Figure 4).

3. to the influence of anesthetized dog coronary flow:

Each dosage group of extraced total timosaponin can increase coronary flow; Increment rate is compared with time point with the normal saline group, and small dose group is from 30min to 90min difference highly significant (P＜0.01); Still there were significant differences during 120min (P＜0.05); Middle and high dosage group is then from the equal highly significant of 30～180min difference (P＜0.01).Positive control drug DIAOXINXUE KANG group also can obviously increase coronary flow, compares difference highly significant (P＜0.01) with the normal saline group with time point from 30～180min increment rate.(the results are shown in: Fig. 5).

4. to the influence of anesthetized dog coronary resistance:

Each dosage group of extraced total timosaponin can reduce coronary resistance, and reduction rate is compared with time point with the normal saline group, and low dose group (20mg/kg) is in 30min significant difference (P＜0.05), 60～180min difference highly significant (P＜0.01); Middle dosage group (60mg/kg) is from 30～180min difference highly significant (P＜0.01); High dose group (120mg/kg) can significantly reduce coronary resistance with the positive control drug DIAOXINXUE KANG from 15min, and the coronary resistance reduction rate is compared difference highly significant (P＜0.01) (the results are shown in Figure 6) with the normal saline group with time point during 30～180min.

Above result of the test explanation, extraced total timosaponin can obviously increase the anesthetized dog coronary flow, reduces coronary resistance, reduces total peripheral resistance, can obviously reduce myocardium oxygen uptake rate, reduces myocardial oxygen consumption.These effects all help prevention and treatment myocardial ischemia, and therefore, extraced total timosaponin can be used for preparing the medicine or the food for the treatment of myocardial ischemia.

The protective effect that embodiment 6, extraced total timosaponin damage rat myocardial ischemia and reperfusion

The present invention tests the protective effect of rat myocardial ischemia and reperfusion damage by gastric infusion extraced total timosaponin.

(1) test method

SD rat (the The 2nd Army Medical College animal center provides) is divided into 6 groups at random: dosage group and extraced total timosaponin high dose group in sham operated rats, normal saline matched group, positive controls, extraced total timosaponin low dose group, the extraced total timosaponin, 10 every group.Positive controls DIAOXINXUE KANG JIAONANG (Chengdu buchu pharmaceutical Co. Ltd, lot number 0402061) as the contrast of extraced total timosaponin curative effect, clinical recommended dose is 0.6g/ days/people (being equivalent to total steroidal saponin 210mg), the people calculates with 60 kg body weight, be 3.5mg/kg/ people (total steroidal saponin amount), conversion is 21mg/kg for the dosage of rat, and design positive control pharmaceutical quantities is equivalent to be subjected to the middle dosage of reagent, so dosage is 63mg/kg.Extraced total timosaponin dog low dosage is 60mg/kg, and middle dosage is 180mg/kg, and high dose is 360mg/kg.Each organizes the coronary ligation back gastric infusion that finishes, and administration volume and ischemia-reperfusion injury model group are consistent to saline volume.

Experimental technique sees pharmacology's part of " Western medicine new drug preclinical study guideline compilation ", bureau of drug policy ﹠ administration of Ministry of Health of the People's Republic of China, 1993 for details; " pharmacological experimental methodology " second edition, People's Health Publisher, 1991).

(2) result of the test

Behind the myocardial ischemia-reperfusion, Serum LDH, the active obviously increase of CK, myocardial infarction area weight obviously increases, and with Sham-operated control group significant differences (P＜0.01) is arranged relatively; Extraced total timosaponin low dose group (60mg/kg) activity of serum CK, myocardial infarction area/percentage by weight is lower than model group whole-heartedly, and difference has significance meaning (P＜0.05); The middle and high dosage group of extraced total timosaponin (120,360mg/kg) Serum LDH, CK activity, myocardial infarction area/percentage by weight is lower than model group whole-heartedly, and difference has highly significant meaning (P＜0.01); DIAOXINXUE KANG JIAONANG group Serum LDH, CK activity, myocardial infarction area/percentage by weight is starkly lower than model group (P＜0.01) (seeing: table 4) whole-heartedly.

Table 4, extraced total timosaponin to the influence of rat heart ischemical reperfusion injury (x ± s, n=10)

^*P＜0.05, ^*Compare with the normal saline group P＜0.01;

^{△ △}Compare with sham operated rats P＜0.01.

Extraced total timosaponin is irritated stomach has protective effect to SD rat myocardium from injury due to the ischemia-reperfusion; LDH, CK overflow in the time of can obviously suppressing due to the ischemia-reperfusion rat myocardium from injury; reduce Serum LDH, CK activity, dwindle myocardial infarction area weight, and certain dose-effect relationship is arranged.Compare with the DIAOXINXUE KANG JIAONANG of test dose, the effect of middle dosage extraced total timosaponin is similar to DIAOXINXUE KANG JIAONANG.

The preparation of embodiment 7, extraced total timosaponin injectable powder

Get extraced total timosaponin 30g, add dextran 30g, add 500ml water for injection, stir and make its dissolving; Add the injection water to 2000ml, add the 3.0g needle-use activated carbon, fully stirred 30 minutes; Decarbonization filtering; With 0.22 μ m filtering with microporous membrane; Be filled in the aseptic cillin bottle, every bottle of 2ml partly rolls plug; Lyophilization, tamponade is rolled lid and is got final product again.

The preparation of embodiment 8, extraced total timosaponin injectable powder

Get extraced total timosaponin 60g, add 500ml water for injection, stir and make its dissolving; Add the injection water to 1000ml, add the 1g needle-use activated carbon, fully stirred 30 minutes; Decarbonization filtering; With 0.22 μ m filtering with microporous membrane; Lyophilization gets sterilized powder, is distributed into 1000 bottles.

The preparation of embodiment 9, extraced total timosaponin injectable powder

Get extraced total timosaponin 40g, add lactose 50g, add 100ml water for injection, stir and make its dissolving; Add the injection water to 1000ml, add the 1.5g needle-use activated carbon, fully stirred 30 minutes; Decarbonization filtering; With 0.22 μ m filtering with microporous membrane; Spray drying gets sterilized powder, is distributed into 1000 bottles.

The preparation of embodiment 10, extraced total timosaponin injectable powder

(1) prescription

(2) preparation technology

Take by weighing the recipe quantity extraced total timosaponin by above prescription, join in an amount of water for injection, stir and make its dissolving; Add recipe quantity mannitol, stir and make dissolving fully, add to the full amount of water for injection; Add 0.1% needle-use activated carbon of amount of liquid, fully stirred 30 minutes; Decarbonization filtering; With 0.22 μ m filtering with microporous membrane; Plug is partly rolled in fill; Lyophilization, lid is rolled in tamponade.Make 9735 bottles altogether, yield rate is 97.35%.

The preparation of embodiment 11, extraced total timosaponin small-volume injection

Get extraced total timosaponin 5g, add 100ml water for injection, stir and make its dissolving; Add the injection water to 1000ml, with 0.22 μ m filtering with microporous membrane; The packing embedding, every bottle of 10ml, sterilization gets final product.

The preparation of embodiment 12, extraced total timosaponin small-volume injection

Get extraced total timosaponin 10g, add propylene glycol 30g, add 200ml water for injection, stir and make its dissolving; Add the injection water to 1000ml, add the 1.5g needle-use activated carbon, fully stirred 30 minutes; Decarbonization filtering; With 0.22 μ m filtering with microporous membrane; The packing embedding, every bottle of 5ml, sterilization gets final product.

The preparation of embodiment 13, extraced total timosaponin glucose infusion liquid

Get extraced total timosaponin 2g, add Polyethylene Glycol 10g, add glucose 500g, add 2000ml water for injection, stir and make its dissolving; Add the injection water to 5000ml; With 0.22 μ m filtering with microporous membrane; The packing embedding, every bottle of 100ml, sterilization gets final product.

The preparation of embodiment 14, extraced total timosaponin glucose infusion liquid

Get extraced total timosaponin 2g, add glucose 250g, add 1000ml water for injection, stir and make its dissolving; Add the injection water to 5000ml; With 0.22 μ m filtering with microporous membrane; The packing embedding, every bottle of 250ml, sterilization gets final product.

The preparation of embodiment 15, the transfusion of extraced total timosaponin sodium chloride

Get extraced total timosaponin 1g, add sodium chloride 90g, add 1000ml water for injection, stir and make its dissolving; Add the injection water to 10000ml; With 0.22 μ m filtering with microporous membrane; The packing embedding, every bottle of 250ml, sterilization gets final product.

The preparation of embodiment 16, extraced total timosaponin tablet

(1) prescription

(2) preparation technology

Take by weighing the principal agent and the adjuvant of recipe quantity respectively by above prescription, progressively increase behind the method mix homogeneously according to making soft material, system granule under the formulation and technology item by equivalent, dry, processes such as granulate, the heavy back of sheet single punch tablet machine and 10.5mm shallow concave punch tabletting have been calculated, control nude film hardness 5～7kg makes 9698 in tablet altogether, and yield rate is 96.98%.Adopt lift-over nebulization coating, art for coating is as follows:

The preparation of coating solution: gastric solubleness thin film dress material: 85G61235 is provided by Shanghai Colorcon Coating Technology Co., Ltd

Art for coating: will treat that (hardness 5～7kg) is put into coating pan to the coating nude film, start agitating device and air blast heater, when treating that the nude film temperature rises to 40 ℃, last 1/3 place that begins to open spray gun alignment tab bed sprays into the coating solution coating, 38～42 ℃ of control strip bed tempertaures, gas pound pressure 6kg, the coating solution flow velocity is 50mL/min, coating membrane heavily account for coated tablet heavy 3%.

The preparation of embodiment 17, extraced total timosaponin tablet

Get extraced total timosaponin 100g, microcrystalline Cellulose 80g, lactose 15g, pregelatinized Starch 60g sieve, and mix homogeneously with an amount of 10%PVP alcoholic solution system soft material, is granulated, and drying adds magnesium stearate 3g, granulate, and tabletting is made 1000.

The preparation of embodiment 18, extraced total timosaponin capsule

(1) prescription

(2) preparation technology gets in crude drug extraced total timosaponin and the prescription other adjuvant respectively by above prescription and crosses 100 mesh sieves respectively, put 60 ℃ of oven dry, take by weighing recipe quantity extraced total timosaponin and microcrystalline Cellulose, the carboxymethyl starch sodium equivalent method mix homogeneously that progressively increases, with an amount of dehydrated alcohol system soft material, 30 mesh sieves are granulated, 50～60 ℃ of dryings 2 hours with 30 mesh sieve granulate, add the Pulvis Talci and the carboxymethyl starch sodium mix homogeneously of recipe quantity.

The preparation of embodiment 19, extraced total timosaponin capsule

Get extraced total timosaponin 100g, microcrystalline Cellulose 100g, carboxymethyl starch sodium 14g, Pulvis Talci 8g, cross 100 mesh sieves respectively, put 60 ℃ of oven dry, get extraced total timosaponin and microcrystalline Cellulose, the carboxymethyl starch sodium equivalent method mix homogeneously that progressively increases, with an amount of dehydrated alcohol system soft material, 30 mesh sieves are granulated, 50～60 ℃ of dryings 2 hours, with 30 mesh sieve granulate, add the Pulvis Talci and the carboxymethyl starch sodium mix homogeneously of recipe quantity.

Claims (7)

1. the application of extraced total timosaponin in the preparation myocardial ischemia treating products,

The preparation method of described extraced total timosaponin comprises the steps:

(1) extract: it is some to get Rhizoma Anemarrhenae raw material, extracts, and gets extracting solution, i.e. Rhizoma Anemarrhenae total extract;

Described extracting method is a solvent extraction method: it is some to get rhizoma ane marrhenae, adds to extract solvent, extracts, and with extracting liquid filtering, concentrated, promptly gets the rhizoma ane marrhenae total extract;

Described solvent is to comprise water or hydrophilic organic solvent;

Described hydrophilic organic solvent is to comprise in ethanol, ethanol water, methanol or the acetone one or more;

Described ethanol water is 10～95% ethanol;

(2) separation and purification:, promptly get extraced total timosaponin with the extracting solution separation and purification;

Described isolation and purification method is to comprise in solvent segregation, solvent extraction, macroporous adsorbent resin method, the sedimentation method, salting out method, column chromatography or crystallization and recrystallization and the Steppecd crystallization one or more.

2. the application of the compositions of extraced total timosaponin according to claim 1 in the preparation myocardial ischemia treating products,

Described active ingredient in pharmaceutical is an extraced total timosaponin, and its content is 5%～90%, and described extraced total timosaponin dosage is 5～20mg/kgd when being used for the patient.

3. application according to claim 1 and 2 is characterized in that, the main component of described extraced total timosaponin is a timosaponin B-II chemical compound.

4. application according to claim 3 is characterized in that, described timosaponin B-II chemical compound content range in extraced total timosaponin is 30%～70%.

5. application according to claim 1 is characterized in that the operation of described macroporous adsorbent resin method comprises the steps:

(1) sample liquid is passed through macroporous adsorptive resins, remove impurity with water wash;

(2), collect the lower alcohol eluent with moisture lower alcohol eluting;

Described lower alcohol is C ₁～C ₅Alcohols, its concentration are 10～40%, and volumetric usage is 5～24 times of resin bed volume;

(3) be concentrated into driedly, and pulverize; Powder leaves standstill with acetone solution, filtration, recovery, separates out crystallization, filters, and wash crystallization promptly gets extraced total timosaponin

6. application according to claim 1 is characterized in that the preparation method of described extraced total timosaponin also comprises exsiccant step.

7. application according to claim 6, it is characterized in that described drying means is to comprise in atmosphere pressure desiccation, hypobaric drying method, boulton process, spray drying method, freeze-drying, far infrared heating drying method or the micro-wave drying method one or more.

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