CN103908571A

CN103908571A - Compound traditional Chinese medicine preparation for treating heart disease

Info

Publication number: CN103908571A
Application number: CN201310006120.8A
Authority: CN
Inventors: 赵宪斌; 肖延斌
Original assignee: Individual
Current assignee: Individual
Priority date: 2013-01-08
Filing date: 2013-01-08
Publication date: 2014-07-09
Anticipated expiration: 2033-01-08
Also published as: CN103908571B

Abstract

In order to meet the clinical needs, better treat cardiovascular and cerebrovascular diseases and improve the health level of the people, the invention provides a new pharmaceutical composition for the treatment of the cardiovascular and cerebrovascular diseases and its preparation method, and the pharmaceutical composition is mainly prepared from hawthorn leaves, allium macrostemon, polygonatum and rhizoma polygonati, and produces previously unimagined effects on the treatment of the cardiovascular and cerebrovascular diseases.

Description

A kind of Chinese traditional compound medicine for the treatment of heart disease

Technical field

The invention belongs to medical technical field, relate to preparation of a kind of main pharmaceutical composition of being made by Folium Crataegi, Bulbus Allii Macrostemonis, Rhizoma Polygonati that is used for the treatment of cardiovascular and cerebrovascular disease and preparation method thereof.

Background technology

Cardiovascular disease, is called again blood circulation diseases, is a series of diseases that relate to blood circulation.Blood circulation refers to hemophoric Organ and tissue in human body, mainly comprises heart, blood vessel (tremulous pulse, vein, blood capillary), can be subdivided into acutely and chronic, is all generally relevant with arteriosclerosis.

Cardiovascular disease is a kind of serious threat mankind, the particularly commonly encountered diseases of more than 50 years old middle-aged and elderly people health, along with the change of growth in the living standard and rhythm of life, " three-hypers disease " (being hypertension, hyperglycemia and hyperlipidemia) that is called as " affluenza " is increasing.With advancing age, Prevalence of Hypertension increases gradually.More than 60 years old in old people, 40%～45% suffer from hypertensive also suffer from hyperglycemia or hyperlipidemia simultaneously, show according to external data, the diabetes patient of 50% left and right is associated with the multiple infirmities of age such as hypertension, hyperlipidemia.The common pathologic basis such as angina pectoris, myocardial infarction, ischemic heart desease are all myocardial ischemia, blood supply of cardiac muscle, confession hypoxgia cause myocardial metabolism disorder, energy supply deficiency, myocardium shrinkage function declines, blood output reduces, and then affect the function of whole body, and even cause cardiomyocyte cell death.Cerebral ischemia re-pouring is the key of cerebrovascular disease therapy clinically.After cardiac-cerebral ischemia, affect energy metabolism, the multiple variations such as secondary lactic acid is piled up, calcium exceeds standard, radical damage; Many target spots reverse or improve these and change, and improving comprehensive therapeutic effect is the important goal of Drug therapy.

Folium Crataegi is the dried leaves of rosaceous plant Fructus Pyri Pashiae Crataegus pinnatifida Bge.var.major N.E.Br. or Fructus Crataegi Crataegus pinnatifida Bge..Folium Crataegi main component is the number of chemical compositions such as vetexin-glucoside, vitexin rhamnoside, rutin, vitexin, hyperin, ursolic acid, Quercetin and vitexin; pharmacological action is mainly reflected in anti-inflammatory and antalgic, blood fat reducing, prevent and treat atherosclerosis, the protection to myocardial ischemia, the protection to cerebral ischemia, to liver protecting, to aspects such as kidney protection, inhibition tumor cells.

Bulbus Allii Macrostemonis is the dry bulb of liliaceous plant Allium macrostemon Allium macrostemon Bge. or Chinese onion Allium chinensis G.Don.Bulbus Allii Macrostemonis main component is fatty acid composition, total sapogenins and the total alkaloids of the compositions such as linoleic acid, Palmic acid, oleic acid and sitosterol.The pharmacological action of Bulbus Allii Macrostemonis mainly contains that inhibiting bacteria and diminishing inflammation, spasmolytic are relievingd asthma, antiplatelet aggregation, antioxidation, blood fat reducing, atherosclerosis, the effects such as antitumor.

Rhizoma Polygonati is the dry rhizome of liliaceous plant Yunnan Rhizoma Polygonati Polygonatum kingianum Coll.et Hemsl., Rhizoma Polygonati Polygonatum sibiricum Red. or Polygonatum cyrtonema Hua Polygonatum cyrtonema Hua.Rhizoma Polygonati main component is polysaccharide, steroidal saponin, flavone, anthraquinone analog compound, aminoacid isoreactivity composition.The pharmacological action of Rhizoma Polygonati mainly contains hypoglycemic, and to the protection of myocardial damage, to the protection of cerebral ischemia, antioxidation, slow down aging, improve memory, antitumor, resisting fatigue, the aspects such as antibacterial and antioxidation.

Utilize at present the interaction of Folium Crataegi, Bulbus Allii Macrostemonis, Rhizoma Polygonati, composition of prescription is used for the treatment of cardiovascular and cerebrovascular disease, have not been reported.

Summary of the invention

In order to meet clinical needs, better treatment cardiovascular and cerebrovascular disease, improve the people's health level, the invention provides a kind of new pharmaceutical composition of cardiovascular and cerebrovascular disease and preparation method thereof that is used for the treatment of, this pharmaceutical composition is mainly prepared from by Folium Crataegi, Bulbus Allii Macrostemonis, Rhizoma Polygonati, aspect treatment cardiovascular and cerebrovascular disease, produce beyond thought effect.

Pharmaceutical composition of the present invention is mainly prepared from by Folium Crataegi, Bulbus Allii Macrostemonis, Rhizoma Polygonati, and the parts by weight of its crude drug are: 1～100 part of Folium Crataegi, 1～100 part of Bulbus Allii Macrostemonis, 1～100 part of Rhizoma Polygonati; Be preferably: 1～10 part of Folium Crataegi, 1～4 part of Bulbus Allii Macrostemonis, 1～2 part of Rhizoma Polygonati; Optimum is: 7 parts of Folium Crataegi, 3.5 parts of Bulbus Allii Macrostemonis, 1.5 parts of Rhizoma Polygonatis.

Folium Crataegi in pharmaceutical composition mentioned above, Bulbus Allii Macrostemonis, Rhizoma Polygonati can be carried or mix to carry with suitable solvent and method list and prepare extract, and total extract is mixed and made into any preparation with pharmaceutically acceptable adjuvant again.Wherein said extraction solvent preferred water or ethanol, extracting method can be infusion process, percolation, decocting method, reflux extraction or continuous extraction.

The invention provides the preferred extraction process of Folium Crataegi, Folium Crataegi extract can be according to being made by following method, but be not limited only to following method:

Take 7 parts of Folium Crataegi, use for the first time after 0.1～10 hour solvent soaking time of 2 times～40 times of medical material gross weight, under the condition of 30 DEG C～100 DEG C, extract 0.1～10 hour, extract 0.1～10 hour the condition of 30 DEG C～100 DEG C with the solvent of 2 times～20 times of medical material gross weight for the second time, extract 0.1～10 hour the condition of 30 DEG C～100 DEG C with 2 times～20 times amount solvents of medical material gross weight for the third time, extract 0.1～10 hour the condition of 30 DEG C～100 DEG C with 2 times～20 times amount solvents of medical material gross weight for the 4th time, merge extractive liquid, distillating recovering solvent, distillate is centrifugal, by the solvent wash precipitation of 0.1 times～100 times of volumes of medical material gross weight three times, merge supernatant after centrifugal and the supernatant of washing precipitation, distillation and concentration is near dry rear dry, pulverize, obtain.

The extract yield of preparing by above-mentioned technique is 10%～35%, and the content of total flavones is with anhydrous rutin (C ₂₁h ₂₀o ₁₂) meter, be not less than 20%; Vitexin rhamnoside (C ₂₇h ₃₄o ₁₄) content be not less than 2%.

The invention provides the preferred extraction process of Bulbus Allii Macrostemonis, Rhizoma Polygonati, Bulbus Allii Macrostemonis, Rhizoma Polygonati extract can be according to being made by following method, but are not limited only to following method:

Take 1.5 parts of 3.5 parts of Bulbus Allii Macrostemonis and Rhizoma Polygonatis, under 50 DEG C～100 DEG C conditions, add for the first time 2 times～40 times solvent extractions 0.1～10 hour of medical material gross weight, add for the second time 2 times～20 times solvent extractions 0.1～10 hour of medical material gross weight, collecting decoction, filtrate is concentrated into 0.1 times～20 times of medical material gross weights, add ethanol make containing alcohol amount be 20%～90%, static or centrifugal, get supernatant, filter Distillation recovery ethanol, continue distillation and concentration and become thick paste, dry, pulverize, to obtain final product.

Bulbus Allii Macrostemonis, the Rhizoma Polygonati extract yield prepared by above-mentioned technique are 25%～45%, contain polysaccharide with anhydrous glucose (C ₆h ₁₂o ₆) meter be not less than 4%.

More than composition is by weight as proportioning, in the time producing, can increase or reduce according to corresponding proportion, as large-scale production can be taking kilogram as raw material, or taking ton as unit, small-scale production also can be in grams, weight can increase or reduce, but the constant rate of weight proportion between each component.

The ratio of above weight proportion obtains through science screening, and for especial patient, ratio that can corresponding adjustment composition, increases or reduce being no more than 100%.

The consumption of medicine components of the present invention through inventor carry out lot of experiments grope sum up draw, each component consumption all has good therapeutic effect within the scope of above-mentioned weight portion.

The invention provides a kind of pharmaceutical composition for the preparation for the treatment of cardiovascular and cerebrovascular disease aspect, be mainly used in treating the aspect diseases such as cerebral thrombosis, coronary heart diseases and angina pectoris, vasculitis, myocardial infarction and hyperlipemia.

Pharmaceutical composition of the present invention can add one or more pharmaceutically acceptable carriers, is applicable to need the patient of this treatment in the mode of oral or parenteral.When oral, can be made into conventional solid preparation, as tablet, capsule, dispersible tablet, oral liquid, granule, chewable tablet, oral cavity disintegration tablet, drop pill, slow releasing tablet, slow releasing capsule, controlled release tablet, controlled release capsule, make liquid preparation if water or oil-suspending agent or other liquid preparations are as syrup etc.; During for parenteral, can be made into solution, water or the oil-suspending agent etc. of injection, as liquid drugs injection, freeze-dried powder, aseptic powder injection, transfusion etc.Preferred form is injection, Tablet and Capsula.

Pharmaceutical composition of the present invention can adopt the conventional method in existing pharmaceutical field to produce, and when needs, can add various pharmaceutically acceptable carriers.Described carrier comprises diluent, excipient, filler, binding agent, wetting agent, disintegrating agent, absorption enhancer, surfactant, absorption carrier, lubricant of pharmaceutical field routine etc.

Pharmaceutical composition of the present invention, in the time making injection, in order to increase its dissolubility, can add the solubilizing agents such as tween 80.In transfusion, can add the isoosmotic adjusting agent for regulating osmotic pressure, for example, sodium chloride, potassium chloride, magnesium chloride, calcium chloride, sodium lactate, glucose, xylitol, sorbitol and dextran etc., preferably sodium chloride or glucose.In powder pin, can add excipient, for example, mannitol, glucose etc.

Pharmaceutical composition of the present invention has the following advantages:

(1) provide a kind of new pharmaceutical composition of cardiovascular and cerebrovascular disease and preparation method thereof that is used for the treatment of, met urgent clinical needs.

(2) interaction to pharmaceutical composition of the present invention and composition of prescription have carried out pharmacodynamic study first, and result is as follows:

1. this pharmaceutical composition can improve S-T section and raises; Can obviously reduce the activity of creatine kinase in serum cardiac enzyme (CK), lactic acid dehydrogenase (LDH), aspartate amino transferase (AST); Atpase activity is reduced, and lactic acid (LD) and free fatty (NEFA) content reduce; Malonaldehyde (MDA) content is reduced, and the activity of total superoxide dismutase (SOD), glutathion peroxidase (GSH-Px) raises; Each administration group whole blood viscosity reduces, plasma viscosity reduces, packed cell volume reduces, erythrocyte sedimentation rate reduces, this pharmaceutical composition energy inhibiting erythrocyte aggregation is described, reduce erythrocyte fragility, strengthen its morphotropism, the heart rate that makes to raise is tending towards normally, left ventricular systolic pressure raises, left chamber diastolic pressure reduces, and the maximum climbing speed in left chamber raises, the maximum fall off rate in left chamber raises, eventually last diastolic pressure reduction.Treatment group pathological changes and model group be obvious alleviates myocardial cell disorder, breaking degree is lighter, karyopycnosis and eosinophilic cytoplasmic apparition reduce, myocardial infarction area obviously reduces, and improvement and therapeutical effect that this pharmaceutical composition has the change of myocardial infarction and ischemia model rat indices are described.

2. this pharmaceutical composition can obviously reduce the content of T-CHOL in serum (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and apolipoprotein B (ApoB), increases HDL-C (HDL-C) and Apolipoprotein A1 (ApoA1) content; Can obviously reduce malonaldehyde (MDA) content, increase the content of nitric oxide (NO), the total superoxide dismutase (SOD) that raises, glutathion peroxidase (GSH-Px) activity; Treatment group fatty degeneration of liver Leukopenia, endochylema lactone drips and reduces or disappear, and cell volume is close to normal group hepatocyte.Illustrate that this pharmaceutical composition has and improves and therapeutical effect the change of Hyperlipemia model rat indices.

3. result of the test shows that medicament composition capsule agent curative effect of the present invention is obviously better than alone Folium Crataegi, Bulbus Allii Macrostemonis, Rhizoma Polygonati.Prompting Folium Crataegi, Bulbus Allii Macrostemonis, the application of Rhizoma Polygonati compatibility have the effect of Synergistic, and consequently those skilled in the art institute is beyond thought.

(3) the each proportioning of the present composition has been carried out to pharmacodynamic study, drawn the optimal proportion of the present composition.

(4) the present invention feeds intake with raw material, and preparation technology is simple, and between different batches medicine, mass discrepancy is little, and drug quality is more uniform and stable.

(5) acute toxicity testing carrying out shows that the maximum tolerated dose of medicament composition capsule agent of the present invention is equivalent to 356 times of 70kg body weight day for human beings research on maximum utilized quantity, has shown pharmaceutical composition low toxicity of the present invention, safe.

(6) stability experiment carrying out shows that medicament composition capsule agent indices of the present invention is all more stable, has ensured the safety of clinical application.

(7) present composition combination drug determined curative effect, and reduced relative dosage, be with a wide range of applications.

Below example is further set forth the beneficial effect of pharmaceutical composition of the present invention by experiment.

Embodiment

The detailed description of the invention of form by the following examples, is described in further detail foregoing of the present invention.But this should be interpreted as to the scope of the above-mentioned theme of the present invention only limits to following embodiment.All technology realizing based on foregoing of the present invention all belong to scope of invention.In above example, the adjuvant of each dosage form can be replaced with pharmaceutically acceptable adjuvant, or reduces, increases.

Embodiment 1: the preparation of Folium Crataegi extract

Take 7 parts of Folium Crataegi, use for the first time after the 45% soak with ethanol time 2 h of 8 times of medical material gross weight, under the condition of 80 DEG C, extract 2 hours, extract 1 hour the condition of 80 DEG C with 45% ethanol of 5 times of medical material gross weight for the second time, extract 0.5 hour the condition of 80 DEG C with 4 times of amount 45% ethanol of medical material gross weight for the third time, extract 0.5 hour the condition of 80 DEG C with 3 times of amount 45% ethanol of medical material gross weight for the 4th time, merge extractive liquid, Distillation recovery ethanol, centrifuge centrifugal 30min under the rotating speed of 3000R per minute for distillate, with the purified water washing precipitation of 2 times of volumes of medical material gross weight three times, merge supernatant after centrifugal and the supernatant of washing precipitation, distillation and concentration is near dry rear dry, pulverize, obtain.

The discriminating of Folium Crataegi extract

Get this product 50mg, add ethanol 5ml, shake up, supersound process 5 minutes, filters, and gets filtrate as need testing solution.Separately get control substance of Rutin, hyperin reference substance, add respectively ethanol and make the solution of every 1ml containing 0.2mg, product solution in contrast.Test according to thin layer chromatography (" Chinese Pharmacopoeia " annex VI B in 2010), draw the each 1 μ l of above-mentioned three kinds of solution, put respectively on same polyamide film, taking ethanol-acetone-water (7: 5: 6) as developing solvent, launch, take out, dry, spray, with aluminum chloride test solution, dries up, place after 1 hour, put under ultra-violet lamp (365nm) and inspect.In test sample chromatograph, with reference substance chromatograph relevant position on, the fluorescence speckle of aobvious same color.

The assay of Folium Crataegi extract

The preparation precision of reference substance solution takes the control substance of Rutin 25mg to constant weight at 120 DEG C of drying under reduced pressure, put in 50ml measuring bottle, add appropriate amount of ethanol, supersound process makes to dissolve, let cool,, shake up to scale with ethanol dilution, precision measures 20ml, put in 50ml measuring bottle, add water to scale, shake up, obtain (every 1ml is containing anhydrous rutin 0.20mg).

The preparation precision of standard curve measures reference substance solution 1ml, 2ml, 3ml, 4ml, 5ml, 6ml, put respectively in 25ml measuring bottle, respectively add water to 6ml, add 5% sodium nitrite solution 1ml, make to mix, place 6 minutes, add 10% aluminum nitrate solution 1ml, shake up, place 6 minutes, hydro-oxidation sodium test solution 10ml, add water to again scale, shake up, place 15 minutes, taking corresponding reagent as blank, according to ultraviolet visible spectrophotometry (" Chinese Pharmacopoeia " annex VA in 2010), wavelength place with 500nm measures absorbance, taking absorbance as vertical coordinate, concentration is abscissa, drawing standard curve.

Algoscopy is got this product 0.15g, accurately weighed, puts in tool plug conical flask, and precision adds Diluted Alcohol 25ml, close plug, shake up, supersound process 5 minutes, places more than 3 hours, filters, and precision measures subsequent filtrate 2ml, put in 25ml measuring bottle, be diluted with water to scale, shake up, as need testing solution.Precision measures need testing solution 2ml, puts in 25ml measuring bottle, and the method under the preparation of sighting target directrix curve, from " adding water to 6ml ", measure absorbance, precision measures need testing solution 2ml simultaneously, puts in 25ml measuring bottle in accordance with the law, add water to scale, shake up, as blank solution.Read the amount of rutin in need testing solution from standard curve, calculate, to obtain final product.

Vitexin rhamnoside

Chromatographic condition and system suitability are taking octadecylsilane chemically bonded silica as filler; Taking oxolane-methanol-acetonitrile-acetic acid-water (38: 3: 3: 4: 152) as mobile phase; Detection wavelength is 330nm.Number of theoretical plate calculates and should be not less than 2500 by vitexin rhamnoside peak.

The preparation precision of reference substance solution takes at the vacuum drying apparatus vitexin rhamnoside reference substance of dry 12 hours appropriate, adds Diluted Alcohol and is diluted to scale and is mixed with every 1mL containing 50 μ g vitexin rhamnoside reference substance solution, to obtain final product.

This product under weight differential item is got in the preparation of need testing solution, and porphyrize is got about 0.25g, accurately weighed, put in 50ml measuring bottle, add Diluted Alcohol 40ml, supersound process 30 minutes, lets cool, and is diluted to scale with Diluted Alcohol, shake up, filter, get subsequent filtrate 5ml, be placed in 10mL volumetric flask, add Diluted Alcohol and be diluted to scale, shake up, to obtain final product.

Algoscopy is accurate reference substance solution and the each 20 μ L of need testing solution of drawing respectively, inject hplc determination, to obtain final product.

Make respectively three batches of Folium Crataegi extract yield and assay the results are shown in Table 1 by above-mentioned technique.

Table 1 Folium Crataegi extract yield and assay result

Batch	Yield (%)	General flavone content (%)	Vitexin rhamnoside content (%)
				1	25.18	29.13	2.78
2	25.52	28.83	2.72
				3	25.4	28.59	2.7
On average	25.37	28.85	2.73

Embodiment 2: the preparation of Bulbus Allii Macrostemonis, Rhizoma Polygonati extract

Take 1.5 parts of 3.5 parts of Bulbus Allii Macrostemonis and Rhizoma Polygonatis, under 100 DEG C of conditions, add for the first time 10 times of water gagings to decoct 2 hours, add for the second time 8 times of water gagings to decoct 1 hour, collecting decoction, filtrate is concentrated into 1: 1, add ethanol make containing alcohol amount be 55%, centrifugal, get supernatant, filter Distillation recovery ethanol, continue distillation and concentration to appropriate, dry, pulverize, to obtain final product.

The assay of Bulbus Allii Macrostemonis, Rhizoma Polygonati extract

The preparation of reference substance solution is learnt from else's experience 105 DEG C and is dried to the anhydrous glucose reference substance 33mg of constant weight, accurately weighed, puts in 100ml measuring bottle, is dissolved in water and is diluted to scale, shakes up, and obtains (in every 1ml, containing anhydrous glucose 0.33mg).

The preparation precision of standard curve measures reference substance solution 0.1ml, 0.2ml, 0.3ml, 0.4ml, 0.5ml, 0.6ml, put respectively in 10ml tool plug scale test tube, respectively add water to 2.0ml, shake up, in ice-water bath, slowly drip 0.2% By Anthrone Sulphuric acid solution to scale, mix, let cool in rearmounted water-bath and be incubated 10 minutes, take out, put immediately in ice-water bath cooling 10 minutes, take out, taking corresponding reagent as blank.According to ultraviolet visible spectrophotometry (" Chinese Pharmacopoeia " annex V A in 2010), measure absorbance at 582nm wavelength place, taking absorbance as vertical coordinate, concentration is abscissa, drawing standard curve.

Algoscopy is got the about 0.5g of this product fine powder, accurately weighed, put in round-bottomed flask, add 80% ethanol 150ml, put in water-bath reflux 1 hour, filter while hot, 80% hot ethanol washing 3 times for residue, each 10ml, residue and filter paper are put in flask, 150ml adds water, put in boiling water bath reflux 1 hour, filter while hot, hot wash 4 times for residue and flask, each 10ml, merging filtrate and washing liquid, let cool, be transferred in 250ml measuring bottle, add water to scale, shake up, precision measures 1ml, put in 10ml tool plug dry test-tube, method under the preparation of sighting target directrix curve, from " adding water to 2.0ml ", measure absorbance in accordance with the law, read the weight (mg) containing anhydrous glucose in need testing solution from standard curve, calculate, obtain.

Make respectively three batches of Bulbus Allii Macrostemonis, Rhizoma Polygonati extract yield and assay by above-mentioned technique and the results are shown in Table 2.

Table 2 Bulbus Allii Macrostemonis, Rhizoma Polygonati extract yield and assay result

Batch	Yield (%)	Polyoses content (%)
			1	36.78	4.51
2	37.05	4.55
			3	37.22	4.63
On average	37.02	4.56

Embodiment 3: the preparation of composition tablet

Prescription

Method for making: Folium Crataegi is extracted to obtain to Folium Crataegi extract according to embodiment 1 method; Bulbus Allii Macrostemonis, Rhizoma Polygonati extract to obtain Bulbus Allii Macrostemonis, Rhizoma Polygonati extract according to embodiment 2.Take the adjuvant of extract and recipe quantity, pulverize, cross respectively 100 mesh sieves, for subsequent use.By Folium Crataegi extract, Bulbus Allii Macrostemonis, Rhizoma Polygonati extract, starch, pregelatinized Starch, microcrystalline Cellulose mix homogeneously, stirs, and makes suitable soft material.Cross 18 mesh sieve granule processed.Granule is dried under the condition of 60 DEG C.Dried granule adds carboxymethyl starch sodium and magnesium stearate, crosses 18 mesh sieve granulate, mix homogeneously.Sampling, semi-finished product chemical examination.According to the definite sheet weight sheet of chemical examination.Finished product is examined entirely, packaging warehouse-in.

Embodiment 4: the preparation of composition capsule

Prescription

Method for making: Folium Crataegi is extracted to obtain to Folium Crataegi extract according to embodiment 1 method; Bulbus Allii Macrostemonis, Rhizoma Polygonati extract to obtain Bulbus Allii Macrostemonis, Rhizoma Polygonati extract according to embodiment 2.Take the adjuvant of extract and recipe quantity, pulverize, cross respectively 100 mesh sieves, for subsequent use.By Folium Crataegi extract, Bulbus Allii Macrostemonis, Rhizoma Polygonati extract, starch mix homogeneously, stirs, and makes suitable soft material.Cross 18 mesh sieve granule processed.Granule is dried under the condition of 60 DEG C.Cross 18 mesh sieve granulate, mix homogeneously.Sampling, semi-finished product chemical examination.Encapsulated according to the weight that chemical examination is definite.Finished product is examined entirely, packaging warehouse-in.

Embodiment 5: the preparation of composition granule

Prescription

Method for making: Folium Crataegi is extracted to obtain to Folium Crataegi extract according to embodiment 1 method; Bulbus Allii Macrostemonis, Rhizoma Polygonati extract to obtain Bulbus Allii Macrostemonis, Rhizoma Polygonati extract according to embodiment 2.Take the adjuvant of extract and recipe quantity, pulverize, cross respectively 100 mesh sieves, for subsequent use.By Folium Crataegi extract, Bulbus Allii Macrostemonis, Rhizoma Polygonati extract, the method mix homogeneously that lactose powder increases progressively with equivalent, adds 2%HPMC60% alcoholic solution appropriate, stirs, and makes suitable soft material.Cross 18 mesh sieve granule processed.Granule is dried under the condition of 55 DEG C.Dry granule is crossed 18 mesh sieve granulate.Sampling, in semi-finished product chemical examination granule, the content of principal agent, determines loading amount.Packaging, finished product is examined entirely, packaging warehouse-in.

Embodiment 6: the preparation of composition soft agent

Prescription

Method for making: Folium Crataegi is extracted to obtain to Folium Crataegi extract according to embodiment 1 method; Bulbus Allii Macrostemonis, Rhizoma Polygonati extract to obtain Bulbus Allii Macrostemonis, Rhizoma Polygonati extract according to embodiment 2.Take the adjuvant of extract and recipe quantity, pulverize, cross respectively 100 mesh sieves, for subsequent use.By the soybean oil of recipe quantity and soybean phospholipid, Cera Flava heating and melting, mix, let cool, add Folium Crataegi extract, Bulbus Allii Macrostemonis, Rhizoma Polygonati extract grind well, and are pressed into soft capsule.

Embodiment 7: the preparation of Folium Crataegi total flavones extract

Get the Folium Crataegi extract of embodiment 1, after defat with petroleum ether by 1/2 amount, discard petroleum ether liquid, be extracted with ethyl acetate again, extract reclaim under reduced pressure ethyl acetate is also concentrated into dry, add suitable quantity of water to make to dissolve, be added on (granularity: 30～60 orders on processed good polyamide column, with 95% ethanol wet method dress post, first use 95% ethanol elution of 3 times of column volumes, use afterwards the water elution of 3 times of column volumes extremely without alcohol taste, for subsequent use), first use the water elution of 2 times of column volumes, discard water lotion, then use 80% ethanol elution of 3 times of column volumes, collect eluent, reclaim ethanol and be concentrated into the concentrated solution of relative density approximately 1.02～1.08 (60 DEG C), dry, obtain Folium Crataegi total flavones extract.

Make respectively three batches of Folium Crataegi total flavones extract yields and assay the results are shown in Table 3 by above-mentioned technique.

Table 3 Folium Crataegi total flavones extract yield and assay result

Batch	Yield (%)	General flavone content (%)	Vitexin rhamnoside content (%)
				1	18.97	85.27	9.21
2	18.74	85.78	9.35
				3	19.02	86.12	9.48
On average	18.91	85.72	9.35

Embodiment 8: the preparation of Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract

Get Bulbus Allii Macrostemonis, the Rhizoma Polygonati extract of embodiment 2, by 95% long-pending ethanol precipitation of diploid, repeatedly carry out 4 times.With the dialyzer small-molecule substance of dialysing, precipitate with ethanol again, finally uses the absolute ether of equal volume and appropriate washing with acetone precipitate, dry Bulbus Allii Macrostemonis, the coarse solomon's seal polysaccharide extract of obtaining.

Make respectively three batches of Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract yield and assay by above-mentioned technique and the results are shown in Table 4.

Table 4 Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract yield and assay result

Batch	Yield (%)	Polyoses content (%)
			1	12.35	14.52
2	12.74	14.66
			3	12.68	14.31
On average	12.59	14.5

Embodiment 9: the preparation of compositions aqueous injection

Prescription

Method for making: take Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract that Folium Crataegi total flavones extract that the Folium Crataegi of recipe quantity prepared according to embodiment 7 and Bulbus Allii Macrostemonis, Rhizoma Polygonati are prepared according to embodiment 8.Carry and process the previous day such as pipeline and the container etc. that dosing is used, rinse with fresh water for injection more before use.Folium Crataegi total flavones extract and Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract are added heated and stirred in the water for injection of dosing amount 70% dissolve completely, benefit adds to the full amount of water for injection.Add the needle-use activated carbon of dosing amount 0.05%, heated and stirred 15 minutes.Through sand stick filtering decarbonization.Measure the also pH value of regulator solution.Through the microporous filter membrane fine straining of 0.45um.Check the clarity of solution, semi-finished product chemical examination.By solution embedding in glass ampule.100 DEG C of flowing steam sterilizations 30 minutes.While hot sample being put into 0.01% methylene blue solution hunts leak.Lamp inspection, finished product is examined entirely, packaging warehouse-in.

Embodiment 10: the preparation of composition powder injection

Prescription

Method for making: take Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract that Folium Crataegi total flavones extract that the Folium Crataegi of recipe quantity prepared according to embodiment 7 and Bulbus Allii Macrostemonis, Rhizoma Polygonati are prepared according to embodiment 8.First container tool and the antibiotic glass bottle dosing used, plug etc. carry out aseptic process.Folium Crataegi total flavones extract and Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract are added heated and stirred in the sterile water for injection of dosing amount 40% dissolve completely.30% the sterile water for injection heated and stirred that mannitol adds dosing amount is dissolved completely, merges above-mentioned solution, adds sterile water for injection to full dose.Add the needle-use activated carbon of dosing amount 0.05%, heated and stirred 15 minutes.Through sand stick filtering decarbonization.Measure the also pH value of regulator solution.Through the microporous filter membrane fine straining of 0.22um.Check the clarity of solution, semi-finished product chemical examination.Be sub-packed in antibiotic glass bottle half tamponade.Sample is put into freeze dryer lyophilization.Carry out lyophilizing by following freeze-dry process: 1. pre-freeze: be cooled to-35 DEG C, keep temperature 5 hours; 2. low-temperature distillation :-35 DEG C of insulations, open vacuum pump evacuation and keep 2 hours, then slowly heat up, about 30 hours, temperature is risen to 0 DEG C; 3. high temperature drying: be warming up to 25 DEG C in 2 hours, insulation was to 2 hours; 4. shutting down lyophilizing finishes.Tamponade, rolls lid.Finished product is examined entirely, packaging warehouse-in.

Embodiment 11: the preparation of compositions sodium chloride injection

Prescription

Method for making: take Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract that Folium Crataegi total flavones extract that the Folium Crataegi of recipe quantity prepared according to embodiment 7 and Bulbus Allii Macrostemonis, Rhizoma Polygonati are prepared according to embodiment 8.Carry and process the previous day such as pipeline and the container etc. that dosing is used, rinse with fresh water for injection more before use.Folium Crataegi total flavones extract and Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract are added heated and stirred in the water for injection of dosing amount 40% dissolve completely, sodium chloride is dissolved completely with the water for injection of dosing amount 20%.Merge above-mentioned solution, benefit adds to the full amount of water for injection.Add the needle-use activated carbon of dosing amount 0.05%, heated and stirred 15 minutes.Through sand stick filtering decarbonization.Measure the also pH value of regulator solution.Through the microporous filter membrane fine straining of 0.45um.Check the clarity of solution, semi-finished product chemical examination.Fill is in the infusion bottle of 250ml.115 DEG C of pressure sterilizings 30 minutes.Lamp inspection, finished product is examined entirely, packaging warehouse-in.

Embodiment 12: the preparation of compositions glucose injection

Prescription

Method for making: take Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract that Folium Crataegi total flavones extract that the Folium Crataegi of recipe quantity prepared according to embodiment 7 and Bulbus Allii Macrostemonis, Rhizoma Polygonati are prepared according to embodiment 8.Carry and process the previous day such as pipeline and the container etc. that dosing is used, rinse with fresh water for injection more before use.Folium Crataegi total flavones extract and Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract are added heated and stirred in the water for injection of dosing amount 40% dissolve completely, glucose is dissolved completely with the water for injection of dosing amount 20%.Merge above-mentioned solution, benefit adds to the full amount of water for injection.Add the needle-use activated carbon of dosing amount 0.05%, heated and stirred 15 minutes.Through sand stick filtering decarbonization.Measure the also pH value of regulator solution.Through the microporous filter membrane fine straining of 0.45um.Check the clarity of solution, semi-finished product chemical examination.Fill is in the infusion bottle of 250ml.115 DEG C of pressure sterilizings 30 minutes.Lamp inspection, finished product is examined entirely, packaging warehouse-in.

Test sample study on the stability

Composition capsule, prescription and preparation method are referring to the preparation of embodiment 4 capsules.

Investigation project: character, moisture, content.

Accelerated test and long-term stable experiment method and result: this product is put under the condition of 40 DEG C ± 2 DEG C of temperature, relative humidity 75% ± 5%, place 6 months and the condition of 25 DEG C ± 2 DEG C of temperature, relative humidity 60% ± 10% under place 12 months, indices has no significant change, and experimental result shows that the long-term placement of composition capsule is basicly stable.

Observation of curative effect

The research of test example 1 present composition acute toxicity test

Get 20 of Kunming kind white mice, male and female half and half.Fasting 12 hours, every gavages 40% (g/mL) present composition medicated powder (preparing by purified water) by 0.4ml/10g.Reach Cmax, maximum administration volume.In one day, gavage 1 time, mice active situation and dead quantity in Continuous Observation 7 days, result mice is by after above-mentioned oral dose administration, and in 7 days, all without death, freely, diet is normal in activity, mao glossy, without loose stool etc.The maximum tolerated dose of mice is 16g/Kg as calculated, is 356 times of clinical adult's consumption.

The research of test example 2 present composition general pharmacologies

1) impact on the performance of Kunming mouse general behavior

After gastric infusion, each administration treated animal and blank group relatively, movable normal, walk with a steady step, without the phenomenon such as sialorrhea, amyostasia, have no generation untoward reaction.

2) impact on Kunming mouse spontaneous activity

After gastric infusion, 0.5h puts animal in mice activity count instrument active box, adapts to record the animal activity number of times in 10min after 5min; Each dosage group and relatively animal activity number of times of blank group.After result administration, each treated animal compares with blank group, and significant difference does not appear in movable number of times, and data are in table 5.

The impact of table 5 medicine on mice autonomic activities

Group	Dosage (mg/kg)	Number of animals	Movable number of times
				Blank group	-	10	236.8±21.76
High dose group	2450	10	233.2±15.68
				Middle dosage group	1220	10	239.9±18.85
Low dose group	409	10	231.4±18.88

Note: * P < 0.05 (with model group comparison).

3) sub-threshold dose pentobarbital sodium is brought out to the impact that Kunming mouse is fallen asleep

1h after each treated animal gastric infusion, lumbar injection pentobarbital sodium 32mg/kg (for the maximum threshold dose that animal 90%～100% mice righting reflex of mensuration does not disappear in advance), records the rear 15min of pentobarbital sodium injection with Mus number more than interior righting reflex loss 1min.Result: respectively organize laboratory animal and giving after corresponding medicine, sub-threshold dose pentobarbital sodium is brought out to mice and fall asleep and do not detect difference, show that sub-threshold dose pentobarbital sodium brings out mice and falls asleep uninfluencedly, data are in table 6.

Table 6 brings out on sub-threshold dose pentobarbital sodium the impact that mice falls asleep

Group	Dosage (mg/kg)	Number of animals	The number of elements of falling asleep	Male and female	The rate of falling asleep (%)
						Blank group	-	10	6	♀2♂4	60
High dose group	2450	10	8	♀5♂3	80
						Middle dosage group	1220	10	6	♀3♂3	60
Low dose group	409	10	7	♀5♂2	70

4) on the pentobarbital sodium impact of the length of one's sleep

Each group Kunming mouse gavage to gastric infusion after 1h, lumbar injection pentobarbital sodium 40mg/kg (for the minimum dose that the animal 100% of mensuration falls asleep in advance), taking righting reflex loss as time for falling asleep, be sleep time (min) from righting reflex loss to recovery time.After result administration, each treated animal compares with blank group, does not occur significant difference the length of one's sleep, and data are in table 7.

Table 7 is on the pentobarbital sodium in mice impact of the length of one's sleep

Group	Dosage (mg/kg)	Number of animals	The number of elements of falling asleep
				Blank group	-	10	30.73±1.35
High dose group	2450	10	31.45±2.26
				Middle dosage group	1220	10	29.13±2.77
Low dose group	409	10	30.66±2.41

Note: * P < 0.05 (with model group comparison).

5) impact on mice coordination exercise

Get surface and twine white glue cloth, diameter 1cm, long 60cm iron staff, vertically fixing.1h after gastric infusion, is positioned over top by mice, and it is creeped naturally, observes mice coordination exercise ability, and records it and climb to the bottom time.The each treated animal of result compares with blank group, all creeps steadily, and the phenomenon that slides and drop, there is not significant difference in the pole-climbing time, data are in table 8.

The impact of table 8 on the mice pole-climbing time

Group	Dosage (mg/kg)	Number of animals	The pole-climbing time (s)
				Blank group	-	10	11.88±0.60
High dose group	2450	10	12.25±0.48
				Middle dosage group	1220	10	12.23±0.52
Low dose group	409	10	11.56±0.83

Note: * P < 0.05 (with model group comparison).

6) impact on Wistar Rat Cardiovascular system

With pentobarbital sodium intraperitoneal injection of anesthesia animal, separate common carotid artery and with System of organism signal linkage record blood pressure and heart rate; Trace electrocardiogram with limb lead (II leads).Before gastric infusion, record once respectively, then after gastric infusion, record These parameters.After result administration, each treated animal compares with blank group, and significant difference does not appear in the every numerical value of electrocardio and blood pressure, and data are in table 9 and table 10.

Table 9 is on the cardiac electrical impact of rat

Group	Dosage (mg/kg)	Number of animals	Heart rate (beat/min)	Maximum (mV)	Minima (mV)
						Blank group	-	8	367.53±21.34	0.41±0.08	-0.26±0.10
High dose group	1701	8	365.70±14.04	0.47±0.10	-0.28±0.14
						Middle dosage group	850.5	8	365.39±43.58	0.48±0.03	-0.32±0.16
Low dose group	283.5	8	365.24±56.27	0.39±0.07	-0.28±0.19

Note: * P < 0.05 (with model group comparison).

The impact of table 10 on rat blood pressure

Group	Dosage (mg/kg)	Number of animals	Systolic pressure (mmHg)	Diastolic pressure (mmHg)
					Blank group	-	8	72.95±12.74	45.52±13.23
High dose group	1701	8	68.92±9.33	44.54±9.18
					Middle dosage group	850.5	8	69.80±21.32	44.44±21.32
Low dose group	283.5	8	86.91±17.36	59.61±15.44

Note: * P < 0.05 (with model group comparison).

7) impact on Wistar rat respiratory system in rats

System of organism signal records the respiratory frequency of animal, on average exhale peak pressure and average suction paddy is pressed.After result administration, each treated animal compares with blank group, and significant difference does not all appear in respiratory frequency, expiration peak value and air-breathing valley, and data are in table 11.

The impact of table 11 on respiratory system in rats

Group	Dosage (mg/kg)	Number of animals	Frequency (beat/min)	Maximum (ml/s)	Minima (ml/s)
						Blank group	-	8	123.20±12.11	0.13±0.14	0.02±0.14
High dose group	1701	8	119.35±6.85	0.24±0.13	0.11±0.13
						Middle dosage group	850.5	8	117.49±17.16	0.24±0.10	0.12±0.11
Low dose group	283.5	8	113.60±1.78	0.26±0.17	0.17±0.19

Note: * P < 0.05 (with model group comparison).

The present composition is learned in indices detection at general pharmacology, and each administration group is organized relatively with blank, unknown significance difference.Illustrate that the present composition is to nervous system, cardiovascular and respiratory system do not produce harmful effect, can be applicable to clinical.

The pharmacodynamics test of research-compositions treatment rats with myocardial ischemia of test example 3 present composition pharmacodynamicss.

Copy myocardial infarction model according to literature method.Adopt intraperitoneal injection of anesthesia with 1% pentobarbital sodium (0.04g/kg), dorsal position is fixed on operating-table.Record standard II lead electrocardiogram, row tracheotomy, be connected with artificial respirator, open breast and expose heart, between aorta circular cone and left auricle, prop up ligation left coronary artery (a sham operated rats not ligation of threading) with surgical thread through the left chamber of arteria coronaria, heart resets, after drum lung, close rapidly thoracic cavity, penicillin is embrocated in wound, prevention traumatic infection.After closing breast, observe rats breathing recovery situation, can off line put back to mouse cage if any autonomous respiration.Occur that with ECG ST section the back of a bow raises as myocardial ischemia modeling success.After surrounding, form chronic myocardial ischemia model.

1) impact on rats with myocardial ischemia S-T section after coronary artery ligation

Rats with myocardial ischemia after screening is evenly divided at random to 5 groups except sham operated rats, and 6 groups of sham operated rats, model group, positive drug control group, high dose group, middle dosage group, low dose group,, 72, raise totally in the lump by 12 every group.Administration group gavage give corresponding dosage containing drug solns, blank group, model group gavage give the distilled water of equivalent, once a day, totally 10 days.After last administration, respectively organize 1% pentobarbital sodium (40mg/kg) intraperitoneal anesthesia for rat, connect biological function experimental system, monitoring limb lead electrocardiogram, measures S-T section.Between group, relatively represent with t inspection, result with represent.The each administration group of result and model group comparison can improve S-T section raises, and data are in table 12.

The impact that table 12 changes ECG ST section

Group	Number of animals (only)	Dosage (mg/kg)	ST section changes (mv)
				Blank group	12	-	0.07±0.03
Model group	12	-	0.23±0.02*
				High dose group	12	1701	0.12±0.02#
Middle dosage group	12	850.5	0.12±0.02#
				Low dose group	12	283.5	0.18±0.01#
Positive control	12	324	0.12±0.02#

Note: with the comparison of blank group, * P < 0.05; With model group comparison, #P < 0.05.

2) impact on serum cardiac enzyme

Laboratory animal grouping and administration are the same.In last administration 1 hour, eyeball is got blood, press test kit description operation, and detect serum creatine kinase (CK), lactic acid dehydrogenase (LDH), aspartate amino transferase (AST) activity, record data with semi-automatic biochemical analyzer.Between group, relatively represent with t inspection, result with represent.The each administration group of result and model group comparison, the present composition can obviously reduce creatine kinase in serum (CK), lactic acid dehydrogenase (LDH), aspartate amino transferase (AST), and data are in table 13.

The impact of table 13 on serum cardiac enzyme

3) impact on myocardial metabolism product

Laboratory animal grouping and administration are the same.In last administration 1 hour, eyeball was got blood, by test kit description operation, and by 756PC type ultraviolet-uisible spectrophotometer detection serum lactic acid (LD), free fatty (NEFA), Na-KATP enzyme index.Record data.Between group, relatively represent with t inspection, result with represent.The each administration group of result and the comparison of model group group, present composition atpase activity reduces, and lactic acid and free fatty acid content reduce, and data are in table 14.

The impact of table 14 on myocardial metabolism product index

4) impact on Plasma free radical damage criterion

Laboratory animal grouping and administration are the same.In last administration 1 hour, eyeball is got blood, press test kit description operation, and detect malonaldehyde (MDA), always superoxide dismutase (SOD), glutathion peroxidase (GSH-Px) index with 756PC type ultraviolet-uisible spectrophotometer.Record data.Between group, relatively represent with t inspection, result with represent.The each administration group of result and model group comparison, the each administration group of the present composition can make malonaldehyde (MDA) content reduce, the activity of total superoxide dismutase (SOD), glutathion peroxidase (GSH-Px) raises, and data are in table 15.

The impact of table 15 on myocardial metabolism product index

5) impact on hemorheology index

Laboratory animal grouping and administration are the same.In last administration 1 hour, abdominal aortic blood, sent with hospital laboratory and checks.Between group, relatively represent with t inspection, result with represent.The each administration group of result and model group comparison, the each administration group of present composition whole blood viscosity reduces, plasma viscosity reduces, packed cell volume reduces, erythrocyte sedimentation rate reduces, and its energy inhibiting erythrocyte aggregation is described, reduces erythrocyte fragility, strengthens its morphotropism, in table 16.

6) impact on hemodynamic index

Laboratory animal grouping and administration are the same.In last administration 1 hour, rat is lain on the back and is fixed on operating-table, cut skin of neck, separate right carotid, row arterial cannulation, inserts right carotid artery by the conduit that contains 0.1% heparin sodium liquid, insert left ventricle through right common carotid artery, judge according to the change of pressure figure shown in display whether intubate enters ventricle, cardiac catheter Bonding pressure transducer, inputs signal in physiograph.And recorded heart rate (HR), left ventricular systolic pressure (LVSP), maximal ascending rate of internal pressure of left ventricle (+dp/dtmax), left ventricular diastolic pressure (LVDp), left ventricular end-diastolic pressure (LVDEp), maximal descending rate of internal (dP/dtmax).Record data.Between group, relatively represent with t inspection, result with represent.The each administration group of result and model group comparison, the each administration group of the present composition can make that heart rate is tending towards normally, left ventricular systolic pressure raises, left chamber diastolic pressure reduces, the maximum climbing speed in left chamber raises, the maximum fall off rate in left chamber raises, eventually last diastolic pressure reduces, in table 17.

7) impact on cardiac muscular tissue's pathomorphology

Laboratory animal grouping and administration are the same.In last administration 1 hour, take out heart, send in hospital pathology department and check.The results are shown in Table 18.

8) to myocardial necrosis area estimation

Laboratory animal grouping and administration are the same.In last administration 1 hour, put to death rat, with the fixing presternum of tweezers, cut off breast bone, cut whole heart, remove blood stains with the normal saline flushing of pre-cooling, filter paper suck dry moisture, weigh whole-heartedly,-30 DEG C of freezing half an hour, under heart ligature, be cut into along left chamber major axis is parallel the myocardium sheet that 1～2mm is thick, evenly be cut into 4～5, be placed in the TTC solution (the PBS buffer of pH value 7.4) of 1% concentration, 37 DEG C of temperature are incubated 15min, water rinses unnecessary dyestuff, filter paper suck dry moisture, visible normal myocardium is red, ischemic myocardium is canescence, cut off infarcted myocardium, weigh, calculate infarction size (infarcted region accounts for heavy whole-heartedly percentage ratio).Record data, relatively represent with t inspection between group, result with represent.The each administration group of result and model group comparison, the each administration group of the present composition can make myocardial infarction area obviously reduce, and difference has significance, in table 19.

Table 16 is on hemorheological impact

Table 17 is on hemorheological impact

The impact of table 18 on cardiac muscular tissue's pathomorphology

The impact of table 19 on myocardial infarction area

Group	Number of animals (only)	Dosage (mg/kg)	Myocardial infarction area (%)
				Model group	12	-	13.01±0.63
High dose group	12	1701	7.00±0.83#
				Middle dosage group	12	850.5	5.89±0.39#
Low dose group	12	283.5	9.50±1.19#
				Positive control	12	324	7.08±0.74#

Conclusion (of pressure testing): improvement and therapeutical effect that the present composition has the change of myocardial infarction and ischemia model rat indices.

The pharmacodynamics test of research-compositions treatment hyperlipidemia rats of test example 4 present composition pharmacodynamicss.

1) impact on hyperlipidemia rats lipid metabolism level

High lipid food composition and preparation: 3% cholesterol, 10% Adeps Sus domestica, 5% dried hen egg yolk, 0.3% sodium cholate, 0.2% methylthiouracil, 81.5% normal feedstuff.10kg high lipid food adds 1～2kg white sugar.In proportion first by cholesterol, dried hen egg yolk, sodium cholate, propylthiouracil, white sugar ground and mixed, then adds in warm Adeps Sus domestica, then mixes thoroughly with normal feedstuff, to obtain final product.

Modeling: 72 SD rat normal diets are fed one week.12 of random selections are organized Mus as blank, and the labelling of weighing gives normal diet and feeds.All the other 60 Mus labelling of weighing, gives high lipid food and feeds, and all freely drinks water, and feeds altogether the foundation of carrying out hyperlipemia model for 4 weeks.The rat body weight of surrounding after recording start modeling respectively, and get blood 1.5～2ml in centrifuge tube in 4th week end eyeground vein clump, leave standstill 3500rmin ^-1centrifugal 15min, separation of serum, the content of measuring T-CHOL in its Diagnostic Value of Fasting Serum (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), HDL-C (HDL-C) with semi-automatic biochemical analyzer, detection method is in accordance with the description of corresponding reagent box.T-CHOL in serum (TC), triglyceride (TG) content during all apparently higher than blank group, illustrate that hyperlipidemia animal model is successfully prepared.Start gastric infusion next day, positive drug is with 0.324g/kg gastric infusion, and the present composition is high, in, low dose group is respectively with 1.701g/kg, 0.8505g/kg, 0.2835g/kg gastric infusion, totally 10 days.During administration, continue to give high lipid food.In last administration, after 1 hour, socket of the eye angular vein clump is got blood 2.5ml, and 4 DEG C leave standstill, 3500rmin ^-1centrifugal 15min, separation of serum, in accordance with corresponding reagent box description, detect T-CHOL (TC), triglyceride (TG) level and Apolipoprotein A1 (ApoA1) and apolipoprotein B (ApoB) level with semi-automatic biochemical analyzer.Between group, relatively represent with t inspection, result represents with X ± S.The each administration group of result is compared with model group and can obviously be reduced the content of T-CHOL in serum (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and apolipoprotein B (ApoB), obviously increase HDL-C (HDL-C) and Apolipoprotein A1 (ApoA1) content, in table 20.

The impact of table 20 on hyperlipemia lipid metabolism level

2) impact on hyperlipidemia rats level of lipid

Laboratory animal grouping and administration are the same.In last administration 1 hour, socket of the eye angular vein clump was got blood, and 4 DEG C leave standstill, 3500rmin ^-1centrifugal 15min, separation of serum, in accordance with corresponding reagent box description, detect total superoxide dismutase (SOD) with ultraviolet-uisible spectrophotometer, malonaldehyde (MDA), nitric oxide (NO) and glutathion peroxidase (GSH-Px) level.Between group, relatively represent with t inspection, result with represent.The each administration group of result and model group comparison can obviously reduce malonaldehyde (MDA) content, obviously increase the content of nitric oxide (NO), raise total superoxide dismutase (SOD), glutathion peroxidase (GSH-Px) activity, in table 21.

The impact of table 21 on hyperlipemia level of lipid

3) impact on hyperlipidemia rats hemorheology index

Laboratory animal grouping and administration are the same.In last administration 1 hour, abdominal aortic blood, sent with hospital laboratory and checks.Between group, relatively represent with t inspection, result with represent.The each administration group of result and model group compare whole blood viscosity reduction, plasma viscosity reduction, packed cell volume reduction, erythrocyte sedimentation rate reduction, and its energy inhibiting erythrocyte aggregation is described, reduction erythrocyte fragility, strengthens its morphotropism, in table 22.

The impact that table 22 becomes hyperlipemia blood flow

4) impact on hyperlipidemia rats liver organization pathomorphism

Laboratory animal grouping and administration are the same.After last administration 1h, hepatic tissue is carried out to routine and draw materials, 10% formalin solution is fixed, conventional H E dyeing.Optical microphotograph Microscopic observation liver organization form.This send in hospital pathology department and checks.The results are shown in Table 23.

The impact of table 23 on cardiac muscular tissue's pathomorphology

Conclusion (of pressure testing): the improvement that the change of Hyperlipemia model rat indices is had and therapeutical effect.

The research of test example 5 present composition long term toxicity tests.

Experiment purpose: observe repetition per os and give the toxic reaction that the present composition produces rat, the symptom and the order of severity that occur, and provide the target organ of toxicity and the degree of reversibility of infringement thereof, determine nontoxic amounts of reactants, to evaluate the safety of present composition long-term prescription, provide reference for drafting clinical trial dosage and observation index.

Experimental animal: cleaning agent SD rat, male and female half and half, 120 of quantity, 80 grams～110 grams of body weight (6～9 week age).

Animal grouping: animal, first the normal raising of laboratory 1 week, is rejected disease Mus and abnormal Mus, gets 120 of rats, is divided at random 4 groups by body weight, 30 every group, male and female half and half.Be respectively Normal group, high, medium and low three the dosage groups of the present composition.

Dosage: the maximum tolerated dose of mice is 16g/Kg, does not occur toxic reaction, therefore, in the case, according to rat high dose group, administration should be greater than clinical 50 times of above principle design dosages.Present composition high dose group rat is pressed 2.7g/kg administration (60 times of clinical plan dosage), middle dosage group rat is pressed 1.35g/kg gastric infusion (30 times of clinical plan dosage), low dose group rat is pressed 0.45g/kg gastric infusion (10 times of clinical plan dosage), volume is 2ml/100g, and Normal group gives same volume distilled water.

Administration time: on every Mondays to 8:30 in the morning left and right administration of Saturday 1 time, not administration on Sunday.Successive administration 6 months.

Observation index

1) general observation of symptoms

1. observing time: before administration,, with the each cage observation of every morning convalescent period, during administration, the upper and lower noon observes simultaneously.

2. observe number of cases: before administration, all buy animal, total Test animal after administration.

3. observe frequency: finish from buying day to convalescent period, once a day.

4. observed content: observe outward appearance, quilt hair, gait, the behavioral activity of animal, to the reaction of sound, have atremia, spasm, whether steadily to breathe, have or not abnormal.When administration and after administration, except noting observing above-mentioned reaction, also want special survey to have or not drowsiness, vomiting, feces shape and color etc.Administration also will be observed the reactivity while arresting animal simultaneously, eye, nose, mouth state around, and hypogastric region, anus place have or not manure contamination, and skin color, has or not wound, tumor etc., the state of breast abdominal part, muscle tonus degree etc.

2) body weight determination

1. assay method: adopt electronic balance to measure.

2. measure number of cases: before administration, all buy animal, total Test animal after administration.

3. measure frequency: buy day and grouping day (before administration) is each surveys once, during administration and convalescent period claim weekly body weight one time, in addition, carrying out when dissected and first measuring the body weight of animal.

3) food ration is measured

1. assay method: every group have altogether supply deduct altogether surplus be every group every day food ration, every group every day food ration be every group every animal food ration every day divided by every treated animal number.

2. measure number of times: before administration, measure once, during administration, measure weekly once with convalescent period.

3) hematology and blood biochemical are learned and are detected

1. detect period: finish each detection once in administration 3 months, 6 months (administration finishes) and convalescent period.

2. detect number of cases: administration 3 months, administration 6 months (administration finishes) and convalescent period finish each group and detect respectively 10.

3. blood-sampling method: take a blood sample front rat fasting more than 14 hours, urethane 0.1g/100g body weight intraperitoneal injection of anesthesia, special blood-drawing pipe and blood taking needle are taken a blood sample from ventral aorta.Hematology's test item and method are in table 24.

Table 24 hematology test item and method

Project	Anticoagulant	Algoscopy	Use instrument
				Red blood cell count(RBC) (RBC)	EDTA-K2	Electric-resistivity method	Animal blood analyser
Hemoglobin (HGB)	EDTA-K2	Photoelectric colorimetry	Animal blood analyser
				Hematid specific volume (HCT)	EDTA-K2	Histogram calculation method	Animal blood analyser
Mean corpuscular volume (MCV) (MCV)	EDTA-K2	Histogram calculation method	Animal blood analyser
				Average hemoglobin content (MCH)	EDTA-K2	Histogram calculation method	Animal blood analyser
Mean corpuscular hemoglobin concentration (MCHC)	EDTA-K2	Histogram calculation method	Animal blood analyser
				Numeration of leukocyte (WBC)	EDTA-K2	Electric-resistivity method	Animal blood analyser
Neutrophilic granulocyte percentage ratio (GRAN%)	EDTA-K2	Histogram calculation method	Animal blood analyser
				Lymphocyte percentage ratio (LYMPH%)	EDTA-K2	Histogram calculation method	Animal blood analyser
Mononuclear cell percentage ratio (MONO%)	EDTA-K2	Histogram calculation method	Animal blood analyser
				Platelet count (PLT)	EDTA-K2	Histogram calculation method	Animal blood analyser

Note: when finding that hemopoietic system is had to the time of impact, should further carry out the inspection of bone marrow.

4. biochemistry detection blood sample processing: after ventral aorta blood sampling, room temperature leaves standstill about 1 hour, centrifugal 3000rpm, 10min, separation of serum.Serum, as do not detected the same day, is put into cryopreservation tube by serum, and-80 DEG C frozen.Concrete test item and method are referring to current edition novel technique guideline.Concrete test item and method are in table 25.

Table 25 blood parameters test item and method

Project	Algoscopy	Use instrument
			Glutamate pyruvate transaminase (ALT)	Continuous monitoring method	Automatic clinical chemistry analyzer
Glutamic oxaloacetic transaminase, GOT (AST)	Continuous monitoring method	Automatic clinical chemistry analyzer
			Alkali phosphatase (ALP)	Continuous monitoring method	Automatic clinical chemistry analyzer
Creatine phosphokinase (CK)	Continuous monitoring method	Automatic clinical chemistry analyzer
			Blood urea nitrogen (BUN)	Dynamic method	Automatic clinical chemistry analyzer
Creatinine (CREA)	Dynamic method	Automatic clinical chemistry analyzer
			Total protein (TP)	End-point method	Automatic clinical chemistry analyzer
Albumin (ALB)	End-point method	Automatic clinical chemistry analyzer
			Globulin (GIB)	End-point method	Automatic clinical chemistry analyzer
Blood glucose (GLU)	End-point method	Automatic clinical chemistry analyzer
			Total bilirubin (TBIL)	End-point method	Automatic clinical chemistry analyzer
T-CHOL (CHOL)	End-point method	Automatic clinical chemistry analyzer
			Triglyceride (TG)	End-point method	Automatic clinical chemistry analyzer
Potassium ion (K ⁺) concentration	Electrode method	Electrolyte analyser
			Sodium ion (Na ⁺) concentration	Electrode method	Electrolyte analyser
Chloride ion (Cl ^-) concentration	Electrode method	Electrolyte analyser

5) processing of death or dying animal

1. processing requirements: find as early as possible, dissect in time, seek as possible reason.

2. dying animal: record dying time, body weight, symptom.

3. dead animal: dead time, body weight, the postmortem finding of naked eye of finding of record.

4. biochemical analysis: get blood before dying sacrifice of animal and carry out hematology and the biochemical detection of blood.

5. pathologic finding: except confirming that other dying and dead animal all will carry out histopathologic examination due to animal dead due to gavage.

6) dissect

1. anatomic method: anaesthetize with urethane 0.1g/100g body weight lumbar injection.Cut open before inspection, the situation of each animal is checked comprehensively; When dissected carries out one by one by system, observes between internal organs color and luster, form, position relationship and internal organs and has or not and be adhered; Organ surface and digestive tube inner membrance have or not congestion, hyperemia, petechia, edema, ulcer; Thoracic cavity, abdominal cavity, pericardial cavity have or not the pathological changes such as hydrops, as note abnormalities, its position of accurate recording and pathological change type in the remarks column of dissecting finding record.

2. between when dissected: same to detection time

3. dissect number of cases: administration 3 months, administration 6 months (administration finishes) and 1 month convalescent period finish each group and dissect respectively 10.

7) organ weights is measured

Assay method: adopt electronic balance to weigh respectively the absolute weight (simultaneously calculating organ coefficient) of 13 internal organs (following table).

1. measure period: when dissected is measured

2. measure number of cases: number of cases is identical with dissecting

3. internal organs table:

Sequence number

1

2

3

4

5

6

7

Internal organs

Brain (brain, cerebellum)

Heart

Liver

Spleen

Lungs

Kidney

Adrenal gland

Internal organs

Thymus

Uterus

Ovary

Testis

Epididymis

Prostate

8) histopathologic examination

All animals all takes out institute's requirement organs and tissues by current edition novel technique guideline, and abnormal tissue and internal organs are thought in perusal.Specimen is fixed on 10% formalin fixative interior one week, repaiies piece and again fixes 12 hours afterwards, and tissue (the first decalcification of skeletal tissue) is with gradient alcohol dehydration, dimethylbenzene is transparent, paraffin embedding, paraffin slicing machine section, H.E. dyeing, neutral gum sealing, light microscopy checking.Record kind and the degree of pathological changes.As a certain tissue generation pathological change, other this tissue of dosage treated animal also should carry out histopathological examination to determine dose-response relationship.

Observe organs and tissues table:

Experimental result

1) ordinary circumstance: each treated animal is during administration and surrounding convalescent period, and outward appearance sign, behavioral activity are normal, and urine, excrement character are without difference, and diet is normal, and each treated animal body weight increases and decreases without significant difference, and group difference is not obvious.Food-intake is in table 26, table 27 and table 28.On the impact of body weight the results are shown in Table 29, table 30 and table 31.

3 months rats eating amounts (g/ days /) of table 26 medication

3～6 months rats eating amounts (g/ days /) of table 27 medication

The quantitative change of table 28 convalescent period rats eating (g/ days/only)

2) hematological indices check result

Rat administration after 90 days and 180 days 24 hours, respectively organize each 10 of rat, ventral aorta blood sampling, collects anticoagulated whole blood, detects hematology's indices with blood counting instrument.After administration 24 hours the last time, respectively organize rat and live and kill 10 rats that retain after 10 and stop administration, normally raise, continue to observe surrounding and all live and kill again.Under the identical condition of situation when with medication 6 months, the index of detection is also identical.

Administration after 90 days result show: with the comparison of blank group, high dose group erythrocyte sum (RBC) significantly reduces, and mean corpuscular volume (MCV) enlarges markedly, and mononuclear cell percentage ratio (MONO%) significantly declines; Low dose group lymphocyte percentage ratio (LYMPH%) significantly declines.All the other indexs: as packed cell volume (HCT), average hemoglobin content (MCH), mean corpuscular hemoglobin concentration (MCHC), platelet count (PLT), hemoglobin (HGB) content, total white blood cells (WBC) and neutrophil cell percentage ratio (GRAN%) differential counting thereof, each administration group and the comparison of blank group, without significant difference, the results are shown in Table 32 and table 33.

Table 32 medication 3 months variation to rat serum routine

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
WBC(×10 ⁹/L)	11.62±2.34	13.37±4.97	13.00±3.87	9.69±1.84
					RBC(×10 ¹²/L)	8.69±1.07	7.33±1.05*	8.65±1.00	8.40±1.45
HGB(g/L)	152.00±13.42	135.50±21.59	147.70±13.62	146.50±17.44
					HCT(％)	42.07±3.95	37.87±5.80	41.56±4.78	40.90±6.86
MCV(fL)	48.61±2.70	51.62±2.34*	48.09±1.96	48.73±1.51
					MCH(pg)	17.57±0.98	18.44±0.88	17.12±0.88	17.60±1.37
MCHC(g/L)	361.70±14.29	357.50±19.48	355.60±18.97	361.50±26.56
					PLT(×10 ⁹/L)	545.30±59.03	570.20±111.71	564.00±72.03	542.00±84.41

Note: with the comparison of blank group, * P < 0.05.

Table 33 medication 3 months variation to rat WBC differential counting (%)

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
GRAN％	19.38±9.39	29.12±20.01	23.94±12.39	23.36±13.18
					LYMPH％	58.71±10.18	48.46±21.78	47.94±18.71	40.51±21.65*
MONO％	21.91±1.99	12.42±6.47***	18.12±7.66	16.13±9.05

Note: with the comparison of blank group, * P < 0.05; * * P < 0.001.

Administration after 180 days result show: with the comparison of blank group, high and low dose group leukocyte (WBC) sum obviously reduces (P < 0.05), and wherein high dose group reduces the most obvious; In, low dose group mononuclear cell percentage ratio (MONO%) significantly raises; High, medium and low dosage group erythrocyte (RBC) sum, hemoglobin (HGB) content and packed cell volume (HCT) significantly reduce, wherein: in, low dose group reduction is the most obvious; The average hemoglobin content of middle dosage group (MCH) and mean corpuscular hemoglobin concentration (MCHC) obviously raise; All the other indexs: as mean corpuscular volume (MCV), platelet count (PLT), total white blood cells (WBC) and differential counting thereof [lymphocyte percentage ratio (LYMPH%) and neutrophil cell percentage ratio (GRAN%)], each administration group and the comparison of blank group, no significant difference between group, the results are shown in Table 34 and table 35.

Table 34 medication 6 months variation to rat serum routine

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
WBC(×10 ⁹/L)	14.65±2.66	8.88±2.79***	12.42±4.08	10.75±3.52*
					RBC(×10 ¹²/L)	10.62±1.46	8.55±1.70**	8.33±0.68***	8.08±0.71***
HGB(g/L)	176.90±14.98	97.80±68.76**	152.80±6.76***	126.00±25.14***
					HCT(％)	52.10±4.87	42.35±8.24**	40.61±3.54*	39.52±2.45***
MCV(fL)	49.19±2.78	49.59±2.23	48.78±2.78	49.11±3.84
					MCH(pg)	16.98±1.29	13.35±6.30	18.41±1.29*	15.63±3.14
MCHC(g/L)	339.60±139.90	266.38±121.87	377.5.00±19.58***	319.10±62.71
					PLT(×10 ⁹/L)	843.80±141.31	726.89±135.95	854.00±110.13	813.90±150.71

Note: with the comparison of blank group, * P < 0.05; * P < 0.01; * * P < 0.001.

Surrounding convalescent period result shows: each administration treated animal hematological indices and the comparison of blank group, no significant difference between group, the results are shown in Table 36 and table 37.

Table 35 medication 6 months variation to rat WBC differential counting (%)

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
GRAN％	22.38±10.95	16.14±12.11	22.31±7.73	21.01±11.05
					LYMPH％	62.21±17.39	61.20±23.35	61.89±8.16	63.94±11.09
MONO％	12.41±2.22	12.66±5.29	15.80±3.02*	15.05±2.58*

Note: with the comparison of blank group, * P < 0.05.

The variation of table 36 convalescent period rat serum routine

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
WBC(×10 ⁹/L)	14.29±4.99	10.19±4.14*	13.14±3.94	11.97±2.87
					RBC(×10 ¹²/L)	9.04±1.24	8.50±0.59	8.73±0.70	8.48±1.15
HGB(g/L)	164.4±15.06	156.7±6.82	159.00±7.39	160.30±15.75
					HCT(％)	43.45±4.41	41.88±2.16	42.43±2.55	41.70±4.64
MCV(fL)	48.27±2.48	49.40±3.13	48.72±1.97	49.38±3.11
					MCH(pg)	18.29±1.26	18.50±1.39	18.27±0.82	19.05±1.61
MCHC(g/L)	378.70±10.18	374.40±11.25	375.10±9.97	385.20±12.60
					PLT(×10 ⁹/L)	820.80±86.69	749.80±145.39	819.80±165.79	794.10±120.85

Note: with the comparison of blank group, * P < 0.05.

The variation of table 37 convalescent period rat WBC differential counting (%)

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
GRAN％	26.90±12.30	31.83±10.98	22.33±9.37	23.07±10.12
					LYMPH％	61.08±11.33	55.85±11.66	65.29±8.75	63.88±9.78
MONO％	12.02±3.24	12.32±1.73	12.33±1.34	13.04±1.57

3) blood parameters testing result

Through the rat administration of getting anticoagulated whole blood above-mentioned each group of 10 rats after 90 days and 180 days, continue to collect blood, separation of serum, with automatic clinical chemistry analyzer, detects every biochemical indicator.After administration 24 hours the last time, respectively organize rat and live and kill 10 rats that retain after 10 and stop administration, normally raise, continue to observe surrounding and all live and kill again.Under the identical condition of situation when with medication 6 months, the index of detection is also identical.

Administration after 90 days result show: with the comparison of blank group, in, low dose group total protein (TP) content significantly raises, wherein: low dose group raises obviously; Low dose group albumin content (ALB) significantly raises; High, medium and low dosage Histaglobin (GIB) content significantly raises, wherein: low dose group raises the most obvious; High, middle dosage group sodium ion (Na ⁺) concentration significantly raises; High dose potassium ion (K ⁺) concentration significantly raises.All the other indexs: as glutamate pyruvate transaminase (ALT), glutamic oxaloacetic transaminase, GOT (AST), blood urea nitrogen (BUN), creatinine (CREA), blood glucose (GLU), total bilirubin (TBIL), T-CHOL (CHOL), triglyceride (TG), chloride ion (Cl ^-) relatively group difference is not obvious for the each administration group of concentration and blank group, does not make significant difference, and the results are shown in Table 38.

Administration after 180 days result show: with the comparison of blank group, in, low dosage aspartate amino transferase (GOT), creatinine (CREA) content significantly reduce, wherein: low dose group reduces the most obvious; In, low dose group total protein (TP) content significantly raises, in, low dose group albumin content (ALB) significantly raises; High and low dose group sodium ion (Na ⁺), chloride ion (Cl ^-) concentration significantly raises, wherein: low dose group raises the most obvious; All the other indexs: relatively group difference is not obvious as alanine aminotransferase (ALT), alkali phosphatase (ALP), blood urea nitrogen (BUN), blood glucose (GLU), total bilirubin (TBIL), T-CHOL (CHOL), the each administration group of triglyceride (TG) and blank group, do not make significant difference, the results are shown in Table 39.

Surrounding convalescent period result shows: with the comparison of blank group, middle dosage group alkali phosphatase (ALP) content significantly reduces; High and low dose group total protein (TP) content significantly raises; High and low dose group albumin (ALB) content significantly raises; High and low dose Histaglobin (GIB) content significantly raises, wherein: low dose group raises the most obvious; High dose group sodium ion (Na ⁺) concentration significantly raises; All the other indexs: as glutamate pyruvate transaminase (ALT), glutamic oxaloacetic transaminase, GOT (AST), blood urea nitrogen (BUN), creatinine (CREA), creatine phosphokinase (CK), blood glucose (GLU), total bilirubin (TBIL), T-CHOL (CHOL), triglyceride (TG), potassium ion (K ⁺), chloride ion (Cl ^-) relatively group difference is not obvious for the each administration group of concentration and blank group, does not make significant difference, and the results are shown in Table 40.

4) system postmortem and histopathologic examination's result

Administration after 90 days result show: each administration group organ coefficient and blank group are substantially close, illustrate that medicine has no significant effect organ weights, in table 41.The internal organs histopathology microscopy of blank group and each administration group rat, by the statistical result of sxemiquantitative scoring.Result shows: except distilled water matched group and medication respectively organize minority example see have the slight heart, lung, kidney interstitial, small intestinal hyperemia and fatty degeneration of liver, and the enlargement of minority example spleen blood stasis, lymph node chronic inflammation, all do not find the pathological changes relevant with drug toxicity, and the each internal organs of the animal changing are dispersed in each group, there is no statistical significance.Result of the test is in table 42.

Administration after 180 days result show: the organ coefficient of three dosage groups and blank group are basically identical, illustrate that medicine has no significant effect organ weights, in table 43.The internal organs histopathology microscopy of blank group and each administration group rat, by the statistical result of sxemiquantitative scoring.Result shows: except distilled water matched group and medication respectively organize minority example see have the slight heart, liver, bladder, lung and the enlargement of minority example spleen blood stasis, lymph node chronic inflammation, all do not find the pathological changes relevant with drug toxicity, and the each internal organs of the animal changing are dispersed in each group, there is no statistical significance.Experimental result is in table 44.

Surrounding convalescent period result shows: internal organs are carried out comprehensively to gross necropsy meticulously, no abnormal organ-tissue.Organ coefficient and the matched group of 10 organs and tissues such as each treated animal internal organs heart, liver, spleen, lung, kidney, adrenal gland, uterus, brain, ovary, thymus are substantially close, organ weights are had no significant effect, in table 45.The internal organs histopathology microscopy of blank group and each administration group rat, by the statistical result of sxemiquantitative scoring.Result shows: except distilled water matched group and medication respectively organize minority example see have the slight heart, small intestinal, liver, kidney, lung and the enlargement of minority example spleen blood stasis, lymph node, stomach, large intestine chronic inflammation, all do not find the pathological changes relevant with drug toxicity, and the each internal organs of the animal changing are dispersed in each group, there is no statistical significance.Experimental result is in table 46.

Table 38 medication 3 months impact on rat blood biochemical indicator

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
ALT(U/L)	80.30±53.66	61.50±18.82	57.70±16.26	67.00±18.97
					AST(U/L)	218.20±143.70	148.20±36.96	151.60±23.32	174.00±38.28
ALP(U/L)	116.2±46.21	186.88±52.59	128.70±64.20	150.70±60.00
					CK(U/L)	724.9±521.27	420.9±127.75	753.30±751.18	835.8±623.10
TP(g/L)	74.08±4.05	80.26±9.01	80.18±5.58*	84.47±6.48***
					ALB(g/L)	32.69±2.29	32.24±5.56	34.21±2.32	35.08±1.62*
GIB(g/L)	41.39±2.74	48.02±8.90*	45.97±4.93*	49.39±5.66***
					BUN(mmol/L)	6.37±1.22	7.78±2.42	7.26±1.44	6.55±1.66
CREA(μmol/L)	39.04±3.83	37.21±6.71	35.50±3.83	39.77±6.08
					GLU(mmol/L)	5.12±0.69	4.41±0.99	4.68±0.86	5.05±0.61
TG(mmol/L)	0.56±0.22	0.87±0.44	0.74±0.86	0.64±0.27
					CHOL(mmol/L)	1.78±0.32	1.63±0.30	1.93±0.35	1.86±0.27
TBIL(μmol/L)	0.60±0.37	0.62±0.40	0.66±0.37	0.45±0.24
					K+(mmol/L)	4.51±0.48	5.84±0.52*	5.02±0.92	5.47±1.37
Na+(mmol/L)	142.70±1.70	145.00±2.45*	146.00±4.62*	145.00±4.83
					Cl-(mmol/L)	101.40±1.96	101.40±2.55	101.50±2.64	101.80±2.39

Note: with the comparison of blank group, * P < 0.05; * * P < 0.001.

Table 39 medication 6 months impact on rat blood biochemical indicator

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
GPT(U/L)	111.6±86.89	100.9±42.01	83.4±35.06	72.90±13.39
					GOT(U/L)	278.3±124.19	243.6±62.93	168.10±36.63	161.00±28.26**
AKP(U/L)	214.3±105.37	197.8±92.24	238.40±112.70	265.40±109.97
					CK(U/L)	2009.38±1301.08	1694.10±941.50	1449.30±712.6	1163.00±402.28
BUN(mmol/L)	6.53±1.17	5.80±1.42	5.73±0.34	5.54±1.13
					CR(μmol/L)	28.93±10.01	36.02±23.18	15.97±6.11**	18.85±8.38*
GLU(mmol/L)	10.15±5.33	10.89±3.96	5.20±0.53	5.08±0.55
					TPR(g/L)	67.2±4.07	63.20±4.10	81.29±6.83	77.97±4.22***
ALB(g/L)	31.84±2.53	31.80±10.54	36.13±3.51	35.38±1.98**
					TBIL(μmol/L)	0.82±0.58	0.55±0.91	1.22±0.67	1.13±0.32
CHO(mmol/L)	1.37±0.33	1.20±0.16	1.70±0.46	1.55±0.32
					TG(mmol/L)	1.09±0.35	1.04±0.41	1.21±0.53	0.82±0.20
K+(mmol/L)	5.13±0.66	4.98±0.20	5.02±0.43	4.65±0.33
					Na+(mmol/L)	137.92±1.70	140.15±2.11*	139.20±1.93	141.70±3.68**
Cl-(mmol/L)	99.77±1.41	100.32±2.57*	99.10±1.79	101.50±3.10**

The variation of table 40 convalescent period rat biochemical indicator

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
ALT(U/L)	88.60±49.49	67.90±15.85	112.00±100.61	63.70±11.66
					AST(U/L)	149.50±34.47	133.40±19.06	208.10±80.48	142.50±37.15
ALP(U/L)	211.70±82.95	184.50±35.35	128.7±29.28**	248.70±99.95
					CK(U/L)	635.30±196.11	873.90±292.15	838.20±305.99	926.30±729.52
TP(g/L)	70.70±3.68	76.22±2.64***	67.92±7.58	80.51±3.25
					ALB(g/L)	31.401±1.68	33.94±2.05***	30.62±3.24	34.61±2.86*
GIB(g/L)	38.77±3.02	42.28±2.74*	37.30±5.38	45.90±3.73***
					BUN(mmol/L)	7.67±1.39	7.65±0.95	6.79±1.44	7.28±0.96
CREA(μmol/L)	29.60±3.88	28.57±4.77	30.86±3.60	33.78±5.64
					GLU(mmol/L)	6.21±1.19	5.03±0.38	6.46±1.57	5.34±0.77
TG(mmol/L)	0.70±0.28	0.99±0.42	0.74±0.41	1.02±0.46
					CHOL(mmol/L)	1.37±0.32	1.45±0.22	1.47±0.32	1.68±0.35
TBIL(μmol/L)	0.23±0.13	0.27±0.19	0.26±0.36	0.32±0.23
					K+(mmol/L)	5.72±0.95	6.10±0.56	5.44±0.73	6.44±0.52
Na+(mmol/L)	143.00±2.21	145.00±0.94*	140.70±1.42	145.70±3.43
					Cl-(mmol/L)	101.70±1.16	102.10±1.29	99.8±1.81	101.70±2.19

Note: with the comparison of blank group, * P < 0.05; * * P < 0.001.

Table 41 medication 3 months is on the impact on Rats Organs and Tissues coefficient

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
Heart percentage of liveweight percent (%)	0.37±0.06	0.35±0.07	0.35±0.09	0.37±0.05
					Liver percentage of liveweight percent (%)	3.23±0.61	2.93±0.57	3.34±0.74	3.04±0.42
Spleen percentage of liveweight percent (%)	0.17±0.07	0.16±0.05	0.16±0.06	0.15±0.03
					Lung percentage of liveweight percent (%)	0.63±0.25	0.57±0.13	0.58±0.10	0.61±0.16
Kidney percentage of liveweight percent (%)	0.30±0.04	0.28±0.08	0.27±0.05	0.28±0.05
					Adrenal gland's percentage of liveweight percent (%)	0.0132±0.0050	0.0116±0.0050	0.01118±0.0031	0.0127±0.0027
Thymus percentage of liveweight percent (%)	0.0707±0.0511	0.682±0.0324	0.0843±0.0202	0.0772±0.0276
					Brain percentage of liveweight percent (%)	0.6748±0.0915	0.6748±0.1308	0.6776±0.1235	0.6935±0.0976
Testis percentage of liveweight percent (%)	0.48±0.10	0.41±0.11	0.41±0.13	0.45±0.03
					Epididymis percentage of liveweight percent (%)	0.23±0.08	0.25±0.05	0.23±0.04	0.23±0.04
Prostate percentage of liveweight percent (%)	0.15±0.06	0.15±0.04	0.15±0.05	0.14±0.04
					Uterus percentage of liveweight percent (%)	0.30±0.072	0.31±0.106	0.31±0.120	0.28±0.150
Ovary percentage of liveweight percent (%)	0.03±0.01	0.03±0.01	0.03±0.01	0.03±0.01

[0239]3 months rat pathological examination results of table 42 administration

Note: in table: "+" represents that internal organs have Minimal change; "-" represents that internal organs do not have obvious pathological change; Numeral (1～10) represents the number of elements of animal.

Table 43 medication 6 months is on the impact on Rats Organs and Tissues coefficient

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
Heart percentage of liveweight percent (%)	0.36±0.09	0.38±0.05	0.37±0.10	0.38±0.05
					Liver percentage of liveweight percent (%)	3.21±0.58	3.36±0.52	3.20±0.73	3.28±0.47
Spleen percentage of liveweight percent (%)	0.17±0.06	0.15±0.03	0.15±0.04	0.18±0.07
					Lung percentage of liveweight percent (%)	0.50±0.07	0.48±0.08	0.47±0.09	0.54±0.08
Kidney percentage of liveweight percent (%)	0.28±0.05	0.30±0.04	0.28±0.05	0.29±0.06
					Adrenal gland's percentage of liveweight percent (%)	0.0121±0.0045	0.0125±0.0054	0.0108±0.0046	0.0120±0.0056
Thymus percentage of liveweight percent (%)	0.0947±0.0330	0.0923±0.0317	0.0903±0.0272	0.1013±0.0412
					Brain percentage of liveweight percent (%)	0.5836±0.0913	0.5914±0.0488	0.5702±0.0791	0.5848±0.0985
Testis percentage of liveweight percent (%)	0.49±0.08	0.51±0.10	0.47±0.05	0.53±0.09
					Epididymis percentage of liveweight percent (%)	0.16±0.03	0.16±0.05	0.14±0.02	0.17±0.05
Prostate percentage of liveweight percent (%)	0.15±0.03	0.15±0.05	0.14±0.05	0.14±0.04
					Uterus percentage of liveweight percent (%)	0.20±0.06	0.18±0.07	0.20±0.02	0.24±0.10
Ovary percentage of liveweight percent (%)	0.02±0.06	0.02±0.01	0.02±0.01	0.03±0.01

6 months rat pathological examination results of table 44 administration

The variation of table 45 convalescent period Rats Organs and Tissues weight

Group	Matched group	High dose group	Middle dosage group	Low dose group
					Number of animals (only)	10	10	10	10
Heart percentage of liveweight percent (%)	0.33±0.08	0.36±0.06	0.33±0.05	0.33±0.04
					Liver percentage of liveweight percent (%)	3.10±0.89	3.26±0.55	3.06±0.56	3.17±0.44
Spleen percentage of liveweight percent (%)	0.17±0.05	0.20±0.06	0.17±0.06	0.19±0.07
					Lung percentage of liveweight percent (%)	0.54±0.12	0.53±0.08	0.54±0.09	0.54±0.09
Kidney percentage of liveweight percent (%)	0.28±0.06	0.32±0.06	0.29±0.04	0.28±0.03
					Adrenal gland's percentage of liveweight percent (%)	0.0101±0.0033	0.0099±0.0035	0.0100±0.0025	0.0104±0.0034
Thymus percentage of liveweight percent (%)	0.0723±0.0322	0.0761±0.0155	0.0771±0.0216	0.0692±0.0416
					Brain percentage of liveweight percent (%)	0.5501±0.0549	0.5724±0.0470	0.5674±0.0520	0.5572±0.0719
Testis percentage of liveweight percent (%)	0.4209±0.05	0.3938±0.07	0.3985±0.13	0.4290±0.07
					Epididymis percentage of liveweight percent (%)	0.1647±0.07	0.1429±0.06	0.1630±0.05	0.1529±0.05
Prostate percentage of liveweight percent (%)	0.1456±0.05	0.1344±0.04	0.1607±0.05	0.1386±0.07
					Uterus percentage of liveweight percent (%)	0.3611±0.03	0.3909±0.06	0.3534±0.07	0.3371±0.08
Ovary percentage of liveweight percent (%)	0.0212±0.008	0.02689±0.01	0.0241±0.01	0.0192±0.01

Table 46 rat long term toxication convalescent period is put to death animal pathological examination results

The present composition is with the clinical dosage (2.7g/kg) for plan of the clinical dosage (1.35g/kg) for plan of 10 times of clinical dosage (0.45g/kg) for plan, 30 times, 60 times, give the continuous gavage of rat 180 days, and test mid-term (administration 90 days) and 1 month convalescent period laboratory observation.Result is:

(1) rat growthing development and general status are not had a significant effect, only in the time of the medicine feed initial stage, have some impact, but after 2 weeks, recover normal.

(2) impact on rat blood index

Successive administration 3 months, with the comparison of blank group, the present composition all has appreciable impact to erythrocyte, leukocyte in rat blood.Wherein: high dose group can obviously reduce mononuclear cell percentage ratio (MONO%) content in Rat Erythrocytes and leukocyte, can enlarge markedly Rat Erythrocytes average external volume (MCV); Low dose group significantly reduces the content of rat leukocyte medium-sized lymphocyte percentage ratio (LYMPH%).

Successive administration 6 months, with the comparison of blank group, the present composition all has appreciable impact to erythrocyte, leukocyte, hemoglobin in rat blood.Wherein: leukocyte in rat blood (WBC) content can obviously fall in high and low dose group; In, the significantly content of mononuclear cell percentage ratio (MONO%) in leukocyte increasing of low dose group; High, medium and low dosage group all can significantly reduce erythrocyte (RBC), hemoglobin (HGB) content and packed cell volume (HCT);

After drug withdrawal January, whole blood is learned index and is all recovered normal.

(3) impact on rat blood biochemical indicator

Successive administration 3 months, with the comparison of blank group, the present composition is more obvious to albumen in rat blood and electrolytical content influence.Wherein: in, low dose group total protein (TPR) content in rat blood that can significantly raise; Low dose group is raising albumin content (ALB) significantly; High, medium and low dosage group globulin (GIB) content that all significantly raises; The remarkable increasing of Na ion of high, middle dosage group (Na ⁺) concentration and the high dose group potassium ion (K that can also significantly raise ⁺) concentration.

Successive administration 6 months, with the comparison of blank group, the present composition except to albumen in rat blood and electrolytical content influence obviously, also larger to aspartate amino transferase (GOT) and creatinine (CREA) content influence.Wherein: in, low dose group can significantly raise total protein (TP) in rat blood, albumin content (ALB) content; High and low dose group is increasing of Na ion (Na significantly ⁺), chloride ion (Cl ^-) concentration.In, low dose group can significantly reduce aspartate amino transferase (GOT), creatinine (CREA) content.

After drug withdrawal 1 month, rat blood Mid-Heaven Gate winter propylhomoserin aminotransferase (GOT), creatinine (CREA) recover normal; Total protein in high and low dose group (TPR), albumin (ALB) and globulin (GIB) content still significantly raise; Middle dosage group can significantly reduce the content of alkali phosphatase (ALP); High dose group sodium ion (Na ⁺) still significantly rising of concentration.

(4) Rats Organs and Tissues weight (heart, liver, spleen, lung, kidney, adrenal gland, thymus, uterus, ovary, testis, epididymis, prostate and brain be totally 13 internal organs) is had no significant effect, respectively organize organ coefficient substantially close.To organs and tissues (except above-mentioned 13 internal organs, also have pancreas, stomach, ileum, colon, hypophysis cerebri, spinal cord, bone marrow, lymph node, bladder, sciatic nerve, thyroid, parathyroid gland, aorta, submaxillary gland, aorta) histopathologic examination, does not find that significantly relevant with drug toxicity pathomorphology changes.

Claims

1. a herbal mixture for the treatment of heart disease, is characterized in that: this pharmaceutical composition is made up of the raw material of Chinese medicine that comprises following composition and parts by weight:

1～100 part of 1～100 part of Rhizoma Polygonati of 1～100 part of Bulbus Allii Macrostemonis of Folium Crataegi.

2. a kind of herbal mixture for the treatment of heart disease according to claim 1, is characterized in that, the parts by weight of described raw material of Chinese medicine are:

1～3 part of 1～5 part of Rhizoma Polygonati of 1～10 part of Bulbus Allii Macrostemonis of Folium Crataegi.

3. a kind of herbal mixture for the treatment of heart disease according to claim 2, is characterized in that, the parts by weight of described raw material of Chinese medicine are:

1.5 parts of 3.5 parts of Rhizoma Polygonatis of 7 parts of Bulbus Allii Macrostemonis of Folium Crataegi.

4. according to a kind of herbal mixture for the treatment of heart disease described in claim 1,2 or 3, it is characterized in that: the dosage form that described herbal mixture is prepared into is clinically any or pharmaceutically acceptable dosage form.

5. a kind of herbal mixture for the treatment of heart disease according to claim 4, is characterized in that: described dosage form is drop pill, tablet, capsule, granule, soft capsule, oral liquid or injection.

6. according to the preparation method of a kind of herbal mixture for the treatment of heart disease described in claims 1 to 3 any one, it is characterized in that comprising the following steps:

Water or any solvent that other is suitable for carrying out Effective Component of Chinese Medicine extraction carry out separately the extracts active ingredients of one or many to each of Folium Crataegi, Bulbus Allii Macrostemonis and Rhizoma Polygonati;

Or water or any solvent that other is suitable for carrying out Effective Component of Chinese Medicine extraction carry out the extracts active ingredients of one or many to the mixture of Folium Crataegi, Bulbus Allii Macrostemonis and Rhizoma Polygonati.

7. according to the preparation method of a kind of herbal mixture for the treatment of heart disease described in claims 1 to 3 any one, it is characterized in that:

(1) take Folium Crataegi by proportional quantity, after 0.1～10 hour solvent soaking time by 2 times～40 times of Folium Crataegi weight, under the condition of 30 DEG C～100 DEG C, extract 0.1～10 hour, described solvent soaking and extraction operation can be carried out 1 time or repeat nearly 6 times, merge extractive liquid,, distillating recovering solvent, distillate is centrifugal, with the solvent wash precipitation of 0.1 times of Folium Crataegi medical material amount～100 times of volumes once or repeated washing nearly five times, merge supernatant after centrifugal and the supernatant of washing precipitation, distillation and concentration near dry after, dry, pulverize, to obtain final product; Described solvent is water or any solvent that other is suitable for carrying out Effective Component of Chinese Medicine extraction.

(2) take Bulbus Allii Macrostemonis and Rhizoma Polygonati by proportional quantity, add for the first time 2 times～40 times solvent extractions of Bulbus Allii Macrostemonis and Rhizoma Polygonati gross weight 0.1～10 hour under 50 DEG C～100 DEG C conditions, leaching process can carry out once or nearly 5 times, united extraction gained decocting liquid, filters and filtrate is concentrated into 0.1 times～20 times of Bulbus Allii Macrostemonis and Rhizoma Polygonati gross weights, and adding ethanol, to make containing alcohol amount be 20%～90%, static or centrifugal, get supernatant, filter Distillation recovery ethanol, continue distillation and concentration and become thick paste, dry, pulverize, to obtain final product; Described solvent is water or any solvent that other is suitable for carrying out Effective Component of Chinese Medicine extraction.

(3) according to after the Bulbus Allii Macrostemonis of the Folium Crataegi extract of above-mentioned steps (1) acquisition and above-mentioned steps (2) acquisition, Rhizoma Polygonati extract mix homogeneously, add suitable additives, make injection, oral solid formulation or oral liquid, described oral solid formulation is tablet, capsule, granule, soft capsule, described oral liquid is oral liquid or syrup, and described additives are one or more in filler, excipient, correctives or antiseptic.

8. a kind of preparation method for the treatment of heart disease herbal mixture according to claim 7, is characterized in that:

(1) take 7 parts of Folium Crataegi, use for the first time after the 45% soak with ethanol time 2 h of 8 times of Folium Crataegi weight, under the condition of 80 DEG C, extract 2 hours, extract 1 hour the condition of 80 DEG C with 45% ethanol of 5 times of Folium Crataegi weight for the second time, extract 0.5 hour the condition of 80 DEG C with 4 times of amount 45% ethanol of Folium Crataegi weight for the third time, extract 0.5 hour the condition of 80 DEG C with 3 times of amount 45% ethanol of Folium Crataegi weight for the 4th time, merge extractive liquid, Distillation recovery ethanol, centrifuge centrifugal 30min under the rotating speed of 3000R per minute for distillate, with the purified water washing precipitation of 2 times of volumes of medical material amount three times, merge supernatant after centrifugal and the supernatant of washing precipitation, distillation and concentration is near dry rear dry, pulverize, obtain.

(2) take 1.5 parts of 3.5 parts of Bulbus Allii Macrostemonis and Rhizoma Polygonatis, under 100 DEG C of conditions, add for the first time 10 times of decoctings of Bulbus Allii Macrostemonis and Rhizoma Polygonati gross weight to boil 2 hours, add for the second time 8 times of decoctings of Bulbus Allii Macrostemonis and Rhizoma Polygonati gross weight to boil 1 hour, collecting decoction, filtrate is concentrated into 1 times of medical material gross weight, add ethanol make containing alcohol amount be 55%, static or centrifugal, get supernatant, filter Distillation recovery ethanol, continue distillation and concentration and become thick paste, dry, pulverize, to obtain final product.

(3) by after the Bulbus Allii Macrostemonis of the Folium Crataegi extract of step (1) acquisition and step (2) acquisition, Rhizoma Polygonati extract mix homogeneously, add suitable additives, make 10 parts of oral solid formulations, soft capsule or oral liquids, described additives are: starch, lactose, sodium carboxymethyl cellulose, magnesium stearate, one or more in sucrose, stevioside, sodium benzoate, soybean oil or soybean phospholipid.

9. a kind of preparation method for the treatment of heart disease herbal mixture according to claim 8, is characterized in that:

(1) Folium Crataegi extract claim 7 step (1) being obtained, after defat with petroleum ether by 1/2 amount, discard petroleum ether liquid, be extracted with ethyl acetate again, extract reclaim under reduced pressure ethyl acetate is also concentrated into dry, add suitable quantity of water to make to dissolve, be added on polyamide column, the water elution of first using 2 times of column volumes, discards water lotion, then uses 80% ethanol elution of 3 times of column volumes, collect eluent, reclaim ethanol and be concentrated into the concentrated solution of 60 DEG C of relative densities approximately 1.02～1.08, dry, obtain Folium Crataegi total flavones extract.

(2) Bulbus Allii Macrostemonis, the Rhizoma Polygonati extract that claim 7 step (2) are obtained, by 95% long-pending ethanol precipitation of diploid, repeatedly carry out 4 times, with the dialyzer small-molecule substance of dialysing, precipitate with ethanol again, finally use the absolute ether of equal volume and appropriate washing with acetone precipitate, dry, obtain Bulbus Allii Macrostemonis, coarse solomon's seal polysaccharide extract.

(3) by after the Bulbus Allii Macrostemonis of the Folium Crataegi total flavones extract of above-mentioned steps (1) acquisition and above-mentioned steps (2) acquisition, coarse solomon's seal polysaccharide extract mix homogeneously, add suitable additives, make 10 parts of injectable powder, small-volume injection or bulk capacity injections.

10. a kind of preparation method for the treatment of heart disease herbal mixture according to claim 9, is characterized in that, in the time making injection, in order to increase its dissolubility, adds tween 80 solubilizing agent; In powder pin, add excipient, described excipient is mannitol or glucose; In infusion preparation, add the isoosmotic adjusting agent for regulating osmotic pressure, described isoosmotic adjusting agent is one or more in sodium chloride, potassium chloride, magnesium chloride, calcium chloride, sodium lactate, glucose, xylitol, sorbitol and dextran, preferably sodium chloride or glucose.