CN102389442A - Lumbricus extract possessing antithrombotic effect - Google Patents
Lumbricus extract possessing antithrombotic effect Download PDFInfo
- Publication number
- CN102389442A CN102389442A CN2011103752073A CN201110375207A CN102389442A CN 102389442 A CN102389442 A CN 102389442A CN 2011103752073 A CN2011103752073 A CN 2011103752073A CN 201110375207 A CN201110375207 A CN 201110375207A CN 102389442 A CN102389442 A CN 102389442A
- Authority
- CN
- China
- Prior art keywords
- pheretima
- lumbricus
- extract
- raw material
- decoction pieces
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Landscapes
- Medicines Containing Plant Substances (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention relates to a lumbricus extract possessing antithrombotic effect, which takes lumbricus decoction pieces as a raw material. The preparation method comprises the following steps: coarse cracking the raw material of the lumbricus decoction pieces, immersing for 4-5 hours by adding 5-15 times water by weight, adding immersion liquid together in a pulverizer to crush by a wet method so as to obtain slurry with average particle fineness of D50<=100 mum, centrifuging and taking a supernatant, passing through a ceramic membrane with average aperture of 0.45 mum, refrigerating a filtrate at the temperature less than or equal to minus 20 DEG C for 2 hours, and then refrigerating and drying the filtrate under the condition of minus 4 DEG C-minus 6 DEG C of temperature and 0.1mbr-0.05mbr of absolute pressure. The lumbricus extract is capable of extracting amino acid and nucleic acid substances, and keeping the activity of effective chemical ingredients of protein and polypeptide with maximum limit. The lumbricus extract has dual effects of anticoagulation and thromboclasis, animal experiments show that the lumbricus extract has the advantages of good antithrombotic effect and simple extraction method with high efficiency.
Description
Technical field
The present invention relates to a kind of Pheretima extract, belong to pharmaceutical field with anti thrombotic action.
Background technology
Pheretima is the dry body of the Annelida hard iron earthworm animal Pheretima aspergillum Pheretima aspergillum of section (E.Perrier), popular Pheretimatschiliensis Pheretima vulgaris Chen, William Pheretimatschiliensis Pheretima.guillelmi (Michaelsen) or comb the region between the heart and the diaphragm hair earthworm Pheretima pectinifera Michaelsen.Before a kind of habit claim " LUMBRICUS ", practise to claim " Shanghai Pheretima " for back three kinds.Pheretima, promptly Lumbricus is cold in nature, and it is little salty to distinguish the flavor of, and is the traditional Chinese medicines material.According to record, before the two thousand years, our ancestors use Pheretima treatment blood stagnancy due to deficiency of QI exactly, apoplexy apoplex involving the channels and collaterals due to the venation block, and diseases such as hemiplegia have been drawn up 40 kinds of Pheretima prescriptions in Compendium of Material Medica.In these prescriptions, Pheretima is many extracts or beats powder direct oral being used as medicine with decocting, and evident in efficacy.
Along with scientific development, the mankind find that gradually Pheretima contains plurality of active ingredients, like xanthine, vitamin B complex, alkaloids etc.Mihara in 1991 finds that Pheretima homogenate extracting solution has solution fibrin and plasminogen activation dual function, and these antithrombotic effective substances of the performance of research proof subsequently are the intravital histone matter of Pheretima, are referred to as Lumbrukinase.These Lumbrukinases improve platelet aggregation for blood viscosity lowering, and are remarkable for prevention and treatment cardiovascular and cerebrovascular disease effect.
Pheretima clinical application at present mainly contains three kinds of modes: first kind is traditional decocting liquid extract, and second kind is direct oral being used as medicine of dozen powder, and the third is the crude extract of Pheretima homogenate, is the class protein composition.These three kinds of extracts all have the significance effect for prevention and treatment thrombotic disease clinically, yet on chemical constituent, bigger difference are arranged but.These three kinds of method for distilling respectively have pluses and minuses.Water decoction extract can extract aminoacid, nucleic acid material effectively, yet is unfavorable for improving the activity of protein, polypeptide constituents; Experiment finds only have blood coagulation resisting function on the water boiling and extraction thing pharmacology, does not have thrombolytic effect.Earthworm decocting pieces beats directly that powder is oral to be used as medicine, and because of un-extracted and purge process, active ingredient is without concentrating, and oral dose is big.Though refining extract can effectively extract protein, and the activity of preserving the polypeptide constituents preferably, leaching process is not through heating, and extraction conditions relaxes, and aminoacid, nucleic acid material can't effectively be extracted; Experiment showed, that the protein material that common homogenate method obtains has thrombolysis activity, but do not have anticoagulating active.
Summary of the invention
To above deficiency, the object of the invention provides a kind of Pheretima extract with anti thrombotic action, and this Pheretima extract is protected protein matter and polypeptide constituents to greatest extent, has anticoagulation and thrombolytic dual function simultaneously.
The present invention has the Pheretima extract of anti thrombotic action, is raw material with the LUMBRICUS decoction pieces, extracts through following method to get:
Raw material LUMBRICUS decoction pieces carries out coarse pulverization, adds 5～15 times of weight water logging bubble 4～5h, goes into the slurry that pulverizer carries out waterproof pulverization to granule average fineness D50≤100 μ m together with soak; The centrifuging and taking supernatant; Cross the ceramic membrane that average pore size is 0.45 μ m, filtered solution≤-20 ℃ freezing 2 hours down, be-4 ℃～-6 ℃ in temperature then; Absolute pressure is lyophilization under 0.1mbr～0.05mbr condition, promptly gets the powdery Pheretima extract.
The coarse pulverization material of said raw material LUMBRICUS decoction pieces is crossed 24 mesh sieves.
The material of said ceramic membrane is ZrO
2, Al
2O
3Or TiO
2
The application of the Pheretima extract of said anti thrombotic action in the antithrombotic reagent of preparation treatment cardiovascular and cerebrovascular disease.
The present invention adopts the case of wet attrition method to reach the crushing level of cell grade; And whole leaching process is controlled under the cryogenic conditions and carries out; The gained Pheretima extract can extract aminoacid, nucleic acid material; Can preserve the activity of protein and polypeptide class component again to greatest extent, fully guarantee the extraction of the effective chemical constituent of Pheretima, the extraction ratio of protein and polypeptide constituents is all apparently higher than common homogenate or boiling water extraction method.Therefore this Pheretima extract has anticoagulation and thrombolytic dual function simultaneously, and zoopery shows that also it has good antithrombotic effect and method for distilling is simple and efficient.
Description of drawings
Fig. 1 is Pheretima chemical constituent high performance liquid chromatogram (HPLC) fingerprint map analyzing figure.
Fig. 2 is the analysis diagram of external anticoagulation experimental result.
Fig. 3 fibrinolytic protein flat-plate experimental result photo.
Fig. 4 is arteriovenous shut thrombosis interpretation figure in the body.
Fig. 5 is that the slurry granularity of common homogenate of Pheretima and waterproof pulverization is with pulverizing the change of time curve chart.
Fig. 6 is in the Pheretima serosity of common homogenate pulverizing and waterproof pulverization, and the protein yield is with pulverizing the change of time curve.
Common homogenate of Fig. 7 and waterproof pulverization extract the chemical constituent finger printing of Pheretima.
The specific embodiment
Embodiment 1: Pheretima waterproof pulverization extracting solution and Pheretima water boiling and extraction liquid main chemical compositions are analyzed.
1. Pheretima waterproof pulverization extracting solution preparation: it is an amount of to take by weighing the LUMBRICUS decoction pieces, after the coarse pulverization, crosses sieve No. two.Accurately take by weighing the Pheretima coarse powder that the sieves 100g that minces, after wash quickly removes surperficial silt, add 1000mL water; After soaking 4h; Feed liquid and Pheretima are together put into vibration type super micron mill (Jinan shellfish power powder company limited is produced, the BL-6 type), pulverize 10min; Get the slurry of average fineness D50≤100 μ m, through 4000rmin
-1Centrifugal, get supernatant, with the supernatant of gained through membrane aperture 0.45 μ m ZrO
2Ceramic membrane promptly gets and contains crude drug amount 0.1gmL
-1Pheretima waterproof pulverization extracting solution.
2. the preparation of Pheretima water boiling and extraction liquid: it is an amount of to take by weighing the LUMBRICUS decoction pieces, after the coarse pulverization, crosses sieve No. two.Accurately take by weighing the Pheretima coarse grain 100g that sieves, wash quickly removes surperficial silt, adds 1000mL water, soak 4h after, heating and refluxing extraction, boil 1h after, stop.With the centrifugal (4000rmin of medicinal liquid
-1), get supernatant, with the supernatant of gained through membrane aperture 0.45 μ m ZrO
2Ceramic membrane is got permeate and is promptly got and be equivalent to contain crude drug amount 0.1gmL
-1Pheretima water boiling and extraction liquid.
3. active component content analysis
(1) protein content analysis:
Adopt Coomassie brilliant blue method research protein content, outcome research shows that Pheretima waterproof pulverization extracting solution protein concentration is 3.09mgml
-1, Pheretima water boiling and extraction liquid has only 0.69mgml
-1Compare with water boiling and extraction, the earthworm protein content in the waterproof pulverization extracting solution has improved several times.
(2) analysis of amino acids:
Adopt FDAC 835 type automatic amino acid analyzers to carry out the free amino acid composition analysis.Adopting analytical column is the stainless steel column of 2.6mm * 150mm, chromatographic column 2622sc
#, 57 ℃ of column temperatures.Buffer solution pump flow velocity 0.225mLmin
-1, the 1,2,3-indantrione monohydrate flow rate pump is 0.30mLmin
-1, each sample size is 20 μ L.
The result sees table 1; The total free amino acid content of every milliliter of Pheretima waterproof pulverization extracting solution is 20.75mg; The total free amino acid content of every milliliter of Pheretima decocting liquid is 30.94mg; The total amino acids content that is water boiling and extraction liquid is a little more than the waterproof pulverization extracting solution, but the two contained aminoacid kind zero difference.
Table 1 Pheretima waterproof pulverization liquid and water boiling and extraction liquid amino acid composition analysis
(3) chemical constituent fingerprint map analyzing:
Reference substance preparation: take by weighing hypoxanthine 4.50mg, xanthine 2.09mg, uracil 2.94mg, put in the 50ml volumetric flask, add 0.05molL-1 (NH4) 2HPO4 solution and make dissolving, and be diluted to scale, shake up, cross 0.45 μ m micro-filtration membrane, get appearance on the subsequent filtrate.
The test sample preparation: Pheretima waterproof pulverization extracting solution and Pheretima water boiling and extraction liquid through 0.45 μ m filter membrane, are got liquid respectively.Carry out chromatography respectively, sample size is 10 μ L.
Chromatographic condition: mobile phase A: 0.05molL
-1(NH
4)
2HPO
4Solution; Mobile phase B: methanol.The detection wavelength is 254nm; Column temperature is 30 ℃; Flow velocity is 0.8mlmin
-1Hedera ODS-2C18 chromatograph (4.6mm * 250mm, 5 μ m).Gradient: 0-7min, 0-2%B; 7-15min, 2-5%B; 15-50min, 5-50%B; Flow velocity is 0.8mlmin
-1Record 60min chromatograph is seen Fig. 1.Among Fig. 1, A is that reference substance collection of illustrative plates, B are micronizing extracting solution collection of illustrative plates, and C is a water boiling and extraction liquid collection of illustrative plates, and 1 is the uracil peak; 2 is the xanthine peak; 3 is the hypoxanthine peak.
Fig. 1 is visible, RRT (t
R) in 0～15min, mainly be the relatively large nucleic acid materials of some polarity, the collection of illustrative plates peak of B and C is more consistent.t
RAfter 15min, be some polar phase to more weak material, 3 obvious characteristics collection of illustrative plates peaks (collection of illustrative plates B) only appear in Pheretima wet method ultra micro extracting solution, large amount of complex collection of illustrative plates peak (collection of illustrative plates C) appears in water boiling and extraction liquid.Explanation is for the extraction of Pheretima chemical constituent, and the waterproof pulverization extraction is boiled with decocting and had certain difference.
It is thus clear that waterproof pulverization has preferably for heat-sensitive substance (for example, protein) etc. and extracts, yet for other chemical constituents, extraction ratio is a little less than water boiling and extraction.
Embodiment 2: Pheretima waterproof pulverization extracting solution and Pheretima decocting liquid antithrombotic drug effect are relatively.
With embodiment 1 gained Pheretima waterproof pulverization extracting solution label is sample 1; Gained Pheretima water boiling and extraction liquid label is a sample 2; Pheretima waterproof pulverization extracting solution is crossed the permeate of the poly (ether sulfone) film of molecular cut off 10000, and label is a sample 3; Pheretima water boiling and extraction liquid is crossed the poly (ether sulfone) film of molecular cut off 10000, gets permeate, and label is a sample 4.
(1) external anticoagulation experiment:
Method: on the ceramic orifice plate of the bacterium of going out; The venous blood 200 μ L that add the healthy SD rat in every hole respectively, each hole adds sample 1, sample 2, sample 3, sample 4, the normal saline (all warm in advance and Mus body temperature consistent temperature) of 200 μ L respectively, and last then every hole adds 4 μ L thrombins (1IU); Mix; Put into 37 ℃ of constant temperature and humidity incubators, the blood coagulation situation of each time period of record in the 1h, experimental result is seen Fig. 2.
Visible by Fig. 2,0-5min is in the time period, and normal saline group part forms blood clot, and a little blood clot appears in the administration group, between group and no significant difference.5-30min is in the time period, and the normal saline group all forms blood clot, and the administration group just partly forms blood clot, and the blood coagulation state between group is no significant difference still.Behind the 60min, the administration group all forms blood clot.More than external anticoagulation result of the test show that Pheretima water boiling and extraction liquid and Pheretima waterproof pulverization extracting solution can both the significant prolongation clotting times, external anticoagulation does not have significant difference.
(2) external thrombolytic experiment:
Method: in diameter is the culture dish of 10cm, add 3mgmL respectively
-1Sanguis Bovis seu Bubali fibrinogen solution 15mL, the agar solution 10mL of mass fraction 1%, 50IUmL
-1Thrombin of beef solution 0.4mL, shake up, leave standstill after 30min solidifies it, use card punch to get the hole of diameter in the above as 5mm, 4 ℃ of refrigerators are preserved subsequent use.Like Fig. 3, survey when living sample thief 1,2,3,4 and positive control (urokinase, 100UmL
-1) 20 μ L, clicking and entering in the aperture, behind 37 ℃ of constant temperature culture 17h, if transparent circle occurs, it is active to explain that medicine has solution fibrin matter, and the transparent circle size is represented the fibrinolytic size.Among Fig. 3, sample 1: Pheretima waterproof pulverization extracting solution; Sample 2: Pheretima water boiling and extraction liquid; Sample 3: remove proteinic Pheretima waterproof pulverization extracting solution; Sample 4: remove proteic Pheretima water boiling and extraction liquid; Positive controls: urokinase.Can know to have only Pheretima waterproof pulverization extracting solution (sample 1) and matched group urokinase the active transparent circle of fibrinolytic protein to occur by Fig. 3, explain that the protein in the Pheretima waterproof pulverization extracting solution has stronger fibrinolytic protein activity.And water boiling and extraction liquid (sample 2); Waterproof pulverization extracting solution (sample 3) behind the removal protein; Transparent circle does not all appear in the no fibrinolytic protein activity of water boiling and extraction liquid (sample 4) behind the removal protein, explains that the protein in the Pheretima water boiling and extraction liquid does not have fibrinolytic.Therefore prove that waterproof pulverization extract of the present invention (liquid) has dissolution in vitro thrombosis drug effect.
(3) antithrombotic experiment in the body:
Method: 80 rats are divided into 8 groups at random by the body constitution amount; Every group 10: blank group (normal saline), aspirin group (100mg/kg), waterproof pulverization extracting solution (0.8,0.4,0.2g crude drug/kg), water boiling and extraction liquid (0.8; 0.4, the 0.2g crude drug/kg).In 1 week of oral administration gavage, behind the last administration 1h, rat is anaesthetized with 3.5% chloral hydrate lumbar injection (1mL/100g); The medisection skin of neck, blunt separation RCCA and left external jugular vein, the polyethylene tube that will be full of heparin-saline (50U/mL) in advance (divides 3 sections bindings; Put into No. 4 surgical threads of long 5cm in the stage casing) an end insert left external jugular vein after, inject heparin 50U/kg from polyethylene tube, clamp tube wall; The RCCA proximal part is clamped with blocking blood flow with hemostatic clamp; Eye scissors one osculum inserts RCCA, ligation with the polyethylene tube other end.Open the open blood flow of hemostatic clamp, middle Herba Clinopodii takes off polyethylene tube rapidly behind the 15min; Take out surgical thread, remove floating blood, be placed on the Cover Glass of claiming quality with wet filter paper; Claim quality with analytical balance, deduct line mass and the Cover Glass quality promptly gets wet weight of thrombus with gross mass.
Result of the test is seen Fig. 4, and with compared with normal, high dose administration group is compared with the aspirin group, can obviously reduce thrombus weight (P＜0.01), and antithrombotic formation is dose dependent.Yet, the anti thrombotic action there was no significant difference (P＞0.05) of Pheretima waterproof pulverization extracting solution and Pheretima water boiling and extraction liquid.Among the figure 4, * P＜0.05, * * P＜0.01 and normal group are relatively; Compare with the aspirin group #P＜0.05, ##P＜0.01.
Can know that to sum up Pheretima waterproof pulverization extract has thrombus and anticoagulation dual function.Antithrombotic can be equal to water boiling and extraction liquid in the body.The Pheretima waterproof pulverization can fully guarantee the extraction of Pheretima antithrombotic acitivity material.
Embodiment 3: the comparative test of common homogenate and waterproof pulverization.
Operation: an amount of Pheretima medical material, through pulverizing sieve No. two, two parts of back Pheretimas of sieving of weight such as get, wash quickly removes Pheretima surface silt respectively, and the water of 10 times of weight of adding adopts Fu Luke tissue refiner (10000rmin respectively
-1) and the vibration type super micron mill pulverize, homogenate pulverize gained material and waterproof pulverization gained material respectively by 1,3,5,10,15, the sampling of 30min different time points, centrifugal immediately, supernatant is subsequent use, the medicinal residues confession grain diameter measurement after centrifugal.
(1) grinding particle size test:
Respectively with the pulverizing time be 1,3,5,10,15, each time point gained sample of 30min centrifugal after the medicinal residues water disperse again after, adopt MICRO RACS 3500 type Particle Size Analyzer determination of laser light scattering medicinal residues particle size distribution, every part of parallel assay 3 times.With meso-position radius D50 value is index, and relatively the particle diameter of two kinds of different breaking methods is with pulverizing change of time curve such as Fig. 5.
Results of grain size analysis sees that Fig. 5 can know, common homogenate of Pheretima and Pheretima waterproof pulverization all reach about 1min, and the two particle diameter all sharply reduces, yet along with the passing of pulverizing the time, change of size all is slow trend, and crush efficiency descends.The pulverizing time is when reaching 30min; The common homogenate microgranule of Pheretima D50 value reaches about 175.85 μ m; And the Pheretima waterproof pulverization is when 1min, and D50 just reaches 158.11 μ m, and the pulverizing time is when reaching 10min; The Pheretima microgranule D50 value of waterproof pulverization reaches 17.53, is with 1/15 of common homogenate microgranule of time at least.Results suggest, the smashing fineness of waterproof pulverization and crush efficiency are apparently higher than common homogenate.
(2) the protein yield is analyzed:
By 1,3,5,10,15, the difference of 30min pulverizes the serosity of time point gained, detects common homogenate and the different pulverizing of waterproof pulverization time protein yield with the Bradford method.
The crude drug amount * 100% of the total protein quality/input of protein yield=extraction.
Fig. 6 result can find out that common homogenate 30min protein yield can be accomplished within the waterproof pulverization 1min, explain that waterproof pulverization can accomplish the extraction of earthworm protein within a short period of time, and the albumen yield is apparently higher than common homogenate.
(3) fingerprint map analyzing:
Draw the 30min centrifugal liquid of above-mentioned two kinds of different breaking methods respectively, remove macro-molecular protein through 70% ethanol precipitation, centrifugal, get supernatant, cross 0.45 μ m micro-filtration membrane, get filtrating and go up appearance, detect record 50min chromatograph.Chromatographic condition is following: chromatographic column is Hedera ODS-2C18 chromatographic column (4.6mm * 250mm, 5 μ m), mobile phase A: 0.05molL
-1(NH4)
2HPO
4Solution.Mobile phase B: methanol.The detection wavelength is 254nm; Column temperature is 30 ℃; Flow velocity is 0.8mLmin
-1Gradient: 0-7min, 0-2%B; 7-15min, 2-5%B; 15-50min, 5-50%B; Flow velocity is 0.8mlmin
-1Write down the 50min chromatograph respectively, chromatograph is seen Fig. 7.
Visible by Fig. 7, the peak shape basically identical of the sample extracting solution of Pheretima waterproof pulverization and common homogenate gained.The material that shows two kinds of breaking methods extraction gained is consistent; Yet in identical crude drug input amount; The chromatographic peak area of waterproof pulverization gained is obviously greater than common homogenate, explain the waterproof pulverization method for the extraction efficiency of other chemical effective ingredient of Pheretima apparently higher than common homogenate method.
Claims (4)
1. the Pheretima extract with anti thrombotic action is characterized in that with the LUMBRICUS decoction pieces be raw material, extracts through following method to get: raw material LUMBRICUS decoction pieces carries out coarse pulverization; Add 5～15 times of weight water logging bubble 4～5h; Go into the slurry that pulverizer carries out waterproof pulverization to granule average fineness D50≤100 μ m together with soak, the centrifuging and taking supernatant, crossing average pore size is the ceramic membrane of 0.45 μ m; Filtered solution≤-20 ℃ freezing 2 hours down; Be-4 ℃～-6 ℃ in temperature then, absolute pressure is lyophilization under 0.1mbr～0.05mbr condition, promptly gets the powdery Pheretima extract.
2. the Pheretima extract with anti thrombotic action according to claim 1 is characterized in that the coarse pulverization material of said raw material LUMBRICUS decoction pieces is crossed 24 mesh sieves.
3. the Pheretima extract with anti thrombotic action according to claim 1 and 2, the material that it is characterized in that said ceramic membrane is ZrO
2, Al
2O
3Or TiO
2
4. the application of the Pheretima extract of the said anti thrombotic action of claim 1 in the antithrombotic reagent of preparation treatment cardiovascular and cerebrovascular disease.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2011103752073A CN102389442B (en) | 2011-11-23 | 2011-11-23 | Lumbricus extract possessing antithrombotic effect |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2011103752073A CN102389442B (en) | 2011-11-23 | 2011-11-23 | Lumbricus extract possessing antithrombotic effect |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102389442A true CN102389442A (en) | 2012-03-28 |
| CN102389442B CN102389442B (en) | 2013-03-13 |
Family
ID=45856907
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2011103752073A Expired - Fee Related CN102389442B (en) | 2011-11-23 | 2011-11-23 | Lumbricus extract possessing antithrombotic effect |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102389442B (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102988418A (en) * | 2012-12-11 | 2013-03-27 | 广东新峰药业股份有限公司 | Preparation method of traditional Chinese medicine aqueous solution for treating bronchial asthma |
| CN104611232A (en) * | 2014-12-16 | 2015-05-13 | 湖南新汇制药股份有限公司 | Bioconversion mycelium and application of bioconversion mycelium extract in preparation of anticoagulant drugs |
| CN113234708A (en) * | 2021-05-10 | 2021-08-10 | 北京中医药大学 | Fibrinolysis active protein, preparation method and application thereof, pharmaceutical composition and nucleic acid for coding protein |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1475266A (en) * | 2003-07-01 | 2004-02-18 | 李振国 | Production method of injection agent used for treating cardio and cerebro vascular diseases and its product |
| CN101584853A (en) * | 2009-06-15 | 2009-11-25 | 南京中医药大学 | Earthworm protein polypeptide preparation |
| CN101590219A (en) * | 2009-06-15 | 2009-12-02 | 南京中医药大学 | A kind of refined earthworm protein polypeptide preparation |
| CN101623302A (en) * | 2009-08-13 | 2010-01-13 | 天津贾立明蚯蚓养殖有限公司 | Method for preparing active earthworm powder |
-
2011
- 2011-11-23 CN CN2011103752073A patent/CN102389442B/en not_active Expired - Fee Related
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1475266A (en) * | 2003-07-01 | 2004-02-18 | 李振国 | Production method of injection agent used for treating cardio and cerebro vascular diseases and its product |
| CN101584853A (en) * | 2009-06-15 | 2009-11-25 | 南京中医药大学 | Earthworm protein polypeptide preparation |
| CN101590219A (en) * | 2009-06-15 | 2009-12-02 | 南京中医药大学 | A kind of refined earthworm protein polypeptide preparation |
| CN101623302A (en) * | 2009-08-13 | 2010-01-13 | 天津贾立明蚯蚓养殖有限公司 | Method for preparing active earthworm powder |
Non-Patent Citations (1)
| Title |
|---|
| 李力等: "复方地龙胶囊组方与制备工艺研究", 《药学进展》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102988418A (en) * | 2012-12-11 | 2013-03-27 | 广东新峰药业股份有限公司 | Preparation method of traditional Chinese medicine aqueous solution for treating bronchial asthma |
| CN104611232A (en) * | 2014-12-16 | 2015-05-13 | 湖南新汇制药股份有限公司 | Bioconversion mycelium and application of bioconversion mycelium extract in preparation of anticoagulant drugs |
| CN113234708A (en) * | 2021-05-10 | 2021-08-10 | 北京中医药大学 | Fibrinolysis active protein, preparation method and application thereof, pharmaceutical composition and nucleic acid for coding protein |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102389442B (en) | 2013-03-13 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP5048673B2 (en) | A kind of extract that prevents or treats thrombotic diseases | |
| CA2787154C (en) | A compound sea cucumber preparation and manufacturing method thereof | |
| CN103257188B (en) | Construction method for compound thrombus clearing preparation bioactivity chromatography finger print | |
| Dandjesso et al. | Phytochemistry and hemostatic properties of some medicinal plants sold as anti-hemorrhagic in Cotonou markets (Benin) | |
| CN104004806A (en) | Lumbricus polypeptide having anticoagulant and thrombolytic effects, and enzymatic hydrolysis preparation method and application thereof | |
| CN103884811B (en) | A kind of biological chromatography compares screening system and application thereof | |
| CN103239706B (en) | Earthworm protein for reducing blood glucose, improving microcirculation and eliminating diabetic complication and application thereof | |
| CN102389442B (en) | Lumbricus extract possessing antithrombotic effect | |
| CN102453106A (en) | Preparation method of bletilla striata polysaccharide | |
| CN109593141A (en) | A kind of preparation method and its usage of motherwort polysaccharide | |
| CN100409890C (en) | Pharmaceutical composition for treating and preventing thrombus and its preparation method | |
| Lin et al. | Chaiqinchengqi decoction regulates necrosis-apoptosis via regulating the release of mitochondrial cytochrome c and caspase-3 in rats with acute necrotizing pancreatitis | |
| Li et al. | Preparation and antithrombotic activity identification of Perinereis aibuhitensis extract: A high temperature and wide pH range stable biological agent | |
| KR100882462B1 (en) | Preparation of anticoagulant sulfated galactan from an enzymatic extract of Red Algae in Jeju | |
| CN110501433B (en) | Method for identifying callicarpa nudiflora medicinal materials in different producing areas | |
| CN108164571A (en) | A kind of mung bean alpha-galactooligosaccharide extract and its extracting method and skin care application | |
| CN102242102A (en) | Preparation method of X factor activator from bothrops atrox venom | |
| CN104498464A (en) | Method for extracting agkistrodon snake venom hemocoagulase from gloydius blomhoffi brevicaudus venom | |
| CN101711793B (en) | Medicinal composition for treating cardiovascular and cerebrovascular diseases and preparation method thereof | |
| CN103610703B (en) | A kind of pharmaceutical composition, preparation method and method of quality control thereof for the treatment of cardiovascular and cerebrovascular disease | |
| CN102813687B (en) | Use of ligustilide in preparing anti-thrombotic medicines | |
| CN104403016B (en) | A kind of mulberry leaf polysaccharide and preparation method for separating and purifying thereof and application | |
| CN102240320A (en) | Motherwort and astragalus capsule used for treating prostatic hyperplasia | |
| CN103041176A (en) | Nervate twayblade herb extracts and application thereof | |
| CN104523758A (en) | Pillbug extract and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20130313 Termination date: 20191123 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |