CN103536621A - Application of holothurian glycosaminoglycan in preparation of medicaments for treating coronary syndromes - Google Patents
Application of holothurian glycosaminoglycan in preparation of medicaments for treating coronary syndromes Download PDFInfo
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Abstract
The invention discloses application of holothurian glycosaminoglycan in preparation of medicaments for treating coronary syndromes. Animal tests prove that more than one holothurian glycosaminoglycan with the weight average molecular weight of 90-130kDa has multiple remarkable physiological activities, such as anti-platelet aggregation, anticoagulation, fibrinolysis promotion, blood viscosity reduction, blood fat regulation and antiviral, can be used for preparing medicaments for treating coronary syndromes, and can be safely and effectively used for preventing and treating coronary syndromes. The holothurian glycosaminoglycan has a remarkable effect of anti-platelet aggregation by intravenous injection in a certain dosage, does not have remarkable effect of anticoagulation, namely the anticoagulation has a smaller probability of causing bleeding, so that the holothurian glycosaminoglycan is safer and more effective for treating coronary syndromes based on the development of platelet activity resistance of holothurian glycosaminoglycan, and has a wide research and developing prospect.
Description
Technical field
The present invention relates to the medical usage of glycosaminoglycan extracted from sea cucumber, the application of the glycosaminoglycan extracted from sea cucumber that is specifically related to molecular weight and is 90kDa to 130kDa in prevention and treatment coronary syndrome.
Background technology
Acute coronary syndrome (Acute Coronary Syndrome, ACS) be to break (rupture) or rotten to the corn (erosion) with Coronary Atherosclerotic Plaque, complete or the incomplete occluding thrombus of secondary forms one group of clinical syndrome of pathologic basis, it is to comprise unstable angina (UA), a series of clinical symptoms of non-ST section elevation myocardial infarction and ST section elevation myocardial infarction.Hematoblastic activation plays an important role in the generation of ACS.In long-term clinical practice, find, many patients' clinical symptoms is different, and its coronary artery but has closely similar pathophysiological change, and Coronary Atherosclerotic Plaque transfers to unstablely by stable, then breaks and causes thrombosis.Therefore the treatment of the thromboembolism preventing of acute coronary syndrome is very important, especially all the more so in non-ST section elevation ACS, and thromboembolism preventing treatment can be divided into Antiplatelet therapy and antithrombin to treat.
Non-ST section is raised acute coronary syndrome: comprise that unstable angina and non-ST section raise heart infarction.In March, 2011 ACC/American Heart Association (ACC/AHA) has issued new edition unstable angina pectoris/non-lifting property of ST section myocardial infarction (UA/NSTEMI) treatment guide, wherein as follows about the suggestion of antiplatelet and anticoagulant therapy:
Drug therapy in 1 early stage institute: antiplatelet and anticoagulant are the foundation stones of UA/NSTEMI treatment.Antiplatelet and anticoagulant therapy are had to detailed regulation:
1) should start rapidly Antiplatelet therapy.First-selected aspirin (Level of Evidence:A)
2) the not tolerant patient of aspirin allergy or gastrointestinal tract illness, should use clopidogrel (Level of Evidence:A)
3) at the inpatient that does not go to Coronary intervention, while being admitted to hospital, except using aspirin, also should use as far as possible clopidogrel, administration time is 1 (Level of Evidence:A)-9 months (Level of Evidence:B)
4) going to the inpatient of intervention, should use clopidogrel 1 month above (Level of Evidence:A), if there is no hemorrhage high risk factor, can use 9 months (Level of Evidence:B)
5) select a time CABG and using the patient of clopidogrel going to, should drug withdrawal 5-7 days (Level of Evidence:B)
6) except using aspirin or using clopidogrel to carry out Antiplatelet therapy, also should use vein unfractionated heparin or subcutaneous LMWH anticoagulant (Level of Evidence:A)
7), for the patient who prepares row cardiac catheterization and PCI, except using aspirin and unfractionated heparin, also should use GPIIb/IIIa receptor antagonist (Level of Evidence:A)
2 discharge medications guides: antiplatelet drug is long-term prescription (secondary prevention).
Antiplatelet drug mainly comprises aspirin, thiophene chloropyridine, dipyridamole, clopidogrel, lamifiban etc. in recent years, and such medicine has anticoagulant effect, and its side effect is different, but common side effect is hemorrhage.Antiplatelet drug anticoagulant, platelet is that initial stage anastalsis is necessary, and it is again the startup thing of artery thrombosis on the other hand, therefore strictly controls dosage, and clinical indication and contraindication are very important.
Therefore, seeking a kind of saferly for preventing and treat the medicine of coronary syndrome, is that people extremely expect.
" Stichopus japonicus, someone is referred to as " extra large Radix Ginseng ", because of the similar Radix Ginseng of benefiting action, gains the name, be the delicacies in feast, be again the treasure of nourishing human body, its medical value is also higher.< < traditional Chinese medical science voluminous dictionary > > records the effect of Stichopus japonicus: " the kidney invigorating and essence nourishing, blood enriching and dryness moistening.Control that essence and blood loss, weak labor are dispelled, sexual impotence, nocturnal emission, dryness of the intestine be just difficult ".< < Bencao Congxin > >: " the kidney invigorating and essence nourishing, flaccidity is treated in tonifying YANG." the < < property of medicine examines > >: " pathogenic fire reducing nourishing kidney, logical intestinal are moisturized." the < < detailed outline > > that picks up any lost article from the road: " raw hundred arteries and veins blood, control chronic dysentery with frequent relapse." the < < book on Chinese herbal medicine > > that looks for novelty: " moistening five ZANG-organs is grown smart diuretic ".
At present, confirmed that the composition in Stichopus japonicus with medical active mainly contains protein, polypeptide, lipid, Saponin and polysaccharide etc.Stichopus japonicus albumen has high nutritive value; In Stichopus japonicus, isolated there is antiinflammatory, the polypeptide of antitumor, raising immunity activity; Ganglioside in Stichopus japonicus lipid has the effect of collaborative nerve growth factor, and holotoxin has very strong antifungic action in addition.
Summary of the invention
The object of this invention is to provide the application of a kind of glycosaminoglycan extracted from sea cucumber in preparation treatment coronary syndrome medicine, the above-mentioned defect existing to overcome prior art, meets people's needs.
A further object of the present invention is to provide a kind of pharmaceutical preparation that molecular weight is 90kDa to 130kDa glycosaminoglycan extracted from sea cucumber that contains.
Inventor's discovery, the aspects such as glycosaminoglycan extracted from sea cucumber antiplatelet aggregation and anticoagulant have stronger activity compared with fucose.The effect of antiplatelet drug prevention and treatment coronary syndrome is very obvious.Glycosaminoglycan extracted from sea cucumber has and at doses, has remarkable anticoagulant effect and do not have anticoagulant active simultaneously, with respect to commercially available similar drugs, has significant security advantages.
Animal experiment proves, weight average molecular weight is more than one in 90kDa to 130kDa glycosaminoglycan extracted from sea cucumber, there is significant antiplatelet aggregation, anticoagulant, promotion fibrinolytic, reduce blood viscosity, regulate the multiple physiologically actives such as blood fat, antiviral, can be for the preparation for the treatment of coronary syndrome medicine;
Described coronary syndrome comprises acute myocardial infarction (AMI), unstable angina pectoris (UA) or arterial thrombus etc.
Preferably, the weight average molecular weight of described glycosaminoglycan extracted from sea cucumber is more than one in 95000Da～125000Da, more preferably more than one in 100000Da～120000Da;
Described " weight average molecular weight is more than one in the glycosaminoglycan extracted from sea cucumber between 90000Da to 130000Da " refers to, can be that weight average molecular weight is in the glycosaminoglycan extracted from sea cucumber between 95000Da to 125000Da, also can be that weight average molecular weight is the combination of the various molecular weight of glycosaminoglycan extracted from sea cucumber between 100000Da to 1200000Da, be a kind of compositions.
The preparation method of glycosaminoglycan extracted from sea cucumber of the present invention, the method that can adopt patent " a kind of extracting method of holothuria leucospilota glycosaminoglycan " document to provide, preferred, comprise the steps:
1) from Stichopus japonicus, extract Stichopus japonicus polysaccharide;
From Stichopus japonicus, extract the step of Stichopus japonicus polysaccharide, comprise to Stichopus japonicus soak, rubbing, enzymolysis precipitation obtain Stichopus japonicus polysaccharide crude product, then, to Stichopus japonicus polysaccharide purifying crude, decolouring, obtains Stichopus japonicus polysaccharide;
Described enzymolysis is to carry out enzymolysis with hydrolysising protease and Cotazym;
Described hydrolysising protease can adopt commercially produced product, as Alcalase, and the product of letter (Shenyang) Bioisystech Co., Ltd of Novi; Described Cotazym, as the product of Wuxi City Xue Mei enzyme preparation Science and Technology Ltd., is avenged prunus mume (sieb.) sieb.et zucc. board
To Stichopus japonicus polysaccharide purifying crude, be to carry out purification with DEAE-cellulose column.
Described Stichopus japonicus is selected from hojothuria leucospilota, Holothuria atra, rough Stichopus japonicus, Thelenota ananas (Jaeger)．, picolour zhuopianshen, actinopyga, is preferably hojothuria leucospilota and rough Stichopus japonicus;
2) adopting is with gel, to cross post to collect weight average molecular weight between the glycosaminoglycan extracted from sea cucumber between 90kDa to 130kDa;
The polydispersity of the glycosaminoglycan extracted from sea cucumber of collecting is less than 2, is preferably less than 1.5, is more preferably less than 1.4;
Described polydispersity refers to the index of the conventional measurement molecular weight distribution in this area, for the width of characterize polymers molecular weight distribution.Polydispersity is being claimed polydispersity index, polydispersity or dispersion of distribution index again herein or in other documents, is the ratio of weight average molecular weight (Mw) and number-average molecular weight (Mn), i.e. Mw/Mn.This ratio changes with molecular weight distribution width.When single dispersion, Mw/Mn equals 1, and along with molecular weight distribution broadens, it is large that Mw/Mn value becomes gradually;
Wherein, step 2), also comprise cryodesiccated step;
The invention still further relates to a kind of pharmaceutical composition, comprise described glycosaminoglycan extracted from sea cucumber and the pharmaceutically acceptable carrier for the treatment of effective dose;
Described carrier refers to the carrier of pharmaceutical field routine, such as: diluent, excipient are as water etc.; Binding agent is as cellulose derivative, gelatin, polyvinylpyrrolidone etc.; Filler is as starch etc.; Burst apart agent as calcium carbonate, sodium bicarbonate; Lubricant is as calcium stearate or magnesium stearate etc.In addition, can also in compositions, add other adjuvant as flavouring agent and sweeting agent, preferably one or more in mannitol, lactose, dextran, glucose, glycine, gelatin hydrolysate, polyvidone and sodium chloride, preferably mannitol;
When described pharmaceutical composition is lyophilized injectable powder, the weight content of described glycosaminoglycan extracted from sea cucumber is 0.5-2%, preferably 1%; Intravenous drip speed is 100ml/h; Intravenous drip dosage is 0.01mg/kg～6mg/kg rat body weight, preferably 0.5～3mg/kg;
The various dosage forms of compositions of the present invention can adopt the method for medical domain routine to be prepared, and wherein the content of active component is 0.1%～99.5%(weight ratio).
Through lot of experiments, show, this pharmaceutical composition can the safe and effective prevention for coronary syndrome and treatment.Its prevention and treatment coronary syndrome are relevant to the adhesion of its antiplatelet aggregation, inhibition platelet and endotheliocyte.And progressively illustrating along with platelet physiology, biochemical function in recent years, novel anticoagulant will constantly occur, glycosaminoglycan extracted from sea cucumber composition intravenous drip under doses has remarkable anticoagulant effect, and do not there is obvious blood coagulation resisting function, both due to anticoagulation, to cause hemorrhage probability less, therefore the exploitation based on its anti-platelet activity, is used for the treatment of coronary syndrome safer, effective, has wide research and development prospect.
Accompanying drawing explanation
Fig. 1 glycosaminoglycan extracted from sea cucumber composition purity of the present invention figure.
The molecular weight distribution collection of illustrative plates of Fig. 2 glycosaminoglycan extracted from sea cucumber composition of the present invention.
The specific embodiment
It is glycosaminoglycan extracted from sea cucumber that Stichopus japonicus mainly contains two kinds of polysaccharide a kind of, and another kind is HF.Stichopus japonicus polysaccharide also has a multiple physiologically active with selenka is the same, and physiologically active main manifestations is: suppress thromboembolism, anticoagulation, raising immunity, promote celloglobulin dissolving, antitumor action, blood sugar lowering etc.Stichopus japonicus of the present invention is selected from hojothuria leucospilota, poor Stichopus japonicus, Holothuria atra, Thelenota ananas (Jaeger)．, picolour zhuopianshen, actinopyga, preferably hojothuria leucospilota.Wherein, hojothuria leucospilota originates in the ground such as the North Sea, China South Sea, and widely distributed, aboundresources, price are suitable, and glycosaminoglycan extracted from sea cucumber rich content, are desirable raw materials.
Of the present invention, prepare in preferred embodiment of glycosaminoglycan extracted from sea cucumber, in order obtaining, to be suitable for intravenous high-purity glycosaminoglycan extracted from sea cucumber composition, can successively dry hojothuria leucospilota body wall to be rubbed, centrifugal after enzymolysis, supernatant precipitates with ethanol, is dried to obtain precipitate I; After this precipitate I is dissolved in water, add calcium chloride, centrifugal, supernatant ethanol precipitate with ethanol, is dried to obtain precipitate II; After precipitate II is dissolved in water, add potassium acetate, get supernatant, add ethanol precipitation, be dried to obtain Stichopus japonicus polysaccharide crude product; Stichopus japonicus polysaccharide crude product is crossed to DEAE-cellulose column, use ethanol precipitate with ethanol, add hydrogen peroxide decolouring place thermal source, can obtain the Stichopus japonicus polysaccharide that molecular weight is 90～130kDa.
The invention provides a kind of drug combination preparation, comprising glycosaminoglycan extracted from sea cucumber composition and pharmaceutically acceptable carrier.The present invention is that the pharmaceutical preparation that contains sea cucumber suger amino sugar is lyophilized injectable powder, pharmaceutically acceptable carrier mannitol,, one or more in lactose, dextran, glucose, glycine, gelatin hydrolysate, polyvidone and sodium chloride.In pharmaceutical composition of the present invention, the content of active component is 0.5～3%,, preferably 1%.
The invention provides the application of a kind of drug combination preparation in the medicine of control coronary syndrome.Preferred non-ST section is raised the dosage of heart infarction administration and generally by doctor, according to patient's concrete condition (as age, body weight, sex, sick time, health etc.), is determined.Generally speaking, in glycosaminoglycan extracted from sea cucumber, the dosage of administration is 0.1～5mg/kg weight in patients, is preferably 0.1～1mg/kg, is preferably 0.l～0.6mg/kg.
For the ease of understanding, below by the drawings and Examples by concrete, present invention is described.It needs to be noted, these descriptions are only exemplary descriptions, do not form limitation of the scope of the invention.
Take hojothuria leucospilota medical material 2kg, water logging is spent the night.Wall of sea cucumber Stichopus japonicus is drenched to solid carbon dioxide and divide, rub, the also moisturizing of weighing is to 6Kg, put in 60 ℃ of water-baths, add sodium hydroxide to adjust pH to 8.0 ± 0.2, add hydrolysising protease Alcalase40ml(inventory 2%) stir enzymolysis 4 hours, it is more than 85 ℃ that deactivation is 10 minutes, be cooled to 50 ℃ ± 2 ℃, add sodium hydroxide to adjust pH to 8.0 ± 0.2, then add Cotazym 10g(inventory 0.2%) stir enzymolysis 4 hours, boil 5 minutes, cooling.4 ℃ centrifugal, collects supernatant, adds hydrochloric acid to adjust pH to 2.5 ± 0.2, and 4 ℃ of cold preservation 2 hours is centrifugal, collects supernatant, adjusts pH to 7.0 ± 0.2, adds 0.8 times of ethanol, 4 ℃ of hold over night.
Centrifugal, collecting precipitation is weighed, and adds 15 times of (v/w) distilled water, be heated to 85 ℃ ± 2 ℃, until completely dissolved, add sodium hydroxide pH to be adjusted to 9.0 ± 0.2, add according to quantity calcium chloride to solution ultimate density to reach 3%, be warming up to 90 ℃ of above maintenances 10 minutes, be cooled to room temperature, 4 ℃ centrifugal, collects supernatant, with saturated sodium carbonate solution, adjusts pH to 11.0 ± 0.2, centrifugal, collect supernatant, with 6mol/LHCl solution, adjust pH to 6.0 ± 0.2, add 0.8 times of ethanol, 4 ℃ of cold preservations are spent the night.
Cold preservation liquid is centrifugal, and collecting precipitation is weighed, and adds 2 times of volume distilled water, and heating is fully dissolved it, and adding potassium acetate, to make its ultimate density be 2mol/L, 4 ℃ of hold over night.Centrifugal, collecting precipitation is weighed, and adds 2 times of volume distilled water, and heating is fully dissolved it, and adding potassium acetate, to make its ultimate density be 2mol/L, 4 ℃ of hold over night.Centrifugal, for precipitating, cold 2mol/L potassium acetate solution washing is three times, then uses successively volumetric concentration 80% ethanol, volume 95% ethanol, absolute ethanol washing, and after ethanol volatilization to the greatest extent, 80 ℃ are dried, and weigh, and obtain crude product III.
Crude product III 35g adds 0.05mol/L, the HAc-NaAc buffer of pH6.0 dissolves the solution upper prop of making weight concentration 2%, solution is by after cellulose chromatography post, HAc-NaAc buffer (pH6.0 ± 0.2) washing with the 0.4mol/LNaCl of 1.5 times of column volumes, use again HAc-NaAc buffer (pH6.0 ± 0.2) eluting of 1mol/LNaCl, according to Ultraviolet Detector, in the pace of change of 220nm place numerical value, collect eluent, put in 60 ℃ of water-baths, with NaOH, adjust pH to 11 ± 0.2, by volume amount adds 3% hydrogen peroxide, keep 4 hours, cooling, centrifugal, collect supernatant, with HCl, adjust pH to 7.0 ± 0.2, add 1 times of amount ethanol, 4 ℃ of hold over night.
Centrifugal, collecting precipitation, uses 95% ethanol, dehydrated alcohol successively, and weigh (weight in wet base), obtains crude product V.
Crude product V is dissolved into 5% solution with distilled water, with the ultrafilter membrane of molecular cut off 10,000, be concentrated into 1/2 of original volume, benefit adds water to original volume, ultrafiltration to 1/2 volume again, then add water and repeat once, ultrafiltrate lyophilization, obtain glycosaminoglycan extracted from sea cucumber, its molecular weight is all at 90kDa～130kDa, D value < 1.5, and purity is more than 98%.
The glycosaminoglycan extracted from sea cucumber that this example obtains, through differential refraction detector (RID-10A, Shimadzu), can obtain purity is 100.0% sterling (see figure 1).
The glycosaminoglycan extracted from sea cucumber obtaining through this example is known the weight average molecular weight 108756 of this product through gel column (TSK gel G4000PWXL) chromatography, D value is shown in Fig. 2 for 1.40(), wherein: Fig. 2 A is chromatogram, Fig. 2 B is graph of molecular weight distribution, and Fig. 2 C is molecular weight cumulative weight distribution curve.
The preparation of the pharmaceutical composition that injection contains glycosaminoglycan extracted from sea cucumber, above 10g glycosaminoglycan extracted from sea cucumber adds water for injection, is made into 1% solution, add again mannitol 20g to dissolve, fine straining, fill, lyophilization, obtains the lyophilized injectable powder of 1000 injection glycosaminoglycan extracted from sea cucumber.
The pharmacodynamic study of glycosaminoglycan extracted from sea cucumber
The impact of 1 glycosaminoglycan extracted from sea cucumber on rat blood coagulation system
1.1 test objective
Impact on rat blood coagulation system after observation sublingual vein injection glycosaminoglycan extracted from sea cucumber.
1.2 test material
1.2.1 test sample:
The glycosaminoglycan extracted from sea cucumber of embodiment 1; Preparation: after precision takes with injection physiological saline solution and be diluted to desired concn.
1.2.2 experimental animal:
Strain: SD rat; Source: Shanghai western pul-Bi Kai laboratory animal company limited; Sex: male; Body weight: 220-250 gram; The animal quality certification number: SCXK(Shanghai) 2008-0016; Raise: animal feeding in positive pressure purification ventilation Animal House, 23 ± 1 ℃ of room temperatures, humidity 50～70%, artificial lighting's simulation changes round the clock, ad lib and drinking-water.
1.2.3 test apparatus
Application of automated coagulation analyzer Sysmex CA-1500
1.3 test method
By 40 of SD rats, be divided into 4 different administration groups, negative control group (sublingual vein injecting normal saline 0.5ml/100g); Basic, normal, high three the dosage groups of GAG (0.3,0.6,1mg/kg) sublingual vein drug administration by injection, volume 0.5ml/100g, injection speed 1min.
Basic, normal, high three the dosage groups of GAG, after blank group sublingual vein drug administration by injection, PT, APTT, TT numerical value are measured in the blood sampling of 30min ventral aorta.
Each treated animal 3% secobarbital intraperitoneal injection of anesthesia (0.1ml/100g body weight) for 10min before operation, the fixedly pneumoretroperitoneum operation of lying on the back, with disposable 3.2% sodium citrate anticoagulant vacuum blood collection blood sampling tube.
1.4 result of the test
GAG all has no significant effect APTT, TT, PT at low, middle dosage (0.3mg/kg, 0.6mg/kg).But when 1mg/kg, produce obvious change, but do not surpass 150%-250% scope.
Table 1GAG rat clotting time
Table 2GAG rat cruor time extending rate
2 glycosaminoglycan extracted from sea cucumber intravenous injections affect dog blood coagulation system
2.1 test objective
Observe the impact of intravenous injection glycosaminoglycan extracted from sea cucumber on dog blood coagulation system.
2.2 test material
2.2.1 test sample:
The glycosaminoglycan extracted from sea cucumber of embodiment 1;
2.2.2 experimental animal
Strain: beagle dog; Source: the dry house pet of first base; Sex: male; Body weight: 8-9kg
The animal quality certification number: SCXK(Shanghai) 2010-0028
2.2.3 test apparatus
Application of automated coagulation analyzer Sysmex CA-1500.
2.3 experimental technique
4 of Beagle dogs, intravenous injection is given and GAG 0.3mg/kg, 0.6mg/kg, 3.0mg/kg respectively, negative control group intravenous injection normal saline, bolus volumes 20ml/, intravenous injection speed 5 minutes.
The front 0min of intravenous injection, finish rear 30min, 60min, 150min blood sampling and measure each dosed administration beagle dog PT, APTT, TT numerical value.Beagle dog rotation dosage group administration blood sampling is measured, and tests 7 times, calculates each dosage group PT, APTT, TT digital average value and self cruor time extending rate.
2.4 experimental result
GAG intravenous injection Beagle dog all has no significant effect APTT, TT, PT at low, middle dosage (0.3mg/kg, 0.6mg/kg).When 3mg/kg, after intravenous injection, 30minAPTT, TT occur significantly to change, but after 60min, basic recovery is normal.
Table 3 dog intravenous injection GAG clotting time
Table 3 continued dog intravenous injection GAG clotting time
Table 4 dog intravenous injection cruor time extending rate
3 glycosaminoglycan extracted from sea cucumber intravenous drips affect dog blood coagulation system
3.1 test objective
Observe the impact of intravenous drip glycosaminoglycan extracted from sea cucumber on beagle dog blood coagulation system.
3.2 test material
3.2.1 test sample:
The glycosaminoglycan extracted from sea cucumber of embodiment 1; Preparation: after accurate absorption with injection normal saline dilution to desired concn.
3.2.2 experimental animal
Strain: beagle dog; Source: the dry house pet of first base; Sex: male; Body weight: the 11-12kg animal quality certification number: SCXK(Shanghai) 2010-0028
3.2.3 test apparatus
Application of automated coagulation analyzer Sysmex CA-1500
3.3 experimental technique
6 of Beagle dogs, are divided into intravenous drip GAG 0.3mg/kg group, 1mg/kg group, normal saline matched group, 2 every group.Intravenous drip volume 100ml, drip velocity 100ml/1h.
Beagle dog is anaesthetized (1ml/1kg body weight) with 3% secobarbital intravenous injection, and 30min, 60min in the front 0min of intravenous drip, administration instil and finish rear 10min, 30min, each dosed administration of 60min blood sampling mensuration beagle dog PT, APTT, TT numerical value.
Beagle dog rotation dosage group administration blood sampling is measured, and tests 3 times, calculates each dosage group PT, APTT, TT digital average value and self cruor time extending rate.
3.4 experimental result
GAG intravenous drip beagle dog is at 0.3mg/kg, 1mg/kg dosage, and on 100ml/1h drip velocity, APTT, PT, TT all find no obvious variation.
Table 5 dog intravenous drip GAG clotting time
Table 5 continued dog intravenous drip GAG clotting time
Table 6 dog intravenous drip cruor time extending rate
The impact of 4 glycosaminoglycan extracted from sea cucumber on rat arterio-venous catheter thrombotic model
4.1 test objective
Impact on rat arterio-venous catheter thrombus model thrombosis quality after observation intravenous injection glycosaminoglycan extracted from sea cucumber.
4.2 test material
4.2.1 test sample:
The glycosaminoglycan extracted from sea cucumber of embodiment 1; Preparation: after accurate absorption with injection normal saline dilution to desired concn.
4.2.2 control sample:
Title: heparin; Source: Chemical Reagent Co., Ltd., Sinopharm Group; Lot number: F20091029
Content: 150U/mg; Preparation: after precision takes with injection physiological saline solution and be diluted to desired concn.
4.2.3 experimental animal
Strain: SD rat; Source: Shanghai western pul-Bi Kai laboratory animal company limited; Sex: male;
Body weight: 250-300 gram; The animal quality certification number: SCXK(Shanghai) 2008-0016; Raise: animal feeding in positive pressure purification ventilation Animal House, 23 ± 1 ℃ of room temperatures, humidity 50～70%, artificial lighting's simulation changes round the clock, ad lib and drinking-water.
4.2.4 test apparatus
BS110s type electronic balance, SARTORIUS company produces, minimum weighing value 0.1mg.
4.3 test method
By 36 of SD rats, be divided into 5 different administration groups, negative control group (normal saline 1ml/kg), basic, normal, high three the dosage groups of GAG (0.1,0.3,2mg/kg), positive control heparin group (0.1mg/kg).All medicines are sublingual vein drug administration by injection, volume 5ml/kg.
Each treated animal before operation 10min with after 12% chloral hydrate intraperitoneal injection of anesthesia (350～400mg/kg), lie on the back fixing, cut skin of neck, separated left carotid artery and right side external jugular vein, with a shunt valve, connect, manage the long 4 trumpeter's art silk threads of a mid-7cm.In administration, after 20 minutes, open respectively blood flow 15 minutes, then take out silk thread and weigh, deduct silk thread weight, be wet weight of thrombus.Calculate wet weight of thrombus meansigma methods and the standard deviation of each test group, with t-check, with normal saline group, compare.And be calculated as follows the wet weight of thrombus suppression ratio of each test group:
4.4 result of the test
Result of the test is in Table 7.Can see, positive drug and tested medicine are tested the formation that can significantly suppress thrombosis after 20 minutes in administration.Tested medicine is proportional to thrombotic inhibitory action and dosage.With the tested medicine comparison of Isodose, positive drug effect is better than tested medicine.
The impact of table 7GAG on rat arterio-venous catheter thrombotic model
Compare with negative group:
*p<0.05,
*p<0.01
The impact that 5 glycosaminoglycan extracted from sea cucumber are assembled ADP induced platelet
5.1 test objective
Observe the impact of intravenous injection glycosaminoglycan extracted from sea cucumber on ADP induction rat platelet aggregation.
5.2 test material
5.2.1 test sample:
The glycosaminoglycan extracted from sea cucumber of embodiment 1; Preparation: after accurate absorption with injection normal saline dilution to desired concn
5.2.2 control sample:
Title: heparin; Source: Chemical Reagent Co., Ltd., Sinopharm Group; Lot number: F20091029
Content: 150U/mg; Preparation: after precision takes with injection physiological saline solution and be diluted to desired concn
5.2.3 experimental animal
Strain: SD rat; Source: Shanghai western pul-Bi Kai laboratory animal company limited; Sex: male;
Body weight: 250-300 gram; The animal quality certification number: SCXK(Shanghai) 2008-0016; Raise: animal feeding in positive pressure purification ventilation Animal House, 23 ± 1 ℃ of room temperatures, humidity 50～70%, artificial lighting's simulation changes round the clock, ad lib and drinking-water.
5.2.4 test apparatus
80-2 type centrifuge, Shanghai Surgical Operation Equipment Factory produces.MK4/HC platelet count instrument (production of U.S. Baker Instruments company).Platelet aggregation instrument (production of CHRONO-LOG Corporation company).
5.2.5 other reagent
ADP, lot number: 109K7019, Sigma company produces, and with pH7.4 phosphate buffer, is made into 1m/l concentration solution for standby.Put-20 ℃ of Refrigerator stores standby.
5.3 experimental technique
By 81 of SD rats, be divided into 5 groups, negative control group (normal saline 1ml/kg), basic, normal, high three the dosage groups of GAG (0.1,0.3,2mg/kg), positive controls heparin (6mg/kg), and each group is divided after administration 10 minutes, 30 minutes, 1 hour three time point.Sublingual vein drug administration by injection, volume is 5ml/kg.
Respectively at administration 10 minutes, 30 minutes, after 1 hour with 12% chloral hydrate intraperitoneal injection of anesthesia (350～400mg/kg), lie on the back fixing, hara kiri skin, separated ventral aorta tremulous pulse intubate, after emitting blood, use 50u/ml heparin in the anticoagulant of 1:9 ratio, with 500rpm centrifugal 5 minutes, get top blood plasma and be platelet rich plasma (PRP), remaining part with 3500rpm centrifugal 15 minutes again, supernatant is platelet poor plasma (PPP), with platelet count instrument counting PRP platelet count, with PPP, adjust platelet count to 6 * 105/mm3 left and right of PRP.In reaction cup, add PRP500 l and add the bar magnet that is numbered #311 to be put in incubation hole, incubation 5 minutes, separately with a PPP pipe, put into reaction cup, do not need to add bar magnet, incubation 5 minutes, regulates monitor to zero point, then adds ADP5l, making ADP final concentration is 10M, and record adds the maximum agglutination rate after ADP.
Calculate average maximum agglutination rate and the standard deviation of each test group, with t-check, with group of solvents, compare.And be calculated as follows the gathering suppression ratio of each test group:
5.4 experimental result
Table 8 result shows, GAG3 dosage group (0.1mg/kg, 0.3mg/kg, 2mg/kg) all has inhibitory action to the rat platelet aggregation of ADP induction, low dose group has remarkable inhibitory action (P<0.05) to the rat platelet aggregation of ADP induction, and middle dosage, high dose group have highly significant inhibitory action (P<0.01) to the rat platelet aggregation of ADP induction; Tested medicine is proportional to anticoagulant effect and dosage.Positive control heparin group, is compared and be there is no notable difference (P>0.05) with negative control normal saline group without obvious inhibitory action the rat platelet aggregation of ADP induction.
The impact of table 8.GAG on the rat platelet aggregation of ADP induction
*p<0.05,
*p<0.01 and NS group are relatively.
Claims (7)
1. the application of glycosaminoglycan extracted from sea cucumber in preparation treatment coronary syndrome medicine, more than one in the glycosaminoglycan extracted from sea cucumber that the weight average molecular weight of described glycosaminoglycan extracted from sea cucumber is 90kDa to 130kDa.
2. application according to claim 1, is characterized in that, preferred, the weight average molecular weight of described glycosaminoglycan extracted from sea cucumber is more than one in 95kDa～125kDa, more preferably more than one in 90kDa～120kDa.
3. application according to claim 1, is characterized in that, described coronary syndrome comprises acute myocardial infarction, unstable angina pectoris or arterial thrombus.
4. a pharmaceutical composition, comprises glycosaminoglycan extracted from sea cucumber and pharmaceutically acceptable carrier described in claim 1～3 any one for the treatment of effective dose.
5. pharmaceutical composition according to claim 4, is characterized in that, described pharmaceutical composition is lyophilized injectable powder.
6. pharmaceutical composition according to claim 5, is characterized in that, the weight content of described glycosaminoglycan extracted from sea cucumber is 0.5-2%.
7. pharmaceutical composition according to claim 6, is characterized in that, intravenous drip speed is 100ml/h; Intravenous drip dosage 6mg/kg～0.01mg/kg rat body weight.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014183467A1 (en) * | 2013-05-13 | 2014-11-20 | 上海开润生物医药有限公司 | Use of black sea cucumber glycosaminoglycan in preparing medicine for prevention and treatment of thromboembolic disease |
| WO2014183466A1 (en) * | 2013-05-13 | 2014-11-20 | 上海开润生物医药有限公司 | Use of sea cucumber glycosaminoglycan in preparing medicine for prevention and treatment of thromboembolic disease |
| CN113018461A (en) * | 2021-03-09 | 2021-06-25 | 南京邮电大学 | Fucoidin embolism microsphere capable of magnetic resonance imaging and preparation method thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1579415A (en) * | 2004-05-20 | 2005-02-16 | 陈任重 | Yuzu Sea-cucumber osamine glycan injecta and its preparation method |
| CN102443077A (en) * | 2011-12-31 | 2012-05-09 | 中国海洋大学 | Isostichopus badionotus fucosylated mucopolysaccharide and application thereof |
| CN103417565A (en) * | 2012-05-17 | 2013-12-04 | 上海开润生物医药有限公司 | Application of depolymerized holothurian glycosaminolycan in preparing drugs for curing and preventing thrombotic diseases |
-
2012
- 2012-07-11 CN CN201210240028.3A patent/CN103536621A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1579415A (en) * | 2004-05-20 | 2005-02-16 | 陈任重 | Yuzu Sea-cucumber osamine glycan injecta and its preparation method |
| CN102443077A (en) * | 2011-12-31 | 2012-05-09 | 中国海洋大学 | Isostichopus badionotus fucosylated mucopolysaccharide and application thereof |
| CN103417565A (en) * | 2012-05-17 | 2013-12-04 | 上海开润生物医药有限公司 | Application of depolymerized holothurian glycosaminolycan in preparing drugs for curing and preventing thrombotic diseases |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014183467A1 (en) * | 2013-05-13 | 2014-11-20 | 上海开润生物医药有限公司 | Use of black sea cucumber glycosaminoglycan in preparing medicine for prevention and treatment of thromboembolic disease |
| WO2014183466A1 (en) * | 2013-05-13 | 2014-11-20 | 上海开润生物医药有限公司 | Use of sea cucumber glycosaminoglycan in preparing medicine for prevention and treatment of thromboembolic disease |
| CN113018461A (en) * | 2021-03-09 | 2021-06-25 | 南京邮电大学 | Fucoidin embolism microsphere capable of magnetic resonance imaging and preparation method thereof |
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