CN107488214A - Its synthesis of the O acetyl LPNISKP of warfarin 4, activity and application - Google Patents
Its synthesis of the O acetyl LPNISKP of warfarin 4, activity and application Download PDFInfo
- Publication number
- CN107488214A CN107488214A CN201610415117.5A CN201610415117A CN107488214A CN 107488214 A CN107488214 A CN 107488214A CN 201610415117 A CN201610415117 A CN 201610415117A CN 107488214 A CN107488214 A CN 107488214A
- Authority
- CN
- China
- Prior art keywords
- pro
- warfarin
- ile
- asn
- lys
- 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
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Genetics & Genomics (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Life Sciences & Earth Sciences (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses the O acetyl Leu Pro Asn Ile Ser Lys Pro of warfarin 4, disclose its preparation method, disclose its anti-arterial thrombus activity, disclose its anti-phlebothrombosis activity, disclose it reduce a (a of II b/ of GP III) of internal platelet membrane glycoprotein Ⅱb/III content activity, the activity of the content of its internal factor II (F II) of reduction is disclosed, discloses the activity that they suppress platelet aggregations.Thus the invention discloses them in the application in preparing anti-arterial thrombus medicine, in the application in preparing anti-phlebothrombosis medicine, application, the application in a antagonists of II b/ of GP III are prepared and the application in prothrombin antagonist is prepared in suppression platelet aggregation drugs are prepared.
Description
Technical field
The present invention relates to warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro, it is related to its preparation side
Method, it is related to its anti-arterial thrombus activity, is related to their anti-phlebothrombosis activity, being related to it reduces the internal a's of II b/ of GP III
The effect of content, it is related to the effect of the content of its internal factor II of reduction, is related to the effect that it suppresses platelet aggregation.Cause
And the present invention relates to it in the application in preparing anti-arterial thrombus medicine, the application in anti-phlebothrombosis medicine is prepared, making
It is standby to suppress the application in platelet aggregation drugs, the application in a antagonists of II b/ of GP III are prepared and preparing prothrombin
Application in antagonist.The invention belongs to biomedicine field.
Background technology
Two kinds of illnesss of arterial thrombus and phlebothrombosis have turned into incidence of disease height and the high disease of the death rate instantly.Its medium sized vein
DVT mainly includes DVT and pulmonary embolism.DVT and patient's number of pulmonary embolism morbidity have exceeded miocardial infarction
The total number of persons fallen ill with apoplexy, and the total number of persons more dead than caused by breast cancer and AIDS is high.Due to arterial thrombosis and
The incidence of disease of Venous Thrombosis increases exponentially state increase with the age, so two kinds of illnesss are to aging country as China
The threat of people's health is especially serious.If population base is taken into account, the absolute negative effect to China's national economy is outstanding
Its is serious.Then, the prevention of arterial thrombosis and Venous Thrombosis and treatment always field of medicaments emphasis of interest.Although
Warfarin is used for clinical practice in nineteen forty-one as medicine is represented, but because the activity of warfarin is by its pharmacokinetics
The common complexing action of the availability of feature, clotting factor metabolism and vitamin K shows.The dose-effect relationship table of warfarin
Reveal very big changeability, also just the anticoagulant therapy of explanation warfarin has a very big dosage range, and show very for this
Big individual difference.Due to the window narrows of warfarin, its underdosage has the possibility for causing pulmonary embolism, and dosage is excessively led
Cause the risk of fatal hemorrhage.A large amount of structure of modification were carried out to warfarin in past more than 50 years, it is but all no to be resisted
The analog that phlebothrombosis activity is strong and hemorrhage side effect is low.Different from traditional thinking, inventor modifies warfarin structure
Target is that warfarin is converted into the analog with anti-arterial thrombus and anti-phlebothrombosis double activity.But, also always not
It can achieve one's goal.Cell adherence take part in the occurrence and development process of a variety of diseases of the mankind.Mediated by cell adhesion molecule, attachment proteins
Adhesion between cell and between cell and extracellular matrix, and they between thrombosis associate to have obtained it is widely studied
The concern of person.The goal in research of polypeptide of the inventor to suppressing cell adhesion activity is, by with suppression cell adhesion activity
Polypeptide Leu-Pro-Asn-Ile-Ser-Lys-Pro is converted into similar with anti-phlebothrombosis double activity with anti-arterial thrombus
Thing.But, also fail to achieve one's goal always.Afterwards, explored again by 5 years, inventor has found 4 of warfarin with acetyl Leu-Pro-
Asn-Ile-Ser-Lys-Pro is modified, and the warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro of generation are in body
Outstanding platelet aggregation inhibitory activity is shown on outer platelet aggregation-against model.Obviously, they are not the prodrugs of warfarin.Hua Fa
Woods -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro anti-arterial thrombus activity is stronger than aspirin 167 times, resists quiet
Arteries and veins thrombus activity is stronger than warfarin 48.7 times.It can be seen that the present invention has significant technological progress.Then, inventor proposes this hair
It is bright.
The content of the invention
First content of the present invention is to provide warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro.
Second content of the present invention is to provide warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro's
Synthetic method, this method include：
1. synthesize warfarin -4-O- benzyl acetates；
2. synthesize warfarin -4-O- acetic acid；
3. using DCC as condensing agent, HOBt is the method for the liquid phase condensations of catalyst, synthesizes HClLeu-Pro-Asn-
Ile-Ser-Lys(Z)-Pro-OBzl
4. using DCC as condensing agent, HOBt is the method for the liquid phase condensations of catalyst, warfarin -4-O- acetic acid and HCl
Leu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl is condensed
5. synthesize warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro
The 3rd content of the present invention is that evaluation warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro resists
The effect of arterial thrombus.
The 4th content of the present invention is that evaluation warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro resists
The effect of phlebothrombosis.
The 5th content of the present invention is the-Leu-Pro-Asn-Ile-Ser-Lys-Pro suppressions of evaluation warfarin -4-O- acetyl
The effect for the platelet aggregation that system is induced by platelet activating factor (PAF), fibrin ferment (TH) and adenosine diphosphate (ADP) (ADP).
The 6th content of the present invention is evaluation warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro drops
The effect of the low a contents of II b/ of GP III in vivo.
The 7th content of the present invention is evaluation warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro drops
The effect of low factor II content in vivo.
Brief description of the drawings
Fig. 1 warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro bromo- 2- the acetic acid of synthetic route (i)
Benzyl ester, acetone, K2CO3,45℃；(ii)CH3OH,Pd/C,H2；(iii)DCC,HOBt,NMM,THF；(iv) second of 4N hydrogen chloride
Acetate solution；(v)2N NaOH,CH3OH。
Fig. 2 warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro anti-arterial thrombus activity, n=10；
The anti-phlebothrombosis activity of Fig. 3 warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro, n=10；
The blood that Fig. 4 warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro are induced PAF, TH and ADP is small
The inhibitory action of plate aggregation, n=3.
Shadows of Fig. 5 warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro to the internal a contents of II b/ of GP III
Ring, n=5；
Shadows of Fig. 6 warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro to the contents of F II in rat body
Ring, n=5.
Embodiment
In order to which the present invention is expanded on further, a series of embodiments are given below.These embodiments be entirely it is illustrative, it
Only be used for the present invention is specifically described, be not construed as limitation of the present invention.
Embodiment 1 prepares warfarin -4-O- benzyl acetates
3.31g (10.00mmol) warfarin is placed in 100mL eggplant bottles, adds about 40mL acetone, is failed it is completely molten
Solution, 45 DEG C of oil bath heatings, which are stirred to warfarin, to be dissolved, and is added 1.73mL (11.00mmol) bromo- 2- benzyl acetates, is continued at 45 DEG C
Reacted under oil bath, find there is white solid to invest in bottle wall after about 1h.After reacting 48h, thin-layer chromatography (TLC, petroleum ether/
Ethyl acetate=2:1) reaction process is monitored, warfarin disappears, colorless solid caused by reaction is filtered off, acetone is removed under reduced pressure,
Pale yellow oil is obtained, (petrol ether/ethyl acetate=8 are purified by silica gel column chromatography:1) 3.02g (65%) title, is obtained
Compound, it is colorless solid.ESI-MS(m/e):457[M+H]+；1H-NMR(300MHz,DMSO-d6) δ/ppm=7.89 (dd,
J1=3.0Hz, J2=9.0Hz, 1H), 7.63 (dt, J1=3.0Hz, J2=9.0Hz, 1H), 7.43~7.31 (m, 9H), 7.24
(t, J=9.0Hz, 2H), 7.15 (tt, J=9.0Hz, 1H), 5.26 (s, 2H), 5.61 (s, 1H), 5.02 (d, J=15.0Hz,
1H), 4.85 (d, J=15.0Hz, 1H), 4.97 (t, J=9.0Hz, 1H), 3.45 (dq, J1=9.0Hz, J2=18.0Hz, 2H),
2.11(s,3H)。
Embodiment 2 prepares warfarin -4-O- acetic acid
2.26g (4.95mmol) warfarin -4-O- benzyl acetates are dissolved in 20mL methanol, add 220mg palladium carbons (Pd/
C), under stirring in water pump vacuumizing, be passed through hydrogen, so operation repeat 3 times, be passed through hydrogen and stir 10h at room temperature.
TLC monitoring reactions are complete, are filtered to remove Pd/C, and filtrate decompression concentration removes solvent, and residue is solidified with petroleum ether, absolute ether
Worn away, obtain 1.72g (93%) title compound, be colorless solid.ESI-MS(m/e):367[M+H]+；1H-NMR
(300MHz,DMSO-d6):δ/ppm=12.86 (s, 1H), 7.90 (d, J=6.0Hz, 1H), 7.63 (t, J=6.0Hz, 1H),
7.43~7.34 (m, 4H), 7.27 (t, J=9.0Hz, 2H), 7.17 (t, J=9.0Hz, 1H), 4.99 (t, J=9.0Hz, 1H),
4.75(q,J1=15.0Hz, J2=30.0Hz, 2H), 3.54~3.47 (m, 2H), 2.14 (s, 3H).
Embodiment 3 prepares Boc-Asn-Ile-OBzl
5.05g (21.76mmol) Boc-Asn is added in 250mL eggplant bottles with 100mL anhydrous tetrahydro furans that its is molten
Solution, 2.91g (21.55mmol) HOBt and 5.33g (25.87mmol) DCC is added under ice bath (0 DEG C), activate 30min.Have big
Dicyclohexylurea (DCU) (DCU) is measured to separate out.8.47g (21.55mmol) TosIle-OBzl is dissolved in 100mL tetrahydrofurans, will
It is added in reaction solution under ice bath, is adjusted pH value to 8~9 with N-methylmorpholine (NMM), is stirred at room temperature after reacting 5h
TLC (methylene chloride/methanol=20:1) reaction process is monitored, raw material point disappears, filters out DCU, filtrate decompression is removed into solvent, residual
Stay thing 100mL ethyl acetate to dissolve, filter off insoluble DCU, filtrate uses saturation Na respectively2CO3Solution (40mL × 3), saturation
NaCl solution (40mL × 3), saturation KHSO4Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation Na2CO3Solution
More than 2h is dried with anhydrous sodium sulfate in (40mL × 3), saturation NaCl solution (40mL × 3) washing, ethyl acetate phase.Filter off
Sodium sulphate, filtrate decompression remove solvent, obtain faint yellow solid 9.02g (96%).ESI-MS(m/e):436[M+H]+。
Embodiment 4 prepares HClAsn-Ile-OBzl
9.00g Boc-Asn-Ile-OBzl are weighed in 250mL eggplant bottles, 15mL anhydrous ethyl acetates is added, makes its complete
Dissolving, the ethyl acetate solution of 90mL 4N hydrogen chloride is added under ice salt bath (- 10 DEG C), continues the stirring reaction under ice bath.
TLC (methylene chloride/methanol=20:1) reaction process is monitored, raw material point disappears, reaction solution is concentrated under reduced pressure with water pump, will be remained
Thing is concentrated under reduced pressure again after being redissolved with anhydrous ethyl acetate, 3 times repeatedly, obtains yellow oil, is depressurized again after being infiltrated with absolute ether
Concentration, 3 times repeatedly, obtains faint yellow oily residue 7.67g (99%).
Embodiment 5 prepares Boc-Pro-Asn-Ile-OBzl
4.45g (20.69mmol) Boc-Pro is added in 250mL eggplant bottles with 100mL anhydrous tetrahydro furans that its is molten
Solution, 2.79g (20.66mmol) HOBt and 5.12g (24.85mmol) DCC is added under ice bath (0 DEG C), activate 30min.Have big
DCU is measured to separate out.7.66g (20.61mmol) HClAsn-Ile-OBzl is dissolved in 100mL tetrahydrofurans, by it in ice bath
In lower addition reaction solution, pH value is adjusted to 8~9 with NMM, TLC (methylene chloride/methanol=20 after reaction 7h are stirred at room temperature:
1) reaction process is monitored, raw material point disappears, filters out DCU, filtrate decompression is removed into solvent, residue is molten with 100mL ethyl acetate
Solution, filters off insoluble DCU, filtrate uses saturation Na respectively2CO3Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation
KHSO4Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation Na2CO3Solution (40mL × 3), saturation NaCl solution
(40mL × 3) are washed, and more than 2h is dried with anhydrous sodium sulfate in ethyl acetate phase.Sodium sulphate is filtered off, filtrate decompression removes molten
Agent, faint yellow solid is obtained, carry out silica gel column chromatography purifying (methylene chloride/methanol=80:1) colorless solid 8.26g, is obtained
(75%).ESI-MS(m/e):533[M+H]+。
Embodiment 6 prepares HClPro-Asn-Ile-OBzl
6.26g (11.76mmol) Boc-Pro-Asn-Ile-OBzl are weighed in 250mL eggplant bottles, add the anhydrous second of 10mL
Acetoacetic ester, it is completely dissolved, the ethyl acetate solution of 60mL 4N hydrogen chloride is added under ice salt bath (- 10 DEG C), is continued in ice
The lower stirring reaction of bath.TLC (methylene chloride/methanol=20:1) reaction process is monitored, raw material point disappears, subtracted reaction solution with water pump
Pressure concentration, is concentrated under reduced pressure, 3 times repeatedly, obtains yellow oil, use absolute ether again after residue is redissolved with anhydrous ethyl acetate
It is concentrated under reduced pressure again after infiltration, 3 times repeatedly, obtains faint yellow oily residue 5.50g (98%).
Embodiment 7 prepares Boc-Leu-Pro-Asn-Ile-OBzl
2.72g (11.78mmol) Boc-Leu is added in 250mL eggplant bottles with 100mL anhydrous tetrahydro furans that its is molten
Solution, 1.59g (11.78mmol) HOBt and 2.91g (14.12mmol) DCC is added under ice bath (0 DEG C), activate 30min.Have big
DCU is measured to separate out.5.49g (11.72mmol) HClPro-Asn-Ile-OBzl is dissolved in 100mL anhydrous tetrahydro furans, will
It is added in reaction solution under ice bath, and pH value is adjusted to 8~9 with NMM, be stirred at room temperature TLC after reaction 4h (dichloromethane/
Methanol=20:1) monitoring reaction is completed, and filters out DCU, and filtrate decompression is removed into solvent, and residue 100mL ethyl acetate dissolves,
Insoluble DCU is filtered off, filtrate uses saturation Na respectively2CO3Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation
KHSO4Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation Na2CO3Solution (40mL × 3), saturation NaCl solution
(40mL × 3) are washed, ethyl acetate phase more than anhydrous sodium sulfate drying 2h.Sodium sulphate is filtered off, filtrate decompression removes solvent, obtained
To faint yellow solid, column chromatography purifying (methylene chloride/methanol=100 are carried out:1) colorless solid 6.37g (84%), is obtained.
ESI-MS(m/e):646[M+H]+。
Embodiment 8 prepares Boc-Leu-Pro-Asn-Ile
5.02g (7.78mmol) Boc-Leu-Pro-Asn-Ile-OBzl are weighed in 250mL eggplant bottles, add 10mL methanol
Dissolved, pH value is adjusted to 12 with the 2N NaOH aqueous solution under ice bath (0 DEG C), stirring reaction about 4h, TLC (dichloromethane/
Methanol=20:1) reaction process is monitored, raw material point disappears, with saturation KHSO4PH value is adjusted to 7, is concentrated under reduced pressure and removes methanol, adjust
PH value is saved to 3, is extracted (50mL × 3) with ethyl acetate, then washed (40mL × 3) with saturation NaCl, ethyl acetate layer use
More than anhydrous sodium sulfate drying 2h, removal of solvent under reduced pressure, worn away with petroleum ether, filter, obtain colorless oil 4.20g
(97%).
Embodiment 9 prepares Boc-Lys (Z)-Pro-OBzl
5.07g (13.34mmol) Boc-Lys (Z) is dissolved in 250mL eggplant bottles with 100mL anhydrous tetrahydro furans,
1.78g (13.18mmol) HOBt and 3.25g (15.77mmol) DCC is added under ice bath (0 DEG C), activates 30min.Have a large amount of
DCU is separated out.3.18g (13.16mmol) HClPro-OBzl is dissolved in 100mL anhydrous tetrahydro furans, by it under ice bath
Add in reaction solution, adjust pH value to 8~9 with NMM, TLC (methylene chloride/methanol=20 after reaction 4h are stirred at room temperature:1)
Monitoring reaction is completed, and filters out DCU, filtrate decompression is removed into solvent, residue 100mL ethyl acetate dissolves, filtered off insoluble
DCU, filtrate use saturation Na respectively2CO3Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation KHSO4Solution (40mL
× 3), saturation NaCl solution (40mL × 3), saturation Na2CO3Solution (40mL × 3), saturation NaCl solution (40mL × 3) washing,
Ethyl acetate phase more than anhydrous sodium sulfate drying 2h.Sodium sulphate is filtered off, filtrate decompression removes solvent, obtains faint yellow solid
7.02g (94%).ESI-MS(m/e):568[M+H]+。
Embodiment 10 prepares HClLys (Z)-Pro-OBzl
6.99g (12.32mmol) Boc-Lys (Z)-Pro-OBzl are weighed in 250mL eggplant bottles, add 30mL anhydrous acetic acids
Ethyl ester, it is completely dissolved, the ethyl acetate solution of 70mL 4N hydrogen chloride is added under ice salt bath (- 10 DEG C DEG C), is continued in ice
The lower stirring reaction of bath.TLC (methylene chloride/methanol=20:1) reaction solution is concentrated under reduced pressure with water pump after the completion of display reaction, will
Residue is concentrated under reduced pressure again after being redissolved with anhydrous ethyl acetate, 3 times repeatedly, obtains yellow oil, after being infiltrated with absolute ether again
It is concentrated under reduced pressure, 3 times repeatedly, obtains faint yellow oily residue 6.15g (98%).
Embodiment 11 prepares Boc-Ser-Lys (Z)-Pro-OBzl
2.53g (12.34mmol) Boc-Ser are dissolved in 250mL eggplant bottles with 100mL anhydrous tetrahydro furans, in
1.67g (12.37mmol) HOBt and 3.05g (14.80mmol) DCC is added under ice bath (0 DEG C), activates 30min.There are a large amount of DCU
Separate out.6.11g (12.13mmol) HClLys (Z)-Pro-OBzl is dissolved in 100mL anhydrous tetrahydro furans, by it in ice
Bath is lower to be added in reaction solution, and pH value is adjusted to 8~9 with NMM, be stirred at room temperature TLC after reaction 6h (methylene chloride/methanol=
20:1) reaction process is monitored, raw material point disappears, filters out DCU, filtrate decompression is removed into solvent, residue 100mL ethyl acetate
Dissolving, filters off insoluble DCU, filtrate uses saturation Na respectively2CO3Solution (40mL × 3), saturation NaCl solution (40mL × 3), satisfy
And KHSO4Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation Na2CO3Solution (40mL × 3), saturation NaCl are molten
Liquid (40mL × 3) washs, ethyl acetate phase more than anhydrous sodium sulfate drying 2h.Sodium sulphate is filtered off, filtrate decompression removes solvent,
Faint yellow solid is obtained, carries out column chromatography purifying (methylene chloride/methanol=100:1) colorless solid 5.17g (62%), is obtained.
ESI-MS(m/e):655[M+H]+。
Embodiment 12 prepares HClSer-Lys (Z)-Pro-OBzl
5.07g (7.75mmol) Boc-Ser-Lys (Z)-Pro-OBzl are weighed in 250mL eggplant bottles, it is anhydrous to add 15mL
Ethyl acetate, it is completely dissolved, the ethyl acetate solution of 50mL 4N hydrogen chloride is added under ice salt bath (- 10 DEG C), is continued
Stirring reaction under ice bath.TLC (methylene chloride/methanol=20:1) reaction process is monitored, raw material point disappears, with water pump by reaction solution
It is concentrated under reduced pressure, is concentrated under reduced pressure again after residue is redissolved with anhydrous ethyl acetate, 3 times repeatedly, obtain yellow oil, with anhydrous second
It is concentrated under reduced pressure again after ether infiltration, 3 times repeatedly, obtains faint yellow oily residue 4.50g (98%).
Embodiment 13 prepares Boc-Leu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl
4.30g (7.75mmol) Boc-Leu-Pro-Asn-Ile are used into 100mL anhydrous tetrahydro furans in 250mL eggplant bottles
Dissolved, 1.05g (7.78mmol) HOBt and 1.92g (9.32mmol) DCC is added under ice bath (0 DEG C), activate 30min.
There are a large amount of DCU to separate out.4.59g (7.78mmol) HClSer-Lys (Z)-Pro-OBzl is dissolved in the anhydrous tetrahydrochysene furans of 100mL
In muttering, it is added in reaction solution under ice bath, pH value is adjusted to 8~9 with NMM, TLC (two after reaction 6h is stirred at room temperature
Chloromethanes/methanol=15:1) monitoring reaction is completed, and filters out DCU, filtrate decompression is removed into solvent, residue 100mL dichloromethanes
Alkane dissolves, and filters off insoluble DCU, filtrate uses saturation Na respectively2CO3Solution (40mL × 3), saturation NaCl solution (40mL × 3),
Saturation KHSO4Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation Na2CO3Solution (40mL × 3), saturation NaCl
Solution (40mL × 3) washs, and dichloromethane mutually uses more than anhydrous sodium sulfate drying 4h.Sodium sulphate is filtered off, filtrate decompression removes molten
Agent, faint yellow solid is obtained, carry out column chromatography purifying (methylene chloride/methanol=20:1) colorless solid 4.17g, is obtained
(49%).ESI-MS(m/e):1092[M+H]+。
Embodiment 14 prepares HClLeu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl
0.91g (0.83mmol) Boc-Leu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl are weighed in 100mL eggplant bottles
In, 3mL anhydrous ethyl acetates are added, are completely dissolved it, the acetic acid of 10mL 4N hydrogen chloride is added under ice salt bath (- 10 DEG C)
Ethyl ester solution, continue the stirring reaction under ice bath.TLC (methylene chloride/methanol=15:1) reaction process is monitored, raw material point disappears
Lose, reaction solution is concentrated under reduced pressure with water pump, is concentrated under reduced pressure again after residue is redissolved with anhydrous ethyl acetate, 3 times repeatedly, is obtained white
Color solid, is concentrated under reduced pressure again after being infiltrated with absolute ether, 3 times repeatedly, obtains white solid 0.84g (99%).
Embodiment 15 prepares warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl
1.02g (2.78mmol) warfarin -4-O- acetic acid is added in 100mL eggplant bottles will with 30mL anhydrous tetrahydro furans
It is dissolved, and 0.37g (2.74mmol) HOBt and 0.68g (3.30mmol) DCC is added under ice bath (0 DEG C), activates 30min.Have
A large amount of DCU are separated out.2.80g (2.73mmol) HClLeu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl is dissolved in
In 30mL anhydrous tetrahydro furans, it is added in reaction solution under ice bath, pH value is adjusted to 8~9 with NMM, is stirred at room temperature
TLC (methylene chloride/methanol=20 after reaction 8h:1) reaction process is monitored, raw material point disappears, filters out DCU, filtrate decompression is removed
Solvent is removed, residue 50mL dichloromethane dissolves, and filters off insoluble DCU, filtrate uses saturation Na respectively2CO3Solution (40mL ×
3), saturation NaCl solution (40mL × 3), saturation KHSO4Solution (40mL × 3), saturation NaCl solution (40mL × 3), saturation
Na2CO3Solution (40mL × 3), saturation NaCl solution (40mL × 3) washing, dichloromethane mutually with anhydrous sodium sulfate drying 2h with
On.Sodium sulphate is filtered off, filtrate decompression removes solvent, obtains faint yellow solid, carries out silica gel column chromatography purifying (dichloromethane/first
Alcohol=8:1), obtain looking for solid 1.21g (67%).ESI-MS(m/e):1362[M+Na]+；1H-NMR(300MHz,DMSO-
d6):δ/ppm=8.42 (d, J=8.1Hz, 1H), 8.24 (d, J=7.2Hz, 1H), 8.13 (d, J=7.5Hz, 1H), 7.87
(d, J=7.8Hz, 2H), 7.63 (td, J1=1.2Hz, J2=7.8Hz, 1H), 7.54 (d, J=8.7Hz, 1H), 7.42~7.14
(m, 18H), 5.10 (m, 2H), 5.00 (s, 2H), 4.92 (m, 1H), 4.64 (s, 2H), 4.53~4.50 (m, 3H), 4.39~
4.30 (m, 3H), 4.23 (t, J=2.1Hz, 1H), 3.65 (m, 1H), 3.56 (m, 2H), 3.46 (m, 4H), 3.44 (m, 3H),
2.92 (m, 2H), 2.60 (m, 1H), 2.42 (m, 1H), 2.13 (s, 3H), 1.92~1.79 (m, 10H), 1.57~1.23 (m,
9H),1.10(m,2H),0.83(m,12H)。
Embodiment 16 prepares warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro (warfarin -4-O- second
Acyl-LPNISKP)
By 1.03g (0.76mmol) warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl
Be dissolved in 50mL methanol, add 100mg Pd/C, under stirring in water pump vacuumizing, be passed through hydrogen, so operation repeat enter
Row 3 times, it is passed through hydrogen stirring reaction about 10h at room temperature.TLC (methylene chloride/methanol=15:1) reaction process, raw material point are monitored
Disappear, Pd/C is filtered to remove, filtrate decompression removes solvent, is solidified into colorless solid with petroleum ether, utilizes C18Column chromatography
Mode is purified, and obtains colorless solid product 231mg (27%).Mp:165-167℃；(c=0.09,
H2O)；ESI-MS(m/e):1116[M+H]+；IR(cm-1):3306,3063,2939,1650,1609,1565,1494,1453,
1396,1354,1272,1227,1198,1165,1146,1102,1070,1019,910,893,756,699,603；1H-NMR
(300MHz,DMSO-d6):δ/ppm=8.46 (dd, J1=7.5Hz, J2=14.1Hz, 1H), 8.28 (m, 1H), 8.10 (d, J=
7.5Hz, 1H), 7.88 (d, J=7.8Hz, 1H), 7.80 (m, 1H), 7.63 (t, J=7.5Hz, 1H), 7.55 (d, J=8.7Hz,
1H), 7.47~7.32 (m, 5H), 7.22 (t, J=7.8Hz, 2H), 7.17 (t, J=7.2Hz, 1H), 7.10 (m, 1H), 5.10
(m, 2H), 4.91 (m, 1H), 4.64~4.61 (m, 2H), 4.51 (m, 2H), 4.38 (m, 1H), 4.30~4.24 (m, 2H),
4.11 (m, 1H), 3.62~3.34 (m, 10H), 2.71 (m, 2H), 2.60 (m, 1H), 2.43 (m, 1H), 2.13 (s, 3H), 2.01
~1.78 (m, 10H), 1.69~1.62 (m, 2H), 1.62~1.42 (m, 4H), 1.42~1.37 (m, 3H), 1.11~1.05
(m, 2H), 0.93~0.76 (m, 12H).
The anti-arterial thrombus that embodiment 17 evaluates warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro is made
With
Experiment material：
Urethane (urethanes, CAS：51-79-6, Chemical Reagent Co., Ltd., Sinopharm Group), liquaemin (CAS：
9041-08-1, lark prestige Science and Technology Ltd.), physiological saline (Shijiazhuang Siyao Co., Ltd).
Experimental animal：
SD strain rats, male, 200 ± 20g, purchased from Beijing Vital River Experimental Animals Technology Co., Ltd..
Experimental method：Experiment forms model using arteriovenous shunt thrombosis.
Bypass the preparation of intubation：
Bypass intubation is formed by three sections, and by internal diameter 1.0mm, it is angle that external diameter 2.0mm polyethylene pipe, which is heated and pulls into one end,
Tubule, fixed length 10.0cm, respectively right jugular vein and left neck artery intubation, positioned at the both ends of bypass intubation；Stage casing is by interior
Footpath 3.5mm polyethylene pipe is formed, fixed length 8.0cm；Be 6.0cm by the crude silk thread fixed length in surface, from weight 4.0 ±
0.1mg and crude degree identical silk thread.
Three sections of Interior Wall of Polythene Pipe carry out silanization with 1% silicon ethereal solution (diethyl ether solution of 1% silicone oil), complete
After drying, silk thread is placed in the polyethylene pipe of stage casing in arteria carotis intubation direction, assembled three sections of polyethylene pipes using sealed membrane
And it is fixed, heparin need to be full of in pipe before intubation.
Packet and dosage：
The compounds of this invention warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro dosage is 0.1 μm of ol/
kg；Positive control aspirin dose is 167 μm of ol/kg and 16.7 μm of ol/kg, and negative control is physiological saline
Agents useful for same is prepared：
Anesthetic is the 20% urethane aqueous solution with normal saline, and anti-coagulants is with normal saline
42mg/100mL heparin sodium water solution.
Experimental implementation：
Gastric infusion, administration 30min pneumoretroperitoneums injection 20% are carried out to rat with the dosage of 0.3mL/100g body weight respectively
Urethane solution anaesthetized (0.7mL/100g).Rat is fixed in rat fixed plate by dorsal position, cuts off skin of neck,
Right common carotid artery and left vena jugularis externa are separated, is respectively tied the distal end of right common carotid artery and left vena jugularis externa with surgical thread
Prick, a V-shape osculum is cut in exposed left vena jugularis externa, the vein end angle that the bypass made above is intubated is inserted outside left neck
The proximal part of vein opening, blood vessel and polyethylene pipe are fixed with surgical thread at intubation, led to the dosage of 0.1mL/100g body weight
The accurate injection heparin sodium water solution of bypass intubation is crossed, syringe does not withdraw polyethylene pipe.It is near that right common carotid artery is clamped with artery clamp
Heart end, a V-shape osculum is cut on exposed artery, the tip of polyethylene pipe is removed from syringe, it is total to insert a tube into right neck
The proximal part of artery, arteries and polyethylene pipe are fixed with surgical thread, unclamp artery clamp, establish extracorporal circulatory system bypass.
Maintain blood flow in rat temperature and bypass intubation unobstructed, first by vein end intubation cutting observation after body circulation 15min
Whether blood circulation is smooth, from the arterial end removal of thromboses line of intubation, weighs and remembers after the floating blood on silk thread is sucked on filter paper
Its weight in wet base is recorded, represents anti-arterial thrombus activity.Data are examined using t and counted.
Experimental result：
Data are included in Fig. 2.As a result 0.1 μm of ol/kg warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser- is shown
Rat suppository weight of the thrombus weight (23.15 ± 5.25mg) of the rat of Lys-Pro treatments significantly less than saline therapy
(30.42 ± 3.49mg, p<0.05) 0.1 μm of ol/kg warfarin -4-O- acetyl-Leu-Pro-Asn-Ile- of compound, is illustrated
Ser-Lys-Pro shows anti-arterial thrombus activity.And the aspirin under 16.7 μm of ol/kg dosage does not have anti-arterial thrombus
Activity, illustrate compound warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro anti-arterial thrombus activity
It is at least stronger 167 times than aspirin.This is unexpected technique effect.
Embodiment 18 evaluates warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro anti-phlebothrombosis shape
Into effect
Experiment material：
Urethane (urethanes, CAS：51-79-6, Chemical Reagent Co., Ltd., Sinopharm Group), physiological saline (stone
The medicine Co., Ltd of the village four of family) warfarin sodium (CAS：129-06-6, lark prestige Science and Technology Ltd.)；
Experimental animal：
SD strain rats, male, 250 ± 20g, purchased from Beijing Vital River Experimental Animals Technology Co., Ltd..
Experimental method：Experiment uses rat Ligation of inferior vena cava model.
Packet and dosage：
Compound warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro the dosage of the present invention is 0.1 μ
Mol/kg, the dosage of positive control warfarin is 4.87 μm of ol/kg, and negative control is physiological saline.
Agents useful for same is prepared：
Anesthetic is 20% urethane solution of normal saline.
Experimental implementation：
Experimental rat adapts to environment simultaneously fasting one day before surgery, and rat is filled with the dosage of 0.3mL/100g body weight
Stomach is administered, and is administered using gavage mode, and administration 30min carries out abdominal cavity after operation consent 2min with 20% urethane solution
Administration anesthesia.Rat is fixed in rat fixed plate, takes blood 2mL from arteria carotis, the measure for blood index of correlation.Will be big
Abdominal cavity is opened along hunter's line after mouse belly preserved skin and sterilization, it is lower to solidification gland, up to expose one jiao of liver.Will be intraperitoneal small
The organs such as intestines are hauled out, and expose inferior caval vein, the organ gauze wrapped for infiltrating physiological saline of hauling-out.Blunt separation blood vessel
Surrounding connective tissue, exposure inferior caval vein and its branch, peel away abdominal aorta and inferior caval vein below renal vein, then
With by the suture that physiological saline soaks in the intersection of inferior caval vein and left renal vein by Ligation of inferior vena cava, by anatomical position
The organs such as intestines are put back into abdominal cavity, with suture layer-by-layer suture abdominal cavity.
Postoperative rat is placed in 25~28 DEG C of environment circulates 4h, after opening abdominal cavity, one by one ligatures its branch, under
Start to take out 2cm inferior caval veins at the ligation of the intersection of vena cave and left renal vein, be taken out thrombus.Calculate blood weight and profit
The mode statistical result examined with t.Operation with every group two only alternately.Experimental data is shown in Fig. 3.
Experimental result：
As a result show, what 0.1 μm of ol/kg warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro was treated
Rat suppository weight (22.93 ± 5.03mg, p of the thrombus weight (14.22 ± 3.90mg) of rat significantly less than saline therapy<
0.01), illustrate that compound warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro shows anti-phlebothrombosis and lived
Property.And controlled in 0.1 μm of ol/kg dosages for Compound warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro
Bolt weight (12.12 ± 3.86mg, the p of the rat of warfarin treatment under the bolt weight of the rat for the treatment of and 4.87 μm of ol/kg dosage>0.05)
It is not significantly different, illustrates compound warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro anti-venous blood
Thrombus activity is at least stronger 48.7 times than warfarin.This is unexpected technique effect.
Embodiment 19 evaluates warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro to PAF, TH and ADP
The inhibitory action of the platelet aggregation of induction
Experiment material：
Sodium citrate (CAS：68-04-2, Chemical Reagent Co., Ltd., Sinopharm Group), (medicine of Shijiazhuang four has physiological saline
Limit company), fibrin ferment (TH, CAS:506-32-1, SIGMA Reagent Company), adenosine diphosphate (ADP) (ADP, CAS:58-64-0,
SIGMA Reagent Companies), platelet activating factor (PAF, CAS:74389-68-7, SIGMA Reagent Company).
Laboratory apparatus：Platelet aggregation instrument：MODEL 700, CHRONO-LOG
Experiment blood：Fresh pig arteria carotis blood
Experiment packet：
Compound warfarin -4-O- acetyl-the Leu-Pro-Asn-Ile-Ser-Lys-Pro of the present invention, negative control are
Physiological saline.
Agents useful for same is prepared：
Anti-coagulants is 3.8% sodium citrate solution of normal saline, and inducer of platelet activation is normal saline
50U/mL TH solution, the 1mM ADP solution of normal saline, the 5 × 10 of normal saline-5MM PAF solution.Induction
The ultimate density of agent be TH be 1U/mL, ADP be 20 μM, PAF be 1 × 10-6mM。
Experimental method：
Carrying the previous day silanization and taking of drying adds 50mL sodium citrate aqueous solutions in blood vial (with blood ratio 1:9).
Gently shaken up after obtaining fresh pig arteria carotis blood, centrifuge 10min in 1000rpm rotating speed rapidly, collect supernatant, obtain rich blood
Platelet-poor plasma (PRP), remaining blood centrifuge 10min in 3500rpm rotating speed, collect supernatant, obtain platelet poor plasma
(PPP)。
500 μ L PPP is added in cuvette on platelet aggregation instrument, adds in PPP holes, 480 μ L PRP is added
Enter in cuvette, add in PRP holes.Measured after PRP used is incubated into 10min in 37 DEG C.Bring into operation in instrument plus
After entering rotor and returning to zero, 10 μ L physiological saline or compound water solution are first added into PRP, then 10 μ L are added into PRP
Platelet activation derivant.The change of light transmittance is recorded, calculates the inhibiting rate of compounds on platelet aggregation.Experimental result
See Fig. 4.
Experimental result：
Test result indicates that warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro can suppress by TH and
The platelet aggregation of ADP inductions.Inhibiting rate to the platelet aggregation of TH inductions is 20.0%；To the platelet aggregation of ADP inductions
The inhibiting rate of collection is 7.0%.Because warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro can suppress in vitro
Platelet aggregation, so it is not prodrug.This is unexpected technique effect.
Embodiment 20, which evaluates warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro, reduces the internal b/ of GP II
The effect of III a contents
Experiment material
Sodium citrate (CAS：68-04-2, Chemical Reagent Co., Ltd., Sinopharm Group), NS (the limited public affairs of the medicine of Shijiazhuang four
Department), distilled water；
Experiment sample
Rat neck after the warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro of embodiment 17 treatment body circulations
Arterial blood
Experimental method
Detected using a of II b/ of rat platelet membrane glycoprotein III (a of II b/ of GP III) enzyme linked immunological kits.
The preparation of standard items
A standard items are taken out from kit, 30s is centrifuged in 6000-10000rpm, is dissolved with 1mL Sample dilutions,
And with pipette tips alignment freeze bottom of the tube and inhale repeatedly and make a call to 5 times with hydrotropy solution, be sufficiently mixed to obtain standard items S7, take 7 1.5mL from
Heart pipe is arranged in order, each to add 250 μ L Sample dilutions, draws 250 μ L standard items S7 in S6 centrifuge tubes, and piping and druming mixes, according to
It is secondary with S6-S1, S0 are Sample dilution.Standard concentration is respectively：S7(400ng/mL)、S6(200ng/mL)、S5
(100ng/mL)、S4(50ng/mL)、S3(25ng/mL)、S2(12.5ng/mL)、S1(6.25ng/mL)、S0(0ng/mL)。
The collection of sample
Sodium citrate solution with 3.8% is anti-coagulants, collection warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-
Arteria carotis blood after the anti-arterial thrombus circulation of Ser-Lys-Pro treatment rats, with centrifuging 15min in 4 DEG C of 1000rpm in 30min,
Supernatant (blood plasma) is taken to be detected as sample.
Pattern detection
Various reagents are moved into (18-25 DEG C) balance at least 30min of room temperature, are marked with quasi- sample wells, warfarin -4-O- second respectively
Acyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro holes.Standard items are separately added into every hole of pre-coated ELISA Plate or treat test sample
This 100 μ L, gently rocks mixing, is covered with plate patch, 37 DEG C of incubation 2h.Liquid is discarded, is dried, biotin labelled antibodies are added per hole
The μ L of working solution 100, it is covered with new plate patch, 37 DEG C of incubation 1h.Liquid is discarded, dries, board-washing 3 times, soaks 2min, 200 μ L/ every time
Per hole, dry.Add the μ L of Horseradish peroxidase-conjugated avidin working solution 100 per hole, be covered with new plate patch, 37 DEG C of incubation 1h.
Liquid is discarded, is dried, board-washing 5 times, the μ L of substrate solution 90 are sequentially added per hole, 37 DEG C of lucifuges colour developing 15-30min, are sequentially added per hole
The μ L of stop bath 50, terminating reaction.Sequentially measure the optical density (OD values) in each hole in 450nm wavelength with ELIASA in 5min.
Standard curve is drawn according to the OD values of standard items, sample is calculated and goes out concentration of specimens, data are included in Fig. 5.
Experimental result
0.1 μm of ol/kg warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser- is can be seen that from Fig. 5 data
The a contents of II b/ of GP III of the rat of Lys-Pro treatments are substantially less than a contents (p of II b/ of GP III of saline therapy rat<
0.05), i.e., warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro can be reduced under 0.1 μm of ol/kg dosage
The a contents of II b/ of GP III in rat body, and with a content phases of II b/ of GP III inside 167 μm of ol/kg aspirin for treatment rats
When.It can be seen that warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro reduces a contents of II b/ of GP III in rat body
Activity it is stronger than aspirin 167 times.This is unexpected technique effect.
Embodiment 21, which is evaluated warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro and reduced in rat body, coagulates
The effect of the content of blood factor II
Experiment material
Sodium citrate (CAS：68-04-2, Chemical Reagent Co., Ltd., Sinopharm Group), (medicine of Shijiazhuang four has physiological saline
Limit company), liquid-transfering gun (Germany general Rand Corporation), distilled water；
Experiment sample
Embodiment 18 gives rat carotid artery blood after warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro
Experimental method
Experiment is detected using the enzyme-linked immunoassay kits of rat F II.
The collection of sample
Sodium citrate solution with 3.8% is anti-coagulants, gathers 0.1 μm of ol/kg warfarin -4-O- acetyl-Leu-Pro-
The arteria carotis blood of the rat of Asn-Ile-Ser-Lys-Pro treatments, with centrifuging 15min in 4 DEG C of 1000rpm in 30min, take supernatant
Liquid (blood plasma) is detected as sample.
The preparation of standard items
Standard items are taken out from kit, the former times standard items of 150 μ L is diluted with 150 μ L standard dilutions, is sufficiently mixed
Standard items S5 is obtained, takes 4 1.5mL centrifuge tube to be arranged in order, it is each to add 150 μ L Sample dilutions, draw 150 μ L standard items
In S4 centrifuge tubes, piping and druming mixes S5, matches somebody with somebody to obtain S5-S1 standard items successively.Standard concentration is respectively S5 (6ng/mL), S4
(3ng/mL), S3 (1.5ng/mL), S2 (0.75ng/mL), S1 (0.375ng/mL) and S0 (0ng/mL).
Blank well, gauge orifice, test serum hole are set respectively.Add 50 μ L standard items in enzyme mark coating plate, in test serum hole
First plus 40 μ L sample dilutions, then add 10 μ L test serums, gently rock mixing.With the rearmounted 37 DEG C of incubations of shrouding film shrouding
30min.Carefully take shrouding film off, discard liquid, board-washing 5 times.50 μ L enzyme marking reagents, 37 DEG C of incubations are added except blank well per hole
30min, washing, per the μ L developer A of Kong Xianjia 50, then add 50 μ L developer B, gently concussion mixes, 37 DEG C of lucifuge colour developings
10min.Add 50 μ L terminate liquid terminating reactions per hole, blank well returns to zero in 15min, the absorbance in each hole of 450nm wavelength measurements
(OD values).Standard curve is drawn according to the OD values of standard items, calculates serum sample concentration, data are included in Fig. 6.
Experimental result
0.1 μm of ol/kg warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser- is can be seen that from Fig. 6 data
The FII contents of the rat of Lys-Pro treatments are substantially less than the FII contents (p of saline therapy rat<0.05), i.e., in 0.1 μ
Warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro can reduce FII in rat body and contain under mol/kg dosage
Amount, and it is suitable with FII contents inside 4.87 μm of ol/kg warfarin treatment rats.It can be seen that warfarin -4-O- acetyl-Leu-
The activity that Pro-Asn-Ile-Ser-Lys-Pro reduces FII contents in rat body is stronger than warfarin 48.7 times.This is unexpected
Technique effect.
Claims (7)
1. the following warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro of structure,
2. claim 1 warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro preparation method, this method bag
Include：
(1) warfarin -4-O- benzyl acetates are synthesized；
(2) warfarin -4-O- acetic acid is synthesized；
(3) dicyclohexylcarbodiimide (DCC) is used as condensing agent, and 1- hydroxy benzo triazoles (HOBt) are the liquid phase of catalyst
The method of condensation, synthesize HClLeu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl；
(4) DCC is used as condensing agent, and HOBt is the method for the liquid phase condensations of catalyst, warfarin -4-O- acetic acid and HCl
Leu-Pro-Asn-Ile-Ser-Lys (Z)-Pro-OBzl is condensed；
(5) warfarin -4-O- acetyl-Leu-Pro-Asn-Ile-Ser-Lys-Pro is synthesized.
3. warfarin -4-O- acetyl-the Leu-Pro-Asn-Ile-Ser-Lys-Pro of claim 1 are preparing anti-arterial thrombus
The application formed in medicine.
4. warfarin -4-O- acetyl-the Leu-Pro-Asn-Ile-Ser-Lys-Pro of claim 1 are preparing anti-phlebothrombosis
The application formed in medicine.
5. warfarin -4-O- acetyl-the Leu-Pro-Asn-Ile-Ser-Lys-Pro of claim 1 are preparing suppression blood platelet
Assemble the application in medicine.
6. warfarin -4-O- acetyl-the Leu-Pro-Asn-Ile-Ser-Lys-Pro of claim 1 are preparing the internal GP of reduction
Application in the medicine of a contents of II b/ III.
7. warfarin -4-O- acetyl-the Leu-Pro-Asn-Ile-Ser-Lys-Pro of claim 1 are preparing factor II
Application in antagonist.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610415117.5A CN107488214B (en) | 2016-06-13 | 2016-06-13 | warfarin-4-O-acetyl-LPNISKP and synthesis, activity and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610415117.5A CN107488214B (en) | 2016-06-13 | 2016-06-13 | warfarin-4-O-acetyl-LPNISKP and synthesis, activity and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN107488214A true CN107488214A (en) | 2017-12-19 |
| CN107488214B CN107488214B (en) | 2021-03-30 |
Family
ID=60642682
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610415117.5A Expired - Fee Related CN107488214B (en) | 2016-06-13 | 2016-06-13 | warfarin-4-O-acetyl-LPNISKP and synthesis, activity and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107488214B (en) |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1552727A (en) * | 2003-12-19 | 2004-12-08 | 复旦大学附属中山医院 | Targetted polypeptide for specificity of liver cancer blood vessel |
| CN104211768A (en) * | 2013-06-05 | 2014-12-17 | 首都医科大学 | Conjugates of beta-carboline-3-carboxylic acid and oligopeptides, preparation, nano structure, and application thereof as antitumor agent |
| CN105218637A (en) * | 2014-06-11 | 2016-01-06 | 首都医科大学 | The indoles quinolizine that LPNISKP modifies, its preparation, nanostructure, active and application |
| CN105218636A (en) * | 2014-06-11 | 2016-01-06 | 首都医科大学 | The β-carboline that LPNISKP modifies, its preparation, nanostructure, active and application |
| CN105294836A (en) * | 2014-06-11 | 2016-02-03 | 首都医科大学 | LPNISKP modified 5-fluorouracil, preparation therefor, nanostructure thereof, activity thereof and application thereof |
-
2016
- 2016-06-13 CN CN201610415117.5A patent/CN107488214B/en not_active Expired - Fee Related
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1552727A (en) * | 2003-12-19 | 2004-12-08 | 复旦大学附属中山医院 | Targetted polypeptide for specificity of liver cancer blood vessel |
| CN104211768A (en) * | 2013-06-05 | 2014-12-17 | 首都医科大学 | Conjugates of beta-carboline-3-carboxylic acid and oligopeptides, preparation, nano structure, and application thereof as antitumor agent |
| CN105218637A (en) * | 2014-06-11 | 2016-01-06 | 首都医科大学 | The indoles quinolizine that LPNISKP modifies, its preparation, nanostructure, active and application |
| CN105218636A (en) * | 2014-06-11 | 2016-01-06 | 首都医科大学 | The β-carboline that LPNISKP modifies, its preparation, nanostructure, active and application |
| CN105294836A (en) * | 2014-06-11 | 2016-02-03 | 首都医科大学 | LPNISKP modified 5-fluorouracil, preparation therefor, nanostructure thereof, activity thereof and application thereof |
Non-Patent Citations (3)
| Title |
|---|
| MARINA ZACCHIGNA等: "Improvement of warfarin biopharmaceutics by conjugation with poly(ethylene glycol)", 《EUROPEAN JOURNAL OF PHARMACEUTICAL SCIENCES》 * |
| 杭燕南: "《当代麻醉学》", 31 August 2013 * |
| 韩瑞发: "《免疫肾脏病学》", 30 March 2016 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN107488214B (en) | 2021-03-30 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN107488212A (en) | The O acetyl RGD tetrapeptides of warfarin 4, it is synthesized, activity and application | |
| CN109134606A (en) | 1,1- dihydroxymethyl-tetrahydro-beta-carboline -3- formyl-GRGDV, synthesis, activity and application | |
| CN109134607A (en) | 1,1- dihydroxymethyl-tetrahydro-beta-carboline -3- formyl-GRGDS, synthesis, activity and application | |
| CN109134605A (en) | 1,1- dihydroxymethyl-tetrahydro-beta-carboline -3- formyl-GYIGSK, synthesis, activity and application | |
| CN109134603A (en) | 1,1- dihydroxymethyl-tetrahydro-beta-carboline -3- formyl-GYIGSR, synthesis, activity and application | |
| CN107686506A (en) | The O acetyl GPRP of warfarin 4, it is synthesized, activity and application | |
| CN103145797B (en) | [(3s)-1,2,3,4-tetrahydroisoquinoline-3-formoxy]-Lys modified RGD tetrapeptide and its synthesis method and use in medical science | |
| CN103450342B (en) | Tetrahydroisoquinoline-3-carboxylic acid modify PARGD seven peptide, its synthesis, antithrombotic acitivity and application | |
| CN107488211A (en) | Warfarin -4-O- acetyl-LDV, it is synthesized, activity and application | |
| CN109134604A (en) | 1,1- dihydroxymethyl-tetrahydro-beta-carboline -3- formyl-GRGDF, synthesis, activity and application | |
| CN103450344B (en) | Tetrahydroisoquinoline-3-carboxylic acid modify LARGD seven peptide, its synthesis, antithrombotic acitivity and application | |
| CN105198960B (en) | Imidazopyridine -6- formyl-Met-AA-OBzl, synthesis, activity and application | |
| CN107488214A (en) | Its synthesis of the O acetyl LPNISKP of warfarin 4, activity and application | |
| CN107488213A (en) | Warfarin -4-O- acetyl-YIGSK, it is synthesized, pharmacological activity and application | |
| CN103450343B (en) | Tetrahydroisoquinoline-3-carboxylic acid modify TARGD seven peptide, its synthesis, antithrombotic acitivity and application | |
| CN108976283A (en) | 1,1- dihydroxymethyl-tetrahydro-beta-carboline -3- formyl-GLDV, synthesis, activity and application | |
| CN107488210A (en) | The O acetyl dipeptides of warfarin 4, it is synthesized, activity and application | |
| CN108976285A (en) | The warfarin of Gly-Pro-Arg-Pro-AA modification, synthesis, activity and application | |
| CN109111502A (en) | 1,1- dihydroxymethyl-tetrahydro-beta-carboline -3- formyl-GGPRP, synthesis, activity and application | |
| CN101318995A (en) | Substituted tetrahydro-isoquinoline isoquinolinium compound, preparation method and application thereof | |
| CN107488157A (en) | The O acetyl Arg/Asp of warfarin 4, it is synthesized, activity and application | |
| CN105294834A (en) | Imidazo-pyridine-6-formyl-AA-OBzl, as well as synthesis, activities and application thereof | |
| CN104211763B (en) | Aspirin Arg Gly Asp Val conjugates, it is synthesized, nanostructured and the application as medicine-carried system | |
| CN113929735B (en) | Gly-Pro-Arg-Pro-NHCH 2 CH 2 NH-warfarin, its synthesis, activity and use | |
| CN113929739B (en) | Gly-Pro-Arg-Pro-NH ethoxycarbonyl warfarin, synthesis, activity and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20210330 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |