CN104876979A - Novel sulfonation pentasaccharides compound with anti-Xa activity - Google Patents
Novel sulfonation pentasaccharides compound with anti-Xa activity Download PDFInfo
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Abstract
The invention relates to a novel sulfonation pentasaccharides compound with anti-Xa activity and a preparing method and application thereof. The compound serves as the ramification of the pentasaccharides compound with the anticoagulation function, and the obvious anti-Xa activity is achieved.
Description
Technical field
The invention belongs to medical art, relate to a kind of new compound, be specifically related to a kind of sulfonated pentasaccharides compound with Anti-Xa factor activity newly.
Technical background
Thrombotic diseases is the disease of serious harm human health, according to thrombosis position, condition and character, is mainly divided into arterial thrombus and phlebothrombosis.Artery thrombosis is from artery vessel wall atherosclerotic lesion and platelet activation, and the bad clinical disease that it causes is mainly acute myocardial infarction, cerebral apoplexy; Phlebothrombosis is brought out by many reasons in vein blood vessel and is formed, and can cause venous thromboembolism, and its main clinical manifestation is venous thrombosis and pulmonary infarction.Clinical trial evidence shows, and anticoagulant therapy can stop spreading of thrombus and recur, the incidence of the low cerebral apoplexy of a step-down of going forward side by side, pulmonary infarction etc. and mortality ratio.Therefore, anticoagulant therapy has become core and the basis of current clinical prevention and treatment thrombotic disease, and the research and development of anticoagulant are also the focus of new drug development all the time.
At present, to research and develop or the novel anticoagulant that gone on the market mainly comprises direct thrombin inhibitor, Xa factor inhibitor, IX factor inhibitors, tissue factor inhibitor and novel vitamin K antagon.Wherein, direct thrombin inhibitor and Xa factor inhibitor are the representational anti-freezing new drugs of most.
Fondaparinux sodium is a kind of Selective activation factor (Xa) inhibitor of synthetic, by the selective binding with AT III, enhance the neutralizing effect that AT III is intrinsic to factor Xa, the neutralizing effect of factor Xa can disturb blood coagulation cascade, and can be formed and thrombus development by Trombin inhibiting, its chemical structural formula following (representing 5 monose from left to right respectively with D, E, F, G, H): simultaneously
Summary of the invention
The object of the invention is to provide a kind of sulfonated pentasaccharides compound (hereinafter referred to as compd A) with Anti-Xa factor activity.
Compd A of the present invention, it has extraordinary Anti-Xa factor activity after measured, and its anti-Xa tires as 612IU/mg, is about 1.5 times of Fondaparinux sodium, has high pharmaceutical use and significant result for the treatment of.
Compd A of the present invention, its chemical structural formula following (representing 5 monose from left to right respectively with D, E, F, G, H):
Another object of the present invention is the preparation method providing compd A.
Concrete, the synthetic route of compd A of the present invention is as follows:
The preparation method of compd A of the present invention, comprises the following steps:
Step 1, the preparation of D:
D1 is dissolved with appropriate anhydrous methylene chloride, adds Trichloroacetonitrile, add DBU, room temperature reaction 0.5h, obtain product D;
Step 2, the preparation of DEF:
, be dissolved in appropriate anhydrous methylene chloride by D and EF, add appropriate dry 4A molecular sieve, stirred at ambient temperature is after 30 minutes, in nitrogen protection, the dichloromethane solution adding Trimethylsilyl trifluoromethanesulfonate at-20 DEG C, reacts 30 minutes, obtains DEF1.DEF1 is dissolved in appropriate diacetyl oxide, in nitrogen protection, the dichloromethane solution adding Trimethylsilyl trifluoromethanesulfonate at 0 DEG C, room temperature reaction 3 hours; Add ammoniacal liquor again, room temperature reaction 4 hours; Finally add appropriate Trichloroacetonitrile and DBU, nitrogen protection, room temperature reaction 3 hours, obtains product DEF;
Step 3, the preparation of GH:
, G, BSP and 4A molecular sieve is dissolved in anhydrous methylene chloride, and stirring at room temperature 30min under nitrogen protection, is cooled to-60 DEG C, slowly drips Tf
2o, drip after finishing, temperature rises to-40 DEG C, adds the H after with anhydrous methylene chloride dissolved dilution, and-40 DEG C of reaction 1h, obtain product G H1.Be dissolved in appropriate anhydrous methylene chloride by GH1, at room temperature add triethylamine, reaction 3h, obtains product G H;
Step 4, the preparation of DEFGH:
, DEF and GH is dissolved in appropriate anhydrous methylene chloride, adds appropriate dry 4A molecular sieve, stirring at room temperature is after 30 minutes, in nitrogen protection, the dichloromethane solution adding appropriate Trimethylsilyl trifluoromethanesulfonate at-20 DEG C, reacts 1 hour, add appropriate triethylamine, stir 30 minutes, filter, filtrate is concentrated into dry, adds q. s. methylene chloride and dissolves, wash with water successively, anhydrous sodium sulfate drying, filter, filtrate is concentrated into dry, cross silicagel column, obtain product DEFGH;
Step 5, the preparation of compd A
, DEFGH is dissolved in THF, adds purified water, drip NaOH solution afterwards, react 18 hours, drip hydrochloric acid neutralization, then add chloroform extraction, get organic phase, be evaporated to dry.Gained sample is dissolved in DMF; add sulfur trioxide trimethylamine complex compound; nitrogen protection; react 18 hours; concentrating under reduced pressure, filtrate loading to sephadex lh-20 chromatography column desalination, by collect the underpressure distillation of product liquid; and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry.Add in hydriding reactor by gained sample, purified water and 10% palladium carbon, in still, hydrogenation is to 1.5MPa, reacts 68 hours, after reaction terminates, filters, by filtrate reduced in volume.Condensate residue is moved in reactor, add purified water, sulfur trioxide pyridine complex is added in reactor in batches, reacts 7 hours, to solution clear, filter, loading to the desalination of sephadex G-25 chromatography column, by the product liquid underpressure distillation of collecting, and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry, after preparing purifying, obtains compd A.
Preferably, the preparation method of the compounds of this invention A, comprises the following steps:
Step 1, the preparation of D:
D1 (18.0g, 42.1mmol) is dissolved in 90ml anhydrous methylene chloride, adds Trichloroacetonitrile (21ml, 209.4mmol), add DBU (1.3ml, 8.7mmol), room temperature reaction 0.5h.Be evaporated to dry, cross silicagel column, obtain product D (22.6g, 39.6mmol).
Step 2, the preparation of DEF:
By D (22.6g; 39.6mmol) with EF (17.5g; 29.2mmol) be dissolved in 200ml anhydrous methylene chloride, add the dry 4A molecular sieve of 40g, stirred at ambient temperature is after 30 minutes; TBSOTf (2.7ml is added in nitrogen protection, at-20 DEG C; dichloromethane solution 11.8mmol), reacts 30 minutes, crosses silicagel column; obtain DEF1 (19.0g, 18.8mmol).DEF1 (19.0g, 18.8mmol) is dissolved in 95ml diacetyl oxide, in nitrogen protection, the dichloromethane solution adding TBSOTf (0.9ml, 3.9mmol) at 0 DEG C, room temperature reaction 3 hours; Add ammoniacal liquor 2ml again, room temperature reaction 4 hours; Finally add Trichloroacetonitrile (11.3ml, 113.0mmol) and DBU (0.4ml, 2.7mmol), nitrogen protection, room temperature reaction 3 hours, cross silicagel column, obtain product DEF (11.1g, 9.2mmol).
Step 3, the preparation of GH:
The 4A molecular sieve of G (30.0g, 41.8mmol), BSP (11.4g, 54.6mmol) and 30g drying is dissolved in 150ml anhydrous methylene chloride, and stirring at room temperature 30min under nitrogen protection, is cooled to-60 DEG C, slowly drips Tf
2o (9.6ml, 58.5mmol), drip after finishing, temperature rises to-40 DEG C, add the H (23.1g, 50.4mmol) after with anhydrous methylene chloride dissolved dilution ,-40 DEG C of reaction 1h, cross silicagel column, obtain product G H1 (16.5g, 15.8mmol).Be dissolved in by GH1 (16.5g, 15.8mmol) in 150ml anhydrous methylene chloride, at room temperature add triethylamine (55ml, 294.7mmol), reaction 3h, crosses silicagel column, obtains product G H (11.8g, 14.0mmol).
Step 4, the preparation of DEFGH:
By DEF (11.1g, 9.2mmol) with GH (9.3g, 11.0mmol) be dissolved in 100ml anhydrous methylene chloride, add the 4A molecular sieve of 20g drying, stirring at room temperature is after 30 minutes, in nitrogen protection, TMSOTf (0.7ml is added at-20 DEG C, dichloromethane solution 3.9mmol), react 1 hour, add appropriate triethylamine, stir 30 minutes, filter, filtrate is concentrated into dry, add q. s. methylene chloride to dissolve, wash with water successively, anhydrous sodium sulfate drying, filter, filtrate is concentrated into dry, cross silicagel column, obtain product DEFGH (12.2g, 6.4mmol).
Step 5, the preparation of compd A:
DEFGH (12.2g, 6.4mmol) is dissolved in 150ml tetrahydrofuran (THF), adds 80ml purified water, drip 1N sodium hydroxide solution 120ml afterwards, room temperature reaction 18 hours, drip the neutralization of 1N hydrochloric acid, then add chloroform extraction, get organic phase, be evaporated to dry.Gained sample is dissolved in 100ml N; in dinethylformamide; add sulfur trioxide trimethylamine complex compound (15.5g, 112.0mmol), nitrogen protection; 60 DEG C are reacted 18 hours; concentrating under reduced pressure, filtrate loading to sephadex lh-20 chromatography column desalination, by collect the underpressure distillation of product liquid; and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry.Add in hydriding reactor by gained sample, 360ml purified water and 10.8g 10% palladium carbon, in still, hydrogenation is to 1.5MPa, and 30 DEG C are reacted 68 hours, after reaction terminates, filters, by filtrate reduced in volume.Condensate residue is moved in reactor, add purified water, by sulfur trioxide pyridine complex (9.5g, 59.7mmol) add in reactor in batches, 10 DEG C are reacted 7 hours, to solution clear, filter, loading is to the desalination of sephadex G-25 chromatography column, by the product liquid underpressure distillation of collecting, and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry, compd A (2.6g, 1.5mmol) is obtained after preparing purifying.
Another object of the present invention is to provide a kind of pharmaceutical composition containing compd A.
Pharmaceutically active substance in pharmaceutical composition of the present invention is compound of the present invention, its in the formulation shared weight percent can be 0.01-99.99%, all the other are medicine acceptable carrier.
Pharmaceutical composition of the present invention, can be prepared into any pharmaceutical dosage forms in use as required, as oral dosage form, and injection form etc.
Pharmaceutical composition of the present invention, when being prepared into pharmaceutical preparation, can add pharmaceutically acceptable carrier as required.
Pharmaceutical composition of the present invention, exists with the dosage form of unitary dose, and described unitary dose refers to the unit of preparation, as every sheet of tablet, and every capsules of capsule, every bottle of oral liquid, granule every bag, often propping up of injection.
Pharmaceutical composition of the present invention can be any pharmaceutically useful formulation, and these formulations comprise: tablet, sugar coated tablet, film coated tablet, enteric coated tablet, capsule, hard capsule, soft capsule, oral liquid, suck agent, granule, electuary, pill, powder, paste, sublimed preparation, suspensoid, pulvis, solution, injection, suppository, ointment, plaster, creme, sprays, drops, pill, patch.
Pharmaceutical composition of the present invention, the preparation of its oral administration can containing conventional vehicle, and such as tackiness agent, weighting agent, thinner, tablet agent, lubricant, disintegrating agent, tinting material, seasonings and wetting agent, can carry out dressing to tablet if desired.
The weighting agent be suitable for comprises Mierocrystalline cellulose, mannitol, lactose and other similar weighting agent.Suitable disintegrating agent comprises starch, polyvinylpyrrolidone and starch derivative, such as sodium starch glycollate.Suitable lubricant comprises, such as Magnesium Stearate.The suitable acceptable wetting agent of medicine comprises sodium lauryl sulphate.
The present invention, by mixing, fills, and the method that compressing tablet etc. are conventional prepares solid oral composition.Repeatedly mix and active substance can be made to be distributed in those compositions of a large amount of weighting agent of whole use.
The form of oral liquid can be such as water-based or oily suspensions, solution, emulsion, syrup or elixir, or can be the composite drying products of a kind of used water before use or other suitable carrier.This liquid preparation can containing conventional additive, such as suspension agent, such as sorbyl alcohol, syrup, methylcellulose gum, gelatin, Natvosol, carboxymethyl cellulose, aluminium stearate gel or hydrogenated edible fats, emulsifying agent, such as Yelkin TTS, anhydro sorbitol monooleate or gum arabic; Non-aqueous carrier (they can comprise edible oil), the oily ester of the such as ester of Prunus amygdalus oil, fractionated coconut oil, such as glycerine, propylene glycol or ethanol; Sanitas, such as para hydroxybenzene methyl esters or propylparaben or Sorbic Acid, and if need, can containing conventional flavouring agent or tinting material.
For injection, the fluid unit dosage form of preparation contains active substance of the present invention and sterile carrier.According to carrier and concentration, this compound can be suspended or dissolve.The preparation of solution is normally by being dissolved in a kind of carrier by active substance, filter-sterilized before being loaded a kind of suitable bottle or ampoule, then seals.Auxiliary material such as a kind of local anesthetic, sanitas and buffer reagent also can be dissolved in this carrier.In order to improve its stability, by freezing for this composition after loading bottle, and under vacuo water can be removed.
Pharmaceutical composition of the present invention, applicable medicine acceptable carrier is optionally added when being prepared into medicament, described medicine acceptable carrier is selected from: N.F,USP MANNITOL, sorbyl alcohol, Sodium Pyrosulfite, sodium bisulfite, Sulfothiorine, cysteine hydrochloride, Thiovanic acid, methionine(Met), vitamins C, EDETATE SODIUM, Ethylenediaminetetraacetic Acid Calcium Salt, the alkali-metal carbonate of monovalence, acetate, phosphoric acid salt or its aqueous solution, hydrochloric acid, acetic acid, sulfuric acid, phosphoric acid, amino acid, sodium-chlor, Repone K, Sodium.alpha.-hydroxypropionate, Xylitol, maltose, glucose, fructose, dextran, glycine, starch, sucrose, lactose, mannitol, silicon derivative, cellulose and its derivates, alginate, gelatin, polyvinylpyrrolidone, glycerine, POLYSORBATE 80, agar, calcium carbonate, Calcium hydrogen carbonate, tensio-active agent, polyoxyethylene glycol, cyclodextrin, beta-cyclodextrin, phospholipid material, kaolin, talcum powder, calcium stearate, Magnesium Stearate etc.
Preferably, pharmaceutical dosage form of the present invention is injection.
Injection of the present invention, in unit formulation, the content of compd A is 1.5mg-6.5mg.
Another object of the present invention is the pharmaceutical use providing compd A.
Compd A of the present invention is preparing the application had in the medicine of Anti-Xa factor activity.
The application of compd A of the present invention in the medicine of preparation treatment thrombotic diseases.
Thrombotic diseases of the present invention is mainly divided into arterial thrombus and Venous Thrombosis.
The present invention also provides the measuring method of compd A chemical structure and purity.
The method for detecting purity of compd A of the present invention, comprises the following steps:
Adopt high performance liquid chromatograph, the polymeric anion exchange column being weighting agent with quaternary ammonium anion shell resin, water and sodium chloride solution are the purity of moving phase, detection compound A.
Concrete detection method of the present invention asks for an interview specification sheets embodiment 2.
The present invention also provides the structure determination of compd A.
The present invention is by the structure of nuclear-magnetism and Mass Spectrometric Identification compd A.From nucleus magnetic hydrogen spectrum, compared with Fondaparinux sodium, 2.74 places are a many methyl peak, the hydrogen chemical shifts of H sugar 2 shifts to 3.93 by the hydrogen that 3.25 shift to low field 3.66, H sugar 3 by 3.62, and other change in location are little.From nuclear-magnetism carbon spectrum, compared with Fondaparinux sodium, 32.76 places are a many carbon peak, the carbon of H sugar 2 also has change in various degree by the carbon that 60.37 shift to 62.85, H sugar 1 and 3, and other change in location are little.Detected by high resolution mass spectrum, infer that the accurate molecular weight of this compound is: 1740.7864, molecular formula is C
32h
45n
3na
10o
49s
8.Can infer that make the chemical shift of the hydrogen of H sugar 2 shift to low field, meanwhile, H sugar 1 also there occurs change with 3 with the hydrogen on H sugar 2 amino be connected by methyl substituted in conjunction with mass spectrum.
The present invention promulgates according to State Food and Drug Administration, and in national drug standards WS1-(the X-147)-2005Z of low molecular weight calcium heparin, Determination method measures Anti-Xa factor (FXa) activity of compd A.The Anti-Xa factor measuring compd A is tired as 612IU/mg, and Fondaparinux sodium is tired as 400IU/mg, and low molecular weight calcium heparin is tired as 105IU/mg.
Compd A belongs to new compound, and current domestic and foreign literature patent is not reported it.By measuring the Anti-Xa factor activity of compd A, its anti-Xa tires as 612IU/mg, is approximately 1.5 times of Fondaparinux sodium, and it is active that it shows better Anti-Xa factor, following upgrading or the Regeneration variety likely becoming Fondaparinux sodium.
In addition, as new compound, compd A also may have the effect of other potential pharmacology aspect, takes a long view, and through more deep research, it also has higher value.
Compd A of the present invention, compared with currently available products, has biological activity strong, the features such as few side effects.As there is stronger biological activity than Fondaparinux sodium and low molecular heparin calcium; Compd A belongs to single compound, than the few side effects of low molecular heparin calcium belonging to mixture.
Compound D 1, EF of the present invention, G, H, belong to currently available products, can commercially buy.
Noun described in literary composition is further explained:
DBU:1,8-diazabicylo 11 carbon-7-alkene
TBSOTf: Trimethylsilyl trifluoromethanesulfonate
TMSOTf: trifluoromethanesulfonic acid trimethylammonium silicone grease
BSP:1-(phenylsulfmyl) piperidines
Above-mentioned raw materials belongs to currently available products, can commercially buy.
Accompanying drawing explanation
Fig. 1: the liquid chromatogram of the compounds of this invention A
Embodiment
By following specific embodiment, the present invention is further explained, but not as restriction of the present invention.
Embodiment 1: the preparation of compd A
Step 1, the preparation of D:
D1 (18.0g, 42.1mmol) is dissolved in 90ml anhydrous methylene chloride, adds Trichloroacetonitrile (21ml, 209.4mmol), add DBU (1.3ml, 8.7mmol), room temperature reaction 0.5h.Be evaporated to dry, cross silicagel column, obtain product D (22.6g, 39.6mmol).
Step 2, the preparation of DEF:
By D (22.6g; 39.6mmol) with EF (17.5g; 29.2mmol) be dissolved in 200ml anhydrous methylene chloride, add the dry 4A molecular sieve of 40g, stirred at ambient temperature is after 30 minutes; TBSOTf (2.7ml is added in nitrogen protection, at-20 DEG C; dichloromethane solution 11.8mmol), reacts 30 minutes, crosses silicagel column; obtain DEF1 (19.0g, 18.8mmol).DEF1 (19.0g, 18.8mmol) is dissolved in 95ml diacetyl oxide, in nitrogen protection, the dichloromethane solution adding TBSOTf (0.9ml, 3.9mmol) at 0 DEG C, room temperature reaction 3 hours; Add ammoniacal liquor 2ml again, room temperature reaction 4 hours; Finally add Trichloroacetonitrile (11.3ml, 113.0mmol) and DBU (0.4ml, 2.7mmol), nitrogen protection, room temperature reaction 3 hours, cross silicagel column, obtain product DEF (11.1g, 9.2mmol).
Step 3, the preparation of GH:
The 4A molecular sieve of G (30.0g, 41.8mmol), BSP (11.4g, 54.6mmol) and 30g drying is dissolved in 150ml anhydrous methylene chloride, and stirring at room temperature 30min under nitrogen protection, is cooled to-60 DEG C, slowly drips Tf
2o (9.6ml, 58.5mmol), drip after finishing, temperature rises to-40 DEG C, add the H (23.1g, 50.4mmol) after with anhydrous methylene chloride dissolved dilution ,-40 DEG C of reaction 1h, cross silicagel column, obtain product G H1 (16.5g, 15.8mmol).Be dissolved in by GH1 (16.5g, 15.8mmol) in 150ml anhydrous methylene chloride, at room temperature add triethylamine (55ml, 294.7mmol), reaction 3h, crosses silicagel column, obtains product G H (11.8g, 14.0mmol).
Step 4, the preparation of DEFGH:
By DEF (11.1g; 9.2mmol) be dissolved in 100ml anhydrous methylene chloride with GH (9.3g, 11.0mmol), add the 4A molecular sieve of 20g drying; stirring at room temperature is after 30 minutes; in nitrogen protection, the dichloromethane solution adding TMSOTf (0.7ml, 3.9mmol) at-20 DEG C, react 1 hour; be evaporated to dry; cross silicagel column, obtain product DEFGH (12.2g, 6.4mmol).
Step 5, the preparation of compd A:
DEFGH (12.2g, 6.4mmol) is dissolved in 150ml tetrahydrofuran (THF), adds 80ml purified water, drip 1N sodium hydroxide solution 120ml afterwards, react 18 hours, drip the neutralization of 1N hydrochloric acid, then add chloroform extraction, get organic phase, be evaporated to dry.Gained sample is dissolved in 100ml N; in dinethylformamide; add sulfur trioxide trimethylamine complex compound (15.5g, 112.0mmol), nitrogen protection; react 18 hours; concentrating under reduced pressure, filtrate loading to sephadex lh-20 chromatography column desalination, by collect the underpressure distillation of product liquid; and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry.Add in hydriding reactor by gained sample, 360ml purified water and 10.8g 10% palladium carbon, in still, hydrogenation is to 1.5MPa, reacts 68 hours, after reaction terminates, filters, by filtrate reduced in volume.Condensate residue is moved in reactor, add purified water, by sulfur trioxide pyridine complex (9.5g, 59.7mmol) add in reactor in batches, react 7 hours, to solution clear, filter, loading is to the desalination of sephadex G-25 chromatography column, by the product liquid underpressure distillation of collecting, and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry, compd A (2.6g, 1.5mmol) is obtained after preparing purifying.
Embodiment 2 is by high performance liquid chromatograph device detection compound A
Chromatographic column is Dionex CarboPac
tMpA1 (250 × 4mm), flow velocity is 1.0ml/min, and determined wavelength is 210nm, column temperature is 35 DEG C, take water as mobile phase A (adding about 10 μ l methyl-sulphoxides in 1000ml water), 116.9g/l sodium chloride solution is Mobile phase B, and it is as shown in the table for gradient:
The purity detecting result of compd A is as shown in Figure 1: retention time is 16.304min, and purity is 99.73%.
Embodiment 3: Structural Identification is carried out to compd A by nuclear-magnetism and mass spectrum
Instrument testing conditions: Agilent 6520 Q-TOF LCMS high-resolution mass spectrometer, flow velocity 1ml/min, sample introduction 1 μ l, ion source ESI; BRUKER 400MHz nuclear magnetic resonance analyser; Solvent: D
2o; Interior mark: TMS.Detection data are as follows:
Table 1 hydrogen spectrum and carbon modal data
Table 2 high resolution mass spectrum data
From nucleus magnetic hydrogen spectrum, as shown in table 1, compared with Fondaparinux sodium, 2.74 places are a many methyl peak, the hydrogen chemical shifts of H sugar 2 shifts to 3.93 by the hydrogen that 3.25 shift to low field 3.66, H sugar 3 by 3.62, and other change in location are little.From nuclear-magnetism carbon spectrum, compared with Fondaparinux sodium, 32.76 places are a many carbon peak, the carbon of H sugar 2 also has change in various degree by the carbon that 60.37 shift to 62.85, H sugar 1 and 3, and other change in location are little.Detected by high resolution mass spectrum, as shown in table 2, infer that the accurate molecular weight of this compound is: 1740.7864, molecular formula is C
32h
45n
3na
10o
49s
8.Can infer that make the chemical shift of the hydrogen of H sugar 2 shift to low field, meanwhile, H sugar 1 also there occurs change with 3 with the hydrogen on H sugar 2 amino be connected by methyl substituted in conjunction with mass spectrum.Determine thus, the structural formula of compd A is as follows:
Embodiment 4: Xa factor (FXa) Activity determination of compd A
Promulgate according to State Food and Drug Administration, in national drug standards WS1-(the X-147)-2005Z of low molecular weight calcium heparin, Determination method measures, and namely externally compares to measure the activity that trial-product accelerates to suppress Xa factor (FXa) with Low molecular heparin (LMWH) standard substance by Antithrombin III (AT III).Concrete grammar is as follows:
One, solution preparation
Tutofusin tris-sodium chloride buffer (pH 7.4): get Tutofusin tris 6.08g and the sodium-chlor 8.77g 500ml that adds water and make dissolving, add bovine serum albumin 10g, with salt acid for adjusting pH value to 7.4, be diluted with water to 1000ml.
Tutofusin tris-Calcium Disodium Versenate damping fluid (pH 8.4): get sodium-chlor 5.12g, Tutofusin tris 3.03g and Calcium Disodium Versenate 1.4g, the 250ml that adds water makes dissolving, with salt acid for adjusting pH value to 8.4, be diluted with water to 500ml.
The preparation of LMWH standard substance and need testing solution: the solution respectively standard substance (S) and trial-product (T) being diluted to 4 different concns with Tutofusin tris-sodium chloride buffer (pH 7.4), the agent distance between each dosage should in the linearity range of log dose-response than being generally this concentration of 1:0.7 ~ 1:0.6.Be generally in every 1ml containing 0.025IU ~ 0.2IU.
Antithrombin (AT III) solution: make the solution containing 1IU in every 1ml with Tutofusin tris-sodium chloride buffer (pH 7.4).
Chromophoric substrate s-2765 (or other FXa specificity chromophoric substrates): solution with water makes the solution of 0.003mol/L, uses Tutofusin tris-Calcium Disodium Versenate damping fluid (pH 8.4) to be diluted to 0.0005mol/L before use.
Xa factor (FXa) solution: with Tutofusin tris-sodium chloride buffer (pH 7.4), preparation FXa solution, debug its concentration, make it replacing in the Anti-Xa factor experiment of LMWH with 0.9% sodium chloride solution, the absorption value at the wavelength place of 405nm is 0.6 ~ 0.7.
Two, assay method
Get small test tube 16, mark T respectively
1, T
2, T
3, T
4and S
1, S
2, S
3, S
4.Each concentration is parallel does two pipes.Often pipe adds trial-product (T) or standard substance (S) the diluent 50 μ l of people's 4 kinds of concentration respectively, and antithrombin solution 50 μ l, mixing (must not produce bubble).By S
1, S
2, S
3, S
4, T
1, T
2, T
3, T
4, T
1, T
2, T
3, T
4, S
1, S
2, S
3, S
4order arrangement, 37 DEG C balance 1 minute.Often pipe adds FXa solution 100 μ l, mixing, and 37 DEG C balance 1 minute, add chromophoric substrate solution 250 μ l, mixing, and 37 DEG C of accurate insulations, after 4 minutes, respectively add 30% acetum 375 μ l, termination reaction.With the semimicro colorimetric pool of 1cm light path, be blank with Tutofusin tris-sodium chloride buffer (pH 7.4), measure the absorbancy of often pipe at the wavelength place of 405nm.Need testing solutions (parallel do two pipes) are replaced with method operation as blank pipe using three light aminomethane-sodium chloride buffer (pH 7.4), when 16 arms start and end ups, the absorbancy of mensuration blank pipe respectively.Both absorbancys must not have significant difference.Take absorbancy as ordinate zou, the logarithmic value of standard solution (or need testing solution) concentration is that X-coordinate does linear regression respectively, by parallel line analysis principle 4 × 4 method experimental design in Bioassay-statistical method (China's coastal port two annex X IV), calculate and tire and experimental error.Average Reliable limit rate (FL%) must not be greater than 15%.Three, test-results
Measure through aforesaid method, the Anti-Xa factor of compd A is tired as 612IU/mg, and Fondaparinux sodium is tired as 400IU/mg, and low molecular weight calcium heparin is tired as 105IU/mg.Compared with Fondaparinux sodium, the Anti-Xa factor of compd A is tired and is about its 1.5 times.
Embodiment 5 compd A injection liquid
Prescription:
Preparation technology:
Step 1, aseptically, gets recipe quantity 80% water for injection, adds recipe quantity sodium-chlor and dissolves, and regulates pH to 5.0-8.0 with 0.1M sodium hydroxide or hydrochloric acid soln;
Step 2, aseptically, gets recipe quantity compd A and adds in above-mentioned solution, stirs, makes dissolving;
Step 3, aseptically, adds water for injection to full dose in above-mentioned gained solution;
Step 4, aseptically, adds 0.1% Medicinal Charcoal to above-mentioned gained solution, and insulation, stirs 30 minutes;
Step 5, aseptically, divides above-mentioned gained solution and is filled in glass ampoule, get product.
Claims (10)
1. have a sulfonated pentasaccharides compd A for Anti-Xa factor activity, its structural formula is as follows:
2. compd A according to claim 1, is characterized in that, this compound Anti-Xa factor is tired as 612IU/mg.
3. the preparation method of compd A according to claim 1, is characterized in that, comprises the following steps:
Step 1, the preparation of D:
D1 is dissolved with appropriate anhydrous methylene chloride, adds Trichloroacetonitrile, add DBU, room temperature reaction 0.5h, obtain product D;
Step 2, the preparation of DEF:
D and EF is dissolved in appropriate anhydrous methylene chloride, add appropriate dry 4A molecular sieve, stirred at ambient temperature is after 30 minutes, in nitrogen protection, the dichloromethane solution adding Trimethylsilyl trifluoromethanesulfonate at-20 DEG C, reacts 30 minutes, obtain DEF1, DEF1 is dissolved in appropriate diacetyl oxide, in nitrogen protection, the dichloromethane solution adding Trimethylsilyl trifluoromethanesulfonate at 0 DEG C, room temperature reaction 3 hours; Add ammoniacal liquor again, room temperature reaction 4 hours; Finally add appropriate Trichloroacetonitrile and DBU, nitrogen protection, room temperature reaction 3 hours, obtains product DEF;
Step 3, the preparation of GH:
, G, BSP and 4A molecular sieve is dissolved in anhydrous methylene chloride, and stirring at room temperature 30min under nitrogen protection, is cooled to-60 DEG C, slowly drips Tf
2o, drip after finishing, temperature rises to-40 DEG C, adds the H after with anhydrous methylene chloride dissolved dilution, and-40 DEG C of reaction 1h, obtain product G H1, be dissolved in appropriate anhydrous methylene chloride by GH1, at room temperature add triethylamine, and reaction 3h, obtains product G H;
Step 4, the preparation of DEFGH:
, DEF and GH is dissolved in appropriate anhydrous methylene chloride, adds appropriate dry 4A molecular sieve, stirring at room temperature is after 30 minutes, in nitrogen protection, the dichloromethane solution adding appropriate Trimethylsilyl trifluoromethanesulfonate at-20 DEG C, reacts 1 hour, add appropriate triethylamine, stir 30 minutes, filter, filtrate is concentrated into dry, adds q. s. methylene chloride and dissolves, wash with water successively, anhydrous sodium sulfate drying, filter, filtrate is concentrated into dry, cross silicagel column, obtain product DEFGH;
Step 5, the preparation of compd A
, DEFGH is dissolved in THF, add purified water, drip NaOH solution afterwards, react 18 hours, dropping hydrochloric acid neutralizes, then chloroform extraction is added, get organic phase, be evaporated to dry, gained sample is dissolved in DMF, add sulfur trioxide trimethylamine complex compound, nitrogen protection, react 18 hours, concentrating under reduced pressure, filtrate loading is to sephadex lh-20 chromatography column desalination, by the product liquid underpressure distillation of collecting, and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry, by gained sample, purified water and 10% palladium carbon add in hydriding reactor, in still, hydrogenation is to 1.5MPa, react 68 hours, after reaction terminates, filter, by filtrate reduced in volume, condensate residue is moved in reactor, add purified water, sulfur trioxide pyridine complex is added in reactor in batches, react 7 hours, to solution clear, filter, loading is to the desalination of sephadex G-25 chromatography column, by the product liquid underpressure distillation of collecting, and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry, compd A is obtained after preparing purifying.
4. the preparation method of compd A according to claim 1, is characterized in that, comprises the following steps:
Step 1, the preparation of D:
By D1 (18.0g, 42.1mmol) be dissolved in 90ml anhydrous methylene chloride, add Trichloroacetonitrile (21ml, 209.4mmol), add DBU (1.3ml, 8.7mmol), room temperature reaction 0.5h, is evaporated to dry, cross silicagel column, obtain product D (22.6g, 39.6mmol)
Step 2, the preparation of DEF:
By D (22.6g, 39.6mmol) with EF (17.5g, 29.2mmol) be dissolved in 200ml anhydrous methylene chloride, add the dry 4A molecular sieve of 40g, stirred at ambient temperature is after 30 minutes, in nitrogen protection, TBSOTf (2.7ml is added at-20 DEG C, dichloromethane solution 11.8mmol), react 30 minutes, cross silicagel column, obtain DEF1 (19.0g, 18.8mmol), by DEF1 (19.0g, 18.8mmol) be dissolved in 95ml diacetyl oxide, in nitrogen protection, TBSOTf (0.9ml is added at 0 DEG C, dichloromethane solution 3.9mmol), room temperature reaction 3 hours, add ammoniacal liquor 2ml again, room temperature reaction 4 hours, finally add Trichloroacetonitrile (11.3ml, 113.0mmol) and DBU (0.4ml, 2.7mmol), nitrogen protection, room temperature reaction 3 hours, cross silicagel column, obtain product DEF (11.1g, 9.2mmol),
Step 3, the preparation of GH:
The 4A molecular sieve of G (30.0g, 41.8mmol), BSP (11.4g, 54.6mmol) and 30g drying is dissolved in 150ml anhydrous methylene chloride, and stirring at room temperature 30min under nitrogen protection, is cooled to-60 DEG C, slowly drips Tf
2o (9.6ml, 58.5mmol), drip after finishing, temperature rises to-40 DEG C, adds the H (23.1g after with anhydrous methylene chloride dissolved dilution, 50.4mmol),-40 DEG C of reaction 1h, cross silicagel column, obtain product G H1 (16.5g, 15.8mmol), GH1 (16.5g, 15.8mmol) is dissolved in 150ml anhydrous methylene chloride, at room temperature adds triethylamine (55ml, 294.7mmol), reaction 3h, crosses silicagel column, obtains product G H (11.8g, 14.0mmol)
Step 4, the preparation of DEFGH:
By DEF (11.1g, 9.2mmol) with GH (9.3g, 11.0mmol) be dissolved in 100ml anhydrous methylene chloride, add the 4A molecular sieve of 20g drying, stirring at room temperature is after 30 minutes, in nitrogen protection, TMSOTf (0.7ml is added at-20 DEG C, dichloromethane solution 3.9mmol), react 1 hour, add appropriate triethylamine, stir 30 minutes, filter, filtrate is concentrated into dry, add q. s. methylene chloride to dissolve, wash with water successively, anhydrous sodium sulfate drying, filter, filtrate is concentrated into dry, cross silicagel column, obtain product DEFGH (12.2g, 6.4mmol),
Step 5, the preparation of compd A:
By DEFGH (12.2g, 6.4mmol) be dissolved in 150ml tetrahydrofuran (THF), add 80ml purified water, drip 1N sodium hydroxide solution 120ml afterwards, room temperature reaction 18 hours, drip the neutralization of 1N hydrochloric acid, then chloroform extraction is added, get organic phase, be evaporated to dry, gained sample is dissolved in 100ml N, in dinethylformamide, add sulfur trioxide trimethylamine complex compound (15.5g, 112.0mmol), nitrogen protection, 60 DEG C are reacted 18 hours, concentrating under reduced pressure, filtrate loading is to sephadex lh-20 chromatography column desalination, by the product liquid underpressure distillation of collecting, and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry, by gained sample, 360ml purified water and 10.8g 10% palladium carbon add in hydriding reactor, in still, hydrogenation is to 1.5MPa, 30 DEG C are reacted 68 hours, after reaction terminates, filter, by filtrate reduced in volume, condensate residue is moved in reactor, add purified water, by sulfur trioxide pyridine complex (9.5g, 59.7mmol) add in reactor in batches, 10 DEG C are reacted 7 hours, to solution clear, filter, loading is to the desalination of sephadex G-25 chromatography column, by the product liquid underpressure distillation of collecting, and loading to 732 sodium form storng-acid cation exchange resin chromatography column, the product liquid of collection is evaporated to dry, compd A (2.6g is obtained after preparing purifying, 1.5mmol).
5. compd A according to claim 1 is preparing the application had in the medicine of Anti-Xa factor activity.
6. the application of compd A according to claim 1 in the medicine of preparation treatment thrombotic diseases, described thrombotic diseases is mainly divided into arterial thrombus and Venous Thrombosis.
7. the pharmaceutical composition containing compd A described in claim 1.
8. pharmaceutical composition according to claim 7, is characterized in that, can be prepared into any pharmaceutical dosage form.
9. pharmaceutical composition according to claim 7, is characterized in that, described pharmaceutical composition is injection, and in unit formulation, the content of compd A is 1.5mg-6.5mg.
10. pharmaceutical composition according to claim 9, is characterized in that, the preparation method of injection comprises the following steps:
Step 1, aseptically, gets recipe quantity 80% water for injection, adds recipe quantity sodium-chlor and dissolves, and regulates pH to 5.0-8.0 with 0.1M sodium hydroxide or hydrochloric acid soln;
Step 2, aseptically, gets recipe quantity compd A and adds in above-mentioned solution, stirs, makes dissolving;
Step 3, aseptically, adds water for injection to full dose in above-mentioned gained solution;
Step 4, aseptically, adds 0.1% Medicinal Charcoal to above-mentioned gained solution, and insulation, stirs 30 minutes;
Step 5, aseptically, divides above-mentioned gained solution and is filled in glass ampoule, get product.
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Citations (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN1989146A (en) * | 2004-03-04 | 2007-06-27 | 普罗吉恩工业有限公司 | Sulfated oligosaccharide derivatives |
US20140336369A1 (en) * | 2011-06-28 | 2014-11-13 | Apicore Us Llc | Process for preparing heparinoids and intermediates useful in the synthesis thereof |
CN104245718A (en) * | 2009-07-31 | 2014-12-24 | 可靠生物医药公司 | Process for preparing fondaparinux sodium and intermediates useful in the synthesis thereof |
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Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN1989146A (en) * | 2004-03-04 | 2007-06-27 | 普罗吉恩工业有限公司 | Sulfated oligosaccharide derivatives |
CN104245718A (en) * | 2009-07-31 | 2014-12-24 | 可靠生物医药公司 | Process for preparing fondaparinux sodium and intermediates useful in the synthesis thereof |
US20140336369A1 (en) * | 2011-06-28 | 2014-11-13 | Apicore Us Llc | Process for preparing heparinoids and intermediates useful in the synthesis thereof |
Non-Patent Citations (1)
Title |
---|
TIEHAI LI,等: "Total Synthesis of Anticoagulant Pentasaccharide Fondaparinux", 《CHEMMEDCHEM》 * |
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CN113461744A (en) * | 2020-03-30 | 2021-10-01 | 鲁南制药集团股份有限公司 | Purification method of fondaparinux sodium intermediate |
CN113461744B (en) * | 2020-03-30 | 2024-03-15 | 鲁南制药集团股份有限公司 | Purification method of fondaparinux sodium intermediate |
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