CN102924629A - Enoxaparin sodium compound and preparation method thereof - Google Patents
Enoxaparin sodium compound and preparation method thereof Download PDFInfo
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- CN102924629A CN102924629A CN2012105082530A CN201210508253A CN102924629A CN 102924629 A CN102924629 A CN 102924629A CN 2012105082530 A CN2012105082530 A CN 2012105082530A CN 201210508253 A CN201210508253 A CN 201210508253A CN 102924629 A CN102924629 A CN 102924629A
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Abstract
The invention provides an enoxaparin sodium compound and a preparation method thereof. Liquaemin and benzethonium chloride are used as raw materials, sodium chloride is used as a neutral reaction medium and a microwave solid-phase synthesis method is carried out to prepare heparin quaternary ammonium salt.
Description
Technical field
The present invention relates to Enoxaparin Sodium and preparation method thereof, belong to medical technical field.
Background technology
Enoxaparin Sodium is a kind of novel antithrombotic reagent of being developed by French Sanofi-Aventis, obtains drugs approved by FDA in October, 2004.Thrombosis and treatment unstable angina pectoris and the non-Q ripple heart stalk etc. of extracorporeal circulation when Enoxaparin Sodium is used for prevention of deep vein thrombosis formation, prevention dvt formation and pulmonary infarction, the established deep venous thrombosis for the treatment of, preclude blood dialysis clinically, be present anticoagulation, antithrombotic first-selected medicine, global marketing volume in 2006 reaches 24.35 Euros.
European Pharmacopoeia 5.3 editions is defined as follows enoxaparin: it is the low molecular weight heparin sodium that is generated by alkaline lysis by the heparin benzyl ester sodio-derivative that the heparin sodium that the chitling mucous membrane extracts obtains after esterification, it is comprised of the not qualitative oligose of many complexity, according to present understanding, most oligose has 4-enol pyrans saccharic acid at the non-reduced end of sugar chain, reduction end at sugar chain has the 15%-25% that 1,6-dehydrated structure accounts for whole sugar.
The Enoxaparin Sodium weight-average molecular weight is between 3800-5000, and the oligose mass percent of molecular weight＜2000 is between 12%-20%, and the oligose mass percent of molecular weight 2000-8000 is at 68%-82%.By tiring at 90-125IU of every milligram of anti-Xa factor of dry product, the ratio of anti-Xa and anti-IIa is between 3.3-5.3.
The present industrial method for preparing Enoxaparin Sodium mainly comprises the steps:
Chondroitin sulfate, dermatan sulfate in the first step, the removal heparin sodium;
Second step, preparation heparin quaternary ammonium salt;
The esterification of the 3rd step, heparin quaternary ammonium salt;
The 4th step, carboxylate obtain Enoxaparin Sodium through alkaline bleach liquor cleavage.
Prior art has been done more research for techniques such as the esterification yield in the Enoxaparin Sodium preparation process, the control that becomes the ring rate and product purifications, and become the technique of quaternary ammonium salt generally to adopt the method for US Patent No. 5389618 for heparin sodium: Teramine is dissolved in the water, join in the aqueous solution of removing the heparin sodium behind the impurity, produce water-fast multiple and salt, filtration obtains multiple and salt, washes rear drying with water.
And the shortcoming of the method is to produce a large amount of contaminated wastewaters, and yield is lower simultaneously.In addition, the more important thing is that the moisture content of this step products heparin quaternary ammonium salt directly affects the height of the esterification yield of subsequent step.And studies show that esterification yield is higher, and the molecular-weight average of depolymerization product is lower, and when esterification yield was 12%-13%, the molecular weight of product just met the standard of European Pharmacopoeia 5.3 editions.And be difficult to accomplish the definitely anhydrous of product in the existing technology.
For the defective of aforesaid method, the contriver becomes the technique of quaternary ammonium salt to improve through large quantity research to heparin sodium.
Summary of the invention
The purpose of this invention is to provide a kind of Enoxaparin Sodium, its structure is as follows:
Wherein, n=1-21, R=H or SO3Na, R1=H or SO3Na or COCH3, R2=H and R3=COONa, or R2=COONa and R3=H.
Another object of the present invention provides a kind of novel preparation method of Enoxaparin Sodium, provide specifically a kind of new synthetic process of heparin quaternary ammonium salt, more specifically, the solid phase synthesis process of heparin quaternary ammonium salt is provided, with overcome the yield that existing aqueous phase synthesis method exists low, produce a large amount of contaminated wastewaters, and affect the defective of subsequent step esterification yield.
Technical scheme of the present invention is: heparin sodium and benzethonium chloride are raw material, take sodium-chlor as the neutral reaction medium, prepare the heparin quaternary ammonium salt by the Microwave Solid synthetic method.
The preferred technical solution of the present invention is as follows:
1, heparin sodium and NaCl, benzyl rope chloramines are mixed;
2, mixture is placed microwave reactor, with the power irradiation of 100-200W, temperature 40-80 ℃, irradiation time is 15-45min;
3, be cooled to room temperature, get the heparin quaternary ammonium salt;
4, the heparin quaternary ammonium salt gets Enoxaparin Sodium through esterification, cracking.
Wherein the weight part of NaCl is 0.002-0.01, preferred 0.005 weight part;
Benzyl rope chloramines weight part is 1-2, preferred 1.5 weight parts;
The preferred 150W of microwave power;
The preferred 30min of microwave irradiating time;
Preferred 60 ℃ of microwave reaction temperature.
Method of the present invention has easy and simple to handle, and the reaction times is short, and yield is higher, does not produce waste water, has avoided simultaneously the moisture defective of product.
Embodiment
Be described in further detail by the following examples, but this should be interpreted as that scope of the present invention only limits to following examples, all technology that realizes based on technical scheme of the present invention all belong to scope of the present invention.
Embodiment 1
Taking heparin sodium 10g, sodium-chlor 50mg, benzyl rope chloramines 25g mixes, and places microwave reactor, and power 150W behind the reaction 30min, cools off to get heparin quaternary ammonium salt 30g under the temperature 60 C;
Adopt the method for the method of US Patent No. 5389618, the heparin quaternary ammonium salt gets enoxaparin 18.5g through esterification, cracking, purifying.
Product test is as follows:
Reference substance source: enoxaparin sodium injection (Ke Sai, the lot number: 110601-J), press Enoxaparin Sodium feed purification method by Suzhou Erye Pharmaceutical Co., Ltd., extract refining of getting import.
One, high resolution mass spectrum
Mass spectrometer system: the LCMS-IT-TOF of Shimadzu company
The off-line positive ion mode, spray voltage 2.0kV, spraying gas velocity 0.5L/min, 200 ℃ of CDL temperature, 200 ℃ of heating module temperature, detector voltage 1.7kV, ion trap zone air pressure 2.4e-002Pa, TOF zone air pressure 1.8e-004Pa, 40 ℃ of TOF temperature.MS sweep limit: 200-1000m/z, ion accumulation time (Ion Accumulation Time) 20msec, multiplicity 2 times.
Reference substance and samples with water are dissolved to 5ug/ul.Regulating sample concentration with acetonitrile and formic acid is 2.5ug/ul.System is 50% acetonitrile/0.1 formic acid.All the other solution examples dilute one times with acetonitrile and formic acid, and mediation body is 50% acetonitrile/0.1 formic acid
1, molecular weight and distribution thereof
Measure by high resolution mass spectrum, according to m/z=(M+nH)/n formula to calculating molecular weight distribution, the molecular weight of the reference substance of Enoxaparin Sodium is from 226-2366, and the molecular weight analyte of Enoxaparin Sodium is from 273-2406, be normal distribution, the molecular weight distribution of the two is basically identical.
2, the spectrum of the disaccharides after the hydrolysis, monose spectrum
In the disaccharides spectrum, sample and reference substance all locate to have identical spectrum peak at m/z=226,301,362,378,453,498,675 etc.; And 301 and 453 places are the highest peak position; In the monose spectrum, sample all has identical spectrum peak at m/z=149,205,279,280,391,445,588,668 places with reference substance; And 149,205,391, locate to be relative highest peak position; Prompting sample and reference substance have essentially identical disaccharides spectrum after hydrolysis and the monose spectrum distributes.
Two, high performance liquid chromatography
Instrument title: Varian Prostar 500 high performance liquid chromatographs
Chromatographic column: 250mm*4.6mm SAX post Machery-Nagel company
Pre-column: 5mm*4.6mm SAX post Machery-Nagel company
Moving phase: (A) 0.28g/L potassium dihydrogen phosphate (pH=3)
(B) 140g/L sodium perchlorate solution (pH=3 is with the preparation of pH=3 potassium dihydrogen phosphate)
Gradient: 0-40 minute A 97%-65% B 3%-35%
40-60 minute A 65%-50% B 35%-50%
60-70 minute A 50% B 50%
70-71 minute A 50%-97% B 50%-3%
71-90 minute A 97% B 3%
HPLC result shows, the monose spectrum behind trifluoroacetic acid hydrolysis, and the peak shape of reference substance and sample is basic identical, and the symmetry between 15-25 minute is better, confirms all to have in the product after reference substance and sample are hydrolyzed the existence of seminose.
Three, chromatography of ions
Ion chromatograph: DIONEX 500 ion chromatographs
Chromatographic column: PA-1 post
Detectability :≤0.02mg/L (Cl-)
Chromatography of ions detects and shows, the monose spectrum peak shape behind the trifluoroacetic acid hydrolysis of enoxaparin sodium sample and reference substance is basic identical.
Four, comprehensive conclusion
Detect through high resolution mass spectrum, high performance liquid chromatography, chromatography of ions, carry out respectively heparinase hydrolysis and trifluoroacetic acid hydrolysis by reference substance and sample to Enoxaparin Sodium, obtain the two disaccharides spectrum and monose spectrum, compare by spectrogram, the sample of Enoxaparin Sodium and reference substance spectrogram structure are basic identical, and tentatively confirmatory sample forms basically identical with reference substance at monose and disaccharides structure.
Claims (4)
1. enoxaparin, its structure is as follows:
Wherein, n=1-21, R=H or SO3Na, R1=H or SO3Na or COCH3, R2=H and R3=COONa, or R2=COONa and R3=H.
2. the preparation method of Enoxaparin Sodium as claimed in claim 1 is characterized in that: take heparin sodium and benzethonium chloride as raw material, take sodium-chlor as the neutral reaction medium, prepare the heparin quaternary ammonium salt by the Microwave Solid synthetic method.
3. 2 method as requested is characterized in that:
1) heparin sodium and NaCl, benzyl rope chloramines are mixed;
2) mixture is placed microwave reactor, with the power irradiation of 100-200W, temperature 40-80 ℃, irradiation time is 15-45min;
3) be cooled to room temperature, get the heparin quaternary ammonium salt;
4) the heparin quaternary ammonium salt gets Enoxaparin Sodium through esterification, cracking.
4. according to claim 3 method is characterized in that:
Wherein the weight part of NaCl is 0.002-0.01, preferred 0.005 weight part;
Benzyl rope chloramines weight part is 1-2, preferred 1.5 weight parts;
The preferred 150W of microwave power;
The preferred 30min of microwave irradiating time;
Preferred 60 ℃ of microwave reaction temperature.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104086674A (en) * | 2014-07-28 | 2014-10-08 | 常州千红生化制药股份有限公司 | Process for preparing enoxaparin sodium |
| WO2019116217A2 (en) | 2017-12-11 | 2019-06-20 | Biological E Limited | Process for the preparation of low molecular weight heparin |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1850865A (en) * | 2006-05-24 | 2006-10-25 | 杭州九源基因工程有限公司 | Production method for purifying enoxaparin sodium |
| CN101165071A (en) * | 2006-10-20 | 2008-04-23 | 江苏江山制药有限公司 | Clexane and preparation method thereof |
-
2012
- 2012-12-03 CN CN2012105082530A patent/CN102924629B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1850865A (en) * | 2006-05-24 | 2006-10-25 | 杭州九源基因工程有限公司 | Production method for purifying enoxaparin sodium |
| CN101165071A (en) * | 2006-10-20 | 2008-04-23 | 江苏江山制药有限公司 | Clexane and preparation method thereof |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104086674A (en) * | 2014-07-28 | 2014-10-08 | 常州千红生化制药股份有限公司 | Process for preparing enoxaparin sodium |
| WO2019116217A2 (en) | 2017-12-11 | 2019-06-20 | Biological E Limited | Process for the preparation of low molecular weight heparin |
| US11299558B2 (en) | 2017-12-11 | 2022-04-12 | Biological E Limited | Process for the preparation of low molecular weight heparin |
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| CN102924629B (en) | 2013-11-13 |
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