CN104558252B - A kind of method that Enoxaparin Sodium is produced by heparin sodium crude - Google Patents
A kind of method that Enoxaparin Sodium is produced by heparin sodium crude Download PDFInfo
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Abstract
The invention discloses a kind of method that Enoxaparin Sodium is produced by heparin sodium crude.The method is with heparin sodium crude as raw material, pre-processed using salt solution technique, then oxidized, ion exchange resin absorption, washing, wash-out, most afterwards through ultrafiltration, it is lyophilized to obtain refined heparin sodium, refined heparin sodium obtains Enoxaparin Sodium through into salt, esterification, depolymerization, oxidation, alcohol precipitation, frozen dried again.The invention has the advantages that controlling Enoxaparin Sodium quality from source and technique, with short production cycle, energy consumption is low, product quality is high, is suitable for large-scale industrial production.
Description
Technical field
The invention belongs to biochemical drug production technical field, and in particular to one kind produces Enoxaparin by heparin sodium crude
The method of sodium.
Background technology
Enoxaparin Sodium is a kind of LMWHs sodium salt, the benzyl ester derivant of the heparin by being originated to pig intestinal mucosa
Carry out alkaline hydrolysis poly- and obtain.Fail completely that qualitatively complex oligosaccharide is constituted by a series of, non-reducing end is mainly by 4- enol form pyrroles
Uronic acid composition is fed, 15%~25% component includes 1,6- anhydro derivatives structures in reducing end.Its weight average molecular weight is
3800~5000.Calculated by dry product, Anti-Xa factor activity potency is 90IU/mg~125IU/mg, and anti-IIa factors potency is
20.0IU/mg~35.0IU/mg, Anti-Xa factor potency is 3.3~5.3 with anti-IIa factors potency ratio.The anti-freezing of Enoxaparin Sodium
Effect is suitable with Natural heparin or more excellent, and the generation of bleeding is less than application Natural heparin, and bioavilability is high in addition, thus
Enjoy clinical favor.Current Enoxaparin Sodium has been used for prevention venous embolism disease (preventing intravenous thrombosis), especially
It is the embolism relevant with some operations；Treatment venae profunda conducted, with or without pulmonary embolism；Treatment unstable angina and non-
Q ripple heart infarctions, with aspirin with use；In for haemodialysis extracorporal circulatory system, thrombosis etc. is prevented.
It is more to the preparation method of Enoxaparin Sodium both at home and abroad at present, mostly using directly from liquaemin by into salt, ester
Change, oxidation, ultrafiltration, drying and other steps obtain [CN102050888B, CN 103342761A].In process of production, liquaemin
Quality has a significant impact to Enoxaparin Sodium finished product.Because liquaemin molecular structure complex, molecular weight is difficult to solid
Fixed, the liquaemin in different process source directly affects the product quality of Enoxaparin.The structure of Enoxaparin Sodium and ratio of components compared with
, there is certain uncertainty in complexity, thus initiation material and production process grind on structural confirmation and quality standard control
Studying carefully just must be strict with control, and raw material has a direct impact to the molecular weight distribution of later stage Enoxaparin Sodium finished product.Therefore develop
The new technology for being directly produced Enoxaparin Sodium from heparin sodium crude can be more effective to ensure from source and technology controlling and process material quality
Product quality.Under the premise of herein, reducing energy consumption, reduction environmental protection pressure are still that those skilled in the art need what is solved to ask as far as possible
Topic.CN102603925B provides a kind of method that Enoxaparin Sodium is directly produced by heparin sodium crude, and heparin sodium crude is first through alcohol
Heavy, oxidation, alcohol precipitation are precipitated thing, and this sediment is used to prepare Enoxaparin Sodium；The sediment through into salt, esterification, degraded, tree
Fat, oxidation, alcohol precipitation, last dissolution filter, are spray-dried to obtain Enoxaparin Sodium finished product.Liquaemin removal step in the method is only
By using and use oxidation for organic solvent, it is unfavorable for obtaining high-quality finished product；Prepared according to promise by liquaemin sediment
It is that can remove sulfate radical in alcohol precipitation step during liquaemin, and sulfate radical is removed using resin in patent, complex steps.
The content of the invention
It is an object of the present invention to overcome the above deficiencies, there is provided one kind is directly produced Enoxaparin from heparin sodium crude
The preparation technology of sodium, the with short production cycle, preparation method that the production low, product quality of power consumption is high.The present invention is with heparin sodium crude
Raw material, is pre-processed using salt solution technique, is subsequently adding hydrogen peroxide, through ion exchange resin absorption, washing, is eluted, most
By ultrafiltration, refined heparin sodium is freezed to obtain, refined heparin sodium is obtained through into salt, esterification, depolymerization, oxidation, alcohol precipitation, frozen dried again
Enoxaparin Sodium.It is simple for process, it is suitable to industrial-scale production.
The inventive method the following specifically describes below：
A kind of method that Enoxaparin Sodium is produced by heparin sodium crude, the method is comprised the following steps：
1) preparation of refined heparin sodium：Sodium chloride is added after heparin sodium crude purifying water dissolves, while adjust pH value being
8.5~9.0, salt solution is carried out, the centrifugation of salt solution liquid obtains liquaemin pretreatment fluid；Gained liquaemin pretreatment fluid temperature is down to 28~
32 DEG C, hydrogen peroxide is added, then add to absorption, washing, wash-out in strong basic type anion-exchange resin post, collect liquaemin wash-out
Liquid；Eluent adjust pH value to after 5.5~6.0 through ultrafiltration, lyophilized obtain refined heparin sodium；
2) into salt：Refined heparin sodium is dissolved in purified water and is made the refined heparin sodium aqueous solution, benzethonium chloride is dissolved in purifying
Water is made the benzethonium chloride aqueous solution, is reacted in room temperature after two kinds of solution are mixed with 1: 2~3: 4 volume ratio, reaction knot
It is centrifuged after beam, washing precipitate, dry heparin quaternary ammonium salt；
3) it is esterified：Benzyl chloride is added after heparin quaternary ammonium salt dichloromethane is dissolved, is esterified at 30~35 DEG C, ester
Change is cooled to room temperature after terminating, and adds methanol extraction, precipitation and separation and washed with methyl alcohol, dry heparin benzyl ester；
4) depolymerization：Heparin benzyl ester water dissolves, to NaOH is added in solution, carry out depolymerization anti-at 50~60 DEG C
Should, reaction terminates to add methanol extraction, precipitation and separation to obtain heparin depolymerization product；
5) aoxidize：Heparin depolymerization product purifying water dissolves, regulation pH is 8.5~10.0, adds hydrogen peroxide to be aoxidized
Decolourize；
6) alcohol precipitation, lyophilized：By the solution methanol extraction after oxidative decoloration, precipitation and separation, precipitation is with after purifying water dissolves
Freeze to obtain Enoxaparin.
Wherein step 1) described in purified water, it is 8~10ml/g that purified water adds the ratio of volume and heparin sodium crude weight；
The ratio of the sodium chloride addition and purifying water consumption volume is 1/100~2/100g/ml；Salt solution temperature is 50 DEG C~60 DEG C,
The salt solution time is 2~3 hours.
Wherein step 1) described in hydrogen peroxide, addition is the 1%~2% of liquaemin pretreatment fluid volume, be volume with
Volume ratio.
Wherein step 1) described in strong basic type anion-exchange resin be OC1074 or FPA-98.
Wherein step 1) described in adsorb, adsorption rate for 0.05~0.06 times of resin column volume per hour (0.05~
0.06BV/h)；The washing, washing solution used is 5.5%~6.0% sodium chloride solution, and washing speed is per hour 0.5
Times resin column volume (0.5BV/h)；The wash-out, wash-out solution used is 12% sodium chloride solution, and elution speed is per small
When 0.05~0.06 times of resin column volume (0.05-0.06BV/h).
Wherein step 2) described in the refined heparin sodium aqueous solution be to add 10-12ml purifying water-soluble by every gram of refined heparin sodium
Solution gained, the benzethonium chloride aqueous solution is to add 8~10ml purifying water dissolves gained by every gram of benzethonium chloride；The salt-forming reaction time is
1~2 hour.
Wherein step 2) described in dry be drying under reduced pressure, temperature 50 C~55 DEG C, vacuum is -0.1~-0.09Mpa,
40~60 hours drying times.
Wherein step 3) in methylene chloride be that every gram of heparin quaternary ammonium salt adds 3~5 milliliters of dichloromethane, benzyl chloride to add
It is that every gram of heparin quaternary ammonium salt adds 1 milliliter of benzyl chloride to enter amount；Esterification time is 25~30 hours；The drying refers to drying temperature
It is 40 DEG C~45 DEG C of drying under reduced pressure, vacuum is -0.1~-0.09Mpa, 10 hours~15 hours drying time.
Wherein step 4) described in heparin benzyl ester with 20 times of water dissolves, i.e., add 20ml water dissolves by every gram of heparin benzyl ester,
Sodium hydroxide concentration is the 8~10% of heparin benzyl ester weight, and the depolymerization time is 1~1.5 hour.
Wherein step 5) described in dioxygen water consumption volume be purify water volume 1%~1.5%.
Wherein step 6) in add methyl alcohol concentration be 75%~80% in the solution.
The present invention with heparin sodium crude as raw material, first by step 1) obtain refined heparin sodium.Heparin sodium crude passes through
Salt solution separates albumen and liquaemin, and the removal of impurity is gone in centrifugation.Addition 1%~2% is double in the liquaemin pretreatment fluid of acquisition
Oxygen water, the simultaneous oxidation in resin adsorption is decolourized simultaneously in purifying resin.Gradient elution technique is used in ion exchange,
The impurity such as protein, nucleic acid, heparan can be removed, and a step completes removal of impurities and decolourizes.Compare with existing Purification of Heparin Sodium,
Repeatedly oxidation, multiple alcohol precipitation step can be exempted, consumed energy low, not use organic solvent, it is economic and environment-friendly.The liquaemin essence for preparing
Product are further used for the production of Enoxaparin Sodium, and the product quality of Enoxaparin Sodium can be effectively ensured.
The process portion of Enoxaparin Sodium is being prepared from refined heparin sodium, heparin depolymerizing substance is being obtained through into salt, esterification, depolymerization
Afterwards, once oxidation, alcohol precipitation are only needed, lyophilized to can obtain Enoxaparin Sodium, product quality meets European Pharmacopoeia standard and U.S.'s medicine
Allusion quotation standard.
The device have the advantages that：
1st, refined heparin sodium preparation technology is succinct.In ion-exchange step, hydrogen peroxide is added in liquaemin pretreatment fluid,
Simultaneous oxidation during resin adsorption, decolourizes simultaneously in purifying resin；Gradient elution technique is used in ion exchange, can be removed
The impurity such as protein, nucleic acid, heparan, and a step completes removal of impurities decolouring.Compare with existing process, reduce repeatedly oxidation, it is many
Secondary alcohol precipitation step, consumes energy low, organic solvent is not used, with important economic worth.
2nd, the refined heparin sodium for preparing is further used for the production of Enoxaparin Sodium, and Enoxaparin can be effectively ensured
The product quality of sodium.
3rd, to prepare the technique of Enoxaparin Sodium by the refined heparin sodium also corresponding succinct.After obtaining heparin depolymerization product, only
Once oxidation, alcohol precipitation are needed, freezing can obtain Enoxaparin Sodium.
4th, the Enoxaparin Sodium product quality for preparing meets European Pharmacopoeia and USP standard.
5th, the present invention obtains refined heparin sodium with heparin sodium crude as raw material, is used further to produce Enoxaparin Sodium, whole system
Standby concise in technology, it is with short production cycle, and the product quality of Enoxaparin Sodium can be effectively ensured from source and technique.
Specific embodiment
Following embodiments are used merely to explain realizes the method for the present invention, should not be construed as limiting the invention, Suo Youji
The modifications and variations made in thinking of the invention all should be attributed to protection scope of the present invention.
Heparin sodium crude used in the present invention is that pharmacy Hua Kun Hebei, North China Bioisystech Co., Ltd is made；Hydrogen peroxide
(commercially available, 30% content), NaOH, sodium chloride, benzethonium chloride, dichloromethane, benzyl chloride, methyl alcohol, ethanol etc. are commercially available；From
Sub-exchange resin OC1074 is U.S.'s ROHM AND HAAS product for German Lanxess Corporation product, FPA-98.
Embodiment 1
1 kilogram of taking heparin sodium crude product, water dissolves are purified with 8L, add 80g sodium chloride, rise high-temperature to 50 DEG C, while with
1mol/L sodium hydroxide solutions regulation pH value is 8.5, with this understanding salt solution 2 hours, and salt solution liquid is centrifuged to obtain liquaemin pretreatment
Liquid.Liquaemin pretreatment fluid temperature is down to 28 DEG C, hydrogen peroxide is added, its volume is the 1% of liquaemin pretreatment fluid, and stirring is equal
Added to after even and adsorbed in the resin column equipped with 15L OC1074 resins, control adsorption rate for 0.05BV/h, absorption is finished
Resin afterwards is first washed with 5.5% sodium chloride solution, flow velocity 0.5BV/h, washing to absorbance A260nm≤0.20、A280nm≤
When 0.20, eluted with 12% sodium chloride solution, elution speed 0.05BV/h, detect lower column liquid refractive power, opened when refractive power is more than 5.5
Begin to collect eluent, about collect 1BV.It is 5.5 that the eluent of collection 2mol/L hydrochloric acid solutions adjust pH, through ultrafiltration, is freezed
Refined heparin sodium 340g.Refined heparin sodium dissolved with 10 times of water (3.4L) after with the benzethonium chloride aqueous solution (680g is dissolved in 6.8L water)
Mixing, room temperature carries out reaction 1 hour, and reaction is centrifuged after terminating, and washes sediment with water, then dry heparin quaternary ammonium salt
1020g (55 DEG C of drying temperature of control, vacuum -0.1Mpa, 40 hours drying times)；Heparin quaternary ammonium salt is with 3 times (3060ml)
1020ml benzyl chlorides are added after dichloromethane dissolving, are esterified at 30 DEG C, esterification terminates after 25 hours, is cooled to room temperature,
Methanol extraction is added, precipitation and separation simultaneously wash with methyl alcohol, dry heparin benzyl ester (it is 40 DEG C to control drying temperature, vacuum-
0.092Mpa, 15 hours drying times)；Heparin benzyl ester adds the NaOH of heparin benzyl ester weight 8% with 20 times of water dissolves,
Depolymerization reaction is carried out at 50 DEG C 1.5 hours, reaction terminates to add methanol extraction, and precipitation and separation is heparin depolymerization product；Heparin solution
Poly- product purifying water dissolves, it is 8.5 to adjust pH with 1mol/L sodium hydroxide solutions, adds 1% hydrogen peroxide to carry out oxidative decoloration；
By the solution methanol extraction after oxidative decoloration, concentration is 75% in the solution to make methyl alcohol to add the amount of methyl alcohol, and it is heavy to separate
Form sediment, precipitation to obtain Enoxaparin Sodium 253.2g with lyophilized after purifying water dissolves.Its testing result is shown in Table 1.
Embodiment 2
1 kilogram of taking heparin sodium crude product, water dissolves are purified with 10L, add 200g sodium chloride, rise high-temperature to 60 DEG C, while
It is 9.0 to adjust pH value with the sodium hydroxide solution of 3mol/L, with this understanding salt solution 3 hours, salt solution liquid be centrifuged liquaemin is pre-
Treatment fluid.Liquaemin pretreatment fluid temperature is down to 32 DEG C, hydrogen peroxide is added, its volume is the 2% of liquaemin pretreatment fluid, is stirred
Mix it is uniform after add to and adsorbed in the resin column equipped with 15L FPA98 resins, it is 0.06BV/h control adsorption rate, has been adsorbed
Resin after finishing first is washed with 6% sodium chloride solution, flow velocity 0.5BV/h, washing to absorbance A260nm≤0.20、A280nm≤
When 0.20, eluted with 12% sodium chloride solution, elution speed 0.06BV/h, detect lower column liquid refractive power, started when refractive power is more than 6
Eluent is collected, 1BV is about collected.It is 6.0 that the eluent of collection 6mol/L hydrochloric acid solutions adjust pH, then through ultrafiltration, lyophilized
Refined heparin sodium 310g.Refined heparin sodium dissolved with 12 times of water (3.72L) after with the benzethonium chloride aqueous solution (620g is dissolved in 8 times of water,
Close 4.96L water) mix, room temperature carries out reaction 1 hour, and reaction is centrifuged after terminating, and washes sediment with water, then dries to obtain heparin
Quaternary ammonium salt 900g (drying under reduced pressure, temperature 50 C, vacuum -0.093Mpa, 45 hours drying times)；Heparin quaternary ammonium salt is with 5 times
900ml benzyl chlorides are added after the dissolving of (4.5L) dichloromethane, is esterified at 30 DEG C, esterification terminates after 28 hours, is cooled to
Room temperature, adds methanol extraction, and precipitation and separation simultaneously wash with methyl alcohol, dry heparin benzyl ester (drying temperature is 45 DEG C, vacuum-
0.1Mpa, 10 hours drying times)；Heparin benzyl ester adds the NaOH of heparin benzyl ester weight 9%, 50 with 20 times of water dissolves
Depolymerization reaction is carried out at DEG C 1 hour, reaction terminates to add methanol extraction, and precipitation and separation is heparin depolymerization product；Heparin depolymerization is produced
Thing purifying water dissolves, pH10.0 is adjusted with the sodium hydroxide solution of 3mol/L, adds 1.0% hydrogen peroxide to carry out oxidative decoloration；
By the solution methanol extraction after oxidative decoloration, concentration is 80% in the solution to add methyl alcohol to make methyl alcohol, and precipitation and separation, precipitation is used
Enoxaparin Sodium 234.4g is freezed to obtain after purifying water dissolves.Its testing result is shown in Table 1.
Embodiment 3
1 kilogram of taking heparin sodium crude product, water dissolves are purified with 10L, add 200g sodium chloride, rise high-temperature to 50 DEG C, while
It is 8.5 to adjust pH value with 6mol/L sodium hydroxide solutions, salt solution 2 hours with this understanding, salt solution liquid be centrifuged liquaemin is located in advance
Reason liquid.Liquaemin pretreatment fluid temperature is down to 30 DEG C, hydrogen peroxide is added, its volume is the 1.5% of liquaemin pretreatment fluid, is stirred
Mix it is uniform after add to and adsorbed in the resin column equipped with 15L FPA98 resins, it is 0.05BV/h control adsorption rate, has been adsorbed
Resin after finishing first is washed with 5.5% sodium chloride solution, flow velocity 0.5BV/h, washing to absorbance A260nm≤0.20、A280nm≤
When 0.20, eluted with 12% sodium chloride solution, elution speed 0.05BV/h, detect lower column liquid refractive power, opened when refractive power is more than 5.5
Begin to collect eluent, about collect 1BV.It is 5.5 that the eluent of collection 8mol/L hydrochloric acid solutions adjust pH, through ultrafiltration, is freezed
Refined heparin sodium 349g.Refined heparin sodium dissolved with 11 times of water (3.84L) after with the benzethonium chloride aqueous solution (698g is dissolved in 9 times of water,
Close 6.28L water) mix, room temperature carries out reaction 1.5 hours, and reaction is centrifuged after terminating, and washes sediment with water, then dries to obtain liver
Plain quaternary ammonium salt 1025g (drying under reduced pressure, temperature 50 C, vacuum -0.09Mpa, 60 hours drying times)；Heparin quaternary ammonium salt uses 4
1025ml benzyl chlorides are added after the dissolving of times dichloromethane (4.1L), is esterified at 30 DEG C, esterification terminates after 25 hours, cooling
To room temperature, methanol extraction, precipitation and separation is added simultaneously to be washed with methyl alcohol, (drying temperature is 43 DEG C of decompression to dry heparin benzyl ester
Dry, vacuum -0.096Mpa, 45 hours drying times)；Heparin benzyl ester adds heparin benzyl ester weight 8% with 20 times of water dissolves
NaOH, depolymerization reaction is carried out at 50 DEG C 1.5 hours, reaction terminate add methanol extraction, precipitation and separation be heparin depolymerization
Product；Heparin depolymerization product purifying water dissolves, pH9.5 is adjusted with 6mol/L sodium hydroxide solutions, adds 1.5% hydrogen peroxide to enter
Row oxidative decoloration；By the solution methanol extraction after oxidative decoloration, concentration is 80% in the solution to add methyl alcohol to make methyl alcohol, is separated
Precipitation, precipitation to obtain Enoxaparin Sodium 261.1g with lyophilized after purifying water dissolves.Its testing result is shown in Table 1.
Embodiment 4
1 kilogram of taking heparin sodium crude product, water dissolves are purified with 9L, add 135g sodium chloride, rise high-temperature to 55 DEG C, while with
4mol/L sodium hydroxide solutions regulation pH value is 9.0, salt solution 2.5 hours with this understanding, salt solution liquid be centrifuged liquaemin is located in advance
Reason liquid.Liquaemin pretreatment fluid temperature is down to 30 DEG C, hydrogen peroxide is added, its volume is the 1% of liquaemin pretreatment fluid, stirring
Added to after uniform and adsorbed in the resin column equipped with 15L OC1074 resins, it is 0.06BV/h control adsorption rate, has been adsorbed
Resin after finishing first is washed with 5.5% sodium chloride solution, flow velocity 0.5BV/h, washing to absorbance A260nm≤0.20、A280nm≤
When 0.20, eluted with 12% sodium chloride solution, elution speed 0.06BV/h, detect lower column liquid refractive power, opened when refractive power is more than 5.5
Begin to collect eluent, about collect 1BV.It is 5.8 that the eluent of collection 1mol/L hydrochloric acid solutions adjust pH, then through ultrafiltration, lyophilized
Obtain refined heparin sodium 325g.With after 10 times of water dissolves, (650g is dissolved in 10 times of water to refined heparin sodium, closes with the benzethonium chloride aqueous solution
6.5L) mix, room temperature carries out reaction 2 hours, reaction is centrifuged after terminating, and washes sediment with water, then dries to obtain heparin quaternary ammonium
Salt 940g (drying under reduced pressure, temperature 50 C, vacuum -0.096Mpa, 50 hours drying times)；Heparin quaternary ammonium salt is with 4 times of dichloros
Methane dissolving (3760ml) adds 940ml benzyl chlorides afterwards, is esterified at 30 DEG C, and esterification terminates after 30 hours, is cooled to room
Temperature, adds methanol extraction, and precipitation and separation simultaneously washs with methyl alcohol, dry heparin benzyl ester (drying temperature is 40 DEG C of drying under reduced pressure,
Vacuum -0.09Mpa, 12 hours drying times)；Heparin benzyl ester adds the hydrogen of heparin benzyl ester weight 10% with 20 times of water dissolves
Sodium oxide molybdena, carries out depolymerization reaction 1 hour at 50 DEG C, reaction terminates to add methanol extraction, and precipitation and separation is heparin depolymerization product；Liver
Plain depolymerization product purifying water dissolves, pH10.0 is adjusted with 4mol/L sodium hydroxide solutions, adds 1.5% hydrogen peroxide to be aoxidized
Decolourize；By the solution methanol extraction after oxidative decoloration, concentration is 75% in the solution to add methyl alcohol to make methyl alcohol, precipitation and separation,
Precipitation to obtain Enoxaparin Sodium 240.0g with lyophilized after purifying water dissolves.Its testing result is shown in Table 1.
The testing result of the gained Enoxaparin Sodium finished product of 1 embodiment of table 1~4
Claims (7)
1. it is a kind of by heparin sodium crude produce Enoxaparin Sodium method, it is characterised in that the method is comprised the following steps：
1) preparation of refined heparin sodium：Heparin sodium crude purifying water dissolves after add sodium chloride, while adjust pH value be 8.5~
9.0, salt solution is carried out, the centrifugation of salt solution liquid obtains liquaemin pretreatment fluid；Gained liquaemin pretreatment fluid temperature is down to 28~32 DEG C,
Hydrogen peroxide is added, absorption, washing, wash-out in strong basic type anion-exchange resin post is then added to, liquaemin eluent is collected, its
In, the simultaneous oxidation in resin adsorption is decolourized simultaneously in purifying resin, and a step completes removal of impurities and decolourizes；Eluent adjusts pH value
After to 5.5~6.0 through ultrafiltration, lyophilized obtain refined heparin sodium；
Wherein described purified water, it is 8~10ml/g that purified water adds the ratio of volume and heparin sodium crude weight；The sodium chloride adds
The ratio for entering amount and purifying water consumption volume is 1/100~2/100g/ml；Salt solution temperature be 50 DEG C~60 DEG C, the salt solution time be 2~
3 hours；
Wherein described hydrogen peroxide, addition is the 1%~2% of liquaemin pretreatment fluid volume, is volume and volume ratio；
Wherein described strong basic type anion-exchange resin is OC1074 or FPA-98；
Wherein described absorption, adsorption rate is 0.05~0.06 times of resin column volume per hour；The washing, washing solution used
It is 5.5%~6.0% sodium chloride solution, washing speed is 0.5 times of resin column volume per hour；The wash-out, wash-out is used
Solution is 12% sodium chloride solution, and elution speed is 0.05~0.06 times of resin column volume per hour；
Wherein, the washing, washing to absorbance A260nm≤0.20、A280nmWhen≤0.20, eluted with 12% sodium chloride solution, inspection
Lower column liquid refractive power is surveyed, starts to collect eluent when refractive power is more than 5.5；
2) into salt：Refined heparin sodium is dissolved in purified water and is made the refined heparin sodium aqueous solution, benzethonium chloride is dissolved in purified water system
Into the benzethonium chloride aqueous solution, reacted in room temperature after two kinds of solution are mixed with 1: 2~3: 4 volume ratio, after reaction terminates
Centrifugation, washing precipitate, dry heparin quaternary ammonium salt；
3) it is esterified：Benzyl chloride is added after heparin quaternary ammonium salt dichloromethane is dissolved, is esterified at 30~35 DEG C, esterification knot
Be cooled to room temperature after beam, add methyl alcohol, precipitation and separation and washed with methyl alcohol, dry heparin benzyl ester；
4) depolymerization：Heparin benzyl ester water dissolves, to NaOH is added in solution, carry out depolymerization reaction, instead at 50~60 DEG C
Should terminate to add methanol extraction, precipitation and separation to obtain heparin depolymerization product；
5) aoxidize：Heparin depolymerization product purifying water dissolves, regulation pH is 8.5~10.0, adds hydrogen peroxide to carry out oxidative decoloration；
6) alcohol precipitation, lyophilized：Solution methanol extraction after oxidative decoloration, precipitation and separation, precipitation are freezed after using purifying water dissolves
Obtain Enoxaparin Sodium.
2. method according to claim 1, wherein step 2) described in the refined heparin sodium aqueous solution be by every gram of liquaemin
Fine work adds 10~12ml purifying water dissolves gained, and the benzethonium chloride aqueous solution is to add 8~10ml to purify by every gram of benzethonium chloride
Water dissolves gained；The salt-forming reaction time is 1~2 hour.
3. method according to claim 1, wherein step 2) described in dry be drying under reduced pressure, temperature 50 C~55 DEG C,
Vacuum is -0.1~-0.09Mpa, 40~60 hours drying times.
4. method according to claim 1, wherein step 3) in methylene chloride be every gram of heparin quaternary ammonium salt add 3~
5 milliliters of dichloromethane, benzyl chloride addition is that every gram of heparin quaternary ammonium salt adds 1 milliliter of benzyl chloride；Esterification time is 25~30 small
When；The drying refers to the drying under reduced pressure that drying temperature is 40 DEG C~45 DEG C, and vacuum is -0.1~-0.09Mpa, drying time
10 hours~15 hours.
5. method according to claim 1, wherein step 4) described in heparin benzyl ester water dissolves, the addition of water is every
Gram heparin benzyl ester adds 20ml water dissolves, and the sodium hydroxide concentration is the 8~10% of heparin benzyl ester weight, and the depolymerization time is 1
~1.5 hours.
6. method according to claim 1, wherein step 5) described in dioxygen water consumption volume be purify water volume 1%
~1.5%.
7. method according to claim 1, wherein step 6) in methyl alcohol concentration is 75%~80% in the solution.
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| CN105131153A (en) * | 2015-08-21 | 2015-12-09 | 苏州融析生物科技有限公司 | Sheep enoxaparin sodium compound preparation method, compound and application of compound |
| CN108219030A (en) * | 2016-12-21 | 2018-06-29 | 鲁南制药集团股份有限公司 | A kind of preparation method of Enoxaparin Sodium crude product |
| CN106967187A (en) * | 2017-05-16 | 2017-07-21 | 苏州二叶制药有限公司 | A kind of macromolecular liquaemin pretreating process |
| CN109467619A (en) * | 2017-09-08 | 2019-03-15 | 山阳县恒瑞肉制品有限公司 | A method of Enoxaparin Sodium is produced by heparin sodium crude |
| US11299558B2 (en) | 2017-12-11 | 2022-04-12 | Biological E Limited | Process for the preparation of low molecular weight heparin |
| CN109485749A (en) * | 2018-10-31 | 2019-03-19 | 江西浩然生物医药有限公司 | A method of chromatography and Ultrafiltration Membrane prepare Enoxaparin Sodium |
| CN116284499A (en) * | 2022-07-28 | 2023-06-23 | 河北常山生化药业股份有限公司 | Preparation method of sheep-derived low-molecular heparin sodium |
| CN115304688A (en) * | 2022-08-25 | 2022-11-08 | 湖北亿诺瑞生物制药有限公司 | Preparation method of dalteparin sodium |
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| CN102585037A (en) * | 2012-02-10 | 2012-07-18 | 麦科罗夫（南通）生物制药有限公司 | Enoxaparin sodium and production purification method thereof |
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