CN112480208A - Industrial preparation method of glycyl-L-glutamine - Google Patents
Industrial preparation method of glycyl-L-glutamine Download PDFInfo
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- CN112480208A CN112480208A CN202011441553.2A CN202011441553A CN112480208A CN 112480208 A CN112480208 A CN 112480208A CN 202011441553 A CN202011441553 A CN 202011441553A CN 112480208 A CN112480208 A CN 112480208A
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- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
- C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
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- C07K5/06008—Dipeptides with the first amino acid being neutral
- C07K5/06017—Dipeptides with the first amino acid being neutral and aliphatic
- C07K5/06026—Dipeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atom, i.e. Gly or Ala
Abstract
The invention relates to an industrial preparation method of glycyl-L-glutamine, belonging to the technical field of organic synthesis, firstly, chloroacetyl chloride and L-glutamine are subjected to acylation reaction under low temperature and alkaline conditions, phase separation is carried out to obtain an N-chloroacetyl-L-glutamine aqueous solution, then electrodialysis is adopted to treat the aqueous solution, ammonia gas is introduced into the aqueous solution obtained by treatment to carry out ammonolysis under pressure, and a glycyl-L-glutamine crude product is obtained through concentration, crystallization, filtration and drying; finally dissolving the crude glycyl-L-glutamine in water, purifying by WA-30 resin, and crystallizing to obtain the high-purity glycyl-L-glutamine. The invention improves the yield and quality of the product, simplifies the process steps, realizes the preparation of the dipeptide crude product by a one-pot method, generates less industrial three wastes, has simple and convenient process operation, is suitable for large-scale industrial production, and can meet the requirement of injection-grade raw materials on the quality of the prepared glycyl-L-glutamine product.
Description
Technical Field
The invention relates to an industrial preparation method of glycyl-L-glutamine, belonging to the technical field of organic synthesis.
Background
Glycyl-L-glutamine (Glycyl-L-glutamine) belongs to artificially synthesized dipeptide amino acid, and is one of main components of compound amino acid (15) dipeptide (2) injection. glycyl-L-glutamine is mainly used to provide glutamine to promote protein synthesis. Compared with L-glutamine, the water solubility and the heat stability of the glutamine are better. glycyl-L-glutamine is used as a carrier for transporting nitrogen sources in vivo, can remove metabolic wastes such as ammonia and the like, provides nitrogen sources for the synthesis of proteins and DNA, and plays a very important role in the stability of proteins in cells. With the rapid development of medical technology, the application of polypeptide amino acids in clinic is increasing.
At present, the preparation methods of glycyl-L-glutamine mainly comprise the following steps:
firstly, taking N-benzyloxycarbonyl glycine and L-glutamine as starting raw materials, preparing active ester from the N-benzyloxycarbonyl glycine and N-Hydroxysuccinimide (HOSU) under the action of N, N-Dicyclohexylcarbodiimide (DCC), condensing the active ester with the L-glutamine to produce a dipeptide intermediate with a protecting group, and finally carrying out palladium catalytic hydrogenation to remove the protecting group to obtain a product (EP0311057B1,1994). The raw materials and the catalyst used in the method are expensive, the material cost is high, the used DCC is difficult to remove, the high-purity product is difficult to prepare, and the requirement of palladium-carbon hydrogenation reaction equipment is strict.
Secondly, phthalic anhydride and glycine are prepared into phthaloyl glycine in a molten state, then active ester or mixed anhydride method is adopted for condensation, and hydrazinolysis is carried out to obtain a final product (CN200410066112, 2004). The method has the problems of complicated process route and high production cost, and is difficult to treat by using triethylamine, hydrazine hydrate and various organic compounds, thus being not beneficial to environmental protection.
Third, WO2012014809(a1) discloses a novel synthesis method of dipeptide amino acids, which comprises the synthesis of Gly-Gln, mixing L-glutamine and tetrabutyl hydrogen phosphine oxide solution, heating under reduced pressure to evaporate water, performing dehydration condensation to obtain L-glutamine ionic liquid (L-Gln-TBP), and adding glycine methyl ester hydrochloride into L-Gln-TBP to form Gly-Gln. The method has simple process, avoids using various organic reagents, but is immature, has low yield and cannot realize industrial production.
Fourthly, chloroacetyl chloride is used as a starting material to react with L-glutamine for acylation reaction to prepare a crude product of the N-chloroacetyl-L-glutamine, the intermediate is refined, and ammonia water and ammonium carbonate are used as ammonolysis agents to react to prepare the product (EP0678501, 1997). In the first step of preparing the chloroacetyl-L-glutamine, the crystalline solid particles are fine, so that the crystalline system is viscous, water is difficult to filter, and the drying is easy to melt; and in the second step, ammonolysis reaction is carried out in ammonia water, the ammonia activity is low, the reaction time is long, the generation of hydrolysis impurity glycyl-L-glutamic acid is difficult to control, a large amount of ammonium salt is introduced, and the recovery energy consumption is high. Although the raw materials are cheap and easy to obtain, the method has the advantages of low total yield of the process, more industrial three wastes, complicated production procedures and longer production period.
Disclosure of Invention
The invention aims to provide an industrial preparation method of glycyl-L-glutamine, which has the advantages of cheap and easily obtained raw materials, simple process operation, single type of used organic solvent, easy recovery and reutilization, environmental protection, high purity of obtained products, capability of meeting the requirements of injection-grade raw materials and suitability for industrial production.
In order to achieve the purpose, the invention adopts the technical scheme that:
an industrialized preparation method of glycyl-L-glutamine comprises the steps of firstly carrying out acylation reaction on chloroacetyl chloride and L-glutamine under low temperature and alkaline conditions, carrying out phase splitting to obtain an N-chloroacetyl-L-glutamine aqueous solution, then treating the N-chloroacetyl-L-glutamine aqueous solution by adopting an electrodialysis membrane separation technology, introducing ammonia gas into the treated N-chloroacetyl-L-glutamine aqueous solution to carry out ammonolysis under pressure, and concentrating, crystallizing, filtering and drying to obtain a glycyl-L-glutamine crude product; finally dissolving the crude glycyl-L-glutamine in water, purifying by WA-30 resin, and crystallizing to obtain high-purity glycyl-L-glutamine, wherein the specific reaction formula is as follows:
the technical scheme of the invention is further improved in that the method comprises the following steps:
(1) preparation of a glycyl-L-glutamine crude product: adding L-glutamine into a mixed solvent of purified water and toluene, simultaneously dropwise adding a toluene solution of chloroacetyl chloride and alkaline water under low-temperature stirring, controlling the pH of a reaction mixture in the dropwise adding process, continuously stirring for acylation reaction after dropwise adding is finished, standing for 0.5h after the reaction is finished, carrying out phase separation, adjusting the pH of an obtained water phase with acid, carrying out electrodialysis treatment, and controlling the conductivity of fresh water to be lower than 10ms/cm as an end point to obtain an N-chloroacetyl-L-glutamine aqueous solution; adding an organic solvent into the N-chloroacetyl-L-glutamine aqueous solution, introducing ammonia gas, carrying out ammonolysis reaction for 2-3 h under pressure at a certain temperature, concentrating under reduced pressure until no moisture is evaporated after the reaction is finished, adding purified water into the residue, heating to 55 ℃, stirring completely, slowly adding the organic solvent in a flowing manner, continuing to keep the temperature and stirring for 0.5h after the adding in the flowing manner is finished, cooling to 20-25 ℃, crystallizing, keeping the temperature and stirring for 3-4 h, centrifuging, filtering and washing after the crystallizing reaction is finished to obtain a wet product, and carrying out vacuum drying on the obtained wet product to obtain a glycyl-L-glutamine crude product;
(2) refining a glycyl-L-glutamine crude product: dissolving the glycyl-L-glutamine crude product obtained in the step (1) in purified water, adding WA-30 resin after dissolving, stirring at room temperature for 2-3 h, filtering and recovering the resin, concentrating the obtained filtrate under reduced pressure to be dry, adding purified water to the remainder for dissolving, performing microfiltration after completely dissolving, adding purified water to wash a container and a pipeline after filtering, combining the washing liquid with the filtrate after microfiltration of washing liquid, stirring and heating to 40-45 ℃, adding an organic solvent in a flowing manner, stirring at a constant temperature for 20min after the end of the flowing manner, cooling to 20-25 ℃, performing crystallization reaction for 1h under constant temperature and stirring, centrifuging, filtering and washing after the completion of the crystallization reaction to obtain a wet product, and drying the wet product in vacuum to obtain the glycyl-L-glutamine with high purity.
The technical scheme of the invention is further improved as follows: the molar ratio of the L-glutamine to the chloroacetyl chloride in the step (1) is 1: 1.05-1: 1.50.
The technical scheme of the invention is further improved as follows: the alkaline water in the step (1) is NaOH aqueous solution, the temperature is controlled to be 8-12 ℃ when the toluene solution of chloroacetyl chloride and the alkaline water are dripped, and the pH of the reaction mixture in the dripping process is controlled to be 10.0-11.0; the temperature of the acylation reaction is 0-5 ℃, and the time is 0.5-1 h.
The technical scheme of the invention is further improved as follows: the acid used for adjusting the pH in the step (1) is hydrochloric acid or sulfuric acid, and the concentration of the acid solution is 0.5-3 mol/L; the membrane used in the electrodialysis treatment is a homogeneous phase ion exchange membrane, and the pH value of the fresh water is controlled to be 7.5-8.0.
The technical scheme of the invention is further improved as follows: the organic solvent added into the N-chloroacetyl-L-glutamine aqueous solution in the step (1) is one of dimethyl sulfoxide, N-dimethylformamide, N-dimethyl-2-imidazolidinone and N-methylpyrrolidone, and the amount of the organic solvent is 1/20 of the weight of the N-chloroacetyl-L-glutamine aqueous solution.
The technical scheme of the invention is further improved as follows: the ammonolysis reaction pressure in the step (1) is 0.6-0.8 Mpa, and the reaction temperature is 38-42 ℃.
The technical scheme of the invention is further improved as follows: in the step (1), purified water with the mass 1.5-2.0 times of that of the remainder is added into the remainder, the crystallization organic solvent is methanol, and the volume ratio of the purified water to the methanol is 1: 2.
The technical scheme of the invention is further improved as follows: the amount of purified water for dissolving the glycyl-L-glutamine crude product in the step (2) is 10 times of the mass of the glycyl-L-glutamine crude product; the dosage of the WA-30 resin is 3.0-4.0 times of the mass of the glycyl-L-glutamine crude product.
The technical scheme of the invention is further improved as follows: the total consumption of the purified water and the purified water washing liquid of the dissolving remainder in the step (2) is 3 times of the mass of the crude glycyl-L-glutamine product; the crystallization organic solvent is methanol, and the ratio of the amount of the methanol to the total volume of the purified water is 2: 1.
Due to the adoption of the technical scheme, the invention has the following technical effects:
the preparation method adopts an electrodialysis membrane separation technology, improves the yield and quality of the product by combining with WA-30 resin purification, simplifies the process steps, realizes the preparation of the dipeptide crude product by a one-pot method, generates less industrial three wastes, has simple and convenient process operation, is suitable for large-scale industrial production, and can prepare the glycyl-L-glutamine product with high quality and meet the requirements of injection-grade raw materials.
The main raw materials used in the invention, such as chloracetyl chloride, L-glutamine and the like, have low price and are easy to be obtained commercially.
The method adopts an electrodialysis membrane separation technology to prepare the N-chloroacetyl-L-glutamine aqueous solution with higher purity, avoids the problems that the solid N-chloroacetyl-L-glutamine prepared by a crystallization method is difficult to filter, dry and easy to melt and the like, reduces material loss caused by intermediate preparation by simplified treatment operation, and has higher total yield, lower production cost and environmental friendliness.
According to the invention, ammonia gas is introduced for ammonolysis under pressure, so that the reaction efficiency is improved, and the generation of a large amount of ammonium salt and waste liquid is reduced compared with the ammonia ammonolysis method; an aprotic polar solvent is introduced, so that the ammonolysis reaction activity is improved, the generation of a hydrolyzed impurity glycyl-L-glutamic acid can be effectively controlled, the subsequent purification difficulty is reduced, and the quality of a crude product is improved.
Detailed Description
The present invention will be described in further detail with reference to specific examples below:
an industrial preparation method of glycyl-L-glutamine comprises the following steps:
(1) preparation of glycyl-L-glutamine crude product
Adding L-glutamine into a mixed solvent of purified water and toluene, dropwise adding a toluene solution of chloroacetyl chloride and alkaline water at the same time under low-temperature stirring, wherein the molar ratio of the L-glutamine to the chloroacetyl chloride is 1: 1.05-1: 1.50, the alkaline water is a NaOH aqueous solution, the pH of the reaction mixture is controlled to be 10.0-11.0 in the dropwise adding process, the temperature is 8-12 ℃, and after the dropwise adding is finished, cooling to 0-5 ℃ and continuously stirring for acylation reaction for 0.5-1 h. And after the reaction is finished and the solution is kept stand for 0.5h, phase separation is carried out, the pH of the obtained water phase is adjusted to 7.5-8.0 by using 0.5-3 mol/L hydrochloric acid or sulfuric acid, and then electrodialysis treatment is carried out, wherein the membrane used in the electrodialysis treatment is a homogeneous phase ion exchange membrane, the pH of fresh water is controlled to be 7.5-8.0, and the conductivity of the fresh water is controlled to be lower than 10ms/cm to be used as an end point, so that the N-chloroacetyl-L-glutamine aqueous solution is obtained.
And adding one of dimethyl sulfoxide, N-dimethylformamide, N-dimethyl-2-imidazolidinone and N-methylpyrrolidone into the N-chloroacetyl-L-glutamine aqueous solution obtained through electrodialysis treatment, introducing ammonia gas, and carrying out pressure ammonolysis reaction for 2-3 hours at 38-42 ℃, wherein the pressure of the ammonolysis reaction is 0.6-0.8 MPa. Concentrating under reduced pressure (the water bath temperature is 60-65 ℃, the vacuum degree is less than or equal to-0.09 Mpa) after the reaction is finished until no moisture is evaporated, adding purified water with the mass of 1.5-2.0 times of that of the residue into the residue, heating to 55 ℃, stirring to be completely dissolved, slowly adding methanol in a flowing manner, wherein the volume ratio of the methanol to the purified water is 2:1, continuously stirring for 0.5h in a heat preservation manner after the feeding is finished, cooling to 20-25 ℃ for crystallization, stirring for 3-4 h in a heat preservation manner, centrifuging, filtering and washing after the crystallization reaction is finished to obtain a wet product, and drying the obtained wet product in vacuum at 60-65 ℃ to obtain a glycyl-L-glutamine crude product.
(2) Refining of glycyl-L-glutamine crude product
Dissolving the glycyl-L-glutamine crude product obtained in the step (1) in purified water with the mass 10 times of that of the glycyl-L-glutamine crude product, adding WA-30 resin with the mass 3.0-4.0 times of that of the glycyl-L-glutamine crude product after the dissolution is clear, stirring at room temperature for 2-3 h, filtering and recovering the resin, concentrating the obtained filtrate under reduced pressure (the water bath temperature is 60-65 ℃, the vacuum degree is less than or equal to-0.09 Mpa) to dryness, adding the purified water into the remainder for dissolution, carrying out microfiltration after the complete dissolution is finished, adding purified water washing liquid for washing a container and a pipeline, mixing the purified water and the purified water washing liquid with the total amount being 3 times of the mass of the glycyl-L-glutamine crude product, mixing the washing liquid with the filtrate after microfiltration, stirring and heating to 40-45 ℃, adding methanol with the total volume ratio of the methanol to the purified water being 2:1, carrying out heat preservation stirring for 20, and (3) carrying out heat preservation and stirring for crystallization reaction for 1h, centrifuging, filtering and washing after the crystallization reaction is finished to obtain a wet product, and carrying out vacuum drying on the wet product at the temperature of 60-65 ℃ until the moisture is 7.0-10.0% to obtain the high-purity glycyl-L-glutamine.
Example 1
(1) Preparation of glycyl-L-glutamine crude product
Adding 7.5kg of purified water and 1.9kg of toluene into a reaction kettle, adding 3.0 kgL-glutamine under the stirring condition, cooling to 10 ℃, dropwise adding a prepared sodium hydroxide solution (5mol/L, about 1.3L) under the stirring condition, keeping the temperature at 8-12 ℃, dropwise adding a mixed solution of 2.48kg of chloroacetyl chloride and 1.6kg of toluene, and dropwise adding a sodium hydroxide solution (5mol/L) at the same time, wherein the pH of the reaction mixture is controlled to be 10.0-11.0. After the dropwise addition, cooling to 0-5 ℃, continuously stirring for 0.5h, standing for 0.5h, carrying out phase separation, recovering toluene, adjusting the pH of a water phase to 7.5-8.0 by using 0.5mol/L hydrochloric acid, adding the water phase into a fresh water chamber of an electrodialysis device, adding equal volume water into a concentrated water chamber, using 100 pairs of homogeneous phase ion exchange membranes, controlling the voltage to be not more than 100V, controlling the pH of the fresh water to be 7.5-8.0, and controlling the conductivity of the fresh water to be lower than 10ms/cm as an end point to obtain the N-chloroacetyl-L-glutamine aqueous solution in the fresh water chamber.
Adding the N-chloroacetyl-L-glutamine aqueous solution (23.5kg) prepared in the previous step into a high-pressure reaction kettle, adding 1.18kg of N, N-dimethylformamide, introducing ammonia gas until the pressure is 0.6-0.8 Mpa, slowly heating to 38-42 ℃, keeping the temperature and pressure and stirring for 2h, concentrating under reduced pressure (vacuum is less than or equal to-0.09 Mpa), heating in a water bath at the temperature of 60-65 ℃, concentrating until no moisture is evaporated, adding 6L of purified water, heating to 55 ℃, stirring until the purified water is completely dissolved, slowly adding 12L of methanol, continuously stirring for 0.5h, cooling to 20-25 ℃, precipitating a large amount of crystals, keeping the temperature and stirring for 3h, centrifugally filtering, washing a filter cake twice with a small amount of methanol, and drying under vacuum at the temperature of 60-65 ℃ to obtain 3.1kg of glycyl-L-glutamine crude product, wherein the molar yield is 74% and.
(2) Refining of glycyl-L-glutamine crude product
Dissolving 3.0kg of glycyl-L-glutamyl crude product in 30L of purified water, adding 10.0kg of WA-30 resin, stirring for 2h at room temperature, filtering and recovering the resin, concentrating the filtrate under reduced pressure (vacuum is less than or equal to-0.09 Mpa), concentrating at the water bath temperature of 60-65 ℃, adding 8L of purified water, completely dissolving, filtering through a 0.22 mu m microporous filter membrane, washing the tank wall and a pipeline filter membrane with 1L of purified water, combining the filtrate and the washing solution, adding into a crystallization tank, stirring and heating to 40-45 ℃, slowly adding 18L of methanol, stirring for 20min at a constant temperature, cooling to 20-25 ℃, stirring for 1h at a constant temperature, centrifuging and filtering, washing the filter cake twice with a small amount of methanol, and drying at the temperature of 60-65 ℃ under vacuum to the moisture content of 7.0-10.0% to obtain 2.59kg of refined glycyl-L-glutamine product with the molar yield of 86..
Example 2
(1) Preparation of glycyl-L-glutamine crude product
Adding 6.25kg of purified water and 1.6kg of toluene into a reaction kettle, adding 2.5 kgL-glutamine under stirring, cooling to 9 ℃, dropwise adding a prepared sodium hydroxide solution (5mol/L, about 1.1L) under stirring, keeping the temperature at 8-12 ℃, dropwise adding a mixed solution of 2.89kg of chloroacetyl chloride and 1.9kg of toluene, and dropwise adding a sodium hydroxide solution (5mol/L) at the same time, wherein the pH of the reaction mixture is controlled to be 10.0-11.0. After the dropwise addition, cooling to 0-5 ℃, continuously stirring for 0.5h, standing for 0.5h, carrying out phase separation, recovering toluene, adjusting the pH of a water phase to 7.5-8.0 by using 0.5mol/L sulfuric acid, adding the water phase into a fresh water chamber of an electrodialysis device, adding equal volume water into a concentrated water chamber, using 100 pairs of homogeneous phase ion exchange membranes, controlling the voltage to be not more than 100V, controlling the pH of the fresh water to be 7.5-8.0, and controlling the conductivity of the fresh water to be lower than 10ms/cm as an end point to obtain the N-chloroacetyl-L-glutamine aqueous solution in the fresh water chamber.
Adding the N-chloroacetyl-L-glutamine aqueous solution (19.2kg) prepared in the previous step and 0.96kg of dimethyl sulfoxide into a high-pressure reaction kettle, introducing ammonia gas to the pressure of 0.6-0.8 Mpa, slowly heating to 38-42 ℃, keeping the temperature and pressure and stirring for 2h, concentrating under reduced pressure (vacuum is less than or equal to-0.09 Mpa), heating the water bath to 60-65 ℃, concentrating until no moisture is evaporated, adding 5L of purified water, heating to 55 ℃, stirring until the purified water is completely dissolved, slowly adding 10L of methanol, continuously stirring for 0.5h, cooling to 20-25 ℃, precipitating a large amount of crystals, keeping the temperature and stirring for 3h, carrying out centrifugal filtration, washing a filter cake twice with a small amount of methanol, and carrying out vacuum drying at the temperature of 60-65 ℃ to obtain 2.46kg of glycyl-L-glutamine crude product, wherein the molar yield is 70.8% and.
(2) Refining of glycyl-L-glutamine crude product
Dissolving 2.4kg of glycyl-L-glutamyl crude product in 24L of purified water, adding 9.6kg of WA-30 resin, stirring for 3h at room temperature, filtering and recovering the resin, concentrating the filtrate under reduced pressure (vacuum is less than or equal to-0.09 Mpa), concentrating at the water bath temperature of 60-65 ℃, adding 6.5L of purified water, completely dissolving, filtering through a 0.22 mu m microporous filter membrane, washing the wall of a tank and a pipeline filter membrane with 0.7L of purified water, then mixing the filtrate and a washing solution, adding into a crystallization tank, stirring and heating to 40-45 ℃, slowly adding 14.4L of methanol, stirring for 20min at a constant temperature, cooling to 20-25 ℃, stirring for 1h at a constant temperature, centrifuging and filtering, washing the filter cake twice with a small amount of methanol, and drying at the temperature of 60-65 ℃ in vacuum until the water content is 7.0-10.0%, thus obtaining 1.98kg of glycyl-L-glutamine refined product with the molar yield.
Example 3
(1) Preparation of glycyl-L-glutamine crude product
Adding 7.5kg of purified water and 1.9kg of toluene into a reaction kettle, adding 3.0 kgL-glutamine under stirring, cooling to 10 ℃, dropwise adding a prepared sodium hydroxide solution (5mol/L, about 1.3L) under stirring, keeping the temperature at 8-12 ℃, dropwise adding a mixed solution of 2.9kg of chloroacetyl chloride and 1.87kg of toluene, and dropwise adding a sodium hydroxide solution (5mol/L) at the same time, wherein the pH of the reaction mixture is controlled to be 10.0-11.0. After the dropwise addition, cooling to 0-5 ℃, continuously stirring for 1h, standing for 0.5h, carrying out phase separation, recovering toluene, adjusting the pH of a water phase to 7.5-8.0 by using 3mol/L hydrochloric acid, adding the water phase into a fresh water chamber of an electrodialysis device, adding equal volume water into a concentrated water chamber, using 100 pairs of homogeneous phase ion exchange membranes, controlling the voltage to be not more than 100V, controlling the pH of the fresh water to be 7.5-8.0, and controlling the conductivity of the fresh water to be lower than 10ms/cm as an end point to obtain the N-chloroacetyl-L-glutamine aqueous solution of the fresh water chamber.
Adding the N-chloroacetyl-L-glutamine aqueous solution (21.8kg) prepared in the previous step and 1.1kg of N-methylpyrrolidone into a high-pressure reaction kettle, introducing ammonia gas until the pressure is 0.6-0.8 Mpa, slowly heating to 38-42 ℃, keeping the temperature and pressure and stirring for 3h, concentrating under reduced pressure (vacuum is less than or equal to-0.09 Mpa), heating in a water bath at the temperature of 60-65 ℃, concentrating until no water is evaporated, adding 6L of purified water, heating to 55 ℃, stirring until the purified water is completely dissolved, slowly adding 12L of methanol, continuously stirring for 0.5h, cooling to 20-25 ℃, precipitating a large amount of crystals, keeping the temperature and stirring for 3h, carrying out centrifugal filtration, washing a filter cake twice with a small amount of methanol, and carrying out vacuum drying at the temperature of 60-65 ℃ to obtain 3.2kg of glycyl-L-glutamine crude product, wherein the molar yield is 76.7.
(2) Refining of glycyl-L-glutamine crude product
Dissolving 3.0kg of crude glycyl-L-glutamyl in 30L of purified water, adding 10.5kg of WA-30 resin, stirring for 2.5h at room temperature, filtering to recover the resin, concentrating the filtrate under reduced pressure (vacuum is less than or equal to-0.09 Mpa), heating the water bath to 60-65 ℃, concentrating to be nearly anhydrous, adding 8L of purified water, completely dissolving, filtering through a 0.22 mu m microporous filter membrane, washing the tank wall and a pipeline filter membrane with 1L of purified water, mixing the filtrate and the washing solution, adding into a crystallization tank, stirring, heating to 40-45 ℃, slowly adding 18L of methanol, stirring for 20min at a constant temperature, cooling to 20-25 ℃, stirring for 1h at a constant temperature, centrifuging, washing the filter cake twice with a small amount of methanol, and drying at 60-65 ℃ under vacuum until the water content is 7.0-10.0%, thus obtaining 2.51kg of refined glycyl-L-glutamine with the molar yield of 83.7%.
In the Chinese pharmacopoeia 2020 edition, the glycyl-L-glutamine standard is as follows: calculated as anhydrate, contains C7H13N3O4Not less than 99.0%; the area of other unknown impurities must not be larger than 1.0% of the area of the main peak of the control solution. The content and the purity of the finished product prepared by the method meet the requirements of Chinese pharmacopoeia standards, and the specific quality conditions of the product of the embodiment are shown in the following table:
Claims (10)
1. an industrial preparation method of glycyl-L-glutamine is characterized in that: firstly, chloracetyl chloride and L-glutamine are subjected to acylation reaction under low temperature and alkaline conditions, an N-chloracetyl-L-glutamine aqueous solution is obtained after phase splitting, then the N-chloracetyl-L-glutamine aqueous solution is treated by adopting an electrodialysis membrane separation technology, ammonia gas is introduced into the treated N-chloracetyl-L-glutamine aqueous solution for ammonolysis under pressure, and a glycyl-L-glutamine crude product is obtained through concentration, crystallization, filtration and drying; finally dissolving the crude glycyl-L-glutamine in water, purifying by WA-30 resin, and crystallizing to obtain the high-purity glycyl-L-glutamine.
2. The industrial production method of glycyl-L-glutamine according to claim 1, characterized in that: the method comprises the following steps:
(1) preparation of a glycyl-L-glutamine crude product: adding L-glutamine into a mixed solvent of purified water and toluene, simultaneously dropwise adding a toluene solution of chloroacetyl chloride and alkaline water under low-temperature stirring, controlling the pH of a reaction mixture in the dropwise adding process, continuously stirring for acylation reaction after dropwise adding is finished, standing for 0.5h after the reaction is finished, carrying out phase separation, adjusting the pH of an obtained water phase with acid, carrying out electrodialysis treatment, and controlling the conductivity of fresh water to be lower than 10ms/cm as an end point to obtain an N-chloroacetyl-L-glutamine aqueous solution; adding an organic solvent into the N-chloroacetyl-L-glutamine aqueous solution, introducing ammonia gas, carrying out ammonolysis reaction for 2-3 h under pressure at a certain temperature, concentrating under reduced pressure until no moisture is evaporated after the reaction is finished, adding purified water into the residue, heating to 55 ℃, stirring completely, slowly adding the organic solvent in a flowing manner, continuing to keep the temperature and stirring for 0.5h after the adding in the flowing manner is finished, cooling to 20-25 ℃, crystallizing, keeping the temperature and stirring for 3-4 h, centrifuging, filtering and washing after the crystallizing reaction is finished to obtain a wet product, and carrying out vacuum drying on the obtained wet product to obtain a glycyl-L-glutamine crude product;
(2) refining a glycyl-L-glutamine crude product: dissolving the glycyl-L-glutamine crude product obtained in the step (1) in purified water, adding WA-30 resin after dissolving, stirring at room temperature for 2-3 h, filtering and recovering the resin, concentrating the obtained filtrate under reduced pressure to be dry, adding purified water to the remainder for dissolving, performing microfiltration after completely dissolving, adding purified water to wash a container and a pipeline after filtering, combining the washing liquid with the filtrate after microfiltration of washing liquid, stirring and heating to 40-45 ℃, adding an organic solvent in a flowing manner, stirring at a constant temperature for 20min after the end of the flowing manner, cooling to 20-25 ℃, performing crystallization reaction for 1h under constant temperature and stirring, centrifuging, filtering and washing after the completion of the crystallization reaction to obtain a wet product, and drying the wet product in vacuum to obtain the glycyl-L-glutamine with high purity.
3. The industrial production method of glycyl-L-glutamine according to claim 2, characterized in that: the molar ratio of the L-glutamine to the chloroacetyl chloride in the step (1) is 1: 1.05-1: 1.50.
4. The industrial production method of glycyl-L-glutamine according to claim 2, characterized in that: the alkaline water in the step (1) is NaOH aqueous solution, the temperature is controlled to be 8-12 ℃ when the toluene solution of chloroacetyl chloride and the alkaline water are dripped, and the pH of the reaction mixture in the dripping process is controlled to be 10.0-11.0; the temperature of the acylation reaction is 0.5-1 h.
5. The industrial production method of glycyl-L-glutamine according to claim 2, characterized in that: the acid used for adjusting the pH in the step (1) is hydrochloric acid or sulfuric acid, and the concentration of the acid solution is 0.5-3 mol/L; the membrane used in the electrodialysis treatment is a homogeneous phase ion exchange membrane, and the pH value of the fresh water is controlled to be 7.5-8.0.
6. The industrial production method of glycyl-L-glutamine according to claim 2, characterized in that: the organic solvent added into the N-chloroacetyl-L-glutamine aqueous solution in the step (1) is one of dimethyl sulfoxide, N-dimethylformamide, N-dimethyl-2-imidazolidinone and N-methylpyrrolidone, and the amount of the organic solvent is 1/20 of the weight of the N-chloroacetyl-L-glutamine aqueous solution.
7. The industrial production method of glycyl-L-glutamine according to claim 2, characterized in that: the ammonolysis reaction pressure in the step (1) is 0.6-0.8 Mpa, and the reaction temperature is 38-42 ℃.
8. The industrial production method of glycyl-L-glutamine according to claim 2, characterized in that: in the step (1), purified water with the mass 1.5-2.0 times of that of the remainder is added into the remainder, the crystallization organic solvent is methanol, and the volume ratio of the purified water to the methanol is 1: 2.
9. The industrial production method of glycyl-L-glutamine according to claim 2, characterized in that: the amount of purified water for dissolving the glycyl-L-glutamine crude product in the step (2) is 10 times of the mass of the glycyl-L-glutamine crude product; the dosage of the WA-30 resin is 3.0-4.0 times of the mass of the glycyl-L-glutamine crude product.
10. The industrial production method of glycyl-L-glutamine according to claim 2, characterized in that: the total consumption of the purified water and the purified water washing liquid of the dissolving remainder in the step (2) is 3 times of the mass of the crude glycyl-L-glutamine product; the crystallization organic solvent is methanol, and the ratio of the amount of the methanol to the total volume of the purified water is 2: 1.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5780677A (en) * | 1994-04-18 | 1998-07-14 | Kyowa Hakko Kogyo Co., Ltd. | Process for producing glutamine derivative |
| CN102993271A (en) * | 2012-12-13 | 2013-03-27 | 山东齐都药业有限公司 | Preparation method of glycyl-L-glutamine |
| CN103694313A (en) * | 2013-12-24 | 2014-04-02 | 济南诚汇双达化工有限公司 | Glycyl-L-glutamine preparation method |
| CN104529807A (en) * | 2014-12-13 | 2015-04-22 | 济南诚汇双达化工有限公司 | Preparation method of N-chloracetyl-L-glutamine |
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- 2020-12-08 CN CN202011441553.2A patent/CN112480208B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5780677A (en) * | 1994-04-18 | 1998-07-14 | Kyowa Hakko Kogyo Co., Ltd. | Process for producing glutamine derivative |
| CN102993271A (en) * | 2012-12-13 | 2013-03-27 | 山东齐都药业有限公司 | Preparation method of glycyl-L-glutamine |
| CN103694313A (en) * | 2013-12-24 | 2014-04-02 | 济南诚汇双达化工有限公司 | Glycyl-L-glutamine preparation method |
| CN104529807A (en) * | 2014-12-13 | 2015-04-22 | 济南诚汇双达化工有限公司 | Preparation method of N-chloracetyl-L-glutamine |
Non-Patent Citations (1)
| Title |
|---|
| 施立钦: "《化工中间体》", 20 November 2005, pages: 21 - 22 * |
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