CN106565821B - N (2) -L-alanyl-L-glutamine novel crystal form and preparation method thereof - Google Patents
N (2) -L-alanyl-L-glutamine novel crystal form and preparation method thereof Download PDFInfo
- Publication number
- CN106565821B CN106565821B CN201610861653.8A CN201610861653A CN106565821B CN 106565821 B CN106565821 B CN 106565821B CN 201610861653 A CN201610861653 A CN 201610861653A CN 106565821 B CN106565821 B CN 106565821B
- Authority
- CN
- China
- Prior art keywords
- alanyl
- glutamine
- temperature
- sterile
- filtering
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- 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
- C07K5/06—Dipeptides
- 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 belongs to the technical field of medicines, and particularly relates to a novel N (2) -L-alanyl-L-glutamine crystal form and a preparation method thereof, which uses Cu-Ka radiation and X-ray powder diffraction expressed by 2 theta angle, has characteristic peaks at 13.76,15.58,18.58,18.72,19.66,20.72, 21.34, 21.42,21.70,22.52,23.28,23.34,24.12,24.86,25.54,26.12,27.66,30.46,30.64,32.56,33.10,34.26,34.62,34.86,36.60,36.90 and 37.48 (+ -0.1 degree), and the novel N (2) -L-alanyl-L-glutamine crystal form obtained by the refining method has good crystal form, higher bulk density, uniform particle size distribution, difficult moisture absorption and convenient storage, and the crystal has high purity, small organic residue and good clarity, is more beneficial to the subsequent processing of products, and simultaneously improves the safety of the N (2) -L-alanyl-L-glutamine for injection.
Description
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to a novel N (2) -L-alanyl-L-glutamine crystal form and a preparation method thereof.
Background
The molecular formula of the N (2) -L-alanyl-L-glutamine is C8H15N3O4, the molecular weight is 217.22, and the structural formula is as follows:
n (2) -L-alanyl-L-glutamine is a component of parenteral nutrition and is suitable for patients in need of glutamine supplementation, including patients in catabolic and hypermetabolic states. Such as: trauma, burns, post-major and minor surgery, bone marrow and other organ transplantation, gastrointestinal syndrome, tumors, severe infections and other ICU patients in a stressful state.
Professor PeterFurst in Germany in the 80 th 20 th century proposed the concept of a dipeptide that is an ideal substitute for human glutamine by synthetic dipeptides as a donor for glutamine, and was developed as a parenteral nutrition in 1995. Because it can be dissolved naturally in water and can endure 121 ℃ heat sterilization without decomposition; thereby effectively overcoming the use defect of the glutamine in the parenteral nutrition. The glutamine dipeptide inputted into vein is hydrolyzed into alanine and glutamine rapidly under the action of glutamyl dipeptidase, and is fully absorbed and utilized. Therefore, the medicine has the functions of promoting muscle protein synthesis and improving clinical and biochemical indexes of critically ill patients; maintaining intestinal function, promoting tissue composition, and maintaining nitrogen balance; enhancing immune system, etc., and is widely used in various countries in the world at present.
The patent CN201110384393.7 provides a high-purity alanylglutamine compound with the following structural formula and a preparation method thereof, wherein the preparation method comprises the following steps of 1, taking N- (α -chlorine) -propionyl-glutamine as a raw material, selecting hydrazine substances as ammoniation reagents, carrying out reaction under normal pressure to prepare alanylglutamine crude products, 2, adding absolute methanol into the alanylglutamine crude products at the temperature of 0-room temperature, stirring, carrying out suction filtration to remove methanol solution, obtaining filter cakes containing alanylglutamine, 3, dissolving the filter cakes containing alanylglutamine into water, heating the obtained water solution to be not higher than 80 ℃, keeping the temperature for certain time for concentration, then adding ethanol, leading the volume ratio of the water solution to the ethanol to be 1: 1-3, carrying out gradient temperature reduction, carrying out recrystallization, obtaining purified alanylglutamine, obtaining a target product with extremely high purity by the method, further optimizing the quality of a preparation product, reducing toxic and side effects, ensuring the safety of clinical medication, and proving that the temperature reduction is carried out after the experiment, the gradient storage of the prepared alanylglutamine has the defects that the crystal preparation method has high hygroscopicity and the low possibility of organic substance content of L-glutamine crystal, and the increase caused by high-purity increase and the following high-purity increase rate increase.
The present invention has been made in view of this situation.
Disclosure of Invention
The technical problem to be solved by the invention is to overcome the defects of the prior art and provide a novel N (2) -L-alanyl-L-glutamine crystal form and a preparation method thereof, wherein the crystal form is good, the bulk density is higher, the particle size distribution is uniform, the crystal purity is high, the organic residue is small, the clarity is good, and the subsequent processing of the product is more facilitated; the hygroscopicity is poor and the storage is convenient.
In order to solve the technical problems, the invention adopts the technical scheme that:
a novel crystal form of N (2) -L-alanyl-L-glutamine has characteristic peaks at 13.76,15.58,18.58,18.72,19.66,20.72, 21.34, 21.42,21.70,22.52,23.28,23.34,24.12,24.86,25.54,26.12,27.66,30.46,30.64,32.56,33.10,34.26,34.62,34.86,36.60,36.90 and 37.48(± 0.1 degree) in X-ray powder diffraction pattern shown in figure 1 when the crystal form is expressed in 2 theta by using Cu-Ka radiation.
In X-ray powder diffraction patterns, the diffraction pattern obtained from a crystalline compound is often characteristic for a particular crystalline form, where the relative intensities of the bands (especially at low angles) may vary due to the dominant orientation effects resulting from differences in crystallization conditions, particle size, and other measurement conditions. Therefore, the relative intensities of the diffraction peaks are not characteristic of the crystal form in question, and when judging whether the diffraction peaks are the same as the known crystal form, the relative positions of the peaks rather than their relative intensities should be noted. The peak position is usually expressed in terms of 2 θ angle in the X-ray powder diffraction pattern.
The crystal form prepared by the invention has a powder X-ray diffraction pattern shown in figure 1.
The method for preparing the novel crystal form of the N (2) -L-alanyl-L-glutamine comprises the following steps:
dissolving in step (1): adding the N (2) -L-alanyl-L-glutamine crude product into water for injection at a certain temperature, and stirring at constant temperature until the N (2) -L-alanyl-L-glutamine crude product is completely dissolved;
decoloring and adsorbing: keeping the temperature unchanged, adding activated carbon into the solution prepared in the step (1), and stirring for a period of time to obtain a decolorized adsorption solution;
filtering in step (3): keeping the temperature unchanged, and filtering the decolorized adsorption solution prepared in the step (2) to obtain sterile fine filtration solution;
and (4) crystallizing: placing the sterile fine filtrate in a crystallization device, taking the filtration temperature as the initial temperature, and adding sterile ethanol into the sterile fine filtrate obtained in the step (3) in batches while performing gradient cooling to crystallize N (2) -L-alanyl-L-glutamine to obtain N (2) -L-alanyl-L-glutamine crystal liquid;
and (5) filtering, washing and drying the crystal liquid under the aseptic condition to obtain dry aseptic N (2) -L-alanyl-L-glutamine crystals.
The method comprises the steps of adding ethanol in batches, and controlling the adding amount and the method of the ethanol, and aims to strictly control the proportion of the ethanol and the water for injection at different temperatures, so that the new crystal form can be prepared under the simultaneous action of the ethanol and the water for injection, the particle size distribution of the new crystal form is more uniform, and the bulk density is higher; the residual quantity of the organic solvent in the crystal is small, and the clarity is good; the new crystal form has poor hygroscopicity and is convenient to store; under the conditions of high temperature, illumination and high humidity, the related substances are increased little, the product content is reduced little, the property is more stable, and the use is safer.
The weight ratio of the N (2) -L-alanyl-L-glutamine crude product to the water for injection in the step (1) is 1Kg: 2 to 2.5 Kg.
When the novel N (2) -L-alanyl-L-glutamine crystal form to be protected is prepared, the quality of injection water is strictly controlled, the ratio of the injection water to ethanol in the crystallization process can be changed to influence the crystal form due to different quality of the injection water, and experiments show that only when the weight ratio of the crude N (2) -L-alanyl-L-glutamine product to the weight of the injection water is 1Kg: when the crystal is 2-2.5 Kg, the novel crystal form of N (2) -L-alanyl-L-glutamine can be prepared by combining the gradient cooling process to be protected.
The volume ratio of the weight of the N (2) -L-alanyl-L-glutamine crude product to the sterile ethanol in the step (4) is 1kg: 8-15L,
preferably, the volume ratio of the weight of the crude N (2) -L-alanyl-L-glutamine to the sterile ethanol is 1kg: 10L.
The N (2) -L-alanyl-L-glutamine in the solution is ensured to be completely precipitated by strictly controlling the adding volume of the ethanol, and the crystal obtained by the proportioning relation of the solvent has high purity and small organic residue.
The gradient cooling process in the step (4), the addition amount of the sterile ethanol and the addition mode are as follows:
the first stage is as follows: pumping the sterile fine filtrate into a crystallization device, then starting to cool, uniformly cooling to 45-50 ℃ at the speed of 7-9 ℃/h by taking the filtration temperature as the starting temperature, and uniformly dropwise adding sterile ethanol in the cooling process, wherein the volume of the added sterile ethanol is 1/3-1/2 of the total volume of the sterile ethanol;
and a second stage: uniformly reducing the temperature from 45-50 ℃ to 35-40 ℃ at the speed of 2-4 ℃/h, uniformly dropwise adding sterile ethanol in the temperature reduction process, wherein the volume of the added sterile ethanol is 1/4-1/3 of the total volume of the sterile ethanol;
and a third stage: and after the second stage is finished, adding the residual sterile ethanol into the crystallization device at one time, uniformly reducing the temperature from 35-40 ℃ to 25-30 ℃ at the speed of 7-9 ℃/h, and then stirring at constant temperature for 2-3 hours.
The temperature reduction process, the addition amount and the addition mode of the sterile ethanol are strictly controlled, the formation process of the new crystal form can be effectively controlled, the crystal form has high bulk crystallinity, uniform particle size distribution and high purity, and the new crystal form prepared by the method has better clarity and meets the requirement of N (2) -L-alanyl-L-glutamine for injection.
In the step (1), the water for injection is heated to 70-80 ℃, preferably 73-75 ℃.
Under the control of the temperature, the N (2) -L-alanyl-L-glutamine can be dissolved faster, the dissolution of impurities in the N (2) -L-alanyl-L-glutamine is limited, and the subsequent impurity removal by activated carbon is facilitated.
The weight of the added active carbon in the step (2) is 0.5-2% of the weight of the crude product of the N (2) -L-alanyl-L-glutamine,
preferably, the added weight of activated carbon is 1% of the weight of the crude N (2) -L-alanyl-L-glutamine.
The adding amount of the active carbon is strictly controlled within the range, so that the impurities in the solution can be effectively removed, and the loss of N (2) -L-alanyl-L-glutamine in the solution can be reduced.
The filtration in the step (3) is divided into three steps: firstly, decarburizing a decolored adsorption solution by using a titanium rod to obtain a crude filtrate; secondly filtering the coarse filtrate by a cylinder filter, and further removing active carbon to obtain fine filtrate; and finally, sterilizing and filtering the fine filtrate by a sterilizing filter to obtain the sterile fine filtrate.
Through twice filtration, the residue of the activated carbon is reduced, after the impurities of the activated carbon are removed, the sterilization operation is carried out while the solution is hot, and when the sterilized solution is crystallized, the obtained new crystal form has better crystal form, better bulk density and less organic residue.
And (5) performing drying operation by firstly adopting compressed air with certain pressure to pre-blow the material, closing the compressed air after blowing for a period of time, drying by using a vacuum system of a filtering, washing and drying machine and an interlayer circulating water system, setting the temperature of circulating water to be 50-60 ℃, starting timing when the temperature of the material reaches the preset temperature, and keeping the temperature for drying for 10-15 hours.
By means of pre-drying and drying, organic residues can be effectively reduced, drying time is shortened, and the obtained crystals are better in clarity.
The pressure of the compressed air is 0.5-1.0MPa, and the air blowing time is 0.5-2.0 hours;
preferably, the compressed air pressure is 0.8 MPa; the air-blowing time was 1.0 hour.
After adopting the technical scheme, compared with the prior art, the invention has the following beneficial effects: the N (2) -L-alanyl-L-glutamine obtained by refining by the method has the advantages of good crystal form, higher bulk density, uniform particle size distribution, high crystal purity, small organic residue and good clarity, is more favorable for subsequent processing of products, and simultaneously improves the safety of the N (2) -L-alanyl-L-glutamine for injection.
The following describes embodiments of the present invention in further detail with reference to the accompanying drawings.
Drawings
The accompanying drawings, which are included to provide a further understanding of the invention, are incorporated in and constitute a part of this specification, illustrate embodiments of the invention and together with the description serve to explain the invention without limiting the invention to the right. It is obvious that the drawings in the following description are only some embodiments, and that for a person skilled in the art, other drawings can be derived from them without inventive effort. In the drawings:
FIG. 1 is the X-ray diffraction spectrum of the new crystal form of N (2) -L-alanyl-L-glutamine prepared by the invention.
It should be noted that the drawings and the description are not intended to limit the scope of the inventive concept in any way, but to illustrate it by a person skilled in the art with reference to specific embodiments.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments will be clearly and completely described below with reference to the drawings in the embodiments of the present invention, and the following embodiments are used for illustrating the present invention and are not intended to limit the scope of the present invention.
The instruments and devices used in the following experiments were sterilized to ensure the sterility of the instruments and devices, and the crude N (2) -L-alanyl-L-glutamine used in the present invention was crude alanyl-glutamine prepared according to the method of example 6 in patent CN201110384393.7, and its alanyl-glutamine content was 88.80% by HPLC.
The first embodiment is as follows:
the preparation method of the novel N (2) -L-alanyl-L-glutamine crystal form comprises the following steps:
dissolving in step (1): adding 162.5kg of water for injection into a cleaned dosing tank, starting the dosing tank to stir, opening a heating system, heating the water for injection to 73-75 ℃, keeping the temperature unchanged, adding 65kg of crude N (2) -L-alanyl-L-glutamine into the water for injection, and stirring to completely dissolve the crude N (2) -L-alanyl-L-glutamine;
decoloring and adsorbing: keeping the temperature unchanged, adding 0.65kg of activated carbon into the solution prepared in the step (1), and stirring for 20 minutes to obtain a decolorized adsorption solution;
(1) and (3) filtering: decarburizing the decolorized adsorption solution prepared in the step (2) by using a 3-micron titanium rod to obtain a crude filtrate; then, the coarse filtrate is filtered for the second time through a cylinder filter with the filtering holes of 0.45 mu m, and the active carbon is further removed to obtain fine filtrate; finally, the refined filtrate is subjected to sterilization filtration through a sterilization filter with the diameter of 0.22 mu m to obtain sterile refined filtrate;
(2) and (3) crystallization: conveying the sterile fine filtrate into a crystallizing tank, preparing 650.0L of sterile ethanol, starting to cool, uniformly cooling to 50 ℃ at the speed of 8 ℃/hour, uniformly dropwise adding the sterile ethanol in the cooling process, and adding the volume of the sterile ethanol to 325L; then, the temperature is uniformly reduced to 40 ℃ from 50 ℃ at the speed of 3 ℃/h, sterile ethanol is uniformly dripped in the temperature reduction process, and 216L of sterile ethanol is added; finally, adding the residual sterile ethanol into a crystallizing tank at one time, after the addition is finished, uniformly reducing the temperature from 40 ℃ to 30 ℃ at the speed of 8 ℃/h, and then stirring for 3 hours at constant temperature to obtain N (2) -L-alanyl-L-glutamine crystal liquid;
opening a bottom valve of a crystallization tank, opening compressed air of the crystallization tank, pressing crystallization liquid in the tank into a filtering, washing and drying machine, filtering to obtain a filter cake, washing the filter cake for three times by using sterile ethanol, opening the filtering, washing and drying machine to pre-dry materials by using the compressed air, adjusting the compressed air pressure to be 0.8MPa, wherein the air blowing time is 1.0 hour, closing the compressed air after the air blowing is finished, opening a vacuum system and an interlayer circulating water system of the filtering, washing and drying machine, setting the circulating water temperature to be 60 ℃, continuing drying for 15 hours after the temperature of the materials reaches 60 ℃, and discharging to obtain 55.8kg of dried sterile N (2) -L-alanyl-L-glutamine crystals. The crystal form of purified N (2) -L-alanyl-L-glutamine was determined by powder X-ray diffraction method, and the results are shown in fig. 1, and X-ray powder diffraction using Cu-Ka radiation at 2 θ angles has characteristic peaks at 13.76,15.58,18.58,18.72,19.66,20.72, 21.34, 21.42,21.70,22.52,23.28,23.34,24.12,24.86,25.54,26.12,27.66,30.46,30.64,32.56,33.10,34.26,34.62,34.86,36.60,36.90,37.48(± 0.1 degrees).
The melting point is 215.5-215.8 ℃, and the purity is 99.98% by HPLC.
Example two: the preparation method of the novel N (2) -L-alanyl-L-glutamine crystal form comprises the following steps:
(1) dissolving: adding 162.5kg of water for injection into a cleaned dosing tank, starting the dosing tank to stir, opening a heating system, heating the water for injection to 73-75 ℃, keeping the temperature unchanged, adding 65kg of crude N (2) -L-alanyl-L-glutamine into the water for injection, and stirring to completely dissolve the crude N (2) -L-alanyl-L-glutamine;
(2) decoloring and adsorbing: keeping the temperature unchanged, adding 0.65kg of activated carbon into the solution prepared in the step (1), and stirring for 20 minutes to obtain a decolorized adsorption solution;
(3) and (3) filtering: decarburizing the decolorized adsorption solution prepared in the step (2) by using a 3-micron titanium rod to obtain a crude filtrate; then, the coarse filtrate is filtered for the second time through a cylinder filter with the filtering holes of 0.45 mu m, and the active carbon is further removed to obtain fine filtrate; finally, the refined filtrate is subjected to sterilization filtration through a sterilization filter with the diameter of 0.22 mu m to obtain sterile refined filtrate;
(4) and (3) crystallization: conveying the sterile fine filtrate into a crystallizing tank, preparing 650.0L of sterile ethanol, starting to cool, uniformly cooling to 50 ℃ at the speed of 8 ℃/hour, uniformly dropwise adding the sterile ethanol in the cooling process, and adding the volume of the sterile ethanol to 216L; then, the temperature is uniformly reduced to 40 ℃ from 50 ℃ at the speed of 3 ℃/h, sterile ethanol is uniformly dripped in the temperature reduction process, and 162.5L of sterile ethanol is added; finally, adding the residual sterile ethanol into a crystallizing tank at one time, after the addition is finished, uniformly reducing the temperature from 40 ℃ to 30 ℃ at the speed of 8 ℃/h, and then stirring for 3 hours at constant temperature to obtain N (2) -L-alanyl-L-glutamine crystal liquid;
(5) opening a bottom valve of a crystallization tank, opening compressed air of the crystallization tank, pressing crystallization liquid in the tank into a filtering, washing and drying machine, filtering to obtain a filter cake, washing the filter cake for three times by using sterile ethanol, opening the filtering, washing and drying machine to pre-dry materials by using the compressed air, adjusting the compressed air pressure to be 0.8MPa, wherein the air blowing time is 1.0 hour, closing the compressed air after the air blowing is finished, opening a vacuum system and an interlayer circulating water system of the filtering, washing and drying machine, setting the circulating water temperature to be 60 ℃, continuing drying for 15 hours after the temperature of the materials reaches 60 ℃, and discharging to obtain 55.2kg of dried sterile N (2) -L-alanyl-L-glutamine crystals. The crystal form of N (2) -L-alanyl-L-glutamine was determined by powder X-ray diffraction method.
The melting point is 215.5-215.8 ℃, and the purity is 99.95% by HPLC.
The crystal modification of N (2) -L-alanyl-L-glutamine made in this example was also subjected to X-ray powder diffraction pattern measurement using Cu-Ka radiation, which is substantially the same as in the example.
Example three:
the preparation method of the novel N (2) -L-alanyl-L-glutamine crystal form comprises the following steps:
(1) dissolving: adding 130kg of injection water into a cleaned dosing tank, starting the dosing tank to stir, opening a heating system, heating the injection water to 80 ℃, keeping the temperature, adding 65kg of N (2) -L-alanyl-L-glutamine crude product into the injection water, and stirring to completely dissolve the N (2) -L-alanyl-L-glutamine crude product;
(2) decoloring and adsorbing: keeping the temperature unchanged, adding 1.3Kg of activated carbon into the solution prepared in the step (1), and stirring for 20 minutes to obtain decolorized adsorption solution;
(3) and (3) filtering: decarburizing the decolorized adsorption solution prepared in the step (2) by using a 3-micron titanium rod to obtain a crude filtrate; then, the coarse filtrate is filtered for the second time through a cylinder filter with the filtering holes of 0.22 mu m, and the active carbon is further removed to obtain fine filtrate; finally, sterilizing and filtering the fine filtrate by two 0.22-micron sterilizing filters connected in series to obtain sterile fine filtrate;
(4) and (3) crystallization: conveying the sterile fine filtrate into a crystallizing tank, preparing 975L of sterile ethanol, starting to cool, uniformly cooling to 45 ℃ at the speed of 9 ℃/hour, uniformly dropwise adding the sterile ethanol in the cooling process, and adding 487.5L of the volume of the sterile ethanol; then, the temperature is uniformly reduced to 35 ℃ from 45 ℃ at a speed of 4 ℃/h, sterile ethanol is uniformly dropwise added in the temperature reduction process, and the volume of the added sterile ethanol is 325L; finally, adding the residual sterile ethanol into a crystallizing tank at one time, after the addition is finished, uniformly reducing the temperature from 35 ℃ to 30 ℃ at the speed of 7 ℃/h, then keeping the temperature at 30 ℃ unchanged, and stirring at constant temperature for 3 hours to obtain N (2) -L-alanyl-L-glutamine crystal liquid;
(5) opening a bottom valve of a crystallization tank, opening compressed air of the crystallization tank, pressing crystallization liquid in the tank into a filtering, washing and drying machine, filtering to obtain a filter cake, washing the filter cake for three times by using sterile ethanol, opening the filtering, washing and drying machine to pre-dry materials by using the compressed air, adjusting the compressed air pressure to be 0.5MPa, keeping the air blowing time to be 0.5 hour, closing the compressed air after the air blowing is finished, opening a vacuum system and an interlayer circulating water system of the filtering, washing and drying machine, setting the circulating water temperature to be 55 ℃, continuing drying for 10 hours after the temperature of the materials reaches 55 ℃, and discharging to obtain 54.5kg of dried sterile N (2) -L-alanyl-L-glutamine crystals.
The melting point is 215.4-216.0 ℃, and the purity is 99.92% by HPLC.
The crystal modification of N (2) -L-alanyl-L-glutamine made in this example was also subjected to X-ray powder diffraction pattern measurement using Cu-Ka radiation, which is substantially the same as in the example.
Example four:
the preparation method of the novel N (2) -L-alanyl-L-glutamine crystal form comprises the following steps:
(1) dissolving: adding 162.5kg of water for injection into a cleaned mixing tank, starting the mixing tank to stir, opening a heating system, heating the water for injection to 70 ℃, keeping the temperature unchanged, adding 65kg of crude N (2) -L-alanyl-L-glutamine into the water for injection, and stirring to completely dissolve the crude N (2) -L-alanyl-L-glutamine;
(2) decoloring and adsorbing: keeping the temperature unchanged, adding 325g of activated carbon into the solution prepared in the step (1), and stirring for 20 minutes to obtain a decolorized adsorption solution;
(3) and (3) filtering: decarburizing the decolorized adsorption solution prepared in the step (2) by using a 3-micron titanium rod to obtain a crude filtrate; then, the coarse filtrate is filtered for the second time through a cylinder filter with the filtering holes of 0.1 mu m, and the active carbon is further removed to obtain fine filtrate; finally, the refined filtrate is subjected to sterilization filtration through a sterilization filter with the diameter of 0.22 mu m to obtain sterile refined filtrate;
(4) and (3) crystallization: conveying the sterile fine filtrate into a crystallizing tank, adding 520L of sterile ethanol, starting to cool, uniformly cooling to 50 ℃ at the speed of 7 ℃/h, uniformly dropwise adding the sterile ethanol in the cooling process, and adding 260L of the sterile ethanol; then, the temperature is uniformly reduced to 35 ℃ from 45 ℃ at the speed of 2 ℃/h, sterile ethanol is uniformly dripped in the temperature reduction process, and the volume of the added sterile ethanol is 173.3L; finally, adding the residual sterile ethanol into a crystallizing tank at one time, after the addition is finished, uniformly reducing the temperature from 35 ℃ to 25 ℃ at the speed of 7 ℃/h, keeping the temperature at 25 ℃ unchanged, and stirring at constant temperature for 2 hours to obtain N (2) -L-alanyl-L-glutamine crystal liquid;
(5) opening a bottom valve of the crystallization tank, opening compressed air of the crystallization tank, pressing crystallization liquid in the tank into a filtering, washing and drying machine, filtering to obtain a filter cake, washing the filter cake for three times by using sterile ethanol, opening the filtering, washing and drying machine to pre-dry the material by using the compressed air, adjusting the pressure of the compressed air to be 1.0MPa, keeping the air blowing time to be 2 hours, closing the compressed air after the air blowing is finished, opening a vacuum system and an interlayer circulating water system of the filtering, washing and drying machine, setting the circulating water temperature to be 50 ℃, continuing drying for 15 hours after the temperature of the material reaches 50 ℃, and discharging to obtain 54.9kg of dry sterile N (2) -L-alanyl-L-glutamine crystals.
The melting point is 215.3-215.8 ℃, and the purity is 99.95% by HPLC.
The crystal modification of N (2) -L-alanyl-L-glutamine made in this example was also subjected to X-ray powder diffraction pattern measurement using Cu-Ka radiation, which is substantially the same as in the example.
Comparative example:
the same crude N (2) -L-alanyl-L-glutamine was purified according to the purification process of example 6 in CN201110384393.7, the melting point was 215.2-216.2 deg.C, and the purity was 99.58% by HPLC.
As can be seen from the above experiments, the crystals prepared by the refining method of N (2) -L-alanyl-L-glutamine of the present invention have a concentrated melting point and a better purity than the crystals prepared by the comparative example, and the safety of N (2) -L-alanyl-L-glutamine for injection is improved.
Experimental example 1: study of hygroscopicity
The present invention examines the hygroscopicity of N (2) -L-alanyl-L-glutamine prepared in examples one, two, three, four and comparative examples, and examines the moisture content of N (2) -L-alanyl-L-glutamine as an index under the conditions of relative humidity of 75% (RH), relative humidity of 95.2% (RH), temperature of 40 ℃, as shown in Table 1.
TABLE 1 Water content in N (2) -L-alanyl-L-Glutamine
Experiments show that the hygroscopicity of the novel N (2) -L-alanyl-L-glutamine crystal form prepared by the invention is obviously lower than that of the N (2) -L-alanyl-L-glutamine crystal form prepared by a comparative example, and the characteristic enables the novel crystal form prepared by the invention to be more convenient to store and more suitable to be used as a raw material of a pharmaceutical preparation.
Experimental example 2 examination of influence factors
In the present invention, N (2) -L-alanyl-L-glutamine purified in examples one, two, three, four and comparative examples was subjected to influence factor examination, and high temperature (60 ℃ C. + -2 ℃ C.), light irradiation (4500Lx + -500 Lx) and high humidity (92.5% RH) were examined to find the contents and contents of the substances, as shown in Table 2.
TABLE 2N (2) -L-alanyl-L-Glutamine content and the content of substances involved
Experimental results show that the novel crystal form of the N (2) -L-alanyl-L-glutamine prepared by the invention is more stable in physical and chemical forms under the conditions of high temperature, illumination and high humidity, and the novel crystal form is difficult to deteriorate and better in stability and is more suitable for preparing injection preparations.
The new crystal form of the N (2) -L-alanyl-L-glutamine prepared by the invention has little increase of related substance content under the conditions, and the possibility of side effect generation of the product is high due to large related substance content and large time change, so that the probability of side effect generation of the product is low when the crystal form prepared by the method is used for preparing an injection preparation, and the use is safer.
Test example 3 examination of clarity, organic residue, and loss on drying
The crystal forms of the crystals prepared in the first to fourth examples and the comparative example were respectively subjected to comparative tests, and the results are shown in table 3:
TABLE 3 clarity, organic residue and loss on drying
| Inspection item | Clarity of the product | Organic residue | Loss on drying |
| Example one | 0.5# | 0.01％ | 0.02％ |
| Example two | 0.5# | 0.02％ | 0.03％ |
| EXAMPLE III | 0.5# | 0.02％ | 0.02％ |
| Example four | 0.5# | 0.02％ | 0.03％ |
| Comparative example | 1# | 0.20％ | 0.09％ |
As can be seen from the table above, the N (2) -L-alanyl-L-glutamine prepared in the first to fourth examples has good crystal form clarity, low organic residue and less drying weight loss, and all the indexes are superior to those of the crystal form of the comparative example.
The clarity is good, and the possibility of adverse drug reactions is reduced; the low organic residue effectively reduces the toxic and side effects on human bodies, so that the injection prepared by the invention is safer.
Although the present invention has been described with reference to a preferred embodiment, it should be understood that various changes, substitutions and alterations can be made herein without departing from the spirit and scope of the invention as defined by the appended claims.
Claims (10)
1. A novel crystal form of N (2) -L-alanyl-L-glutamine, characterized by: an X-ray powder diffraction method using Cu-Ka radiation and expressed by 2 theta angle has characteristic peaks at +/-0.1 degrees of the following angles of 13.76,15.58,18.58,18.72,19.66,20.72, 21.34, 21.42,21.70,22.52,23.28,23.34,24.12,24.86,25.54,26.12,27.66,30.46,30.64,32.56,33.10,34.26,34.62,34.86,36.60,36.90 and 37.48, and the characteristic peaks have a powder X-ray diffraction pattern shown in figure 1, and the novel preparation method of the N (2) -L-alanyl-L-glutamine crystal form comprises the following steps:
dissolving in step (1): adding the N (2) -L-alanyl-L-glutamine crude product into water for injection at a certain temperature, and stirring at constant temperature until the N (2) -L-alanyl-L-glutamine crude product is completely dissolved;
decoloring and adsorbing: keeping the temperature unchanged, adding activated carbon into the solution prepared in the step (1), and stirring for a period of time to obtain a decolorized adsorption solution;
filtering in step (3): keeping the temperature unchanged, and filtering the decolorized adsorption solution prepared in the step (2) to obtain sterile fine filtration solution;
and (4) crystallizing: placing the sterile fine filtrate in a crystallization device, taking the filtration temperature as the initial temperature, and adding sterile ethanol into the sterile fine filtrate obtained in the step (3) in batches while performing gradient cooling to crystallize N (2) -L-alanyl-L-glutamine to obtain N (2) -L-alanyl-L-glutamine crystal liquid;
and (5) filtering, washing and drying the crystal liquid under the aseptic condition to obtain dry aseptic N (2) -L-alanyl-L-glutamine crystals.
2. A process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 1, characterized in that: the preparation method comprises the following steps:
dissolving in step (1): adding the N (2) -L-alanyl-L-glutamine crude product into water for injection at a certain temperature, and stirring at constant temperature until the N (2) -L-alanyl-L-glutamine crude product is completely dissolved;
decoloring and adsorbing: keeping the temperature unchanged, adding activated carbon into the solution prepared in the step (1), and stirring for a period of time to obtain a decolorized adsorption solution;
filtering in step (3): keeping the temperature unchanged, and filtering the decolorized adsorption solution prepared in the step (2) to obtain sterile fine filtration solution;
and (4) crystallizing: placing the sterile fine filtrate in a crystallization device, taking the filtration temperature as the initial temperature, and adding sterile ethanol into the sterile fine filtrate obtained in the step (3) in batches while performing gradient cooling to crystallize N (2) -L-alanyl-L-glutamine to obtain N (2) -L-alanyl-L-glutamine crystal liquid;
filtering, washing and drying the crystal liquid under the aseptic condition to obtain dry aseptic N (2) -L-alanyl-L-glutamine crystal;
the weight ratio of the N (2) -L-alanyl-L-glutamine crude product to the water for injection in the step (1) is 1Kg: 2-2.5 Kg; heating water for injection to 70-80 ℃ in the step (1);
the volume ratio of the weight of the N (2) -L-alanyl-L-glutamine crude product to the sterile ethanol in the step (4) is 1kg: 8-15L;
the gradient cooling process in the step (4), the addition amount of the sterile ethanol and the addition mode are as follows:
the first stage is as follows: pumping the sterile fine filtrate into a crystallization device, then starting to cool, uniformly cooling to 45-50 ℃ at the speed of 7-9 ℃/h by taking the filtration temperature as the starting temperature, and uniformly dropwise adding sterile ethanol in the cooling process, wherein the volume of the added sterile ethanol is 1/3-1/2 of the total volume of the sterile ethanol;
and a second stage: uniformly reducing the temperature from 45-50 ℃ to 35-40 ℃ at the speed of 2-4 ℃/h, uniformly dropwise adding sterile ethanol in the temperature reduction process, wherein the volume of the added sterile ethanol is 1/4-1/3 of the total volume of the sterile ethanol;
and a third stage: and after the second stage is finished, adding the residual sterile ethanol into the crystallization device at one time, uniformly reducing the temperature from 35-40 ℃ to 25-30 ℃ at the speed of 7-9 ℃/h, and then stirring at constant temperature for 2-3 hours.
3. The process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 2, characterized in that: the volume ratio of the weight of the crude N (2) -L-alanyl-L-glutamine to the sterile ethanol is 1kg to 10L.
4. The process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 2, characterized in that: in the step (1), the water for injection is heated to 73-75 ℃.
5. The process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 2, characterized in that: the adding weight of the activated carbon in the step (2) is 0.5-2% of the weight of the crude product of the N (2) -L-alanyl-L-glutamine.
6. The process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 5, characterized in that: the added weight of the activated carbon is 1 percent of the weight of the crude product of the N (2) -L-alanyl-L-glutamine.
7. The process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 2, characterized in that: the filtration in the step (3) is divided into three steps: firstly, decarburizing a decolored adsorption solution by using a titanium rod to obtain a crude filtrate; secondly filtering the coarse filtrate by a cylinder filter, and further removing active carbon to obtain fine filtrate; and finally, sterilizing and filtering the fine filtrate by a sterilizing filter to obtain the sterile fine filtrate.
8. The process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 2, characterized in that: and (5) performing drying operation by firstly adopting compressed air with certain pressure to pre-blow the material, closing the compressed air after blowing for a period of time, drying by using a vacuum system of a filtering, washing and drying machine and an interlayer circulating water system, setting the temperature of circulating water to be 50-60 ℃, starting timing when the temperature of the material reaches the preset temperature, and keeping the temperature for drying for 10-15 hours.
9. The process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 8, characterized in that: the pressure of the compressed air is 0.5-1.0 MPa; the air blowing time is 0.5-2.0 hours.
10. The process for preparing a novel crystalline form of N (2) -L-alanyl-L-glutamine according to claim 9, characterized in that: the pressure of the compressed air is 0.8 MPa; the air-blowing time was 1.0 hour.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610861653.8A CN106565821B (en) | 2016-09-28 | 2016-09-28 | N (2) -L-alanyl-L-glutamine novel crystal form and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610861653.8A CN106565821B (en) | 2016-09-28 | 2016-09-28 | N (2) -L-alanyl-L-glutamine novel crystal form and preparation method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106565821A CN106565821A (en) | 2017-04-19 |
| CN106565821B true CN106565821B (en) | 2020-04-10 |
Family
ID=58532556
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610861653.8A Active CN106565821B (en) | 2016-09-28 | 2016-09-28 | N (2) -L-alanyl-L-glutamine novel crystal form and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106565821B (en) |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104211757B (en) * | 2014-09-03 | 2016-11-02 | 山东金城医药股份有限公司 | N (2)-Ala-Gln novel crystal forms and preparation method thereof |
-
2016
- 2016-09-28 CN CN201610861653.8A patent/CN106565821B/en active Active
Also Published As
| Publication number | Publication date |
|---|---|
| CN106565821A (en) | 2017-04-19 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| WO2017050171A1 (en) | Crystal form of pyrroloquinoline quinone sodium salt and preparation method and use thereof | |
| CN105440057B (en) | A kind of method for preparing cefoperazone sodium | |
| CN110771892B (en) | High-quality curcumin product, production method and application thereof | |
| CN106687470A (en) | Sofosbuvir in crystalline form and process for its preparation | |
| JPS62277348A (en) | Spray drying process for amino acid composition | |
| CN106360718A (en) | Extraction technology of aloe vera gels | |
| CA3199800A1 (en) | Method for refining glycolide and glycolide obtained therefrom | |
| CN106565821B (en) | N (2) -L-alanyl-L-glutamine novel crystal form and preparation method thereof | |
| CN101947208B (en) | Fludarabine phosphate composition for injection and preparation method thereof | |
| CN106810583B (en) | A kind of fructose sodium diphosphate compound and preparation method thereof | |
| CN102643255B (en) | Andrographolide compound | |
| CN108685843B (en) | Aminocaproic acid injection and preparation method thereof | |
| JPH05271241A (en) | Production of crystal | |
| CN102367229B (en) | Ethylenediamine diaceturate compound and pharmaceutical composition thereof | |
| WO2015067130A1 (en) | (s)-oxiracetam crystal form iii, preparation method therefor, and application thereof | |
| CN106543124A (en) | dapagliflozin compound | |
| CN102358721B (en) | More stable aceglutamide compound and medicinal composition thereof | |
| CN110627792A (en) | Pentoxifylline compound | |
| CN109232707B (en) | Method for removing solvent residues of diammonium glycyrrhizinate bulk drug | |
| CN106565552A (en) | Belinostat crystal and preparation method therefor | |
| CN106831926B (en) | Drying method of rocuronium bromide bulk drug for injection | |
| CN104610385A (en) | Refining method of D-glucosamine hydrochloride | |
| TWI680983B (en) | The l-proline complex, monohydrate and crystal of a sodium-glucose contransporter 2 inhibitor | |
| CN110225918B (en) | Preparation method of azacitidine crystal form | |
| CN110862344A (en) | 1/20 aquo-vitamin B6 compound |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |