Method for recovering cyclosporin A crystallization mother liquor

Abstract

The invention relates to a recovery method of cyclosporin A crystallization mother liquor, which comprises the following steps: carrying out previous treatment and impurity removal on the cyclosporin A crystallization mother liquor, carrying out primary crystallization with a polar solvent, carrying out secondary crystallization with a non-polar solvent, and finally carrying out crystal transformation and drying with the polar solvent to obtain a cyclosporin A finished product. According to the method, the residual cyclosporin A in the crystallization mother liquor is effectively purified, waste of the residual cyclosporin A in the crystallization mother liquor is avoided, the production cost of the cyclosporin A can be remarkably reduced, the method is suitable for industrial production, the product content reaches 98.5% or above, and the maximum single impurity content is within 0.7%.

Description

Technical Field

[0001] The present invention belongs to the field of biological medicine technology, and particularly relates to a method for recovering the crystallization mother liquor of cyclosporin A. Background Art

[0002] Cyclosporin A is a cyclic polypeptide containing 11 amino acids isolated from the culture of Beauveria bassiana, a white fungus. It is a very strong immunosuppressant. In addition to cyclosporin A, there are more than 20 homologues such as B, C, and D. Cyclosporin A is used for immunosuppressive treatment in kidney, liver, heart and other transplantation surgeries. It can also be used to treat some autoimmune diseases, such as lupus erythematosus, rheumatoid arthritis, idiopathic thrombocytopenic purpura, ulcerative colitis, myasthenia gravis, chronic nephritis, chronic active hepatitis and nephrotic syndrome and other autoimmune diseases. The compound information of cyclosporin A is as follows:

[0003] Chinese Name: Cyclosporin A

[0004] English Name: Ciclosporin A

[0005] CAS Registry Number: 59865-13-3

[0006] The chemical structural formula of cyclosporin A is as follows:

[0007]

[0008] The main impurities in the fermentation products of cyclosporin A are cyclosporin A homologues with similar structures, including B, C, D, etc. In the prior art, the main process route for the extraction and purification of cyclosporin A is to first extract with organic solvents and then separate and purify through different types of column chromatography. The initial content of the cyclosporin A intermediate obtained by the fermentation method is only about 15%. Later, through extraction, purification, macroporous adsorption resin chromatography, and silica gel column chromatography, the crude product of cyclosporin A is obtained. The crude product of cyclosporin A can finally obtain the medicinal standard cyclosporin A through recrystallization or preparative liquid phase technology. Whether it is recrystallization or high-performance liquid chromatography preparative technology, a certain amount of mother liquor will be generated during the refining, purification, and crystal transformation of cyclosporin A. Usually, the content of this mother liquor is 35% - 80%, the maximum single impurity is 1% - 3%, and the yield accounts for 10% - 40% of the total product. How to make full use of this mother liquor, improve the product yield, and reduce the production cost has become an urgent problem for each production enterprise.

[0009] Patent US597638 discloses a method for obtaining cyclosporin A through silica gel column chromatography using high-pressure carbon dioxide as a mobile carrier, but this method has high requirements for equipment and a small sample processing capacity, and cannot achieve large-scale production.

[0010] Patent US2002 / 0162789A1 discloses a method of separating by multi-step silica gel chromatography with mobile phases of toluene and ethyl acetate. The main disadvantages are that toluene is highly toxic, the chromatographic steps are numerous, and the solvent consumption is large.

[0011] WO01 / 64935A1 describes a method for separating and purifying cyclosporin A by extraction with organic solvents, gel filtration, and silica gel column chromatography. The main disadvantages of this method are that the gel filtration packing is expensive, and the solvent from the previous step needs to be removed between each step, resulting in a large workload and high solvent consumption.

[0012] WO20010064935 discloses that after extracting the fermentation broth with methanol, alum is added to the concentrated solution and stirred for 3 hours, followed by filtration. The filter cake is dissolved in cyclohexane, and after concentration, a crude product of cyclosporin A with a purity of 59.8% is obtained. The crude product is further purified by gel filtration, column chromatography, and crystallization to obtain the cyclosporin A product.

[0013] The above patent documents have studied the separation and purification of cyclosporin A. However, none of the above technical solutions involve the recovery of residual cyclosporin A in the secondary crystallization mother liquor. Therefore, it is necessary to develop an effective method for recovering cyclosporin A crystallization mother liquor, which will greatly reduce the production cost of cyclosporin A and also reduce waste and environmental pollution during the production process. Summary of the Invention

[0014] The purpose of the present invention is to provide a method for recovering cyclosporin A crystallization mother liquor. By pre-treating the cyclosporin A crystallization mother liquor to remove impurities and combining two crystallization and crystal transformation methods, the technical problem of waste of residual cyclosporin A in the cyclosporin A crystallization mother liquor is solved, the residual cyclosporin A in the crystallization mother liquor is effectively purified, and the production cost of cyclosporin A is significantly reduced.

[0015] The technical solution adopted to achieve the above invention purpose is as follows:

[0016] 1. A method for recovering cyclosporin A crystallization mother liquor, comprising the following steps:

[0017] 1) Remove the solvent from the cyclosporin A crystallization mother liquor, dissolve it in acetonitrile, and perform solid-liquid separation to obtain a filtrate;

[0018] 2) Perform activated carbon decolorization treatment on the filtrate obtained in step 1) to obtain a decolorized solution;

[0019] 3) Recover the solvent from the decolorized solution obtained in step 2), add a polar solvent for primary crystallization, and perform solid-liquid separation to obtain Crystal I;

[0020] 4) Add a non-polar solvent to Crystal I obtained in step 3) for secondary crystallization, and perform solid-liquid separation to obtain Crystal II;

[0021] 5) Transfer the crystalline II obtained in step 4) to a polar solvent for recrystallization, perform solid-liquid separation, and conduct vacuum drying to obtain the finished product of cyclosporin A.

[0022] Among them, the volume ratio of acetonitrile added in step 1) to the amount of the product in the mother liquor is 10 - 20:1 (w / v).

[0023] Among them, the decolorization conditions of the activated carbon in step 2): the decolorization temperature is 40°C - 50°C, and the amount of activated carbon added is 1% - 3% (w / w) of the weight of cyclosporin A.

[0024] Among them, the solvent recovery in step 3) is carried out by vacuum concentration under the conditions of a temperature of 50°C - 60°C and a vacuum degree of -0.09 to -0.1 Mpa.

[0025] Among them, step 3) further includes adding purified water accounting for 15% - 20% of the volume of the crystallization system before primary crystallization.

[0026] Among them, the primary crystallization conditions in step 3): the crystallization temperature is -5°C - 0°C, the crystallization holding time is 2 h, and the filtration temperature is -15°C - -10°C.

[0027] Among them, the non-polar solvent in step 4) is at least one of ethyl acetate, hexane, and petroleum ether, and the amount of the non-polar solvent added is 10 - 50 times the amount of the product (w / v).

[0028] Among them, the secondary crystallization conditions in step 4): the crystallization temperature during purification is -10°C - -5°C, the crystallization holding time is 2 h, and the filtration temperature is -20°C - -15°C.

[0029] Among them, the recrystallization conditions in step 5): the concentration of cyclosporin A is 2% - 5%, the crystallization temperature during purification is -5°C - 0°C, the holding time is 2 h, and the filtration temperature is -10°C - -5°C.

[0030] Among them, the drying conditions in step 5): the temperature is 40 - 70°C, the vacuum degree is -0.09 MPa - -0.1 Mpa, and the drying time is 40 h.

[0031] Among them, the polar solvent in step 3) and step 5) is at least one of ethanol, methanol, and acetone, and the polar solvent is 10 - 50 times the amount of the product (w / v).

[0032] The technical solution of the present invention has at least the following beneficial technical effects:

[0033] 1. The process route of the method for recovering the crystallization mother liquor of cyclosporin A disclosed in the present invention is short, the investment is small, the solvent can be recycled, the yield of cyclosporin A is improved, and the production cost of cyclosporin A is significantly reduced.

[0034] 2. The process of the present invention effectively purifies the residual cyclosporin A in the crystallization mother liquor, avoids the waste of the residual cyclosporin A in the crystallization mother liquor, can significantly reduce the production cost of cyclosporin A, is suitable for industrial production, and the product content reaches more than 98.5%, with the maximum single impurity within 0.7%. Detailed Embodiments

[0035] The following will clearly and completely describe the implementation schemes of the present invention in combination with the embodiments. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. The following description of at least one exemplary embodiment is actually only illustrative and in no way limits the present invention and its application or use. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts belong to the scope of protection of the present invention.

[0036] The "cyclosporin A crystallization mother liquor" described in the present invention can be the mother liquor obtained after the first crystallization treatment of the cyclosporin A fermentation broth, as well as the mother liquor after the second and third crystallizations or their mixture. This crystallization treatment can be the method commonly used in the art.

[0037] The HPLC detection method for the following examples and comparative examples: Chromatographic conditions: Octadecylsilane-bonded silica gel is used as the filler (a stainless steel tube with a diameter of φ0.25 mm × 1000 mm is connected in front of the column); the mobile phase is acetonitrile - water - tert-butyl methyl ether - phosphoric acid (430:520:50:1), the stainless steel tube and the column temperature are 70°C; the detection wavelength is 210 nm; the injection volume is 20 μl. Calculated by the peak area normalization method.

[0038] Example 1

[0039] Take 500 L of the acetone mother liquor of cyclosporin A (concentration 2.1 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 38%, and the maximum single impurity is 2.0%. Concentrate it to dryness at 60 °C and -0.09 Mpa to obtain 2.8 kg of concentrate. Add 45 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -15 °C, stir for 2 h, and filter. Add 80 g of activated carbon to the filtrate, stir and heat to 45 °C, perform decolorization treatment for 1 h, and filter. Concentrate the filtrate to dryness at 55 °C and -0.09 Mpa. Add 40 L of acetone to the concentrate, heat and stir at 40 °C to completely dissolve it to obtain a clear and transparent solution. Perform acetone crystallization. Add 6 L of purified water, which is 15% of the volume of the crystallization system, to the system at 5 °C, keep the temperature at 0 °C for crystal precipitation for 2 h, filter at -15 °C, and collect the crystals. After completely dissolving the cyclosporin A crystals in 20 L of a mixed solution of ethyl acetate and n-hexane (80:20) by heating to 40 °C, perform recrystallization. Start crystal precipitation at -3 °C, keep the temperature for 2 h, filter at -20 °C, and collect the crystals. Add 40 L of acetone to the cyclosporin A crystals, heat and stir at 40 °C to completely dissolve them to obtain a clear and transparent solution. Perform acetone crystallization. Keep the temperature at 0 °C for crystal precipitation for 2 h, filter at -10 °C, and collect the crystals. Dry at 70 °C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 380 g of cyclosporin A product, and the yield is 36.2%. In this experiment, after one-time acetone purification, one-time crystallization with a mixed solution of ethyl acetate and n-hexane, and acetone crystal conversion, cyclosporin A meeting the pharmaceutical standard with a maximum single impurity of 0.52% and a content of 98.9% can be obtained.

[0040] Example 2

[0041] Take 500 L of cyclosporin A acetone mother liquor (concentration 1.6 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 70%, and the maximum single impurity is 1.9%. Concentrate it to dryness at 60 °C and -0.09 Mpa to obtain 1.15 kg of concentrate. Add 25 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -15 °C, stir for 2 h, and filter. Add 100 g of activated carbon to the filtrate, stir and heat to 45 °C for 1 h of decolorization treatment, and filter. Concentrate the filtrate to dryness at 60 °C and -0.09 Mpa. Add 40 L of ethanol to the concentrate, heat and stir at 40 °C to completely dissolve it to obtain a clear and transparent solution. Perform ethanol crystallization. Add 8 L of purified water, which is 20% of the volume of the crystallization system, to the system at 5 °C, keep the temperature at 0 °C for crystal precipitation for 2 h, filter at -15 °C, and collect the crystals. After completely dissolving the cyclosporin A crystals in 20 L of a mixed solution of ethyl acetate and petroleum ether (80:20) by heating to 40 °C, perform recrystallization. Start crystal precipitation at -5 °C, keep the temperature for 2 h, filter at -18 °C, and collect the crystals. Add 40 L of acetone to the cyclosporin A crystals, heat and stir at 40 °C to completely dissolve them to obtain a clear and transparent solution. Perform acetone crystallization. Keep the temperature at -2 °C for crystal precipitation for 2 h, filter at -10 °C, and collect the crystals. Dry at 70 °C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 336 g of cyclosporin A product with a yield of 42.0%. In this experiment, after one-time ethanol purification, one-time crystallization with a mixed solution of ethyl acetate and petroleum ether, and acetone crystal conversion, cyclosporin A meeting the pharmaceutical standard with a maximum single impurity of 0.60% and a content of 98.6% can be obtained.

[0042] Example 3

[0043] Take 500 L of the acetone mother liquor of cyclosporin A (concentration 1.8 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 78%, and the maximum single impurity is 1.5%. Concentrate it to dryness at 60 °C and -0.09 Mpa to obtain 1.15 kg of concentrate. Add 21 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -20 °C, stir for 2 h, and filter. Add 100 g of activated carbon to the filtrate, stir and heat to 45 °C for 1 h for decolorization treatment, and then filter. Concentrate the filtrate to dryness at 55 °C and -0.09 Mpa. Add 40 L of acetone to the concentrate, heat and stir at 40 °C to completely dissolve it to obtain a clear and transparent solution. Perform acetone crystallization. Add 6 L of purified water, which is 15% of the volume of the crystallization system, to the system at 3 °C, keep the temperature at -1 °C for crystal precipitation for 2 h, filter at -15 °C, and collect the crystals. Heat the cyclosporin A crystals to 40 °C and completely dissolve them in 20 L of a mixed solution of ethyl acetate and n-hexane (80:20), then perform recrystallization. Start crystal precipitation at -2 °C, keep the temperature for 2 h, filter at -15 °C, and collect the crystals. Add 40 L of acetone to the cyclosporin A crystals, heat and stir at 40 °C to completely dissolve them to obtain a clear and transparent solution. Perform acetone crystallization. Keep the temperature at 0 °C for crystal precipitation for 2 h, filter at -10 °C, and collect the crystals. Dry at 70 °C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 370 g of cyclosporin A product, and the yield is 41.1%. In this experiment, after one-time acetone refining, one-time crystallization with ethyl acetate and n-hexane, and acetone crystal transformation, cyclosporin A meeting the pharmaceutical standard with a maximum single impurity of 0.55% and a content of 98.7% can be obtained.

[0044] Example 4

[0045] Take 500 L of the acetone mother liquor of cyclosporin A (concentration 2.0 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 58%, and the maximum single impurity is 1.8%. Concentrate it to dryness at 60 °C and -0.09 Mpa to obtain 1.8 kg of concentrate. Add 36 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -15 °C, stir for 2 h, and filter. Add 72 g of activated carbon to the filtrate, stir and heat to 45 °C for 1 h for decolorization treatment, and then filter. Concentrate the filtrate to dryness at 55 °C and -0.09 Mpa. Add 40 L of ethanol to the concentrate, heat and stir at 40 °C to completely dissolve it to obtain a clear and transparent solution. Perform ethanol crystallization. Add 6 L of purified water, which is 15% of the volume of the crystallization system, to the system at 5 °C, keep the temperature at 0 °C for crystal precipitation for 2 h, filter at -15 °C, and collect the crystals. After completely dissolving the cyclosporin A crystals in 20 L of petroleum ether by heating to 40 °C, perform recrystallization. Start crystal precipitation at 10 °C, keep the temperature for 2 h, filter at -10 °C, and collect the crystals. Add 40 L of acetone to the cyclosporin A crystals, heat and stir at 40 °C to completely dissolve them to obtain a clear and transparent solution. Perform acetone crystallization. Keep the temperature at 0 °C for crystal precipitation for 2 h, filter at -10 °C, and collect the crystals. Dry at 70 °C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 318 g of cyclosporin A product, with a yield of 31.8%. In this experiment, after one-time acetone refining, one-time petroleum ether crystallization, and acetone crystal conversion, cyclosporin A meeting the pharmaceutical standard with a maximum single impurity of 0.68% and a content of 98.5% can be obtained.

[0046] Example 5

[0047] Take 500 L of cyclosporin A acetone mother liquor (concentration 1.7 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 55%, and the maximum single impurity is 1.9%. Concentrate it to dryness at 60 °C and -0.09 Mpa to obtain 1.55 kg of concentrate. Add 31 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -15 °C, stir for 2 h, and filter. Add 90 g of activated carbon to the filtrate, stir and heat to 45 °C for 1 h for decolorization treatment, and filter. Concentrate the filtrate to dryness at 60 °C and -0.09 Mpa. Add 40 L of methanol to the concentrate, heat and stir at 40 °C to completely dissolve it to obtain a clear and transparent solution. Perform methanol crystallization. Add 8 L of purified water, which is 20% of the volume of the crystallization system, to the system at 5 °C, keep the temperature at 0 °C for crystal precipitation for 2 h, filter at -17 °C, and collect the crystals. After completely dissolving the cyclosporin A crystals in 20 L of n-hexane by heating to 40 °C, perform recrystallization. Start crystal precipitation at -5 °C, keep the temperature for 2 h, filter at -15 °C, and collect the crystals. Add 40 L of acetone to the cyclosporin A crystals, heat and stir at 40 °C to completely dissolve them to obtain a clear and transparent solution. Perform acetone crystallization. Keep the temperature at -2 °C for crystal precipitation for 2 h, filter at -10 °C, and collect the crystals. Dry at 70 °C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 256 g of cyclosporin A product, with a yield of 30.1%. In this experiment, after one-time ethanol refining, one-time n-hexane crystallization, and acetone crystal conversion, cyclosporin A meeting the pharmaceutical standard with a maximum single impurity of 0.69% and a content of 98.6% can be obtained.

[0048] Example 6

[0049] Take 500 L of cyclosporin A acetone mother liquor (concentration 1.7 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 68%, and the maximum single impurity is 1.5%. Concentrate it to dryness at 60 °C and -0.09 Mpa to obtain 1.25 kg of concentrate. Add 25 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -20 °C, stir for 2 h, and filter. Add 50 g of activated carbon to the filtrate, stir and heat to 45 °C for 1 h of decolorization treatment, and filter. Concentrate the filtrate to dryness at 55 °C and -0.09 Mpa, add 40 L of acetone to the concentrate, heat and stir at 40 °C to completely dissolve it to obtain a clear and transparent solution. Perform acetone crystallization. Add 6 L of purified water, which is 15% of the crystallization system volume, to the system at 3 °C, keep the temperature at -1 °C for crystal precipitation for 2 h, filter at -15 °C, and collect the crystals. After completely dissolving the cyclosporin A crystals in 20 L of ethyl acetate by heating to 40 °C, perform recrystallization. Start crystal precipitation at -10 °C, keep the temperature for 2 h, filter at -25 °C, and collect the crystals. Add the cyclosporin A crystals to 40 L of acetone, heat and stir at 40 °C to completely dissolve it to obtain a clear and transparent solution. Perform acetone crystallization. Keep the temperature at 0 °C for crystal precipitation for 2 h, filter at -10 °C, and collect the crystals. Dry at 70 °C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 236 g of cyclosporin A product, with a yield of 27.8%. In this experiment, after one acetone purification and one ethyl acetate crystallization and acetone crystal conversion, cyclosporin A meeting the pharmaceutical standard with a maximum single impurity of 0.58% and a content of 98.7% can be obtained.

[0050] Comparative Example 1

[0051] Take 500 L of cyclosporin A acetone mother liquor (concentration 1.7 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 69%, and the maximum single impurity is 1.5%. Concentrate it to dryness at 60 °C and -0.09 Mpa to obtain 1.30 kg of concentrate. Add 26 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -20 °C, stir for 2 h, and filter. Add 50 g of activated carbon to the filtrate, stir and heat to 45 °C for 1 h of decolorization treatment, and filter. Concentrate the filtrate to dryness at 55 °C and -0.09 Mpa, add 20 L of a mixed solution of ethyl acetate and petroleum ether (80:20) to the concentrate, heat to 40 °C to completely dissolve it, then perform recrystallization. Start crystal precipitation at -10 °C, keep the temperature for 2 h, filter at -25 °C, and collect the crystals. Add the cyclosporin A crystals to 40 L of acetone, heat and stir at 40 °C to completely dissolve it to obtain a clear and transparent solution. Perform acetone crystallization. Keep the temperature at -5 °C for crystal precipitation for 2 h, filter at -15 °C, and collect the crystals. Dry at 70 °C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 367 g of cyclosporin A product, with a yield of 43.2%. In this experiment, after one crystallization with a mixed solution of ethyl acetate and petroleum ether and acetone crystal conversion, cyclosporin A with a maximum single impurity of 0.75% and a content of 98.0% can be obtained.

[0052] Comparative Example 2

[0053] Take 500 L of the acetone mother liquor of cyclosporin A (concentration 1.6 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 70%, and the maximum single impurity is 1.9%. Concentrate it to dryness at 60°C and -0.09 Mpa to obtain 1.15 kg of concentrate. Add 23 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -15°C, stir for 2 h, and filter. Add 60 g of activated carbon to the filtrate, stir and heat to 45°C for decolorization treatment for 1 h, and filter. Concentrate the filtrate to dryness at 60°C and -0.09 Mpa, add 40 L of acetone to the concentrate, heat and stir at 40°C to completely dissolve it to obtain a clear and transparent solution. Perform acetone crystallization. Add 8 L of purified water, which is 20% of the volume of the crystallization system, to the system at 5°C, keep the temperature at 0°C for crystal precipitation for 2 h, filter at -15°C, and collect the crystals. Add the cyclosporin A crystals to 40 L of acetone, heat and stir at 40°C to completely dissolve them to obtain a clear and transparent solution. Perform acetone crystallization, keep the temperature at -3°C for crystal precipitation for 2 h, filter at -12°C, and collect the crystals. Dry at 70°C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 348 g of cyclosporin A product, with a yield of 43.5%. In this experiment, after one-time acetone refining and acetone crystal conversion, cyclosporin A with a maximum single impurity of 0.88% and a content of 96.6% can be obtained.

[0054] Comparative Example 3

[0055] Take 500 L of the acetone mother liquor of cyclosporin A (concentration 1.7 g / l) for vacuum concentration. The content of cyclosporin A in the mother liquor is about 68%, and the maximum single impurity is 1.7%. Concentrate it to dryness at 60°C and -0.09 Mpa to obtain 1.25 kg of concentrate. Add 25 L of acetonitrile to the concentrate, stir to completely dissolve the product, place the solution at -20°C, stir for 2 h, and filter. Add 100 g of activated carbon to the filtrate, stir and heat to 45°C for decolorization treatment for 1 h, and filter. Concentrate the filtrate to dryness at 55°C and -0.09 Mpa, add 40 L of acetone to the concentrate, heat and stir at 40°C to completely dissolve it to obtain a clear and transparent solution. Perform acetone crystallization. Add 6 L of purified water, which is 15% of the volume of the crystallization system, to the system at 3°C, keep the temperature at -1°C for crystal precipitation for 2 h, filter at -15°C, and collect the crystals. After heating the cyclosporin A crystals to 40°C and completely dissolving them with 20 L of a mixed solution of ethyl acetate and n-hexane (80:20), perform recrystallization. Start crystal precipitation at -2°C, keep the temperature for 2 h, filter at -15°C, and collect the crystals. Dry at 70°C under a vacuum of -0.09 MPa to -0.1 Mpa for 40 h to obtain 384 g of cyclosporin A product, with a yield of 45.2%. In this experiment, after one-time acetone refining and one-time crystallization with ethyl acetate and n-hexane, cyclosporin A with a maximum single impurity of 0.71% and a content of 98.1% can be obtained.

Claims

1. A method for recovering the mother liquor of cyclosporin A, comprising the following steps: 1) Remove the solvent from the mother liquor of cyclosporin A, dissolve it with acetonitrile, and perform solid-liquid separation to obtain a filtrate; 2) Decolorize the filtrate obtained in step 1) with activated carbon to obtain a decolorized solution; 3) Recover the solvent from the decolorized solution obtained in step 2), perform primary crystallization with a polar solvent, and perform solid-liquid separation to obtain Crystal I; 4) Perform secondary crystallization on Crystal I obtained in step 3) with a non-polar solvent, and perform solid-liquid separation to obtain Crystal II; 5) Perform crystal conversion on Crystal II obtained in step 4) with a polar solvent, perform solid-liquid separation, and perform vacuum drying to obtain the finished product of cyclosporin A.

2. The purification method according to claim 1, characterized in that the activated carbon decolorization conditions in step 2) are: the decolorization temperature is 40°C - 50°C, and the amount of activated carbon added is 1% - 3% (w / w) of the weight of cyclosporin A.

3. The purification method according to claim 1, characterized in that the solvent recovery in step 3) is carried out by vacuum concentration under the conditions of a temperature of 50°C - 60°C and a vacuum degree of -0.09 to -0.1 Mpa.

4. The purification method according to claim 1, characterized in that step 3) further includes adding 15% - 20% of purified water based on the volume of the crystallization system before primary crystallization.

5. The purification method according to claim 4, characterized in that the primary crystallization conditions are: the crystallization temperature is -5°C - 0°C, the crystallization holding time is 2 h, and the filtration temperature is -15°C - -10°C.

6. The purification method according to claim 1, characterized in that the non-polar solvent in step 4) is selected from at least one of ethyl acetate, hexane, and petroleum ether, and the amount of non-polar solvent added is 10 - 50 times (w / v) of the product amount.

7. The purification method according to claim 1, characterized in that the secondary crystallization conditions in step 4) are: the crystallization temperature during purification is -10°C - -5°C, the crystallization holding time is 2 h, and the filtration temperature is -20°C - -15°C.

8. The purification method according to claim 1, characterized in that the crystal conversion conditions in step 5) are: the concentration of cyclosporin A is 2% - 5%, the crystallization temperature during purification is -5°C - 0°C, the holding time is 2 h, and the filtration temperature is -10°C - -5°C.

9. The purification method according to claim 1, characterized in that the drying conditions in step 5) are: the temperature is 40 - 70°C, the vacuum degree is -0.09 Mpa - -0.1 Mpa, and the drying time is 40 h.

10. The purification method according to claim 1, characterized in that the polar solvents in step 3) and step 5) are selected from at least one of ethanol, methanol, and acetone, and the polar solvent is 10 - 50 times (w / v) of the product amount.

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