Freeze-drying process of indocyanine green for injection

Abstract

The invention belongs to the technical field of pharmaceutical preparation processes, and particularly relates to a freeze-drying process of indocyanine green for injection, which comprises the following steps: placing the indocyanine green for injection in a freeze dryer, quickly cooling and freezing to-45 to-35 DEG C, preserving heat and freeze-drying, tempering to a temperature higher than the eutectic point temperature of the indocyanine green for injection, and continuously preserving heat to obtain the indocyanine green for injection. Freezing to-45 DEG C to-35 DEG C at a rapid cooling rate, and then carrying out heat preservation and freeze-drying to finish pre-freezing; and slowly heating the pre-frozen indocyanine green for injection to 0-5 DEG C, preserving heat, heating to 15-25 DEG C at a rate of 0.8-1.2 DEG C / min, and preserving heat to obtain the indocyanine green freeze-dried preparation for injection. According to the freeze-drying process, the water content in the indocyanine green freeze-drying preparation for injection can be reduced to 2.0% or below within a short time, the content of degraded impurities is lower than 2.0%, the product stability is remarkably improved, and the freeze-drying process is convenient to operate, shorter in time consumption and higher in economic benefit.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical preparation processes, and in particular relates to a freeze-drying process for indocyanine green for injection. Background Art

[0002] Indole cyanine dyes are a very important class of near-infrared dyes. Most of the cyanine dyes that have been synthesized and applied are indole cyanine dyes. Compared with other types of cyanine dyes, indole cyanine dyes have excellent optical properties such as higher fluorescence quantum yield, larger molar extinction coefficient, no fluorescence background, good fluorescence performance and a wide tunable absorption wavelength range. At the same time, they have relatively high chemical stability and photostability. Therefore, they are widely used in tumor targeted therapy, protein labeling and trace metal ion detection.

[0003] Indocyanine green (ICG) is a water-soluble tricarbocyanine dye developed by Kodak Research Laboratories in 1955 and approved by the U.S. Food and Drug Administration for medical diagnosis in 1956. With the continuous iteration and update of diagnostic instruments, cyanine dyes such as indocyanine green combined with probes are being used in a wider range of medical and biological fields. As the first wave of indocyanine dyes developed for tumor and vascular contrast agents, the quality standards of its raw materials and indocyanine green for injection (United States Pharmacopoeia and Pharmacopoeia of the People's Republic of China) do not include clear process impurities and degradation impurities, and there are no clear reports on impurity detection in relevant literature. However, process impurities and degradation impurities in drugs will directly affect the effectiveness and safety of drugs. There are relevant literature reports on ICG that after ICG decomposition by singlet oxygen or light decomposition, its decomposition products are cytotoxic. Due to the different doses and conditions of use of ICG during surgery, the precise safety threshold of ICG application has not yet been determined. Therefore, studying ICG degradation products is conducive to the study of clinically related side effects of ICG and ensures a more scientific use of the drug.

[0004] In order to make the indocyanine green preparation for injection as stable as possible during storage, it is prepared as a freeze-dried preparation for storage to ensure its stability. However, the current freeze-drying process is time-consuming and complicated, and the freeze-dried preparation has high degradation impurities and uneven crystal size. Summary of the invention

[0005] In view of this, the present invention provides a freeze-drying process for indocyanine green for injection, which can reduce the moisture content in the indocyanine green freeze-dried preparation for injection to below 2.0% in a short time, and the content of degradation impurities is lower than 2.0%, which significantly improves the product stability. In addition, the freeze-drying process is easy to operate, takes less time, and has greater economic benefits.

[0006] In order to solve the above technical problems, the first aspect of the present invention provides a freeze-drying process of indocyanine green for injection, comprising the following steps:

[0007] S1, pre-freezing: placing the indocyanine green for injection in a freeze dryer, freezing to -45 to -35°C at a cooling rate of 0.8 to 1.2°C / min, then keeping warm for freeze-drying, tempering to a temperature 8 to 12°C higher than the eutectic point temperature of the indocyanine green for injection, and continuing to keep warm for 110 to 130 minutes, and then freezing to -45 to -35°C at a cooling rate of 0.8 to 1.2°C / min, and then keeping warm for freeze-drying, completing the pre-freezing;

[0008] S2, sublimation drying: heating the pre-frozen indocyanine green for injection to 0-5°C at a rate of 0.5-1.5°C / min and then keeping warm;

[0009] S3. analytical drying: heating the indocyanine green for injection after sublimation drying to 15-25° C. at a rate of 0.8-1.2° C. / min and then keeping the temperature to obtain a lyophilized indocyanine green for injection.

[0010] The freeze-drying process of indocyanine green for injection provided by the present invention adopts a rapid cooling method in the pre-freezing process to quickly freeze the indocyanine green to form fine freeze-dried crystals, and then appropriately heats up and tempers to promote the recrystallization of the indocyanine green. When the freeze-drying temperature is higher than the eutectic point temperature of the indocyanine green, smaller ice crystal molecules can migrate to larger ice crystal molecules, thereby providing more sublimation channels for internal water molecules, improving sublimation efficiency, and shortening the freeze-drying time; after tempering for a period of time, the temperature is quickly lowered again for freezing, so that the sample in the glassy state tends to form a more thermodynamically stable state and forms more crystalline forms.

[0011] In combination with the first aspect, in step S1, before tempering, freeze-drying is carried out at -35 to -40°C for 75 to 85 minutes. The freeze-drying temperature and freeze-drying time can make the indocyanine green freeze quickly to form fine freeze-dried crystals, thereby ensuring the uniform size of the crystals and being more conducive to the sublimation of water molecules.

[0012] Preferably, before the tempering, freeze-drying is performed at -40°C for 80 minutes.

[0013] In combination with the first aspect, in step S1, after the tempering, the sample is freeze-dried at -45 to -35°C for 85 to 95 minutes, and the sample is rapidly cooled again after the tempering to make the sample in the glassy state more thermodynamically stable and form more crystalline forms.

[0014] Preferably, after the tempering, the mixture is freeze-dried at -40°C for 90 minutes.

[0015] In combination with the first aspect, the tempering is to increase the temperature to -18 to -12°C at a rate of 0.8 to 1.2°C / min. The tempering temperature is about 10°C higher than the eutectic temperature of indocyanine green, which can make the crystal size more uniform.

[0016] Preferably, the tempering is to increase the temperature to -15°C at a rate of 1.0°C / min and keep the temperature for 120 minutes.

[0017] In combination with the first aspect, during the sublimation drying, the vacuum degree in the freeze dryer is controlled to be 15±5Pa, preferably 15±2Pa.

[0018] In combination with the first aspect, the insulation time of the sublimation drying step is 5 to 7 hours, preferably 6 hours.

[0019] In combination with the first aspect, during the analytical drying, the vacuum degree in the freeze dryer is controlled to be 15±5Pa, preferably 15±2Pa.

[0020] In combination with the first aspect, the insulation time of the analytical drying step is 1 to 3 hours, preferably 2 hours.

[0021] The vacuum degree and holding time in the above drying process can make water molecules sublime as much as possible, thereby reducing the moisture content in the resulting freeze-dried preparation as much as possible, providing a guarantee for product stability.

[0022] A second aspect of the present invention provides a lyophilized indocyanine green preparation for injection, which is prepared by the lyophilization process according to any one of claims 1 to 8.

[0023] Beneficial effects obtained by the present invention: The freeze-drying process of indocyanine green for injection provided by the present invention obtains a freeze-dried indocyanine green for injection preparation having a moisture content and a total amount of impurities equivalent to or even lower than that of a reference preparation (moisture content less than 2.0%, total amount of impurities less than 2.0%) through pre-freezing, sublimation drying and analytical drying processes with a specific cooling rate or heating rate. Moreover, the freeze-drying process has simple steps, is more convenient to operate, and is conducive to large-scale application. DETAILED DESCRIPTION

[0024] In order to make the purpose, technical solution and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present invention and are not used to limit the present invention.

[0025] Indocyanine green lyophilized preparation for injection is a sterile lyophilized product of indocyanine green, which is a dark green loose solid, easily decomposed when exposed to moisture and heat, and the decomposition product has cytotoxicity, which brings side effects to the user. Therefore, the lower the moisture content contained in its lyophilized preparation, the more stable it is during storage, the less impurity content produced by decomposition, and the less side effects during use. However, the current lyophilization process has the problems of long process time and complicated operation, and the moisture content in the lyophilized preparation is high, the crystal size is uneven, resulting in a high content of degradation impurities, and stability is difficult to ensure. In view of this, the present invention provides a lyophilization process for indocyanine green for injection, and the drug crystal size of the lyophilized preparation obtained by the lyophilization process is uniform, the moisture content is lower, the time consumption is shorter, the degradation impurities are less, and the product is more stable.

[0026] In the following examples and comparative examples, the indocyanine green preparation for injection was prepared as follows: 10 g of indocyanine green was weighed into water for injection, stirred for 30 to 60 min, and after it was completely dissolved, the volume was made up to 1000 mL with water for injection, and filled into 400 preparations with a specification of 2.5 mL / tube.

[0027] Example 1

[0028] This embodiment provides a freeze-drying process for indocyanine green for injection, and the preparation steps include:

[0029] S1. Prefreezing: half-stopper the filled ICG for injection and transfer it into the freeze dryer. First, freeze it to -40°C at a cooling rate of 1°C / min and keep it for 80 min. Then, heat it to -15°C at a rate of 1°C / min and keep it for 120 min. Continue to freeze it to -40°C at a cooling rate of 1°C / min and keep it for 90 min to complete the prefreezing.

[0030] S2. Sublimation drying: raise the temperature to 2.5°C at a rate of 0.5°C / min, control the vacuum degree at 15±2Pa, and keep warm for 6h.

[0031] S3. Desorption and drying: Raise the temperature to 20°C at a rate of 1°C / min, control the vacuum degree to 15±2Pa, and keep warm for 2h to obtain a lyophilized indocyanine green preparation for injection.

[0032] Example 2

[0033] This embodiment provides a freeze-drying process for indocyanine green for injection, and the preparation steps include:

[0034] S1. Prefreezing: half-stopper the filled ICG for injection and transfer it into the freeze dryer. First, freeze it to -45°C at a cooling rate of 1.2°C / min and keep it for 75min. Then, heat it to -12°C at a rate of 0.8°C / min and keep it for 125min. Continue to freeze it to -45°C at a cooling rate of 1.2°C / min and keep it for 85min to complete the prefreezing.

[0035] S2. Sublimation drying: heat to 0°C at a rate of 1°C / min, control the vacuum degree at 18±2Pa, and keep warm for 5h.

[0036] S3. Desorption and drying: raise the temperature to 15°C at a rate of 0.8°C / min, control the vacuum degree to 18±2Pa, and keep warm for 3 hours to obtain a lyophilized indocyanine green preparation for injection.

[0037] Example 3

[0038] This embodiment provides a freeze-drying process for indocyanine green for injection, and the preparation steps include:

[0039] S1. Prefreezing: half-stopper the filled ICG for injection and transfer it into the freeze dryer. First, freeze it to -35°C at a cooling rate of 0.8°C / min and keep it for 85 minutes. Then, heat it to -18°C at a rate of 1.2°C / min and keep it for 115 minutes. Continue to freeze it to -35°C at a cooling rate of 0.8°C / min and keep it for 95 minutes to complete the prefreezing.

[0040] S2. Sublimation drying: raise the temperature to 5°C at a rate of 1.5°C / min, control the vacuum degree at 12±2Pa, and keep warm for 7h.

[0041] S3. Desorption and drying: raise the temperature to 25°C at a rate of 1.2°C / min, control the vacuum degree to 12±2Pa, and keep warm for 1 hour to obtain a lyophilized indocyanine green preparation for injection.

[0042] Comparative Example 1

[0043] This comparative example provides a freeze-drying process for indocyanine green for injection, and the preparation steps include:

[0044] S1. Prefreezing: half-stopper the filled ICG for injection and move it into the freeze dryer. First, freeze it to -40°C at a cooling rate of 0.16°C / min and keep it for 80 min. Then, heat it to -15°C at a rate of 1°C / min and keep it for 120 min. Continue to freeze it to -40°C at a cooling rate of 0.16°C / min and keep it for 90 min to complete the prefreezing.

[0045] Steps S2 and S3 are the same as steps S2 and S3 of embodiment 1, respectively.

[0046] Comparative Example 2

[0047] This comparative example provides a freeze-drying process for indocyanine green for injection, and the preparation steps include:

[0048] S1. Prefreezing: half-stopper the filled ICG for injection and transfer it into the freeze dryer. First, freeze it to -40°C at a cooling rate of 1°C / min and keep it for 80 minutes. Then, heat it to -15°C at a rate of 1°C / min and keep it for 120 minutes to complete the prefreezing.

[0049] Steps S2 and S3 are the same as steps S2 and S3 of embodiment 1, respectively.

[0050] Comparative Example 3

[0051] This comparative example provides a freeze-drying process for indocyanine green for injection, and the preparation steps include:

[0052] S1. Same as step S1 in Example 1.

[0053] S2. Same as step S2 in Example 1.

[0054] S3. Desorption and drying: raise the temperature to 20°C at a rate of 0.3°C / min, control the vacuum degree to 15±2Pa, and keep warm for 2h to obtain a lyophilized indocyanine green preparation for injection.

[0055] Comparative Example 4

[0056] This comparative example provides a freeze-drying process for indocyanine green for injection, and the preparation steps include:

[0057] S1. Same as step S1 in Example 1.

[0058] S2. Same as step S2 in Example 1.

[0059] S3. Desorption and drying: Raise the temperature to 40°C at a rate of 1°C / min, control the vacuum degree to 15±2Pa, and keep warm for 2h to obtain a lyophilized indocyanine green preparation for injection.

[0060] Test Example 1

[0061] The pH, moisture and impurity content of the indocyanine green lyophilized preparations for injection prepared in Examples 1 to 3 and Comparative Examples 1 to 4 and the reference preparation (Daiichi Sankyo Co., Ltd., QHS0165 batch) were measured, and the results are shown in Table 1.

[0062] Table 1

[0063]

[0064]

[0065] It can be seen from Table 1 that the pH, water content and impurity content of the indocyanine green lyophilized preparations for injection prepared in Examples 1 to 3 of the present invention are comparable to or even lower than those of the reference preparations.

[0066] Test Example 2

[0067] The indocyanine green lyophilized preparations for injection prepared in Examples 1 to 3 and Comparative Examples 1 to 4 and the reference preparation (Daiichi Sankyo Co., Ltd., QHS0165 batch) were subjected to accelerated tests and / or medium- and long-term stability studies, wherein the test results of the accelerated tests are shown in Table 2. The conditions for the accelerated test were: temperature 40°C ± 2°C, humidity 75% RH ± 5% RH, and the conditions for the long-term stability test were: temperature 25°C ± 2°C, humidity 60% RH ± 5% RH.

[0068] Table 2

[0069]

[0070]

[0071] As can be seen from Table 2, the indocyanine green freeze-dried preparation prepared by the freeze-drying process provided by the present invention has good stability during the accelerated experiment and / or the medium- and long-term stability investigation, the quality of the freeze-dried preparation has not changed significantly, and the moisture content of the prepared freeze-dried preparation is significantly lower than that of the reference preparation, the total impurity content is lower, and the purity is higher.

[0072] What is described above is only a preferred specific implementation manner of the present invention, but the protection scope of the present invention is not limited thereto. Any technician familiar with the technical field can make equivalent replacements or changes according to the technical scheme and inventive concept of the present invention within the technical scope disclosed by the present invention, which should be covered by the protection scope of the present invention.

Claims

1. A freeze-drying process for indocyanine green for injection, characterized in that the steps include: S1, pre-freezing: placing the indocyanine green for injection in a freeze dryer, freezing to -45 to -35°C at a cooling rate of 0.8 to 1.2°C / min, then keeping warm for freeze-drying, tempering to a temperature 8 to 12°C higher than the eutectic point temperature of the indocyanine green for injection, and continuing to keep warm for 110 to 130 minutes, and then freezing to -45 to -35°C at a cooling rate of 0.8 to 1.2°C / min, and then keeping warm for freeze-drying, completing the pre-freezing; S2, sublimation drying: heating the pre-frozen indocyanine green for injection to 0-5°C at a rate of 0.5-1.5°C / min and then keeping warm; S3. analytical drying: heating the indocyanine green for injection after sublimation drying to 15-25° C. at a rate of 0.8-1.2° C. / min and then keeping the temperature to obtain a lyophilized indocyanine green for injection.

2. The freeze-drying process of indocyanine green for injection according to claim 1, characterized in that: In step S1, before tempering, freeze-drying is performed at -45 to -35°C for 75 to 85 minutes.

3. The freeze-drying process of indocyanine green for injection according to claim 1, characterized in that: In step S1, after the tempering, the product is freeze-dried at -45 to -35°C for 85 to 95 minutes.

4. The freeze-drying process of indocyanine green for injection according to any one of claims 1 to 3, characterized in that: The tempering is to increase the temperature to -18 to -12°C at a rate of 0.8 to 1.2°C / min.

5. The freeze-drying process of indocyanine green for injection according to claim 1, characterized in that: During the sublimation drying, the vacuum degree in the freeze dryer is controlled to be 15±5Pa.

6. The freeze-drying process of indocyanine green for injection according to claim 1, characterized in that: The heat preservation time of the sublimation drying step is 5 to 7 hours.

7. The freeze-drying process of indocyanine green for injection according to claim 1, characterized in that: During the analytical drying, the vacuum degree in the freeze dryer is controlled to be 15±5Pa.

8. The freeze-drying process of indocyanine green for injection according to claim 1, characterized in that: The heat preservation time of the analytical drying step is 1 to 3 hours.

9. A freeze-dried indocyanine green preparation for injection, characterized in that: The product is prepared by the freeze-drying process described in any one of claims 1 to 8.

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