A lyophilization process for indocyanine green for injection

Abstract

The application belongs to the technical field of pharmaceutical preparation, 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, freezing at a rapid cooling rate to-45 to-35 DEG C, and then freeze-drying after heat preservation; tempering to a temperature higher than the eutectic point of the indocyanine green for injection, and then continuing heat preservation; freezing at a rapid cooling rate to-45 to-35 DEG C again, and then freeze-drying after heat preservation, to complete pre-freezing; slowly heating the indocyanine green for injection after pre-freezing to 0 to 5 DEG C, and then heat preserving; and heating at a rate of 0.8 to 1.2 DEG C / min to 15 to 25 DEG C, and then heat preserving, to obtain the freeze-dried preparation of the indocyanine green for injection. The freeze-drying process can reduce the water content in the freeze-dried preparation of the indocyanine green for injection to less than 2.0% within a short time, and the content of degradation impurities is less than 2.0%, so that the stability of the product is significantly improved; and the freeze-drying process is convenient to operate, takes less time, and has greater economic benefits.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of pharmaceutical preparation, and particularly relates to a lyophilization process of indocyanine green for injection. BACKGROUND

[0002] Indocyanine dyes are very important near-infrared dyes, and most of the synthesized and applied cyanine dyes are indocyanine dyes. Compared with other heterocyanine dyes, indocyanine dyes have high fluorescence quantum yield, large molar extinction coefficient, no fluorescence background, good fluorescence performance, and adjustable absorption wavelength, and have excellent optical properties, and have relatively high chemical stability and light stability, and are widely used in tumor targeted therapy, protein labeling, and trace metal ion detection.

[0003] Indocyanine green (ICG) is a water-soluble tricarbocyanine dye, which was developed by Kodak Research Laboratories in 1955, and was approved by the U.S. Food and Drug Administration for medical diagnosis in 1956. With the continuous iteration and update of diagnostic instruments, indocyanine green and other cyanine dyes combined with probes are being applied to more extensive medical and biological fields. As the first wave of indocyanine dyes developed for tumor and angiography, the quality standards (U.S. Pharmacopoeia and Chinese Pharmacopoeia) of the drug substance and indocyanine green for injection do not collect explicit process impurities and degradation impurities, and there is no explicit impurity detection report in the related literature. However, the process impurities and degradation impurities in the drug will directly affect the effectiveness and safety of the drug. Related literature has reported that the decomposition products of ICG have cytotoxicity after singlet oxygen decomposition or photodecomposition of ICG. Since the dose and use conditions of ICG during surgery are different, the accurate safety threshold of ICG application has not been determined. Therefore, the research on ICG degradation products is beneficial to the research on the related side effects of ICG in clinic, and ensures more scientific use of the drug.

[0004] In order to make the preparation of indocyanine green for injection as stable as possible during storage, the preparation is stored in the form of a lyophilized preparation, so as to ensure the stability. However, the current lyophilization process has the problems of long time consumption and complexity, and the prepared lyophilized preparation has high degradation impurities and uneven crystal size. SUMMARY

[0005] In view of this, the application provides a lyophilization process of indocyanine green for injection, which can reduce the water content in the lyophilized preparation of indocyanine green for injection to below 2.0% within a short time, and the content of degradation impurities is less than 2.0%, which significantly improves the stability of the product, and the lyophilization process is convenient to operate, takes shorter time, and has greater economic benefits.

[0006] To solve the above technical problems, the first aspect of the present application provides a lyophilization 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 at a cooling rate of 0.8-1.2 ℃ / min to-45 to-35 ℃, and then freeze-drying after holding, tempering to 8-12 ℃ higher than the eutectic point temperature of the indocyanine green for injection, and continuing to hold for 110-130 min, and then freeze-drying after freezing at a cooling rate of 0.8-1.2 ℃ / min to-45 to-35 ℃ and holding, completing pre-freezing;

[0008] S2, sublimation drying: warming the indocyanine green for injection after pre-freezing to 0-5 ℃ at a rate of 0.5-1.5 ℃ / min, and holding;

[0009] S3, desorption drying: warming the indocyanine green for injection after sublimation drying to 15-25 ℃ at a rate of 0.8-1.2 ℃ / min, and holding, to obtain a lyophilized preparation of indocyanine green for injection.

[0010] The lyophilization process of indocyanine green for injection provided by the present application adopts rapid cooling in the pre-freezing process, quickly freezes the indocyanine green to form fine lyophilized crystals, and then appropriately warms and tempers to promote the recrystallization of the indocyanine green. When the lyophilization temperature is higher than the eutectic point temperature of the indocyanine green, the ice crystal molecules with smaller volume can migrate to the ice crystal molecules with larger volume, thereby providing more sublimation channels for the internal water molecules, improving the sublimation efficiency, and shortening the lyophilization time. After tempering for a period of time, the sample in the glass state tends to form a more thermodynamically stable state, and more crystalline forms are formed.

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

[0012] Preferably, before the tempering, the freeze-drying is performed at-40 ℃ for 80 min.

[0013] In combination with the first aspect, after the tempering in step S1, the freeze-drying is performed at-45 to-35 ℃ for 85-95 min. Rapid cooling after tempering can make the sample in the glass state form a more thermodynamically stable state, and more crystalline forms are formed.

[0014] Preferably, after the tempering, the freeze-drying is performed at-40 ℃ for 90 min.

[0015] In combination with the first aspect, the tempering is heating at a rate of 0.8-1.2℃ / min to -18 to -12℃, which is about 10℃ higher than the eutectic point of indocyanine green, so that the crystal size is more uniform.

[0016] Preferably, the tempering is heating at a rate of 1.0℃ / min to -15℃ and holding for 120min.

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

[0018] In combination with the first aspect, the holding time of the sublimation drying step is 5-7h, preferably 6h.

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

[0020] In combination with the first aspect, the holding time of the desorption drying step is 1-3h, preferably 2h.

[0021] The vacuum degree and holding time in the above drying process can make water molecules sublimate as much as possible, so as to reduce the water content in the obtained lyophilized preparation as much as possible, thereby providing guarantee for the stability of the product.

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

[0023] The indocyanine green lyophilization process provided by the present application can obtain an indocyanine green lyophilized preparation for injection with a water content and total impurity content equivalent to or even lower than those of a reference preparation (water content <2.0%, total impurity content <2.0%) through the pre-freezing, sublimation drying and desorption drying processes with specific cooling or heating rates. Moreover, the lyophilization process is simple in steps and convenient to operate, which is conducive to the realization of large-scale application. DETAILED DESCRIPTION

[0024] In order to make the purpose, technical solutions and advantages of the present application clearer, the present application will be further described in detail below in combination with specific embodiments. It should be understood that the specific embodiments described herein are only used to explain the present application and do not limit the present application.

[0025] The indocyanine green for injection is a sterile lyophilized product of indocyanine green, which is a dark green loose solid, and is easily decomposed by heat and moisture, and the decomposition product has cytotoxicity, which brings side effects to the user. Therefore, the lower the water content in the lyophilized preparation, the more stable it is during storage, the less impurities are generated by decomposition, and the smaller the side effects when used. However, the current lyophilization process has the problems of long process time and complex operation, and the water content in the lyophilized preparation is high, the crystal size is uneven, which leads to high degradation impurities and difficult to guarantee the stability. In view of this, the present application provides a lyophilization process for indocyanine green for injection, and the lyophilized preparation obtained by using the lyophilization process has uniform drug crystal size, lower water content, shorter time, fewer degradation impurities, and more stable product.

[0026] In the following examples and comparative examples, the indocyanine green for injection is prepared according to the following method: 10g of indocyanine green is weighed in water for injection, stirred for 30-60min, and after complete dissolution, the volume is adjusted to 1000mL with water for injection, and 400 preparations are filled into 2.5mL / branch.

[0027] Example 1

[0028] The present embodiment provides a lyophilization process for indocyanine green for injection, and the preparation steps include:

[0029] S1, pre-freezing: the filled indocyanine green for injection is moved into the freeze dryer after half-casing, first frozen to-40℃ at a cooling rate of 1℃ / min, and kept for 80min; then warmed to-15℃ at a rate of 1℃ / min, and kept for 120min; continue to freeze to-40℃ at a cooling rate of 1℃ / min, and keep for 90min, complete the pre-freezing.

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

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

[0032] Example 2

[0033] The present embodiment provides a lyophilization process for indocyanine green for injection, and the preparation steps include:

[0034] S1, pre-freezing: the filled indocyanine green for injection was moved into the freeze dryer after half-casing, first frozen to -45℃ at a cooling rate of 1.2℃ / min, kept for 75min; then warmed to -12℃ at a rate of 0.8℃ / min, kept for 125min; continue to freeze to -45℃ at a cooling rate of 1.2℃ / min, keep for 85min, complete the pre-freezing.

[0035] S2, sublimation drying: warm to 0℃ at a rate of 1℃ / min, control the vacuum degree to be 18±2Pa, keep for 5h.

[0036] S3, analytical drying: warm to 15℃ at a rate of 0.8℃ / min, control the vacuum degree to be 18±2Pa, keep for 3h, get the indocyanine green for injection freeze-dried preparation.

[0037] Example 3

[0038] The example provides a freeze-drying process of indocyanine green for injection, the preparation steps comprising:

[0039] S1, pre-freezing: the filled indocyanine green for injection was moved into the freeze dryer after half-casing, first frozen to -35℃ at a cooling rate of 0.8℃ / min, kept for 85min; then warmed to -18℃ at a rate of 1.2℃ / min, kept for 115min; continue to freeze to -35℃ at a cooling rate of 0.8℃ / min, keep for 95min, complete the pre-freezing.

[0040] S2, sublimation drying: warm to 5℃ at a rate of 1.5℃ / min, control the vacuum degree to be 12±2Pa, keep for 7h.

[0041] S3, analytical drying: warm to 25℃ at a rate of 1.2℃ / min, control the vacuum degree to be 12±2Pa, keep for 1h, get the indocyanine green for injection freeze-dried preparation.

[0042] Comparative example 1

[0043] The comparative example provides a freeze-drying process of indocyanine green for injection, the preparation steps comprising:

[0044] S1, pre-freezing: the filled indocyanine green for injection was moved into the freeze dryer after half-casing, first frozen to -40℃ at a cooling rate of 0.16℃ / min, kept for 80min; then warmed to -15℃ at a rate of 1℃ / min, kept for 120min; continue to freeze to -40℃ at a cooling rate of 0.16℃ / min, keep for 90min, complete the pre-freezing.

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

[0046] Comparative example 2

[0047] The present comparative example provides a lyophilization process of indocyanine green for injection, the preparation steps comprising:

[0048] S1, pre-freezing: the filled indocyanine green for injection was moved into the freeze dryer after half-casing, first frozen to -40℃ at a cooling rate of 1℃ / min, and kept for 80 min; then warmed to -15℃ at a rate of 1℃ / min, and kept for 120 min, to complete the pre-freezing.

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

[0050] Comparative Example 3

[0051] The present comparative example provides a lyophilization process of indocyanine green for injection, the preparation steps comprising:

[0052] S1, the same as step S1 of Example 1.

[0053] S2, the same as step S2 of Example 1.

[0054] S3, desiccation: warmed to 20℃ at a rate of 0.3℃ / min, controlled the vacuum degree to be 15±2 Pa, and kept for 2 h, to obtain the lyophilized preparation of indocyanine green for injection.

[0055] Comparative Example 4

[0056] The present comparative example provides a lyophilization process of indocyanine green for injection, the preparation steps comprising:

[0057] S1, the same as step S1 of Example 1.

[0058] S2, the same as step S2 of Example 1.

[0059] S3, desiccation: warmed to 40℃ at a rate of 1℃ / min, controlled the vacuum degree to be 15±2 Pa, and kept for 2 h, to obtain the lyophilized preparation of indocyanine green for injection.

[0060] Test Example 1

[0061] The lyophilized preparations of indocyanine green for injection prepared in Examples 1-3 and Comparative Examples 1-4, and a reference preparation (QHS0165 batch, Japan's first three companies) were respectively determined for pH, moisture content and impurity content, and the determination results are shown in Table 1.

[0062] Table 1

[0063]

[0064]

[0065] As shown in Table 1, the pH, moisture content and impurity content of the indocyanine green freeze-dried preparations prepared in Examples 1-3 are equivalent to or even lower than those of the reference preparation.

[0066] Test Example 2

[0067] The accelerated test and / or long-term stability test were performed on the indocyanine green freeze-dried preparations prepared in Examples 1-3 and Comparative Examples 1-4, and the reference preparation (QHS0165 batch, Sanwa Kagaku Co., Ltd.). The test results of the accelerated test are shown in Table 2. The accelerated test condition was 40℃±2℃ and 75%RH±5%RH, and the long-term stability test condition was 25℃±2℃ and 60%RH±5%RH.

[0068] Table 2

[0069]

[0070]

[0071] As shown in Table 2, the indocyanine green freeze-dried preparations prepared by the freeze-drying process provided in the present application have good stability during the accelerated test and / or long-term stability test, and the quality of the freeze-dried preparations does not change significantly. The moisture content of the prepared freeze-dried preparations is significantly lower than that of the reference preparation, and the total impurity content is lower and the purity is higher.

[0072] The above description is merely preferred embodiments of the present application, but the protection scope of the present application is not limited to this. Any person skilled in the art, according to the technical range disclosed in the present application and the inventive concept, can make equivalent replacements or changes, which should be covered in the protection scope of the present application.

Claims

1. A lyophilization process for indocyanine green for injection, characterized in that the steps of Comprise: S1, pre-freezing: the injection indocyanine green is placed in the freeze dryer, frozen to -45 to -35℃ at a cooling rate of 0.8 to 1.2℃ / min, and then kept for freeze drying for 75 to 85 min, tempered to 8 to 12℃ higher than the eutectic point temperature of the injection indocyanine green and kept for 110 to 130 min, then frozen to -45 to -35℃ at a cooling rate of 0.8 to 1.2℃ / min and kept for freeze drying for 85 to 95 min, and the pre-freezing is completed; S2, sublimation drying: the injection indocyanine green after pre-freezing is warmed to 0 to 5℃ at a rate of 0.5 to 1.5℃ / min and kept for 5 to 7h, and the vacuum degree is controlled to be 15±5Pa; S3, analytical drying: the injection indocyanine green after sublimation drying is warmed to 15 to 25℃ at a rate of 0.8 to 1.2℃ / min and kept for 1 to 3h, the vacuum degree is controlled to be 15±5Pa, and the injection indocyanine green freeze-dried preparation is obtained.

2. The lyophilization process of indocyanine green for injection as claimed in claim 1, wherein, The tempering is warming to -18 to -12℃ at a rate of 0.8 to 1.2℃ / min.

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