Carbetocin injection and preparation method thereof

Abstract

The invention relates to a carbetocin injection and a preparation method thereof, and relates to the technical field of pharmaceutical preparations, the carbetocin injection is added with a stabilizing auxiliary agent, based on 10L of the carbetocin injection, the adding amount of the stabilizing auxiliary agent is 0.25-1.0 g, the adding amount of the stabilizing auxiliary agent is 0.1-1.0 g, and the adding amount of the stabilizing auxiliary agent is 0.1-1.0 g. The stabilizing aid is selected from at least one of apigenin, catechin, luteolin, troxerutin, puerarin and baicalin. It is found that by introducing a proper amount of specific natural substances such as puerarin into the carbetocin injection as the stabilizing additive, the increase of the impurity content in the storage process of the carbetocin injection can be effectively slowed down, and the variety and dosage of auxiliary materials in a prescription can be reduced, so that the storage stability of the carbetocin injection is improved; the risk of increasing the microbial level and the risk of compatibility with raw materials caused by introduction of excessive auxiliary materials are also avoided, so that the medication safety is improved.

Description

Technical Field

[0001] The present invention relates to the technical field of pharmaceutical preparations, and particularly to a carbetocin injection and a preparation method thereof. Background Art

[0002] Carbetocin, chemically named deamino-2-oxo-methyltyrosine-1-κ-oxytocin, has significant long-acting characteristics. It can be administered intravenously as a single dose immediately after cesarean section under epidural or spinal anesthesia and after vaginal delivery, effectively preventing uterine hypotonia and postpartum hemorrhage, and greatly facilitating clinical use. Due to the convenient use, rapid onset, and long half-life of its injection dosage form, it has been widely used clinically.

[0003] The structural formula of carbetocin is as follows: 。

[0004] However, as a synthetic polypeptide drug, carbetocin poses certain challenges in terms of stability. Especially when stored for a long time in an environment with high temperature, oxidation, and high microbial levels, the peptide bonds of carbetocin are prone to reactions such as cleavage and condensation, resulting in an increase in degradation impurities and affecting its drug formation and clinical efficacy. Therefore, it is necessary to improve its stability through fine formulation design and pharmaceutical process optimization to ensure the quality of the drug during storage and use.

[0005] In addition, carbetocin cannot tolerate high temperature for a long time, so it is not suitable to be sterilized by terminal sterilization process. At the same time, no obvious antibacterial property has been found in the aqueous solution of carbetocin, which makes the risk of microbial contamination higher during the production process, thus bringing potential hidden dangers to clinical use. How to effectively control microbial contamination and ensure the sterility of the drug is a key problem to be solved in the preparation process of carbetocin injection.

[0006] On the other hand, puerarin and the like, as widely distributed reducing secondary metabolites in plants, have various biological activities, such as immunity and metabolic regulation, anti-inflammatory, antibacterial, antiviral, antioxidant, etc. Moreover, compared with synthetic drugs, they have a wider range of indications, fewer side effects, and better tolerance.

[0007] At present, injection products such as puerarin injection have been industrialized and used clinically for a long time, and their clinical safety has been effectively confirmed. However, there is no relevant report in the prior art on adding natural substances such as puerarin as components to carbetocin injection. Summary of the Invention

[0008] To solve the above problems, the present invention provides a carbetocin injection and its preparation method. It is first discovered that applying an appropriate amount of specific natural substances such as puerarin as a stabilizing adjuvant in the preparation of carbetocin injection can effectively slow down the increase in the impurity content during the storage of carbetocin injection, so as to improve the storage stability of carbetocin injection.

[0009] In the first aspect, the present invention provides a carbetocin injection, which contains carbetocin, a stabilizing adjuvant and water for injection; Based on 10 L of the carbetocin injection, the addition amount of the stabilizing adjuvant is 0.25 g to 1.0 g; The stabilizing adjuvant is selected from at least one of apigenin, catechin, luteolin, troxerutin, puerarin and baicalin.

[0010] Further, based on 10 L of the carbetocin injection, the addition amount of the stabilizing adjuvant is 0.25 g to 0.5 g; the stabilizing adjuvant is puerarin.

[0011] Further, the carbetocin injection is composed of carbetocin, the stabilizing adjuvant, an osmotic pressure regulator, a pH regulator and water for injection; Based on 10 L of the carbetocin injection, the addition amount of carbetocin is 0.9 g to 1.1 g; Based on 10 L of the carbetocin injection, the addition amount of the osmotic pressure regulator is 88 g to 92 g; The dosage of the pH regulator is such that the pH value of the carbetocin injection is 4.0 to 6.5; The dissolved oxygen content of the water for injection ≤ 3.0 mg / L.

[0012] Further, based on 10 L of the carbetocin injection, the addition amount of carbetocin is 1.0 g; based on 10 L of the carbetocin injection, the addition amount of the osmotic pressure regulator is 90 g; the dosage of the pH regulator is such that the pH value of the carbetocin injection is 5.0 to 6.0; the dissolved oxygen content of the water for injection ≤ 1.0 mg / L.

[0013] Further, the osmotic pressure regulator includes sodium chloride.

[0014] Further, the pH regulator includes at least one of sodium hydroxide, triethylamine, hydrochloric acid and glacial acetic acid.

[0015] In the second aspect, based on the same inventive concept, the present invention provides a preparation method of the carbetocin injection according to any one of the first aspect, and the preparation method includes the following steps: Introduce a protective gas into water for injection to obtain water for injection with dissolved oxygen content ≤ 3.0 mg / L; Add other components except the pH regulator to the water for injection with dissolved oxygen content ≤ 3.0 mg / L for dissolution, then make up the volume to 95 vol% of the formulated volume, adjust the pH of the system to 4.0 - 6.5 with the pH regulator, and then make up the volume to the full amount to obtain the carbetocin injection.

[0016] Further, in terms of volume percentage, the protective gas is a sterile inert gas with purity > 90.0%, and the sterile inert gas includes at least one of nitrogen and helium.

[0017] Further, the preparation method further includes: Subject the carbetocin injection to sterile filtration and perform sealing under an inert gas atmosphere, and then perform leak detection and lamp inspection to obtain the finished product of carbetocin injection.

[0018] Further, in terms of volume percentage, the residual oxygen content in the reagent bottle containing the carbetocin injection after sealing is ≤ 5%, preferably ≤ 2%; the sterile filtration is carried out using two - stage 0.22 μm filters, and the filter material includes at least one of polyethersulfone, polyvinylidene fluoride, and polytetrafluoroethylene.

[0019] The above - mentioned technical solutions provided by the embodiments of the present invention have at least the following advantages compared with the prior art: The embodiments of the present invention provide a carbetocin injection and its preparation method. The present invention discovers that by introducing an appropriate amount of specific natural substances such as puerarin as a stabilizing adjuvant into the carbetocin injection, it can effectively slow down the increase in the content of impurities such as 9 impurity Gly - OH, impurity sulfoxide I, impurity sulfoxide II, impurity D - Asn 5 and other impurities, and reduce the types and dosages of excipients in the prescription. This not only improves the storage stability of the carbetocin injection, but also avoids the risks of increased microbial levels and risks of incompatibility with raw materials brought about by excessive introduction of excipients, thereby enhancing the drug safety of its use. Detailed implementation manners

[0020] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are some, rather than all, of the embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts shall fall within the protection scope of the present invention.

[0021] Unless otherwise specified, various raw materials, reagents, instruments, and equipment used in the present invention can be obtained through market purchases or can be prepared by existing methods.

[0022] In a first aspect, the present invention provides a carbetocin injection, and the carbetocin injection contains carbetocin, a stabilizing adjuvant, and water for injection; Based on 10 L of the carbetocin injection, the addition amount of the stabilizing adjuvant is 0.25 g to 1.0 g; The stabilizing adjuvant is selected from at least one of apigenin, catechin, luteolin, troxerutin, puerarin, and baicalin.

[0023] An embodiment of the present invention provides a carbetocin injection. The present invention discovers that by introducing an appropriate amount of specific natural substances such as puerarin as a stabilizing adjuvant into the carbetocin injection, it can effectively slow down the increase in the content of impurities such as impurity Gly 9 -OH, impurity sulfoxide I, impurity sulfoxide II, impurity D-Asn 5 and other impurities, and reduce the types and dosages of excipients in the formulation, which not only improves the storage stability of the carbetocin injection, but also avoids the risks of increased microbial levels and compatibility with raw materials brought about by excessive introduction of excipients, thereby enhancing the safety of its use.

[0024] In the present invention, the CAS number of apigenin is 520-36-5, the CAS number of catechin is 7295-85-4, the CAS number of luteolin is 491-70-3, the CAS number of troxerutin is 7085-55-4, the CAS number of puerarin is 3681-99-0, and the CAS number of baicalin is 21967-41-9. Among the above natural substances, injection products such as troxerutin injection and puerarin injection are existing industrialized and long-term clinically used injection products, and their clinical safety has been effectively proven.

[0025] In some specific embodiments, based on 10 L of the carbetocin injection, the addition amount of the stabilizing adjuvant is 0.25 g to 0.5 g; the stabilizing adjuvant is puerarin.

[0026] In some specific embodiments, the carbetocin injection is composed of carbetocin, the stabilizing adjuvant, an osmotic pressure regulator, a pH regulator, and water for injection; Based on 10 L of the carbetocin injection, the addition amount of carbetocin is 0.9 g to 1.1 g; Based on 10 L of the carbetocin injection, the addition amount of the osmotic pressure regulator is 88 g to 92 g; The dosage of the pH regulator is such that the pH value of the carbetocin injection is 4.0 - 6.5; The dissolved oxygen content of the water for injection ≤ 3.0 mg / L.

[0027] After adding carbetocin, a stabilizing aid, an osmotic pressure regulator, and a pH regulator to the carbetocin injection provided by the present invention according to the above parameter requirements, the balance is water for injection.

[0028] In some specific embodiments, based on 10 L of the carbetocin injection, the addition amount of carbetocin is 1.0 g; based on 10 L of the carbetocin injection, the addition amount of the osmotic pressure regulator is 90 g; the dosage of the pH regulator is such that the pH value of the carbetocin injection is 5.0 - 6.0; the dissolved oxygen content of the water for injection ≤ 1.0 mg / L.

[0029] In some specific embodiments, the osmotic pressure regulator includes sodium chloride.

[0030] In some specific embodiments, the pH regulator includes at least one of sodium hydroxide, triethylamine, hydrochloric acid, and glacial acetic acid.

[0031] In a second aspect, based on the same inventive concept, the present invention provides a preparation method for the carbetocin injection according to any one of the first aspect, and the preparation method includes the following steps: Introduce a protective gas into the water for injection to obtain water for injection with a dissolved oxygen content ≤ 3.0 mg / L; Add other components except the pH regulator to the water for injection with a dissolved oxygen content ≤ 3.0 mg / L for dissolution, and then adjust the pH of the system to 4.0 - 6.5 with the pH regulator to obtain the carbetocin injection.

[0032] In the preparation method of the carbetocin injection provided by the present invention, by strictly controlling the oxygen content in the preparation process, the chemical stability and the sterile guarantee level of this product are further ensured. At the same time, this preparation method is implemented based on the carbetocin injection according to any one of the first aspect, so it has at least the beneficial effects described in any one of the first aspect, which will not be elaborated here.

[0033] In some specific embodiments, by volume percentage, the protective gas is a sterile inert gas with a purity > 90.0%, and the sterile inert gas includes at least one of nitrogen and helium.

[0034] In some specific embodiments, the preparation method further includes: Subject the carbetocin injection to sterile filtration and perform filling and sealing under an inert gas atmosphere, and then perform leak detection and lamp inspection to obtain the finished product of the carbetocin injection.

[0035] In some specific embodiments, in terms of volume percentage, the residual oxygen content in the reagent bottle containing carbetocin injection after potting is ≤5%, preferably ≤2%. In the present invention, the sterilized and filtered liquid medicine is potted into a medium-borosilicate glass ampoule bottle; during the potting process, the empty ampoule bottle is first purged and filled with a protective gas, and then the liquid medicine is potted into the ampoule bottle. The potting volume is 1-1.6 ml. Before sealing, an inert gas is used to fill the ampoule bottle containing the potted liquid medicine so that the residual oxygen content reaches a preset value; during the potting process, a laminar flow of the protective gas is maintained.

[0036] In some specific embodiments, the sterilization filtration is performed using a two-stage 0.22 μm filter, and the filter material includes at least one of polyethersulfone, polyvinylidene fluoride, and polytetrafluoroethylene.

[0037] The two-stage 0.22 μm filter in the present invention is a precision filtration device commonly used in liquid or gas purification. It is composed of two filtration units with a pore size of 0.22 microns connected in series, which can provide a higher level of filtration effect, and commercially available equipment can be directly used.

[0038] It should be noted that for the component raw materials involved in the carbetocin injection and its preparation method provided in the embodiments of the present invention, if there is no special limitation or specific description, commercially available products can be directly used or self-made using existing publicly disclosed preparation methods; at the same time, for the steps and parameters involved, if there is no special limitation or specific description, they can be carried out according to the existing processing technology of carbetocin injection or directly using existing equipment, and the present invention document will not elaborate one by one.

[0039] The present invention will be further described below in conjunction with specific embodiments. It should be understood that these embodiments are only used to illustrate the present invention and not to limit the scope of the present invention. The experimental methods without specific conditions noted in the following embodiments are usually determined according to national standards. If there is no corresponding national standard, they are carried out according to general international standards, conventional conditions, or conditions recommended by the manufacturer.

[0040] The impurity information in the following examples and comparative examples is as follows: Impurity Gly 9 -OH (or called [Gly 9 -OH] carbetocin) has the following chemical structure: .

[0041] The chemical structure of impurity sulfoxide I (or called carbetocin sulfoxide I) is as follows: .

[0042] The chemical structural formula of impurity sulfoxide II (or carbetocin sulfoxide II) is as follows: .

[0043] Impurity D-Asn 5 (or D-Asn 5 carbetocin) has the following chemical structural formula: .

[0044] The chemical structural formula of unknown impurity 1 is shown as follows: .

[0045] Example 1 This example provides a carbetocin injection, which is composed of carbetocin, a stabilizing adjuvant, an osmotic pressure regulator, a pH regulator, and water for injection; Based on 10 L of the carbetocin injection, the addition amount of carbetocin is 1.0 g; Based on 10 L of the carbetocin injection, the addition amount of the stabilizing adjuvant is 0.25 g, and the stabilizing adjuvant is troxerutin; Based on 10 L of the carbetocin injection, the addition amount of the osmotic pressure regulator is 90 g, and the osmotic pressure regulator is sodium chloride; The dosage of the pH regulator is such that the pH value of the carbetocin injection is 5.3.

[0046] The preparation method of the above carbetocin injection includes the following steps: Introduce a protective gas (nitrogen) into water for injection to obtain water for injection with a dissolved oxygen content of 1.48 mg / L; Add 0.25 g of troxerutin, 90 g of sodium chloride, and 1 g of carbetocin to the water for injection with a dissolved oxygen content of 1.48 mg / L, stir and dissolve, make up the water to 95 vol% of the preparation amount, then adjust the pH of the system to 5.3 with 1 mol / L glacial acetic acid and 2 mol / L sodium hydroxide, and make up the water to the full volume to obtain 10 L of the carbetocin injection; Filter the carbetocin injection through a secondary 0.22 μm PVDF filter element for sterilization and seal it under a nitrogen inert gas atmosphere. The filling volume is 1.2 ml / branch, and the residual oxygen content detection is 3.22%. Then, conduct leak detection and lamp inspection to obtain qualified finished products of the carbetocin injection.

[0047] On the basis of Example 1, Examples 2 to 6 provide a carbetocin injection and its preparation method. The differences between Examples 2 to 6 and Example 1 are only as follows: the types of stabilizing aids are different; among them, the stabilizing aid in Example 2 is baicalin, the stabilizing aid in Example 3 is puerarin, the stabilizing aid in Example 4 is apigenin, the stabilizing aid in Example 5 is catechin, and the stabilizing aid in Example 6 is luteolin.

[0048] On the basis of Example 1, Examples 7 to 8 provide a carbetocin injection and its preparation method. The differences between Examples 7 to 8 and Example 1 are only as follows: the dosages of the stabilizing aid are different; among them, based on 10 L of the carbetocin injection, the addition amount of the stabilizing aid in Example 7 is 0.5 g (that is, the addition amount of the stabilizing aid in the preparation process of Example 1 is increased from 0.25 g to 0.5 g), and based on 10 L of the carbetocin injection, the addition amount of the stabilizing aid in Example 8 is 1.0 g (that is, the addition amount of the stabilizing aid in the preparation process of Example 1 is increased from 0.25 g to 1.0 g).

[0049] Comparative Example 1 This example provides a carbetocin injection and its preparation method. The difference from Example 1 is only as follows: no stabilizing aid is added (that is, the addition amount of the stabilizing aid in the preparation process of Example 1 is adjusted from 0.25 g to 0 g); the remaining steps and parameters are the same.

[0050] Comparative Example 2 This example provides a carbetocin injection and its preparation method. The difference from Example 1 is only as follows: based on 10 L of the carbetocin injection, the addition amount of the stabilizing aid is 0.1 g (that is, the addition amount of the stabilizing aid in the preparation process of Example 1 is adjusted from 0.25 g to 0.1 g); the remaining steps and parameters are the same.

[0051] Comparative Example 3 This example provides a carbetocin injection and its preparation method. The difference from Example 1 is only as follows: the stabilizing aid is adjusted to bamboo leaf flavonoids (that is, troxerutin in the example is replaced by bamboo leaf flavonoids); the remaining steps and parameters are the same.

[0052] Comparative Example 4 This example provides a reported carbetocin injection, and its preparation method includes the following steps: Add 10 g of methionine, 470 g of mannitol, 11.9 g of succinic acid, and 1 g of carbetocin to 10 L of the water for injection without filling protective gas (nitrogen), stir and dissolve, and then adjust the pH of the system to 5.3 with 1 mol / L glacial acetic acid and 2 mol / L sodium hydroxide to obtain the carbetocin injection; The carbetocin injection is sealed through a secondary 0.22 μm PVDF filter element, with a filling volume of 1.2 ml per vial. After sealing, leak detection and lamp inspection are carried out to obtain carbetocin injection samples.

[0053] Comparative Example 5 This example provides a carbetocin injection and its preparation method, which is only different from Example 1 in that: no protective gas is used to control the dissolved oxygen content and residual oxygen content; the remaining steps and parameters are the same.

[0054] Test Example In this example, the carbetocin injection samples obtained from the above examples and comparative examples are subjected to storage stability investigation for 6 months under the conditions of 40°C ± 2°C and relative humidity of 75% ± 5%. At the investigation time points of 0 month (i.e., when the experiment starts), 1 month, 3 months, and 6 months, high performance liquid chromatography is used to detect the impurity content. The detection results are shown in Table 1, Table 2, and Table 3; among them, Table 1 and Table 2 show the comparative results of the influence of different types of stabilizing agents on the storage stability of carbetocin injection (the stabilizing agent in Example 1 is troxerutin, the stabilizing agent in Example 2 is baicalin, the stabilizing agent in Example 3 is puerarin, the stabilizing agent in Example 4 is apigenin, the stabilizing agent in Example 5 is catechin, the stabilizing agent in Example 6 is luteolin, and the stabilizing agent in Comparative Example 3 is bamboo leaf flavonoid; Comparative Example 4 is a reported carbetocin injection), and Table 3 shows the comparative results of the influence of different addition amounts of stabilizing agents on the storage stability of carbetocin injection (the stabilizing agent in Example 7 is added at 0.5 g, the stabilizing agent in Example 8 is added at 1 g, the stabilizing agent in Comparative Example 1 is added at 0 g, and the stabilizing agent in Comparative Example 2 is added at 0.1 g); Table 4 shows the comparative results of the influence of protective gas on the storage stability of carbetocin injection (nitrogen is used as the protective gas in Example 1, Comparative Example 4 is a reported carbetocin injection, and Comparative Example 5 is without using protective gas).

[0055] Table 1 Comparative Results of the Influence of Different Types of Stabilizing Agents on the Storage Stability of Carbetocin Injection - I Table 2 Comparative Results of the Influence of Different Types of Stabilizing Agents on the Storage Stability of Carbetocin Injection - II Table 3 Comparative Results of the Influence of Different Addition Amounts of Stabilizing Agents on the Storage Stability of Carbetocin Injection Table 4 Comparative Results of the Influence of Protective Gas on the Storage Stability of Carbetocin Injection As can be seen from Table 1, Table 2, Table 3 and Table 4: 1) Different types of stabilizing agents with the same addition amount have different stabilizing effects on carbetocin injection. Among them, in Example 3, when the stabilizer is puerarin, the stabilizing effect is better than that of other types of stabilizers. The total impurity growth in Examples 1 - 6 is less than that in Comparative Example 3 and Comparative Example 4. When the stabilizers are troxerutin, baicalin, puerarin, apigenin, catechin, and luteolin, they have good stabilizing effects.

[0056] 2) When the dosage of the stabilizer increases from 0.25 g / 10 L to 0.5 g / 10 L, the growth of the total impurities in 6 - month stability decreases from 1.14% to 0.80%. When it continues to increase to 1.0 g / 10 L, the growth of the total impurities in 6 - month stability is 0.77%, showing no obvious change compared with 0.5 g / 10 L. When the dosage of the stabilizer is reduced to 0.1 g / 10 L, the growth of the total impurities in 6 - month stability is 2.55%, which is significantly increased compared with Example 1. When the dosage of the stabilizer continues to be reduced to 0 g / 10 L, the growth of the total impurities in 6 - month stability is 3.11%. When the addition amount of the stabilizer is in the range of 0.25 g - 1.0 g, it has good stability.

[0057] 3) When no protective gas is added to control dissolved oxygen and residual oxygen, sulfoxide I and sulfoxide II increase significantly. In 6 - month stability, the growth of the total impurities is 4.94%, which is significantly increased compared with Example 1 and Comparative Example 4. Therefore, adding a protective gas to control dissolved oxygen and residual oxygen can effectively improve the stability of this product.

[0058] In addition, in the present invention, the minimum clinical dose of troxerutin is 240 mg / time, and the maximum addition amount in this experiment is 0.1 mg, which is 1 / 2400 of the minimum clinical use dose. Its dosage is lower than 1 / 1000 of the minimum dose required to evaluate drug cross - contamination, and the dosage is safe with no obvious medication risk in clinical use. And to further confirm the safety of carbetocin injection after adding the stabilizer, a rabbit vascular irritation test, a guinea pig systemic active anaphylaxis test, a rat passive cutaneous anaphylaxis test, and a hemolysis test were carried out using the carbetocin injection with puerarin as the stabilizer in Example 3. The test results showed no irritation, no anaphylaxis, and no hemolysis, indicating that the carbetocin injection prepared by the present invention has safety.

[0059] (1) Rabbit vascular irritation test Seven rabbits that passed quarantine and adaptability observation were selected. First, the two lightest rabbits (1 female and 1 male) were eliminated. Then, 5 rabbits with remaining body weights of 2.2669 - 2.7233 kg for females and 2.4232 - 2.7370 kg for males were selected, including 3 females and 2 males, with no pregnancy in females. They were sorted by body weight from light to heavy by gender. The study used the self - comparison method on the left and right sides of the same body, that is, the test article or the marketed preparation at a concentration of 100 μg / ml was injected into the left ear marginal vein of the rabbit, and an equal volume of negative control (i.e., 0.9% sodium chloride injection) was injected into the right ear marginal vein. The drug was administered once a day for 7 consecutive days, and whether there were irritant reactions in the blood vessels at the administration site was observed.

[0060] After 96 hours of general condition observation after the last administration, 3 rabbits (2 females and 1 male) were euthanized by carbon dioxide. The blood vessels at the administration site and the surrounding tissues were taken for histopathological examination. The remaining 2 rabbits were observed for 14 days for recovery. After the observation, macroscopic observation and histopathological examination of the administration site were carried out in the same way as described above to understand the reversibility of the stimulant reaction.

[0061] Table 5 Results of rabbit vascular irritation test No irritant reactions were found in the experimental group during the administration period, after the last administration, and during the recovery period. The results of the rabbit vascular irritation test of this product showed no vascular irritation.

[0062] (2) Guinea pig active systemic anaphylaxis test Thirty - three SPF - grade guinea pigs that passed quarantine and adaptability observation were selected, including 18 females and 15 males, with no pregnancy in females. First, the lightest (3 females and 2 males) and the heaviest (1 male) animals were selected as reserve animals. The remaining 27 animals, with body weights of 335.4 - 376.9 g for females and 341.6 - 378.6 g for males, were sorted by body weight from small to large by gender and divided into a negative control group, a positive control group, and a test article group according to the randomized block method, with 3 groups in total, 9 animals in each group, including 5 females and 4 males. The test article group was intraperitoneally injected with the test article at a concentration of 100 μg / ml × 0.5 ml / animal for sensitization, the positive control group was sensitized with ovalbumin at a concentration of 5 mg / ml × 0.5 ml / animal, and the negative control group was sensitized with 0.5 ml / animal of 0.9% sodium chloride injection. Sensitization was carried out once every other day for 3 consecutive times. Three animals (2 females and 1 male) from each group were challenged by a one - time rapid intravenous injection with 2 - fold volume of the sensitization dose on the 14th day after the last sensitization injection; if the challenge result of the test article group was negative on the 14th day after the last sensitization, the remaining animals were challenged on the 21st day after the last sensitization injection; if the challenge result of the test article group was positive on the 14th day after the last sensitization, all 9 animals in each group were challenged on the 14th day after the last sensitization injection. The allergic reaction symptoms of the guinea pigs were observed, and the degree of allergic reaction was judged and the incidence of allergic reaction was calculated.

[0063] Table 6 Results of body weight examination of guinea pigs in each test substance group during sensitization and challenge Table 7 Results of systemic anaphylaxis test after challenge on the 14th day after the last sensitization Table 8 Results of systemic anaphylaxis test after challenge on the 21st day after the last sensitization Conclusion: The results showed that: ① During the sensitization period, no obvious abnormalities were observed in the general conditions of guinea pigs in each group. ② Compared with the negative control group, there were no significant differences in the body weights of guinea pigs in the test substance group during the three sensitizations and at the challenges on the 14th and 21st days after the last sensitization (p>0.05). ③ No anaphylactic reaction symptoms occurred within 3 hours after the challenges on the 14th and 21st days after the last sensitization in the negative control group and the test substance group, and their anaphylactic reaction incidences were both 0%, and the anaphylactic reactions were all judged as negative. The positive control group immediately showed anaphylactic reaction symptoms after the challenge, and its anaphylactic reaction incidence was 100%, and the anaphylactic reaction was judged as extremely strongly positive. The results of the guinea pig systemic active anaphylaxis test of this product showed no active anaphylaxis.

[0064] (3) Rat passive cutaneous anaphylaxis test Fourteen rats that passed quarantine and adaptation observation were selected, with 7 males and 7 females, and the females were not pregnant. First, the animals with the lightest (1 female) and heaviest (1 male) weights were eliminated as spare animals. The remaining 12 rats, with female weights ranging from 181.4 to 194.3 g and male weights ranging from 206.5 to 218.6 g, were sorted by weight from lightest to heaviest by sex and divided into a negative control group, a positive control group, and a test article group, a total of 3 groups, with 4 rats in each group. The rats in the test article group were sensitized by intraperitoneal injection of 100 μg / ml × 0.5 ml / rat of the test article. The rats in the positive control group were sensitized by intraperitoneal injection of 5 mg / ml × 0.5 ml of ovalbumin plus 0.5 ml of 4% aluminum hydroxide gel. The rats in the negative control group were sensitized by intraperitoneal injection of 0.5 ml / rat of 0.9% sodium chloride injection. Sensitization was performed once every other day for 3 consecutive times. Sensitized serum was prepared on the 14th day after the last sensitization. Another 22 SPF-grade SD rats were selected, with 11 males and 11 females, and the females were not pregnant. First, the animals with the heaviest weights (2 females and 2 males) were eliminated as spare animals. The remaining 18 rats, with female weights ranging from 183.5 to 207.6 g and male weights ranging from 210.7 to 219.8 g, were sorted by weight from lightest to heaviest by sex and divided into a negative control group, a positive control group, and a test article group, a total of 3 groups, with 6 rats in each group, 3 males and 3 females. Each group was passively sensitized by intradermal injection of 0.1 ml of antiserum diluted 1:2, 1:4, and 1:8. 48 hours later, the animals were challenged by intravenous injection of the same antigen dose as the sensitization dose for preparing the antiserum plus 0.5 ml / rat of 1% Evans blue solution. 30 minutes later, the animals in each group were sacrificed, the back skin was excised, and whether blue spots appeared in the inner layer of the corresponding injection site skin was observed. The diameter of the blue reaction spots was measured with an electronic digital caliper to observe whether the rats had passive cutaneous anaphylaxis reactions.

[0065] Table 9 Blue Spot Determination of Carbetocin Injection in Rat Passive Cutaneous Anaphylaxis Test Conclusion: The test results showed that ① after challenge, the number of rats with blue spot diameters greater than 5 mm in the inner layer of the skin of the rats in the negative control group and the test article group was zero, and the positive rate was 0.0%. ② In the positive control group, typical blue spots were formed. The number of rats with blue spot diameters greater than 5 mm in the inner layer of the skin of the 1:2, 1:4, and 1:8 antiserum groups were 4, 6, and 5 respectively, and the positive rates were 66.7%, 100.0%, and 83.3% respectively. The results of the rat passive cutaneous anaphylaxis test of this product showed that this product has no passive anaphylaxis.

[0066] (4) Hemolysis Test During the test, take 7 clean test tubes and number them. Tubes 1 - 5 are the test article test tubes, tube 6 is the negative control tube, and tube 7 is the positive control tube. Add 2% rabbit red blood cell suspension, 0.9% sodium chloride injection or purified water, and the test article to each tube as required. Gently shake each tube and incubate in a water bath at 37°C ± 0.5°C. Observe the hemolysis or aggregation situation 15 min - 3 h after adding 2% rabbit red blood cell suspension, and observe at 0 min and 3 h. To further observe the hemolysis situation, centrifuge the solutions in all test tubes at 1500 rpm for 15 min at 4°C, observe the color of the supernatant, and confirm the hemolysis situation as shown in Table 10.

[0067] Table 10 Determination Results of Hemolytic Test of Carbetocin Injection Note: “+” indicates hemolysis or aggregation, “-” indicates no hemolysis or aggregation Conclusion: The test results show that ① all hemolysis occurred in the positive control tube (tube 7) within 15 min after adding the liquid, and no hemolysis or aggregation occurred in the negative control tube (tube 6) within 3 h. ② No aggregation and hemolysis occurred in the test article tubes (tubes 1 - 5) within 3 h. ③ After centrifugation and observation at 3 h, the supernatants of the test article tubes (tubes 1 - 5) and the negative control tube (tube 6) were colorless and clear. Therefore, the test article does not cause hemolytic reaction.

[0068] The various embodiments of the present invention may exist in the form of a range; it should be understood that the description in the form of a range is only for convenience and brevity, and should not be construed as a rigid limitation on the scope of the present invention; therefore, it should be considered that the described range description has specifically disclosed all possible sub - ranges and single values within that range. For example, it should be considered that the range description from 1 to 6 has specifically disclosed sub - ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., and single numbers within the range, such as 1, 2, 3, 4, 5, and 6, regardless of the range. Additionally, whenever a numerical range is indicated herein, it means including any cited number (fraction or integer) within the indicated range.

[0069] The above - mentioned are only specific embodiments of the present invention, enabling those skilled in the art to understand or implement the present invention. Various modifications to these embodiments will be obvious to those skilled in the art. The general principles defined herein can be implemented in other embodiments without departing from the spirit or scope of the present invention. Therefore, the present invention will not be limited to these embodiments shown herein, but rather will conform to the broadest scope consistent with the principles and novel features claimed herein.

Claims

1. A carbetocin injection, characterized in that: The carbetocin injection contains carbetocin, a stabilizing agent and water for injection; Based on 10L of the carbetocin injection, the amount of the stabilizing agent added is 0.25g to 1.0g; The stabilizing aid is selected from at least one of apigenin, catechin, luteolin, troxerutin, puerarin and baicalin.

2. The carbetocin injection according to claim 1, characterized in that: Based on 10L of the carbetocin injection, the added amount of the stabilizing agent is 0.25g-0.5g; the stabilizing agent is puerarin.

3. The carbetocin injection according to claim 1 or 2, characterized in that: The carbetocin injection is composed of carbetocin, the stabilizing agent, an osmotic pressure regulator, a pH regulator and water for injection; Based on 10L of the carbetocin injection, the amount of carbetocin added is 0.9g-1.1g; Based on 10L of the carbetocin injection, the amount of the osmotic pressure regulator added is 88g-92g; The pH regulator is used in an amount such that the pH value of the carbetocin injection is 4.0-6.5; The dissolved oxygen content of the injection water is ≤3.0 mg / L.

4. The carbetocin injection according to claim 3, characterized in that: Based on 10L of the carbetocin injection, the amount of carbetocin added is 1.0g; based on 10L of the carbetocin injection, the amount of the osmotic pressure regulator added is 90g; the amount of the pH regulator is such that the pH value of the carbetocin injection is 5.0-6.0; the dissolved oxygen content of the water for injection is ≤1.0mg / L.

5. The carbetocin injection according to claim 3, characterized in that: The osmotic pressure regulator includes sodium chloride.

6. The carbetocin injection according to claim 3, characterized in that: The pH adjuster includes at least one of sodium hydroxide, triethylamine, hydrochloric acid and glacial acetic acid.

7. A method for preparing the carbetocin injection according to any one of claims 1 to 6, characterized in that: The preparation method comprises the following steps: Introduce protective gas into water for injection to obtain water for injection with dissolved oxygen content ≤3.0mg / L; The components other than the pH adjuster are added to the injection water with a dissolved oxygen content of ≤3.0 mg / L for dissolution, and then water is added to 95 vol% of the preparation amount. The pH of the system is adjusted to 4.0-6.5 with a pH adjuster, and then water is added to the full amount to obtain the carbetocin injection.

8. The method for preparing carbetocin injection according to claim 7, characterized in that: The protective gas includes at least one of carbon dioxide, nitrogen and helium; the purity of the protective gas is greater than 95 vol%.

9. The method for preparing carbetocin injection according to claim 7, characterized in that: The preparation method further comprises: The carbetocin injection is sterilized and filtered and sealed under an inert gas atmosphere, and then leak tested and inspected by light to obtain a finished carbetocin injection.

10. The method for preparing carbetocin injection according to claim 9, characterized in that: Measured in volume percentage, the residual oxygen content in the ampoule containing carbetocin injection after filling is ≤5%; the sterilization filtration adopts a two-stage 0.22μm filter, and the material of the filter includes at least one of polyethersulfone, polyvinylidene fluoride and polytetrafluoroethylene.

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