Somatostatin powder injection and preparation method thereof

Abstract

The invention relates to a somatostatin powder injection and a powder injection preparation method, and belongs to the technical field of somatostatin, the powder injection comprises the following components by weight: 3-5% of somatostatin, 1-5% of sodium carboxymethyl cellulose, 1-5% of sodium carboxymethyl cellulose, 1-5% of sodium carboxymethyl cellulose, and the balance of water. Mannitol with a weight percentage of 45%-55%; 5%-10% by weight of a cyclodextrin derivative; 4%-8% by weight of a pH regulator; the pH value of the final system is 3.5-5.0 due to the addition amount of the additive; the weight part of the antioxidant is 0.01%-0.05%; the total weight of the components is 100%. The powder injection is packaged in a PTFE (Polytetrafluoroethylene) lined glass bottle after being subjected to 0.22 mu m filtration, semi-stoppered filling and vacuum landslide type freeze-drying, and is sealed under a nitrogen filling condition. A new half-stoppered filling freeze-drying process is adopted, and under the action of a cyclodextrin derivative eutectic matrix and a trace amount of antioxidant, the freeze-dried somatostatin powder injection can be stored for 12 months or more. The somatostatin freeze-dried powder injection has a remarkable effect of prolonging the storage time of the freeze-dried somatostatin powder injection.

Description

Technical Field

[0001] This invention belongs to the field of somatostatin technology, and specifically relates to a somatostatin powder injection and a method for preparing the powder injection. Background Technology

[0002] Somatostatin is a cyclic polypeptide (14-peptide) composed of 14 amino acids, first isolated from the hypothalamus of sheep in 1973. It has a potent inhibitory effect on the pituitary gland's secretion of growth hormone. Its clinical indications include: 1. Severe acute esophageal variceal bleeding; 2. Severe acute gastric or duodenal ulcer bleeding, or complicated by acute erosive gastritis or hemorrhagic gastritis; 3. Adjunctive treatment of pancreatic, biliary, or intestinal fistulas; 4. Prevention and treatment of complications after pancreatic surgery; 5. Adjunctive treatment of diabetic ketoacidosis. Because the peptide bonds of this polypeptide are easily broken in aqueous solution, leading to decreased activity and impurities, lyophilized powder injection is currently the most mature commercial formulation method.

[0003] Most domestically marketed lyophilized somatostatin formulations use a single volatile acid such as acetic acid or hydrochloric acid as a pH adjuster. For example, the preparation method of lyophilized somatostatin powder for injection with patent number CN103417499A mainly involves mixing mannitol (2g / 100mL) with somatostatin, adjusting the pH to 4.5-5.0 with a pH adjuster, followed by 0.22μm filtration and freeze-drying. Another example is the lyophilized somatostatin powder for injection with patent number CN201410747747.3, which consists of somatostatin, mannitol, tert-butanol, and a pH adjuster. The method provided in this invention involves first dissolving mannitol in water for injection, then adding tert-butanol and somatostatin, adding water for injection to the total volume, adjusting the pH value, filling ampoules, freeze-drying, and then sealing the ampoules. Inert gas is used for protection during the drug preparation and freeze-drying processes. The somatostatin lyophilized powder injection prepared by this invention has advantages such as low moisture content, good appearance, low levels of related substances, and suitability for long-term storage.

[0004] However, the lactic acid or glacial acetic acid used in existing technologies are prone to volatilization, adsorption, and migration during freeze-drying and room-temperature storage, leading to significant increases or decreases in the pH of the formulation system. Acidity fluctuations still occur because the pH exceeds the quality standard range of 3.0-6.0, promoting the opening of disulfide bonds and generating high-molecular-weight impurities. Simultaneously, related substances accumulate because pH imbalance increases degradation products such as polypeptide chain breakage and deamidation, significantly increasing the area of ​​impurity peaks. Furthermore, decreased clarity upon reconstitution occurs because pH imbalance causes protein / peptide aggregation, resulting in turbidity and precipitation after reconstitution. Summary of the Invention

[0005] Based on this, the present invention provides a somatostatin powder injection to solve the technical problems existing in the prior art, such as significant increase or decrease in the pH value of the formulation system, resulting in acidity fluctuations, accumulation of related substances, and decreased clarity after reconstitution.

[0006] The technical solution of the present invention to solve the above-mentioned technical problems is as follows:

[0007] A somatostatin powder for injection, the powder for injection comprising the following components, by weight ratio:

[0008] Somatostatin, 3%–5% by weight;

[0009] Mannitol, in parts by weight 45%–55%;

[0010] Cyclodextrin derivatives, in parts by weight of 5%–10%;

[0011] pH adjuster, 4%–8% by weight; its addition amount makes the final pH of the system 3.5–5.0;

[0012] Antioxidant, in parts by weight of 0.01% to 0.05%;

[0013] Add water for injection to bring the total weight to 100%.

[0014] The powder injection is filtered through 0.22μm, partially stoppered, and freeze-dried under vacuum slipway conditions. It is then packaged in PTFE-lined glass bottles and sealed under nitrogen-filled conditions.

[0015] Preferably, the pH adjuster is one of acetic acid and lactic acid.

[0016] Preferably, the cyclodextrin derivative is hydroxypropyl-β-cyclodextrin, and its weight is 7% to 9%.

[0017] Preferably, the antioxidant is thioacetamide, and the amount added is 0.02% to 0.04% by weight.

[0018] Preferably, the mass ratio of mannitol to somatostatin is 15:1 to 30:1.

[0019] A method for preparing a powder injection, comprising:

[0020] S1: Dissolve somatostatin in a 0.5%–2.0% pH adjuster solution to obtain the first solution;

[0021] S2: Add mannitol, cyclodextrin derivative, and antioxidant to water for injection and stir until completely dissolved to obtain the second solution;

[0022] S3: Slowly add the first solution dropwise to the second solution, while using a pH adjuster to adjust the pH of the system to 3.7–4.8.

[0023] S4: Use 0.22μm sterile filtration, then partially stopper the glass bottle with PTFE lining, each bottle contains 0.5mL;

[0024] After bottling, the solution was placed in a freeze dryer and pre-frozen at -45°C for 3 hours. Then, it was sublimated once at -10°C under a vacuum of 160 μbar for 12 hours. It was then sublimated again at -5°C under a vacuum of 120 μbar for 4 hours. Finally, it was desorbed and dried at -25°C under a vacuum of 80 μbar for 3 hours.

[0025] S5: After freeze-drying, nitrogen is introduced under vacuum, followed by full-pressing and capping.

[0026] Preferably, the cooling rate for pre-freezing is 0.5℃ / min, the heating rate for primary sublimation is 5℃ / h, and the heating rate for secondary sublimation is 3℃ / h.

[0027] Preferably, when stored at 25℃±2℃ and relative humidity of 60%±5% for 12 months, the pH change is ≤0.7 pH units and the high molecular weight impurities are ≤0.1%.

[0028] A powder composition for the treatment or prevention of severe acute esophageal variceal bleeding, severe acute gastric or duodenal ulcer bleeding, adjunctive treatment of pancreatic, biliary, or intestinal fistulas, prevention and treatment of postoperative pancreatic complications, and indications for diabetic ketoacidosis.

[0029] Compared with the prior art, the present invention has at least the following advantages:

[0030] (1) The semi-stoppered filling and new freeze-drying process (pre-freezing-heat preservation-first sublimation (slide vacuum)-second sublimation) are adopted. The semi-stoppered filling can first prevent the re-adsorption of external water vapor and volatile acids. The new freeze-drying process can achieve uniform water removal and reduce pH fluctuations, achieving a pH change of ≤0.4 pH units, thereby reducing acidity fluctuations. Compared with the blank group, it is significantly better than the conventional freeze-drying process.

[0031] (2) Through the cyclodextrin derivative eutectic matrix, hydroxypropyl-β-cyclodextrin, somatostatin, and mannitol are mixed and then subjected to a new freeze-drying process. After pre-freezing, heat preservation, first sublimation (slope vacuum), and second sublimation, hydroxypropyl-β-cyclodextrin, somatostatin, and mannitol will form a low-viscosity, low-hygroscopic eutectic structure. The freeze-drying process can achieve uniform water removal, avoid local pressure differences, and reduce pH fluctuations. Compared with Experimental Example 1 and Experimental Example 2, the addition of hydroxypropyl-β-cyclodextrin and the freeze-drying process can significantly reduce moisture and inhibit the "scaling" phenomenon of somatostatin powder injection in the bottle.

[0032] (3) By adding trace amounts of antioxidants and introducing 0.02%-0.04% thioacetamide, the risk of oxidation of somatostatin disulfide bonds in a slightly alkaline environment can be effectively reduced. Compared with Experimental Example 3, the related impurities (≤0.5%) and polymeric impurities (≤0.1%) are significantly reduced, and the clarity of the solution after reconstitution is improved.

[0033] By employing a novel semi-stoppered freeze-drying process, a cyclodextrin derivative eutectic matrix, and trace amounts of antioxidants, the freeze-dried somatostatin powder for injection can be stored for 12 months or more. This demonstrates a significant effect on extending the shelf life of the freeze-dried somatostatin powder for injection. Detailed Implementation

[0034] It should be noted that, unless otherwise specified, the embodiments and features described in the present invention can be combined with each other. The technical solutions of the present invention will be further described below, but the present invention is not limited to the following specific embodiments.

[0035] A somatostatin powder for injection, the powder for injection comprising the following components, by weight ratio:

[0036] Somatostatin, also known as growth hormone-releasing inhibitory hormone, is a chemically synthesized 14-amino acid cyclic polypeptide (14-peptide) that inhibits the release of pituitary hormones, with a weight percentage of 3%–5% and a purity of 87%.

[0037] Mannitol, also known as wood molasses alcohol, has the chemical formula C6H. 14 O6 is a white crystalline solid belonging to the hexanediol class of substances, with a weight percentage of 45%–55% and a purity of 99%.

[0038] Cyclodextrin derivatives, in parts by weight of 5%–10%, such as hydroxypropyl-β-cyclodextrin (HP-β-CD) and hydroxypropyl-γ-cyclodextrin. During the preparation of somatostatin powder for injection, in the later stages of the first drying process, uneven evaporation of moisture at the bottom of the vial causes the powder cake to be drawn away from the bottom, resulting in a "lifting" phenomenon, leading to decreased yield and substandard appearance. However, by adding hydroxypropyl-β-cyclodextrin (HP-β-CD) to the traditional mannitol matrix, a cyclodextrin derivative eutectic matrix is ​​formed, possessing a low-viscosity, low-hygroscopic eutectic structure, thus suppressing the lifting phenomenon.

[0039] pH adjusters, such as acetic acid, lactic acid, hydrochloric acid, and citric acid, are added at a weight of 4% to 8%. The pH adjuster mainly adjusts the pH value of the mixture, and its addition amount makes the final pH of the system 3.5 to 5.0.

[0040] Antioxidants help capture and neutralize free radicals, preventing the harmful effects of oxygen. The concentration is 0.01%–0.05% by weight; purity is 99%. It should be noted that although a pH adjuster is added, oxidation can still occur if acidity fluctuates, especially as the disulfide bonds of somatostatin approach alkalinity. Introducing trace amounts of antioxidants (0.01%–0.05%) protects against the oxidation of somatostatin's disulfide bonds and significantly reduces high-molecular-weight impurities.

[0041] Add water for injection to bring the total weight to 100%.

[0042] The powder for injection is filtered through a 0.22μm filter, partially stoppered and filled, and then freeze-dried using a vacuum slip-flow method. It is then packaged in PTFE-lined glass vials and sealed under nitrogen-filled conditions. The filtration process uses a PVDF membrane with a pore size of 0.22μm. The partially stoppered filling, also known as semi-closed filtration, is performed in a sterile environment and the powder is added to the PTFE-lined glass vials. This process prevents the re-adsorption of external moisture and volatile acids, ensuring pH stability and no increase in impurities for 12 months at 25℃ / 60% humidity.

[0043] Specifically, the somatostatin is 3% by weight;

[0044] Specifically, the somatostatin is 4% by weight;

[0045] Specifically, the somatostatin is 5% by weight;

[0046] In a preferred embodiment, the pH adjuster is either acetic acid or lactic acid. Acetic acid is another name for acetic acid, with the chemical formula CH3COOH; lactic acid is a carboxyl compound (carboxylic acid) containing a hydroxyl group, with the chemical formula C3H6O3.

[0047] Specifically, the pH adjuster is lactic acid, which has a higher boiling point than acetic acid, resulting in more stable acidity fluctuations.

[0048] In a preferred embodiment, the cyclodextrin derivative is hydroxypropyl-β-cyclodextrin, and its weight is 7% to 9%.

[0049] Specifically, the hydroxypropyl-β-cyclodextrin is 8% by weight.

[0050] In a preferred embodiment, the antioxidant is thioacetamide, which is added in an amount of 0.02% to 0.04% by weight.

[0051] Specifically, the thioacetamide is present in a weight fraction of 0.03%.

[0052] In a preferred embodiment, the mass ratio of mannitol to somatostatin is 15:1 to 30:1.

[0053] A method for preparing a powder injection, comprising:

[0054] S1: Dissolve somatostatin in a 0.5%–2.0% pH adjuster solution to obtain the first solution; wherein, the 99% pure pH adjuster is diluted with water to obtain a 0.5%–2.0% pH adjuster solution; for example, take 0.6g of somatostatin and add it to 5mL of 0.5%–2.0% lactic acid solution, and stir magnetically for 30min at 25℃ until completely dissolved, so that the somatostatin solution is acidic.

[0055] S2: Add mannitol, cyclodextrin derivative, and antioxidant to water for injection and stir until completely dissolved to obtain a second solution; for example, take 50 ml of water, add 0.8 mL of 10% lactic acid and 0.006 mL of 10% thioacetamide, stir until completely dissolved, and until the solution is clear. Adjust the pH in real time, using a pH meter to adjust to 3.8-4.2 to obtain a second solution.

[0056] S3: Slowly add the first solution dropwise to the second solution at a rate of 0.5 mL / min, while adding the solution dropwise, using a pH adjuster to adjust the pH of the system to 3.7–4.8; for example, to adjust the pH of the system to 3.9–4.0.

[0057] S4: Use 0.22μm sterile filtration, then partially stopper the glass bottle with PTFE lining, each bottle contains 0.5mL;

[0058] After bottling, the solution was placed in a freeze dryer and pre-frozen at -45°C for 3 hours. Then, it was sublimated once at -10°C under a vacuum of 160 μbar for 12 hours. It was then sublimated again at -5°C under a vacuum of 120 μbar for 4 hours. Finally, it was desorbed and dried at -25°C under a vacuum of 80 μbar for 3 hours.

[0059] S5: After freeze-drying, nitrogen is introduced under vacuum, followed by full-pressing and capping.

[0060] In a preferred embodiment, the pre-freezing cooling rate is 0.5℃ / min, the pre-freezing temperature is -45℃ to -40℃, the vacuum degree is 200μbar, and the duration is 3h, forming a uniform ice crystal structure and avoiding uneven porosity caused by excessively large local ice crystals. Subsequently, the ice crystals are held at -40℃ and a vacuum degree of 200μbar for 1h to completely solidify the ice crystals and reduce the thermal shock during subsequent sublimation.

[0061] The temperature for the first sublimation is raised from -40℃ to -10℃ at a rate of 5℃ / h, and the vacuum level is reduced from 160μbar to 80μbar for 12 hours. The first sublimation mainly removes most of the free water and forms a microporous structure. It is important to note that a slip-slope vacuum is used, that is, the vacuum level is reduced from 160μbar to 80μbar. The vacuum level drops significantly, i.e., a slip-slope vacuum. The vacuum gradient is synchronized with the temperature gradient, so that the vapor can form a uniform "vapor front" on the bottle wall before sublimation, effectively suppressing the "lifting" phenomenon of the powder cake being pulled away from the bottom of the bottle.

[0062] The secondary sublimation temperature was raised from -10℃ to 0℃ at a rate of 3℃ / h, with a vacuum of 80μbar for 4 hours, to further remove "bound water" and improve the uniformity of residual moisture at the drying endpoint.

[0063] In a preferred embodiment, after storage for 12 months at 25℃±2℃ and 60%±5% relative humidity, the pH change is ≤0.7 pH units, and the polymeric impurities are ≤0.1%. The polymeric impurities are large polymer molecules, such as dimer-SOMA. Related impurities and polymeric impurities are existing technologies and will not be described further here.

[0064] The following describes various preparation methods for powder injections and provides further analysis.

[0065] Blank group

[0066] Experimental materials:

[0067] Somatostatin with a purity of 87%, mannitol with a purity of 99%, acetic acid with a purity of 99%, sterile water for injection, and nitrogen gas with a purity of 99.99%.

[0068] formula:

[0069] Somatostatin 0.6g; mannitol 10g; acetic acid 0.8g; and a certain amount of water for injection;

[0070] Preparation steps:

[0071] (1) Add 0.6g of somatostatin and 10g of mannitol to 50mL of water for injection, stir until clear, then add 0.8mL of 10% acetic acid, monitor the pH in real time, and adjust to 4.0 using a pH meter.

[0072] (2) Filter the mixture through a 0.22μm filtration (PVDF membrane) and add it to a 3mL glass bottle in a sterile operating table, injecting 0.5mL into each bottle.

[0073] (3) Freeze-drying. The freeze-drying process adopts the conventional freeze-drying method, namely, pre-freezing at -35℃ with a vacuum of 160μbar for 3 hours; heating up to -5℃ and holding for 2 hours with a vacuum of 160μbar; then cooling down to -20℃ and holding for 3 hours.

[0074] (4) Drying is carried out at a temperature of 0 to 25°C (heating rate of 2°C / h) and a vacuum of 80 μbar to 30 μbar for 3 hours; nitrogen filling is carried out by directly filling inert nitrogen under vacuum to prevent oxygen from re-entering; sealing is done by full-pressure plugging and capping to complete aseptic sealing.

[0075] A total of 30 blank control groups were prepared and stored at 25℃ and 60% relative humidity. Among them, 24 groups were tested for reconstitution, clarity, and stability, and 6 groups were observed after static observation. All experimental data below are average values.

[0076] Reconstitution and Clarity Testing

[0077] The reconstitution time was 2.5 min, which was carried out in a 37℃ water bath. The reconstitution time was less than the standard time by 3 min, and the composite reconstitution standard was used.

[0078] The particle size, after dynamic light detection, is 0.46µm, which is less than 0.50µm, meeting the composite particle size standard.

[0079] The color appears as a light yellow to the naked eye, conforming to the composite color standard.

[0080] After UV-Vis testing, the clarity value was 0.054, which is greater than the standard of 0.02, indicating that the clarity does not meet the standard.

[0081] Stability testing

[0082] Time: Initial; pH: 3.51; Related impurities: 0.36; Polymer impurities: 0.13; Moisture: 2.22.

[0083] Time: January; pH: 3.63; Related impurities: 0.40; Polymer impurities: 0.16; Moisture: 2.25.

[0084] Time: March; pH: 4.02; Related impurities: 0.52; Polymer impurities: 0.28; Moisture: 2.39.

[0085] Time: June; pH: 4.21; Related impurities: 0.68; Polymer impurities: 0.29; Moisture: 2.50.

[0086] Time: September; pH: 4.36; Related impurities: 0.72; Polymer impurities: 0.32; Moisture: 2.61.

[0087] Time: December; pH: 4.52; Related impurities: 0.80; Polymer impurities: 0.33; Moisture: 2.72.

[0088] "Pull-up" test

[0089] Time: December. Visual inspection of the somatostatin powder injection in the still bottle revealed that the somatostatin powder injection at the bottom of the bottle was slightly floating, while the somatostatin powder injection at the top of the bottle showed a "lifting" phenomenon.

[0090] In conclusion, the somatostatin powder injection showed high clarity and turbidity after reconstitution, and under normal storage conditions for 12 months, the pH change was ≥0.4 pH units, the relevant impurities and macromolecular impurities exceeded the thresholds of 0.5% and 0.1% respectively, the moisture content was not maintained below 1.5%, the acidity fluctuated greatly, and the content of relevant impurities and macromolecular impurities was high. In addition, the somatostatin powder injection in the bottle showed a "sinking" phenomenon.

[0091] Experimental Example 1

[0092] Experimental materials:

[0093] Somatostatin with a purity of 87%, mannitol with a purity of 99%, acetic acid with a purity of 99%, sterile water for injection, and nitrogen gas with a purity of 99.99%.

[0094] formula:

[0095] Somatostatin 0.6g; mannitol 10g; acetic acid 0.8g; and a certain amount of water for injection;

[0096] Preparation steps:

[0097] (1) Add 0.6g of somatostatin and 10g of mannitol to 50mL of water for injection, stir until clear, then add 0.8mL of 10% acetic acid, monitor the pH in real time, and adjust to 4.0 using a pH meter.

[0098] (2) Filter the mixture through a 0.22μm PVDF membrane and add it to a 3mL glass bottle with a PTFE liner that has been partially sealed (partially closed) in a sterile operating table. Inject 0.5mL into each bottle.

[0099] (3) Freeze-drying: First, pre-freeze at -45℃ to -40℃ (cooling rate of 0.5℃ / min), vacuum of 200μbar, for 3 hours; then hold at -40℃ under vacuum of 200μbar for 1 hour; then sublimate at -40℃ to -10℃ at a rate of 5℃ / h, vacuum of 160μbar to 80μbar (slope vacuum), for 12 hours; then sublimate at -10℃ to 0℃ at a rate of 3℃ / h, vacuum of 80μbar, for 4 hours.

[0100] (4) Drying is carried out at a temperature of 0 to 25°C (heating rate of 2°C / h) and a vacuum of 80 μbar to 30 μbar for 3 hours; nitrogen filling is carried out by directly filling inert nitrogen under vacuum to prevent oxygen from re-entering; sealing is done by full-pressure plugging and capping to complete aseptic sealing.

[0101] Experiment 1 consisted of 30 groups, which were stored at 25℃ and 60% relative humidity. 24 groups underwent reconstitution, clarification, and stability testing, while 6 groups were observed under static conditions. All experimental data below are averages.

[0102] Reconstitution and Clarity Testing

[0103] The reconstitution time was 2.5 min, which was carried out in a 37℃ water bath. The reconstitution time was less than the standard time by 3 min, and the composite reconstitution standard was used.

[0104] The particle size, after dynamic light detection, is 0.46µm, which is less than 0.50µm, meeting the composite particle size standard.

[0105] The color appears as a light yellow to the naked eye, conforming to the composite color standard.

[0106] After UV-Vis testing, the clarity value was 0.032, which is greater than the standard of 0.02, indicating that the clarity does not meet the standard.

[0107] Stability testing

[0108] Time: Initial; pH: 3.81; Related impurities: 0.35; Polymer impurities: 0.10; Moisture: 1.86.

[0109] Time: January; pH: 3.89; Related impurities: 0.39; Polymer impurities: 0.12; Moisture: 1.89.

[0110] Time: March; pH: 3.95; Related impurities: 0.50; Polymer impurities: 0.15; Moisture: 2.05.

[0111] Time: June; pH: 3.98; Related impurities: 0.65; Polymer impurities: 0.18; Moisture: 2.12.

[0112] Time: September; pH: 4.01; Related impurities: 0.68; Polymer impurities: 0.26; Moisture: 2.33.

[0113] Time: December; pH: 4.12; Related impurities: 0.72; Polymer impurities: 0.29; Moisture: 2.45.

[0114] "Pull-up" test

[0115] Time: December. Visual inspection of the somatostatin powder injection in the still bottle revealed that the somatostatin powder injection at the bottom of the bottle was slightly floating, while the somatostatin powder injection at the top of the bottle showed a "lifting" phenomenon.

[0116] In conclusion, under normal storage conditions in December, the pH change was ≤0.4 pH units. Because the glass bottles were lined with a PTFE membrane and filled with a semi-stopper, the re-adsorption of external moisture and volatile acids was prevented. Secondly, the new freeze-drying process, involving pre-freezing, heat preservation, primary sublimation (slide vacuum), and secondary sublimation, achieved uniform water removal and reduced pH fluctuations, thus lowering acidity fluctuations. However, the somatostatin powder injection showed high clarity and turbidity after reconstitution, with related impurities and high molecular weight impurities exceeding the thresholds of 0.5% and 0.1%, respectively. Although the moisture content decreased compared to the control group, it still did not remain below 1.5%, indicating a high content of related impurities and high molecular weight impurities. Furthermore, the somatostatin powder injection showed a "pumping" phenomenon within the bottle.

[0117] Experiment Example 2

[0118] Experimental materials:

[0119] Somatostatin with a purity of 87%, mannitol with a purity of 99%, lactic acid with a purity of 99%, sterile water for injection, and nitrogen gas with a purity of 99.99%.

[0120] formula:

[0121] Somatostatin 0.6g; mannitol 10g; lactic acid 0.8g; and a certain amount of water for injection;

[0122] Preparation steps:

[0123] (1) Add 0.6g of somatostatin and 10g of mannitol to 50mL of water for injection, stir until clear, then add 0.8mL of 10% lactic acid, monitor the pH in real time, and adjust to 4.0 using a pH meter.

[0124] (2) Filter the mixture through a 0.22μm PVDF membrane and add it to a 3mL glass bottle with a PTFE liner that has been partially sealed (partially closed) in a sterile operating table. Inject 0.5mL into each bottle.

[0125] (3) Freeze-drying: First, pre-freeze at -45℃ to -40℃ (cooling rate of 0.5℃ / min), vacuum of 200μbar, for 3 hours; then hold at -40℃ under vacuum of 200μbar for 1 hour; then sublimate at -40℃ to -10℃ at a rate of 5℃ / h, vacuum of 160μbar to 80μbar (slope vacuum), for 12 hours; then sublimate at -10℃ to 0℃ at a rate of 3℃ / h, vacuum of 80μbar, for 4 hours.

[0126] (4) Drying is carried out at a temperature of 0 to 25°C (heating rate of 2°C / h) and a vacuum of 80 μbar to 30 μbar for 3 hours; nitrogen filling is carried out by directly filling inert nitrogen under vacuum to prevent oxygen from re-entering; sealing is done by full-pressure plugging and capping to complete aseptic sealing.

[0127] Experiment 2 consisted of 30 groups, which were stored at 25℃ and 60% relative humidity. Of these, 24 groups underwent reconstitution, clarification, and stability testing, while 6 groups were observed under static conditions. All experimental data below are averages.

[0128] Reconstitution and Clarity Testing

[0129] The reconstitution time was 2.5 min, which was carried out in a 37℃ water bath. The reconstitution time was less than the standard time by 3 min, and the composite reconstitution standard was used.

[0130] The particle size, after dynamic light detection, is 0.46µm, which is less than 0.50µm, meeting the composite particle size standard.

[0131] The color appears as a light yellow to the naked eye, conforming to the composite color standard.

[0132] After UV-Vis testing, the clarity value was 0.034, which is greater than the standard of 0.02, indicating that the clarity does not meet the standard.

[0133] Stability testing

[0134] Time: Initial; pH: 3.84; Related impurities: 0.37; Polymer impurities: 0.12; Moisture: 1.84.

[0135] Time: January; pH: 3.86; Related impurities: 0.40; Polymer impurities: 0.13; Moisture: 1.90.

[0136] Time: March; pH: 3.93; Related impurities: 0.45; Polymer impurities: 0.15; Moisture: 2.01.

[0137] Time: June; pH: 3.96; Related impurities: 0.53; Polymer impurities: 0.20; Moisture: 2.08.

[0138] Time: September; pH: 4.00; Related impurities: 0.59; Polymer impurities: 0.23; Moisture: 2.25.

[0139] Time: December; pH: 4.05; Related impurities: 0.65; Polymer impurities: 0.28; Moisture: 2.36.

[0140] "Pull-up" test

[0141] Time: December. Visual inspection of the somatostatin powder injection in the still bottle revealed that the somatostatin powder injection at the bottom of the bottle was slightly floating, while the somatostatin powder injection at the top of the bottle showed a "lifting" phenomenon.

[0142] In conclusion, under normal storage conditions in December, the pH change was ≤0.4 pH units. Because the glass bottles were lined with a PTFE membrane and partially stoppered, the re-adsorption of external moisture and volatile acids was prevented. Secondly, the new freeze-drying process, involving pre-freezing, heat preservation, primary sublimation (slide vacuum), and secondary sublimation, achieved uniform dehydration and reduced pH fluctuations, thus lowering acidity fluctuations. It should be noted that the effects of using lactic acid and acetic acid were similar, but lactic acid resulted in more stable pH fluctuations. However, the somatostatin powder injection showed high clarity and turbidity after reconstitution, with related impurities and high molecular weight impurities exceeding the thresholds of 0.5% and 0.1%, respectively. Although the moisture content decreased compared to the control group, it still did not remain below 1.5%, indicating a high content of related impurities and high molecular weight impurities. Furthermore, the somatostatin powder injection showed a "pumping" phenomenon within the bottle.

[0143] Experimental Example 3

[0144] Somatostatin with a purity of 87%, mannitol with a purity of 99%, lactic acid with a purity of 99%, hydroxypropyl-β-cyclodextrin with a purity of 99%, sterile water for injection, and 99.99% nitrogen.

[0145] formula:

[0146] Somatostatin 0.6g; mannitol 10g; lactic acid 0.8g; hydroxypropyl-β-cyclodextrin 1.6g; and a certain amount of water for injection;

[0147] Preparation steps:

[0148] (1) Add 0.6g somatostatin, 10g mannitol and 1.6g hydroxypropyl-β-cyclodextrin to 50mL of water for injection, stir until clear, then add 0.8mL of 10% lactic acid, monitor the pH in real time, and adjust to 4.0 using a pH meter.

[0149] (2) Filter the mixture through a 0.22μm PVDF membrane and add it to a 3mL glass bottle with a PTFE liner that has been partially sealed (partially closed) in a sterile operating table. Inject 0.5mL into each bottle.

[0150] (3) Freeze-drying: First, pre-freeze at -45℃ to -40℃ (cooling rate of 0.5℃ / min), vacuum of 200μbar, for 3 hours; then hold at -40℃ under vacuum of 200μbar for 1 hour; then sublimate at -40℃ to -10℃ at a rate of 5℃ / h, vacuum of 160μbar to 80μbar (slope vacuum), for 12 hours; then sublimate at -10℃ to 0℃ at a rate of 3℃ / h, vacuum of 80μbar, for 4 hours.

[0151] (4) Drying is carried out at a temperature of 0 to 25°C (heating rate of 2°C / h) and a vacuum of 80 μbar to 30 μbar for 3 hours; nitrogen filling is carried out by directly filling inert nitrogen under vacuum to prevent oxygen from re-entering; sealing is done by full-pressure plugging and capping to complete aseptic sealing.

[0152] Experiment 3 consisted of 30 groups, which were stored at 25℃ and 60% relative humidity. Of these, 24 groups underwent reconstitution, clarification, and stability testing, while 6 groups were observed under static conditions. All experimental data below are averages.

[0153] Reconstitution and Clarity Testing

[0154] The reconstitution time was 2.5 min, which was carried out in a 37℃ water bath. The reconstitution time was less than the standard time by 3 min, and the composite reconstitution standard was used.

[0155] The particle size, after dynamic light detection, is 0.46µm, which is less than 0.50µm, meeting the composite particle size standard.

[0156] The color appears as a light yellow to the naked eye, conforming to the composite color standard.

[0157] After UV-Vis testing, the clarity value was 0.031, which is greater than the standard of 0.02, indicating that the clarity does not meet the standard.

[0158] Stability testing

[0159] Time: Initial; pH: 3.85; Related impurities: 0.38; Polymer impurities: 0.11; Moisture: 1.12.

[0160] Time: January; pH: 3.87; Related impurities: 0.39; Polymer impurities: 0.13; Moisture: 1.13.

[0161] Time: March; pH: 3.92; Related impurities: 0.42; Polymer impurities: 0.15; Moisture: 1.24.

[0162] Time: June; pH: 3.96; Related impurities: 0.49; Polymer impurities: 0.21; Moisture: 1.30.

[0163] Time: September; pH: 4.01; Related impurities: 0.56; Polymer impurities: 0.24; Moisture: 1.37.

[0164] Time: December; pH: 4.04; Related impurities: 0.60; Polymer impurities: 0.27; Moisture: 1.45.

[0165] "Pull-up" test

[0166] Time: December. Visual inspection of the somatostatin powder injection in the still bottle revealed that the somatostatin powder injection at the bottom of the bottle was located at the bottom without any slight floating, and the somatostatin powder injection at the top of the bottle did not show any "lifting" phenomenon.

[0167] In conclusion, under normal storage conditions in December, the pH change was ≤0.7 pH units. Because the glass bottles were lined with a PTFE membrane and partially capped, the re-adsorption of external moisture and volatile acids was prevented. Secondly, the new freeze-drying process, involving pre-freezing, heat preservation, primary sublimation (slide vacuum), and secondary sublimation, achieved uniform water removal and reduced pH fluctuations, thus lowering acidity fluctuations. However, the clarity and turbidity of the somatostatin powder injection after reconstitution were relatively high. The clarity of Example 3 was lower than that of the blank group, Example 1, and Example 2. Related impurities and polymeric impurities exceeded the thresholds of 0.5% and 0.1%, respectively. The concentration was kept below 1.5%. Based on Experiment 2, the addition of hydroxypropyl-β-cyclodextrin resulted in a low-viscosity, low-hygroscopic eutectic structure formed by hydroxypropyl-β-cyclodextrin, somatostatin, and mannitol after pre-freezing, heat preservation, first sublimation (slope vacuum), and second sublimation. The freeze-drying process also achieved uniform water removal and avoided local pressure differences, thus significantly reducing moisture content and inhibiting the "scaling" phenomenon of somatostatin powder injection in the bottle. This is more conducive to improving the storage time after freeze-drying. However, it still cannot solve the problem of related impurities and high molecular weight impurities.

[0168] Experiment Example 4

[0169] Somatostatin with a purity of 87%, mannitol with a purity of 99%, lactic acid with a purity of 99%, hydroxypropyl-β-cyclodextrin with a purity of 99%, thioacetamide with a purity of 99%, sterile water for injection, and nitrogen gas with a purity of 99.99%.

[0170] formula:

[0171] Somatostatin 0.6g; mannitol 10g; lactic acid 0.8g; hydroxypropyl-β-cyclodextrin 1.6g; thioacetamide 0.024g; and a certain amount of water for injection;

[0172] Preparation steps:

[0173] (1) Add 0.6g somatostatin, 10g mannitol, 1.6g hydroxypropyl-β-cyclodextrin and 0.024g thioacetamide to 50mL of water for injection, stir until clear, then add 0.8mL of 10% lactic acid, monitor the pH in real time, and adjust to 4.0 using a pH meter.

[0174] (2) Filter the mixture through a 0.22μm PVDF membrane and add it to a 3mL glass bottle with a PTFE liner that has been partially sealed (partially closed) in a sterile operating table. Inject 0.5mL into each bottle.

[0175] (3) Freeze-drying: First, pre-freeze at -45℃ to -40℃ (cooling rate of 0.5℃ / min), vacuum of 200μbar, for 3 hours; then hold at -40℃ under vacuum of 200μbar for 1 hour; then sublimate at -40℃ to -10℃ at a rate of 5℃ / h, vacuum of 160μbar to 80μbar (slope vacuum), for 12 hours; then sublimate at -10℃ to 0℃ at a rate of 3℃ / h, vacuum of 80μbar, for 4 hours.

[0176] (4) Drying is carried out at a temperature of 0 to 25°C (heating rate of 2°C / h) and a vacuum of 80 μbar to 30 μbar for 3 hours; nitrogen filling is carried out by directly filling inert nitrogen under vacuum to prevent oxygen from re-entering; sealing is done by full-pressure plugging and capping to complete aseptic sealing.

[0177] Experiment 4 consisted of 30 groups, stored at 25℃ and 60% relative humidity. Of these, 24 groups underwent reconstitution, clarification, and stability testing, while 6 groups were observed under static conditions. All experimental data below are averages.

[0178] Reconstitution and Clarity Testing

[0179] The reconstitution time was 2.5 min, which was carried out in a 37℃ water bath. The reconstitution time was less than the standard time by 3 min, and the composite reconstitution standard was used.

[0180] The particle size, after dynamic light detection, is 0.46µm, which is less than 0.50µm, meeting the composite particle size standard.

[0181] The color appears as a light yellow to the naked eye, conforming to the composite color standard.

[0182] After UV-Vis testing, the clarity value was 0.016, which is greater than the standard of 0.02, indicating that the clarity does not meet the standard.

[0183] Stability testing

[0184] Time: Initial; pH: 3.84; Related impurities: 0.24; Polymer impurities: 0.06; Moisture: 1.11.

[0185] Time: January; pH: 3.86; Related impurities: 0.27; Polymer impurities: 0.06; Moisture: 1.13.

[0186] Time: March; pH: 3.93; Related impurities: 0.34; Polymer impurities: 0.07; Moisture: 1.23.

[0187] Time: June; pH: 3.96; Related impurities: 0.39; Polymer impurities: 0.08; Moisture: 1.29.

[0188] Time: September; pH: 4.00; Related impurities: 0.42; Polymer impurities: 0.09; Moisture: 1.35.

[0189] Time: December; pH: 4.03; Related impurities: 0.46; Polymer impurities: 0.10; Moisture: 1.40.

[0190] "Pull-up" test

[0191] Time: December. Visual inspection of the somatostatin powder injection in the still bottle revealed that the somatostatin powder injection at the bottom of the bottle was located at the bottom without any slight floating, and the somatostatin powder injection at the top of the bottle did not show any "lifting" phenomenon.

[0192] In conclusion, under the normal storage conditions in December, (1) the use of semi-stoppered filling and new freeze-drying process (pre-freezing-heat preservation-first sublimation (slide vacuum)-second sublimation) and semi-stoppered filling can firstly prevent the re-adsorption of external water vapor and volatile acids. The new freeze-drying process can achieve uniform water removal and reduce pH fluctuations, achieving a pH change of ≤0.4 pH units, thereby reducing acidity fluctuations. Compared with the blank group, it is significantly better than the conventional freeze-drying process. (2) Through the cyclodextrin derivative eutectic matrix, hydroxypropyl-β-cyclodextrin, somatostatin, and mannitol are mixed and then subjected to a new freeze-drying process. After pre-freezing, heat preservation, first sublimation (slide vacuum), and second sublimation, hydroxypropyl-β-cyclodextrin, somatostatin, and mannitol form a low-viscosity, low-hygroscopic eutectic structure. The freeze-drying process can achieve uniform water removal, avoid local pressure differences, and reduce pH fluctuations. Compared with Experimental Example 1 and Experimental Example 2, the addition of hydroxypropyl-β-cyclodextrin and the freeze-drying process can significantly reduce moisture and inhibit the "scaling" phenomenon of somatostatin powder injection in the bottle. (3) By adding a trace amount of antioxidant and introducing 0.02%-0.04% thioacetamide, the oxidation risk of somatostatin disulfide bonds in a slightly alkaline environment can be effectively reduced. Compared with Experimental Example 3, the related impurities (≤0.5%) and high molecular impurities (≤0.1%) are significantly reduced, and the clarity of the solution after reconstitution is improved. By employing a novel semi-stoppered freeze-drying process, a cyclodextrin derivative eutectic matrix, and trace amounts of antioxidants, the freeze-dried somatostatin powder for injection can be stored for 12 months or more. This demonstrates a significant effect on extending the shelf life of the freeze-dried somatostatin powder for injection.

[0193] A powder composition for the treatment or prevention of severe acute esophageal variceal bleeding, severe acute gastric or duodenal ulcer bleeding, adjunctive treatment of pancreatic, biliary, or intestinal fistulas, prevention and treatment of postoperative pancreatic complications, and indications for diabetic ketoacidosis.

[0194] Obviously, the above embodiments of the present invention are merely examples for clearly illustrating the present invention, and are not intended to limit the implementation of the present invention. Those skilled in the art can make other variations or modifications based on the above description. It is neither necessary nor possible to exhaustively describe all embodiments here. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A somatostatin powder for injection, characterized in that, The powder for injection comprises the following components, in the following weight ratio: Somatostatin, 3%–5% by weight; Mannitol, in parts by weight 45%–55%; Cyclodextrin derivatives, in parts by weight of 5%–10%; pH adjuster, 4%–8% by weight; its addition amount makes the final pH of the system 3.5–5.0; Antioxidant, in parts by weight of 0.01% to 0.05%; Add water for injection to bring the total weight to 100%. The powder injection is filtered through 0.22μm, partially stoppered, and freeze-dried under vacuum slipway conditions. It is then packaged in PTFE-lined glass bottles and sealed under nitrogen-filled conditions.

2. The somatostatin powder injection as described in claim 1, characterized in that, The pH adjuster is either acetic acid or lactic acid.

3. The somatostatin powder injection as described in claim 1, characterized in that, The cyclodextrin derivative is hydroxypropyl-β-cyclodextrin, with a weight of 7% to 9%.

4. The somatostatin powder injection as described in claim 1, characterized in that, The antioxidant is thioacetamide, which is added at a weight of 0.02% to 0.04%.

5. The somatostatin powder injection as described in claim 1, characterized in that, The mass ratio of mannitol to somatostatin is 15:1 to 30:

1.

6. A method for preparing a powder injection, using a somatostatin powder injection as described in any one of claims 1-4, characterized in that, include: S1: Dissolve somatostatin in a 0.5%–2.0% pH adjuster solution to obtain the first solution; S2: Add mannitol, cyclodextrin derivative, and antioxidant to water for injection and stir until completely dissolved to obtain the second solution; S3: Slowly add the first solution dropwise to the second solution, while using a pH adjuster to adjust the pH of the system to 3.7–4.

8. S4: Use 0.22μm sterile filtration, then partially stopper the glass bottle with PTFE lining, each bottle contains 0.5mL; After bottling, the solution was placed in a freeze dryer and pre-frozen at -45°C for 3 hours. Then, it was sublimated once at -10°C under a vacuum of 160 μbar for 12 hours. It was then sublimated again at -5°C under a vacuum of 120 μbar for 4 hours. Finally, it was desorbed and dried at -25°C under a vacuum of 80 μbar for 3 hours. S5: After freeze-drying, nitrogen is introduced under vacuum, followed by full-pressing and capping.

7. The method for preparing a powder injection as described in claim 1, characterized in that, The cooling rate for pre-freezing is 0.5℃ / min, the heating rate for primary sublimation is 5℃ / h, and the heating rate for secondary sublimation is 3℃ / h.

8. The method for preparing a powder injection as described in claim 1, characterized in that, When stored at 25℃±2℃ and 60%±5% relative humidity for 60 days for 12 months, the pH change is ≤0.7 pH units and the high molecular weight impurities are ≤0.1%.

9. A pharmaceutical powder composition, comprising a somatostatin powder for injection as described in any one of claims 1-4, characterized in that, Indications for use in the treatment or prevention of severe acute esophageal variceal bleeding, severe acute gastric or duodenal ulcer bleeding, adjunctive treatment of pancreatic, biliary, or intestinal fistulas, prevention and treatment of postoperative pancreatic complications, and diabetic ketoacidosis.