4-boron-L-phenylalanine freeze-dried preparation and preparation method thereof

Abstract

The invention relates to the technical field of medicine, in particular to a 4-boron-L-phenylalanine freeze-dried preparation and a preparation method thereof.The preparation method comprises the steps that firstly, 4-boron-L-phenylalanine and hydroxypropyl-beta-cyclodextrin are mixed, the solubility of the 4-boron-L-phenylalanine is increased through the inclusion effect of the hydroxypropyl-beta-cyclodextrin, and then the 4-boron-L-phenylalanine freeze-dried preparation is obtained; meanwhile, the hydroxypropyl-beta-cyclodextrin is included on the 4-boron-L-phenylalanine, so that the stability of the 4-boron-L-phenylalanine can be improved, 4-boron-L-phenylalanine drug molecules are protected, the degradation of the 4-boron-L-phenylalanine drug molecules in the storage or use process is reduced, the stability of the 4-boron-L-phenylalanine is improved, and the validity period of the 4-boron-L-phenylalanine is prolonged; then adding the soluble compound into a mixed solution of 1, 2-propylene glycol and mannitol, uniformly mixing to form a freeze-drying solution, taking mannitol as a freeze-drying protective agent, protecting the activity of the medicine in the freeze-drying process, reducing the formation of ice crystals and avoiding the damage to the structure of the medicine, finally filling the freeze-drying solution into a sterile container, and carrying out a freeze-drying process to obtain the freeze-dried powder. And preparing to obtain the 4-boron-L-phenylalanine freeze-dried preparation.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to a 4-boron-L-phenylalanine freeze-dried preparation and a preparation method thereof. Background Art

[0002] Boron neutron capture therapy (BNCT) is a binary radiotherapy method that combines neutron irradiation and boron-containing targeted drugs. It is mainly used to treat locally invasive malignant tumors such as malignant melanoma, malignant brain tumors and recurrent head and neck cancer. It first uses a boron carrier to transport boron-10 ( 10 B) Atoms selectively accumulate in tumor cells and are then irradiated with thermal neutrons, using low-energy thermal neutrons to 10 B undergoes neutron capture reaction 10 B(n,α) 7 Li, thereby releasing α and 7 Li particles. The range of these particles in tissues is similar to the diameter of a cell, 5 to 9 μm. Therefore, their killing power is limited to selective enrichment of sufficient amounts. 10 B atoms in tumor cells, while those that do not take up 10 B has little radiation damage to normal tissues. Therefore, the successful implementation of BNCT in targeted cancer treatment depends on three aspects, which are also the challenges currently faced. (1) The key to implementing BNCT is to use an ideal boron carrier. 10 B is enriched in tumor tissue (20-50 μg 10 (B / g), and have a high tumor / normal tissue and tumor / blood boron concentration ratio (>3), as well as low systemic toxicity and high bioavailability of the boron carrier; (2) A reasonable neutron source. Currently, most countries use neutron sources based on reactors and accelerators to develop BNCT, but the cost of reactors is high, and the operation procedures are complicated and the safety is low. The neutron energy generated by accelerators is high, making it difficult for its stability to meet actual medical requirements, and there may be contamination by fast neutrons and gamma rays, which cannot fully meet medical needs. A reasonable neutron source is being further developed; (3) An accurate dose measurement system is performed before treatment. 10 Aggregation detection of B, 10 The distribution and content of B in tumor tissue determine the dose of neutron irradiation.

[0003] 4-borono-L-phenylalanine (BPA) is an important boron-containing compound, mainly used in boron neutron capture therapy (BNCT). The structure, physicochemical properties, cellular uptake mechanism and human biodistribution of BPA are crucial for the clinical treatment of BNCT. 4-borono-L-phenylalanine is insoluble in water and needs to be dissolved by adding a cosolvent under the action of strong acid and strong base, and then the pH value is adjusted. The solution needs to be prepared and used immediately, and the cumbersome preparation process limits its convenient use. Chinese patent CN103100094A provides a freeze-drying process for L-BPA, which discloses a method of freezing the L-BPA solution at low temperature, then sublimating and drying under vacuum conditions, and then desorbing and drying after the sublimation is completed to remove most of the bound water. This technology avoids the cumbersome preparation process of BPA injection, and at the same time, the dosage is accurate and easy to store. Patent Chinese patent CN116270504A provides a BPA freeze-dried preparation and a preparation method thereof. The preparation method comprises dissolving BPA and a polyol in an aqueous solution with an alkali to obtain a clear solution, and then freeze-drying to obtain a freeze-dried preparation. The freeze-dried preparation obtained by the method has a low impurity content and remains stable for a long time.

[0004] In the above-mentioned prior art, since BPA itself is insoluble in water, it is necessary to add fructose as a co-solvent to BPA, and then dissolve it with a strong alkali or strong acid solution, and adjust the pH value of the solution to a physiological pH of about 7.4 after dissolution, and then prepare a lyophilized preparation. 4-boron-L-phenylalanine is generally administered by injection, so during the preparation process of its preparation, it is necessary to ensure complete dissolution and solve the problem of low yield and easy generation of impurities in the preparation process of protecting the BPA preparation. Summary of the invention

[0005] In view of this, the object of the present invention is to provide a 4-boron-L-phenylalanine freeze-dried preparation and a preparation method thereof, so as to at least solve the problems existing in the prior art that BPA is poorly soluble in water, has a low yield, and is prone to produce impurities.

[0006] The present invention solves the above technical problems by the following technical means:

[0007] In a first aspect, an embodiment of the present invention provides a lyophilized preparation of 4-boron-L-phenylalanine, comprising a readily soluble complex, 1,2-propylene glycol and mannitol, wherein the readily soluble complex is formed by encapsulating 4-boron-L-phenylalanine with hydroxypropyl-β-cyclodextrin, and the mass ratio of 4-boron-L-phenylalanine, hydroxypropyl-β-cyclodextrin, 1,2-propylene glycol and mannitol is 1:(1-3):(40-50):(30-50).

[0008] Preferably, the mass ratio of 4-boron-L-phenylalanine, hydroxypropyl-β-cyclodextrin, 1,2-propylene glycol and mannitol is 1:2:44:48.

[0009] In a second aspect, an embodiment of the present invention provides a method for preparing a lyophilized preparation of 4-boron-L-phenylalanine, comprising the following steps:

[0010] Weigh 4-boron-L-phenylalanine and hydroxypropyl-β-cyclodextrin respectively and mix them to obtain a readily soluble complex;

[0011] The easily soluble complex is added to a mixed solution containing 1,2-propylene glycol and mannitol, heated at 60-80°C for 4-6 hours and refluxed, activated carbon is added and refluxed for 1-3 hours, and then an organic filter membrane is used to obtain a lyophilized solution;

[0012] The lyophilized liquid is divided into sterile containers, and the sterile containers are then installed on a freeze dryer, and are successively subjected to pre-freezing, sublimation drying, and analytical drying treatments to obtain a 4-boron-L-phenylalanine lyophilized preparation.

[0013] Preferably, the pre-freezing conditions are set as follows: temperature is -30 to -60°C, pressure is 10 to 20 Pa, and time is 2 to 4 hours.

[0014] Preferably, the sublimation drying conditions are set as follows: temperature is -15 to -35°C, pressure is 5 to 10 Pa, and time is 5 to 10 hours.

[0015] Preferably, the analytical drying conditions are set as follows: temperature is 18-35° C., pressure is 5-10 Pa, and time is 10-15 h.

[0016] Preferably, the mass ratio of 4-boron-L-phenylalanine, hydroxypropyl-β-cyclodextrin, 1,2-propylene glycol and mannitol is 1:(1-3):(40-50):(30-50).

[0017] Preferably, the specific surface area of ​​the activated carbon is 1000-1500m 2 / g.

[0018] Preferably, the organic filter membrane is filtered 3 to 5 times.

[0019] The invention discloses a 4-boron-L-phenylalanine freeze-dried preparation and a preparation method thereof. Firstly, 4-boron-L-phenylalanine is mixed with hydroxypropyl-β-cyclodextrin, and the solubility of 4-boron-L-phenylalanine is increased by the inclusion effect of hydroxypropyl-β-cyclodextrin. Meanwhile, hydroxypropyl-β-cyclodextrin is included in 4-boron-L-phenylalanine, so that the stability of 4-boron-L-phenylalanine can be improved, the 4-boron-L-phenylalanine drug molecules can be protected, the degradation of 4-boron-L-phenylalanine drug molecules during storage or use can be reduced, and the stability and shelf life of 4-boron-L-phenylalanine can be improved. Then, the soluble complex is added to a mixed solution of 1,2-propylene glycol and mannitol, and the mixture is evenly mixed to form a freeze-dried liquid. Mannitol is used as a freeze-dried protective agent to protect the activity of the drug during the freeze-drying process, reduce the formation of ice crystals, and avoid damage to the drug structure. Finally, the freeze-dried liquid is filled into a sterile container, and subjected to a freeze-drying process to prepare the 4-boron-L-phenylalanine freeze-dried preparation.

[0020] The preparation method of the 4-boron-L-phenylalanine freeze-dried preparation of the present invention uses cheap and easily available raw material components, the entire preparation method has a simple process, is easy to operate, and is suitable for large-scale production; the prepared 4-boron-L-phenylalanine freeze-dried preparation product has good resolubility, dissolves quickly in water, has high purity, good stability, no visible foreign matter, and good impurity control effect. After testing, the 4-boron-L-phenylalanine freeze-dried preparation of the present invention was left at room temperature for 20 hours after being re-dissolved, and the solution remained clear, with a light transmittance greater than 99.6%, and the impurity content could be as low as 0.01% DETAILED DESCRIPTION

[0021] In order to make the purpose, technical scheme and advantages of the embodiments of the present application clearer, the technical scheme in the embodiments of the present application will be clearly and completely described below in combination with the specific embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of this application.

[0022] In the description of the embodiments of the present application, unless otherwise specified, "multiple" means two or more, for example, multiple processing units refer to two or more processing units, etc., multiple elements refer to two or more elements, etc.

[0023] In the embodiments of the present application, words such as "exemplary" or "for example" are used to indicate examples, illustrations or descriptions. Any embodiment or design described as "exemplary" or "for example" in the embodiments of the present application should not be interpreted as being more preferred or more advantageous than other embodiments or designs. Specifically, the use of words such as "exemplary" or "for example" is intended to present related concepts in a specific way.

[0024] The 4-boron-L-phenylalanine freeze-dried preparation of the present application is first mixed with 4-boron-L-phenylalanine and hydroxypropyl-β-cyclodextrin, and the solubility of 4-boron-L-phenylalanine is increased by the inclusion complex of hydroxypropyl-β-cyclodextrin. At the same time, hydroxypropyl-β-cyclodextrin can also improve the stability of 4-boron-L-phenylalanine, protect the 4-boron-L-phenylalanine drug molecules, reduce its degradation during storage or use, and improve the stability and shelf life of 4-boron-L-phenylalanine; then, the soluble complex is added to a mixed solution of 1,2-propylene glycol and mannitol, mixed evenly to form a freeze-dried solution, and mannitol is used as a freeze-dried protective agent to protect the activity of the drug during the freeze-drying process, reduce the formation of ice crystals, and avoid damage to the drug structure. Finally, the freeze-dried solution is placed in a sterile container, and a 4-boron-L-phenylalanine freeze-dried preparation is prepared through a freeze-drying process.

[0025] Specifically, the preparation method of the 4-boron-L-phenylalanine freeze-dried preparation of the present application comprises the following steps:

[0026] S1: Weigh 4-boron-L-phenylalanine and hydroxypropyl-β-cyclodextrin respectively and mix them to obtain a readily soluble complex;

[0027] S2: adding the easily soluble complex to a mixed solution containing 1,2-propylene glycol and mannitol, heating at 60-80°C for 4-6 hours and refluxing, then adding activated carbon and refluxing for 1-3 hours, and then using an organic filter membrane to obtain a lyophilized solution;

[0028] S3: The freeze-dried liquid is divided into sterile containers, and the sterile containers are installed on a freeze dryer, and are subjected to pre-freezing, sublimation drying, and analytical drying treatments to obtain a 4-boron-L-phenylalanine freeze-dried preparation.

[0029] Among them, the pre-freezing conditions are set as follows: temperature is -30~-60℃, pressure is 10~20Pa, and time is 2~4h; the sublimation drying conditions are set as follows: temperature is -15~-35℃, pressure is 5~10Pa, and time is 5~10h; the analytical drying conditions are set as follows: temperature is 18~35℃, pressure is 5~10Pa, and time is 10~15h. The freeze-drying process using pre-freezing, sublimation drying, and analytical drying treatments can greatly avoid the incomplete removal of moisture during the freeze-drying process, and avoid the adverse effects of poor temperature control on the product morphology, and control the product moisture at the lowest level; during the freeze-drying process, the temperature is evenly transmitted, the product is loose and full, and the particles are uniform.

[0030] The mass ratio of 4-boron-L-phenylalanine, hydroxypropyl-β-cyclodextrin, 1,2-propylene glycol and mannitol is 1:(1-3):(40-50):(30-50). The specific surface area of ​​the activated carbon used for reflux is 1000-1500m2 / g, filter with organic filter membrane 3 to 5 times.

[0031] In order to better understand the above technical solution, the above technical solution will be described in detail below in conjunction with specific implementation methods.

[0032] Example 1

[0033] The preparation method of the 4-boron-L-phenylalanine lyophilized preparation of this embodiment is as follows:

[0034] Step S1, weigh 500 g of 4-boron-L-phenylalanine and 500 g of hydroxypropyl-β-cyclodextrin, mix them, stir them at 1000 rpm in the dark, and obtain a readily soluble complex.

[0035] Step S2, weigh 20 kg of 1,2-propylene glycol and 15 kg of mannitol respectively, mix them evenly to obtain a mixed solution, heat it under reflux at 60° C. for 4 h to fully dissolve it, and then add a 1000 m 2 / g of activated carbon was refluxed for 1h, and finally filtered three times using an organic filter membrane to obtain a freeze-dried liquid.

[0036] Step S3, the prepared lyophilized liquid is packaged into sterile brown ampoules according to standard weight, and the ampoules are then placed on a freeze dryer, first pre-frozen at a pressure of 10Pa and a temperature of -30°C for 2h, then sublimated and dried at a pressure of 5Pa and a temperature of -15°C for 5h, and finally desorbed and dried at a pressure of 5Pa and a temperature of 18°C ​​for 10h to obtain a 4-boron-L-phenylalanine lyophilized preparation.

[0037] Example 2

[0038] The preparation method of the 4-boron-L-phenylalanine lyophilized preparation of this embodiment is as follows:

[0039] Step S1, weigh 500 g of 4-boron-L-phenylalanine and 1500 g of hydroxypropyl-β-cyclodextrin, mix them, stir them at 1000 rpm in the dark, and obtain a readily soluble complex.

[0040] Step S2, weigh 25 kg of 1,2-propylene glycol and 25 kg of mannitol respectively, mix them evenly to obtain a mixed solution, heat it under reflux at 80° C. for 6 h to fully dissolve it, and then add a 1500 m 2 / g of activated carbon was refluxed for 3h, and finally filtered 5 times with an organic filter membrane to obtain a freeze-dried liquid.

[0041] Step S3, the prepared lyophilized liquid is packaged into sterile brown ampoules according to standard weight, and the ampoules are then placed on a freeze dryer, first pre-frozen at a pressure of 20Pa and a temperature of -60°C for 4 hours, then sublimated and dried at a pressure of 10Pa and a temperature of -35°C for 10 hours, and finally desorbed and dried at a pressure of 10Pa and a temperature of 35°C for 15 hours to obtain a lyophilized preparation of 4-boron-L-phenylalanine.

[0042] Example 3

[0043] The preparation method of the 4-boron-L-phenylalanine lyophilized preparation of this embodiment is as follows:

[0044] Step S1, weigh 500 g of 4-boron-L-phenylalanine and 1000 g of hydroxypropyl-β-cyclodextrin, mix them, stir them at 1000 rpm in the dark, and obtain a readily soluble complex.

[0045] Step S2, weigh 22 kg of 1,2-propylene glycol and 24 kg of mannitol respectively, mix them evenly to obtain a mixed solution, heat it under reflux at 70° C. for 5 h to fully dissolve it, and then add a 1200 m 2 / g of activated carbon was refluxed for 2h, and finally filtered four times using an organic filter membrane to obtain a freeze-dried liquid.

[0046] Step S3, the prepared lyophilized liquid is packaged into sterile brown ampoules according to standard weight, and the ampoules are then placed on a freeze dryer, first pre-frozen at a pressure of 15Pa and a temperature of -50°C for 3h, then sublimated and dried at a pressure of 8Pa and a temperature of -20°C for 6h, and finally desorbed and dried at a pressure of 8Pa and a temperature of 20°C for 12h to obtain a 4-boron-L-phenylalanine lyophilized preparation.

[0047] The 4-boron-L-phenylalanine freeze-dried preparations prepared in Examples 1 to 3 were re-dissolved in water, and the boron content, impurity content and light transmittance thereof were tested by conventional methods. The test results of the boron content after re-dissolution are shown in Table 1:

[0048] name Theoretical boron content (mg / ml) Actual boron content (mg / ml) Example 1 4 3.92 Example 2 4 3.95 Example 3 4 3.94

[0049] Table 1

[0050] It can be seen from the data in Table 1 that the actual boron content of the 4-boron-L-phenylalanine freeze-dried preparation prepared by the preparation method of the present application after reconstitution is not much different from the theoretical boron content, the reconstitution effect is good, and the problem of the insolubility of 4-boron-L-phenylalanine is effectively solved.

[0051] The test results of impurity content and transmittance after re-dissolution are shown in Table 2:

[0052] name Standing time (h) Clarity Light transmittance (%) Impurity content (%) Example 1 20 clarify 99.8 0.02 Example 2 20 clarify 99.6 0.01 Example 3 20 clarify 99.8 0.03

[0053] Table 2

[0054] It can be seen from Table 2 that after re-dissolution, the solution remained clear after standing at room temperature for 20 hours, with a transmittance greater than 99.6% and an impurity content as low as 0.01%.

[0055] It should be noted that the impurities in Table 2 mainly include organic impurities, inorganic impurities, residual solvents and moisture. Organic impurities include starting material impurities such as iodide residues, alkyl impurities, intermediate impurities such as N-Boc-4-boric acid-L-phenylalanine, reaction by-products such as boric acid diesters, and isomer impurities. Inorganic impurities include metal ion impurities such as magnesium, lithium, activated carbon, etc. If the freeze-drying conditions are not ideal, the moisture content in the product may also exceed the standard.

[0056] The above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the present invention. Although the present invention is described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the purpose and scope of the technical solutions of the present invention, which should be included in the scope of the claims of the present invention. The techniques, shapes, and structural parts not described in detail in the present invention are all known technologies.

Claims

1. A lyophilized preparation of 4-boron-L-phenylalanine, characterized in that: The invention comprises a soluble complex, 1,2-propylene glycol and mannitol. The soluble complex is formed by encapsulating 4-boron-L-phenylalanine by hydroxypropyl-β-cyclodextrin. The mass ratio of 4-boron-L-phenylalanine, hydroxypropyl-β-cyclodextrin, 1,2-propylene glycol and mannitol is 1:(1-3):(40-50):(30-50).

2. A lyophilized preparation of 4-boron-L-phenylalanine according to claim 1, characterized in that: The mass ratio of 4-boron-L-phenylalanine, hydroxypropyl-β-cyclodextrin, 1,2-propylene glycol and mannitol is 1:2:44:

48.

3. A method for preparing a lyophilized preparation of 4-boron-L-phenylalanine, characterized in that: The following steps are involved: Weigh 4-boron-L-phenylalanine and hydroxypropyl-β-cyclodextrin respectively and mix them to obtain a readily soluble complex; The easily soluble complex is added to a mixed solution containing 1,2-propylene glycol and mannitol, heated at 60-80°C for 4-6 hours and refluxed, activated carbon is added and refluxed for 1-3 hours, and then an organic filter membrane is used to obtain a lyophilized solution; The lyophilized liquid is divided into sterile containers, and the sterile containers are then installed on a freeze dryer, and are successively subjected to pre-freezing, sublimation drying, and analytical drying treatments to obtain a 4-boron-L-phenylalanine lyophilized preparation.

4. The method for preparing a lyophilized 4-boron-L-phenylalanine preparation according to claim 3, characterized in that: The pre-freezing conditions are set as follows: temperature is -30 to -60°C, pressure is 10 to 20 Pa, and time is 2 to 4 hours.

5. The method for preparing a lyophilized 4-boron-L-phenylalanine preparation according to claim 4, characterized in that: The sublimation drying conditions are set as follows: temperature is -15 to -35°C, pressure is 5 to 10 Pa, and time is 5 to 10 hours.

6. The method for preparing a lyophilized 4-boron-L-phenylalanine preparation according to claim 5, characterized in that: The analytical drying conditions are set as follows: temperature is 18-35° C., pressure is 5-10 Pa, and time is 10-15 h.

7. The method for preparing a lyophilized 4-boron-L-phenylalanine preparation according to claim 3, characterized in that: The mass ratio of the 4-boron-L-phenylalanine, hydroxypropyl-β-cyclodextrin, 1,2-propylene glycol and mannitol is 1:(1-3):(40-50):(30-50).

8. The method for preparing a lyophilized 4-boron-L-phenylalanine preparation according to claim 3, characterized in that: The specific surface area of ​​the activated carbon is 1000-1500m 2 / g.

9. The method for preparing a lyophilized 4-boron-L-phenylalanine preparation according to claim 3, characterized in that: The organic filter membrane is filtered 3 to 5 times.