A method for reducing the osmotic pressure of a boron carrier injection

Abstract

The present invention relates to a method for reducing the osmotic pressure of a boron carrier injection. The method includes using nanofiltration technology. This method is simple and easy to implement, has a good effect on reducing the osmotic pressure, good stability, and high safety, and is suitable for industrial production.

Description

Technical Field

[0001] The present invention relates to a method for reducing the osmotic pressure of a boron carrier injection, and specifically relates to a method for reducing the osmotic pressure of a boron carrier injection by using nanofiltration technology, belonging to the field of pharmaceutical technology. Background Art

[0002] The chemical name of L-BPA is 4-dihydroxyboranyl-L-phenylalanine, and its structural formula is as follows:

[0003]

[0004] L-BPA is a boron carrier used in boron neutron capture therapy. Boron neutron capture therapy requires that 10 9 boron atoms reach tumor cells. However, due to the low boron loading amount of L-BPA and limited targeting ability, a large dose of 100-500 mg / kg is required. Since L-BPA has poor water solubility, a large amount of sodium hydroxide is needed to dissolve it during the preparation process. After forming a complex with polyhydroxy sugars, hydrochloric acid is used to adjust the pH back to an acceptable range. Under such process conditions, a large amount of excess sodium chloride is generated, resulting in a high osmotic pressure of the preparation and a greater risk during use.

[0005] US6169076B1 patent provides a method for reducing osmotic pressure, that is, after preparing the L-BPA-sugar complex, instead of using hydrochloric acid for callback, an H + type cation exchange resin is used to exchange Na + in the solution, and at the same time, the pH of the solution is reduced. However, this method uses a large amount of resin, the concentration of the prepared L-BPA solution is low, and it is difficult to control the end point. It is necessary to monitor the pH in real time. Once the pH is too low, precipitation will occur. Therefore, the practicality of this method is limited.

[0006] Currently, when using L-BPA preparations clinically, they are all prepared and used immediately. The main reason is that the L-BPA-fructose solution is sensitive to temperature and is not easy to store. Patent CN103100094B provides a freeze-drying method, but this method is only applicable to low concentrations and small batches, and is not suitable for large-scale production. Summary of the Invention Overview of the Invention

[0008] The present invention aims to provide a method for reducing the osmotic pressure of L-BPA injection that is safe, non-toxic, simple to operate, and has good controllability. The present invention applies nanofiltration technology to the preparation process of the injection, removes the excess sodium chloride in the injection, and while reducing the osmotic pressure, can achieve a concentration effect, facilitating the subsequent treatment of the injection. This method is easy to control during the production process, has a good effect on reducing the osmotic pressure, and is suitable for large-scale production.

[0009] Since the L-BPA solution is unstable at room temperature and difficult to store, it is prepared into a freeze-dried powder injection, which avoids the cumbersome preparation process during clinical use and is convenient for storage and transportation.

[0010] The present invention provides a method for reducing the osmotic pressure of L-BPA injection, including using nanofiltration technology to obtain an L-BPA injection with an osmotic pressure of 280 mosm / kg - 330 mosm / kg. This method is simple and easy to implement, has good stability and high safety, and is suitable for industrial production.

[0011] Term Definition

[0012] The term "comprising" or "including" is an open expression, that is, it includes the content specified in the present invention, but does not exclude other aspects of the content.

[0013] In the context of the present invention, all the numbers disclosed herein are approximate values whether or not words such as "about" or "approximately" are used. The numerical value of each number may have a difference of less than 10% or a reasonable difference considered by those skilled in the art, such as a difference of 1%, 2%, 3%, 4% or 5%.

[0014] In the present invention, min represents minute, h represents hour, mg represents milligram, g represents gram, mL represents milliliter, Mpa represents megapascal, rpm represents revolutions per minute, rps represents revolutions per second, mg / kg represents milligram per kilogram, μl represents microliter, M represents mole per liter, L / min represents liter per minute, ml / min represents milliliter per minute, mm represents millimeter, nm represents nanometer, mg / kg / d represents milligram per kilogram per day, w represents watt, °C represents degree Celsius, W represents weight, V represents volume, and mosm / kg represents milliosmole per kilogram.

[0015] In the present invention, the concentration of the active ingredient L-BPA or the cosolvent concentration is the ratio of the weight of the active ingredient or the cosolvent to the total volume of the injection, that is, W / V, mg / mL. For example, 25 mg / mL of L-BPA means that 1 mL of the injection contains 25 mg of L-BPA.

[0016] In the examples of the present invention, "-" represents no content. Detailed Description of the Invention

[0018] Based on the deficiencies of the prior art, through in-depth investigation and research, the present invention applies nanofiltration technology to the preparation process of the injection. While removing excess sodium chloride in the injection and reducing the osmotic pressure, it can achieve a concentration effect, which is convenient for the subsequent treatment of the injection. This method is easy to control during the production process, has a good effect of reducing the osmotic pressure, and is suitable for large-scale production.

[0019] The present invention provides a method for reducing the osmotic pressure of L-BPA injection, including using nanofiltration technology to obtain an L-BPA injection with an osmotic pressure of 280 mosm / kg - 330 mosm / kg. In some embodiments, nanofiltration technology is used to obtain an osmotic pressure of 309 mosm / kg - 323 mosm / kg. In some embodiments, nanofiltration technology is used to obtain an L-BPA injection with an osmotic pressure of 310 mosm / kg - 318 mosm / kg. This method is simple and easy to implement, has good stability and high safety, and is suitable for industrial production. In some embodiments, nanofiltration technology is used to obtain an L-BPA injection with an osmotic pressure of 323 mosm / kg, 309 mosm / kg, 310 mosm / kg, 317 mosm / kg, 318 mosm / kg, or 312 mosm / kg.

[0020] In some embodiments, the pressure used in the nanofiltration technology is 2.0 Mpa - 5.0 Mpa. In some embodiments, the pressure used in the nanofiltration technology is 2.0 Mpa - 4.0 Mpa. In some embodiments, the pressure used in the nanofiltration technology is 2.0 Mpa - 3.0 Mpa; in some embodiments, the pressure used in the nanofiltration technology is 3.0 Mpa - 4.0 Mpa; in some embodiments, the pressure used in the nanofiltration technology is 3.0 Mpa - 5.0 Mpa; in some embodiments, the pressure used in the nanofiltration technology is 4.0 Mpa - 5.0 Mpa. In some embodiments, the pressure used in the nanofiltration technology is 2.0 Mpa, 3.0 Mpa, or 4.0 Mpa. A certain pressure range can improve the filtration efficiency of sodium chloride and will not cause the filter membrane to rupture.

[0021] In some embodiments, the rotation speed used in the nanofiltration technology is 10 rpm - 50 rpm. In some embodiments, the rotation speed used in the nanofiltration technology is 10 rpm - 30 rpm. In some embodiments, the rotation speed used in the nanofiltration technology is 10 rpm - 15 rpm; in some embodiments, the rotation speed used in the nanofiltration technology is 10 rpm - 20 rpm; in some embodiments, the rotation speed used in the nanofiltration technology is 15 rpm - 20 rpm; in some embodiments, the rotation speed used in the nanofiltration technology is 15 rpm - 30 rpm; in some embodiments, the rotation speed used in the nanofiltration technology is 20 rpm - 30 rpm; in some embodiments, the rotation speed used in the nanofiltration technology is 20 rpm - 50 rpm. In some embodiments, the rotation speed used in the nanofiltration technology is 10 rpm, 15 rpm, or 20 rpm. A certain pressure range can improve the filtration efficiency of sodium chloride and shorten the time required for nanofiltration.

[0022] In some embodiments, the frequency at which the nanofiltration technology is employed is 5 Hz - 50 Hz, or 20 Hz - 50 Hz. In some examples, the frequency at which the nanofiltration technology is employed is 50 Hz.

[0023] In some embodiments, according to the ratio of the weight of L-BPA to the total volume of the injection, before the nanofiltration technology is adopted, the concentration range of L-BPA is 10.0 mg / mL - 50.0 mg / mL (W / V). Before the nanofiltration technology is adopted, in some embodiments, the L-BPA concentration range is 15.0 mg / mL - 30.0 mg / mL; in some embodiments, the L-BPA concentration range is 10.0 mg / mL - 20.0 mg / mL; in some embodiments, the L-BPA concentration range is 10.0 mg / mL - 30.0 mg / mL; in some embodiments, the L-BPA concentration range is 10.0 mg / mL - 40.0 mg / mL; in some embodiments, the L-BPA concentration range is 20.0 mg / mL - 30.0 mg / mL; in some embodiments, the L-BPA concentration range is 20.0 mg / mL - 40.0 mg / mL; in some embodiments, the L-BPA concentration range is 20.0 mg / mL - 50.0 mg / mL; in some embodiments, the L-BPA concentration range is 30.0 mg / mL - 40.0 mg / mL; in some embodiments, the L-BPA concentration range is 30.0 mg / mL - 50.0 mg / mL; in some embodiments, the L-BPA concentration range is 40.0 mg / mL - 50.0 mg / mL. Before the nanofiltration technology is adopted, in some examples, the L-BPA concentration range is 25.0 mg / mL or 50.0 mg / mL.

[0024] Before the nanofiltration technology is adopted, the osmotic pressure of the L-BPA injection is greater than 330 mosm / kg.

[0025] In some embodiments, the preparation method of the L-BPA injection includes: adding L-BPA to an appropriate amount of water for dispersion, adding sodium hydroxide to completely dissolve it, then adding sugar alcohol substances, and after complete dissolution, adding hydrochloric acid to adjust the pH to obtain the injection before the nanofiltration technology is adopted.

[0026] In some embodiments, the sugar alcohol substances include at least one selected from fructose, sorbitol, and mannitol. In some examples, the sugar alcohol substance includes fructose; in some examples, the sugar alcohol substance includes sorbitol; in some examples, the sugar alcohol substance includes mannitol.

[0027] In some embodiments, the concentration of the sugar alcohol substance is 0.80 to 1.30 times the concentration of L-BPA. In some embodiments, the concentration of the sugar alcohol substance is 0.85 to 1.10 times the concentration of L-BPA; in some embodiments, the concentration of the sugar alcohol substance is 0.80 to 0.90 times the concentration of L-BPA; in some embodiments, the concentration of the sugar alcohol substance is 0.80 to 1.00 times the concentration of L-BPA; in some embodiments, the concentration of the sugar alcohol substance is 0.80 to 1.10 times the concentration of L-BPA; in some embodiments, the concentration of the sugar alcohol substance is 0.90 to 1.00 times the concentration of L-BPA; in some embodiments, the concentration of the sugar alcohol substance is 0.90 to 1.10 times the concentration of L-BPA; in some embodiments, the concentration of the sugar alcohol substance is 0.90 to 1.30 times the concentration of L-BPA; in some embodiments, the concentration of the sugar alcohol substance is 1.00 to 1.10 times the concentration of L-BPA. In some examples, the concentration of the sugar alcohol substance is 0.78 times, 0.86 times, 0.95 times, 1.12 times, or 1.30 times the concentration of L-BPA.

[0028] In some examples, the method for preparing the L-BPA injection includes: adding the prescribed amount of L-BPA to an appropriate amount of water for dispersion, adding sodium hydroxide to completely dissolve it, then adding fructose, stirring until completely dissolved, and adding hydrochloric acid to adjust the pH to 7.8 to obtain the injection before using the nanofiltration technology.

[0029] In some embodiments, after using the nanofiltration technology, the L-BPA injection is further prepared into a freeze-dried powder injection.

[0030] In some embodiments, the preparation steps of the freeze-dried powder injection include:

[0031] (1) Pre-freezing at -50°C to -35°C and performing rewarming at -15°C to -5°C;

[0032] (2) Performing primary sublimation at -20°C to -10°C and then cooling to -25°C to -15°C for sublimation;

[0033] (3) Heating to 30°C to 50°C for drying.

[0034] In some embodiments, the preparation steps of the freeze-dried powder injection include:

[0035] (1) Pre-freezing at -50°C to -35°C and performing rewarming at -15°C to -5°C;

[0036] (2) After sublimation at -20°C to -10°C for 50 h to 70 h, the temperature is then decreased to -25°C to -15°C for sublimation;

[0037] (3) The temperature is raised to 30°C to 50°C for drying for 10 h to 40 h.

[0038] The nanofiltration technology adopted in the present invention is simple and easy to implement, has good stability and high safety, and is suitable for industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 It is the low-temperature DSC spectrum of the solution before nanofiltration.

[0040] Figure 2 It is the low-temperature DSC spectrum of the solution after nanofiltration. DETAILED DESCRIPTION OF THE INVENTION

[0041] In order to enable those skilled in the art to better understand the technical solutions of the present invention, some non-limiting embodiments are further disclosed below to further elaborate on the present invention in detail.

[0042] All the reagents used in the present invention can be purchased from the market or can be prepared by the methods described in the present invention.

[0043] Example 1 (Nanofiltration Process of L-BPA Injection)

[0044] The prescription is shown in Table 1.

[0045] Table 1

[0046]

[0047] Preparation process:

[0048] (1) Weigh the prescribed amount of the active ingredient and disperse it in an appropriate amount of water;

[0049] (2) Add an appropriate amount of sodium hydroxide and stir until the active ingredient is completely dissolved;

[0050] (3) Add the prescribed amount of fructose and stir until completely dissolved;

[0051] (4) Adjust the pH to 7.8 with hydrochloric acid, make up the volume, and filter;

[0052] (5) Dilute the solution to the following concentration (i.e., the initial nanofiltration concentration) and perform nanofiltration: control the nanofiltration pressure at 2.0 to 4.0 Mpa and the rotation speed at 10 to 20 rpm (the total amount of the drug added during nanofiltration with different processes is the same. Adjust the rotation speed to the corresponding value at the beginning of nanofiltration. After exhausting air at low pressure, then slowly increase the pressure to the corresponding value);

[0053] (6) During the nanofiltration process, the flow rate was recorded every 5 minutes. When the recorded flow rate was lower than 10 ml / min, the nanofiltration was stopped, the nanofiltration permeate was discarded, the nanofiltration circulating liquid was collected, and the nanofiltration circulating liquid was tested. The test items included: L-BPA content, Na + The content, osmotic pressure corresponding to 30mg / ml and 50mg / ml L-BPA concentration. The nanofiltration process is shown in Table 2.

[0054] Table 2

[0055]

[0056]

[0057] The experimental results are shown in Table 3.

[0058] Table 3

[0059]

[0060] In the process of studying the L-BPA prescription, it was found that L-BPA and fructose are solubilized by complexation at a molar ratio of 1:1, that is, L-BPA forms a sodium salt under alkaline conditions and then complexes with sugar alcohols. Since the amount of NaOH added when L-BPA is completely dissolved is excessive, hydrochloric acid needs to be added to neutralize the excess NaOH and adjust the pH to near neutrality, which will produce a large amount of excess NaCl, causing excessive osmotic pressure. In clinical practice, intravenous infusion is required to be as close to the isotonic range as possible (280-330mosm / kg), especially when the administration volume is large, otherwise it is easy to cause adverse reactions. At present, the concentration of L-BPA in clinical trials is about 30mg / ml, so it is necessary to ensure that the product is in an isotonic state within this concentration range.

[0061] According to the above experimental scheme and its results, we can know that:

[0062] (1) Processes 1 to 4 are single factors that investigate the effect of initial nanofiltration concentration on the product after nanofiltration. From the test results, the higher the initial nanofiltration concentration, the shorter the time required for nanofiltration, and the higher the concentration after nanofiltration, but the osmotic pressure is also relatively high, and the sodium ion removal effect is poor. Among them, process 3 and process 4 basically meet the requirements, that is, when the initial nanofiltration concentration is below 50 mg / ml, they can both meet the effect of removing sodium chloride, so that the osmotic pressure of 30 mg / ml L-BPA solution is in the range of 280-330mosm / kg. However, process 4 is more preferred. When investigating the initial nanofiltration concentration, only four concentrations between 25-100 mg / ml were investigated, and lower concentration conditions were not investigated. This is because in actual production, the lower the initial nanofiltration concentration, the larger the sample volume that needs to be processed, which may exceed the volume of the production preparation tank or intermediate storage tank, and prolonging the nanofiltration time has no practical significance;

[0063] (2) Processes 4 / 5 / 6 were used to investigate the effect of pressure on the product after nanofiltration in a single-factor study. From the test results, the higher the pressure during nanofiltration, the faster the treatment, the shorter the time consumed, and the treatment effects of Processes 4 / 5 / 6 were basically the same. However, in the production process, the one with the shortest time consumption was preferably selected on the basis of achieving the effect: because in the production process of injectables, the aseptic risk needs to be considered, and the longer the time consumption, the greater the risk. Therefore, the nanofiltration pressure was preferably 4.0 Mpa. Only the pressures of 2 / 3 / 4 Mpa were studied here, and higher pressure conditions were not studied. The reason is that the maximum pressure that the nanofiltration membrane element can withstand is 4.1 Mpa. Under higher pressure conditions, quality problems may occur, so the upper limit of pressure study was set at 4 Mpa; lower pressures were not studied mainly because the treatment speed at low pressures is too slow, which will prolong the treatment time and consume too much time, and it is not necessary in the actual production process;

[0064] (3) Processes 4 / 7 / 8 were used to investigate the effect of rotation speed on the product after nanofiltration in a single-factor study. From the test results, the greater the rotation speed during nanofiltration, the greater the feeding power, the faster the treatment, and the shorter the time consumed. The effects of Processes 4 / 7 / 8 in removing sodium chloride were basically the same. However, in the actual production process, the one with the shortest time consumption was preferably selected on the basis of achieving the effect. Therefore, the rotation speed can be preferably the maximum rotation speed, that is, operating at the maximum rotation speed within the range allowed by the equipment to achieve the effect of rapid treatment.

[0065] Detection method for L-BPA content and related substances (common in examples):

[0066] Chromatographic conditions:

[0067] High-performance liquid chromatograph (Agilent HPLC-DAD)

[0068] Chromatographic column: Welch Ultimate AQ-C18(D), 4.6 * 250 mm, 5 μm

[0069] Detector: DAD detector, 210 nm Flow rate: 1.0 ml / min

[0070] Mobile phase A: 10 mmol / L pH2.8 KH2PO4 Buffer Mobile phase B: Methanol

[0071] Diluent: 0.1M HCl Column temperature: 35 °C

[0072] Gradient elution conditions for related substances are shown in Table 4, the running time is 55 min, and the injection volume is 10 μl.

[0073] Table 4

[0074]

[0075]

[0076] Isocratic elution of content, mobile phase A: mobile phase B = 95:5, running time is 20 min, injection volume is 10 μl

[0077] L-BPA osmotic pressure detection method: Dilute the L-BPA solution with known content to 30 mg / ml and 50 mg / ml respectively according to the concentration, take 100 μl and detect it with a freezing point osmometer (osmometer: Loser OM819).

[0078] Na + Content detection method (common in examples): ICP-MS: 7800 ICP-MS, Agilent; method parameters are shown in Table 5.

[0079] Table 5

[0080]

[0081] Example 2 (Effect of nanofiltration of L-BPA injection on freeze-drying process)

[0082] Freeze-drying liquid drugs can improve the stability of some unstable drugs and is conducive to transportation and storage. The freeze-drying technology is mainly divided into three steps: pre-freezing, primary sublimation (primary drying) and secondary sublimation (desorption drying). Pre-freezing is an important step in the freeze-drying process. Only after the liquid product is completely frozen can primary sublimation and secondary sublimation be carried out to complete the freeze-drying. Pre-freezing means placing the drug solution at a low temperature to completely freeze it and form a solid matrix. This temperature is generally below the eutectic point or its solid glass transition temperature (Tg') (related to the solid crystalline state: crystalline substances are below the eutectic point, and amorphous substances correspond to Tg'. Tg' is the temperature at which the amorphous solid matrix changes from the glassy state to the high elastic state, generally a temperature range). Therefore, for the pre-freezing process, the lower the Tg' of the solution, the more difficult the pre-freezing process, the more likely it is to collapse during the primary sublimation process, and the greater the difficulty of freeze-drying. Increasing the Tg' of the solution is beneficial to freeze-drying. The Tg' of the solution can be measured by differential scanning calorimetry (DSC).

[0083] The prescription is shown in Table 6.

[0084] Table 6

[0085]

[0086] Preparation process:

[0087] (1) Weigh the prescription amount of the active ingredient and disperse it in purified water,

[0088] (2) Add an appropriate amount of sodium hydroxide and stir until the active ingredient is completely dissolved;

[0089] (3) Add the prescribed amount of fructose and stir until completely dissolved;

[0090] (4) Adjust the pH to 7.8 with hydrochloric acid, make up the volume to the full volume, and filter;

[0091] (5) Take a part of the sample for nanofiltration. The nanofiltration parameters are the same as those in Process 4 of the example. Measure the low-temperature DSC of the sample before and after nanofiltration;

[0092] (6) Respectively fill the solutions before and after nanofiltration (adjust the concentration to be the same) into medium-borosilicate glass injection bottles, with 15 ml filled in each bottle, and carry out freeze-drying. The freeze-drying process is as follows. Examine the appearance and reconstitution of the product after freeze-drying. The freeze-drying process is shown in Table 7.

[0093] Table 7

[0094]

[0095]

[0096] Experimental results

[0097] Figure 1 is the low-temperature DSC spectrum of the solution before nanofiltration, Figure 2 is the low-temperature DSC spectrum of the solution after nanofiltration. The spectrum results show that the Tg’ of the solution before nanofiltration is between -42.54 °C and -39.75 °C, and the Tg’ of the solution after nanofiltration is between -31.74 °C and -29.31 °C. That is, the Tg’ temperature of the solution after nanofiltration will rise, the difficulty of pre-freezing is reduced, and the difficulty of freeze-drying is also reduced.

[0098] (2) The comparison of the products after freeze-drying of the solution with or without nanofiltration is shown in Table 8. The results show that the appearance of the product after freeze-drying of the solution after nanofiltration is better, the reconstitution time is shorter, and it is more convenient to use.

[0099] Table 8

[0100]

[0101] Stability of Example 3 (stability of solution and freeze-dried powder)

[0102] The prescription is shown in Table 9.

[0103] Table 9

[0104]

[0105] Preparation process:

[0106] (1) Weigh the prescribed amount of the active ingredient and disperse it in an appropriate amount of purified water;

[0107] (2) Add an appropriate amount of sodium hydroxide and stir until the active ingredient is completely dissolved;

[0108] (3) Add the prescribed amount of fructose and stir until completely dissolved;

[0109] (4) Adjust the pH to 7.8 with hydrochloric acid, make up the volume, and filter;

[0110] (5) Fill the solution in Prescription 1 into vials, 5 ml per vial, stopper, and crimp the cap; subject the solution in Prescription 2 to nanofiltration according to Process 4 in Example 1, and lyophilize the product after nanofiltration according to the lyophilization process in Example 2;

[0111] (6) Examine the stability of the products obtained from Prescription 1 and Prescription 2 at 60 °C respectively.

[0112] The experimental results are shown in Table 10. According to the results, for Prescription 1, i.e., the L-BPA-fructose solution, after being placed at 60 °C for 5 days, the content decreased by about 46%, and the color changed from light yellow to dark yellow; for Prescription 2, after being placed at 60 °C for 10 days, the content showed no obvious change, the total impurities increased by about 1%, and it had good stability and was more suitable for long-term storage.

[0113] Table 10

[0114]

[0115] Toxicity experiment in Example 4 (improvement of toxicity after adding the nanofiltration process)

[0116] The prescriptions are shown in Table 11.

[0117] Table 11

[0118]

[0119] Preparation process: same as in Example 2, measure the osmotic pressure of the unnanofiltrated samples and nanofiltrated samples under different concentration conditions respectively.

[0120] The experimental results are shown in Table 12. According to the results, the osmotic pressure of the samples after nanofiltration treatment decreased significantly.

[0121] Table 12

[0122]

[0123]

[0124] Conduct toxicity experiments on rats and dogs with the unnanofiltrated and nanofiltrated samples respectively.

[0125] Experimental conditions and procedures:

[0126] Single-day administration toxicity in rats: Sprague-Dawley (SD) rats were used in the experiment; infusion was performed at a concentration of 100 mg / ml, the administration dose was 20 ml / kg / time, administered in two doses with an interval of 5 - 6 h, and the total administration dose was 4000 mg / kg / d. During the administration period, the death situation and clinical symptoms of the animals were observed, body weight and food intake were measured, and the observation period was 14 days.

[0127] Repeated administration toxicity in rats: Sprague-Dawley (SD) rats were used in the experiment; infusion was performed at a concentration of 100 mg / ml, the administration dose was 20 ml / kg / time, administered in two doses with an interval of 5 - 6 h, and the total administration dose was 4000 mg / kg / d. During the administration period, the death situation and clinical symptoms of the animals were observed, body weight and food intake were measured. The group administered with the non-ultrafiltered solution was continuously administered for 4 days, and the group administered with the ultrafiltered solution was continuously administered for 7 days.

[0128] Single-day administration toxicity in dogs: Beagle dogs were used in the experiment; for the non-ultrafiltered solution, intravenous infusion was performed at a concentration of 100 mg / ml, 10 mg / kg / time, administered in two doses with a total dose of 2000 mg / kg; for the ultrafiltered solution, administration was carried out in two groups. ① Intravenous infusion was performed at a concentration of 100 mg / ml, 20 mg / kg / time, with a total dose of 2000 mg / kg. ② Intravenous infusion was performed at a concentration of 75 mg / ml, 40 ml / kg / time, with a total dose of 3000 mg / kg. During the administration period, the death situation and clinical symptoms of the dogs were observed, and body weight and food intake were measured.

[0129] The rats and Beagle dogs used in the experiment were quarantined regularly, and the breeding environment met the specifications of GB 14925-2010 Laboratory Animal Environment and Facilities.

[0130] Experimental results:

[0131] Single-day administration toxicity in rats: In the non-ultrafiltered solution group, after a single administration, the animals showed reduced activity and unsteady gait, and individual animals died 5 min after the second administration; in the ultrafiltered solution group, no obvious changes in activity status and obvious toxic reactions were observed after two administrations. The experimental results indicate that the toxicity of the ultrafiltered solution has decreased.

[0132] Repeated administration toxicity in rats: In the non-ultrafiltered solution group, continuous administration was carried out for 4 days, and during the clinical observation period, the animals showed reduced activity and increased urination; in the ultrafiltered solution group, continuous administration was carried out for 7 days, and the animals had good tolerance and no obvious toxic reactions were observed. The experimental results indicate that the tolerance of the animals to the solution has been improved after ultrafiltration.

[0133] Canine single-dose administration toxicity: After two doses of the non-nanofiltration solution group, the animals showed vomiting, scleral congestion, and yellow eye discharge; in the two dosing groups after nanofiltration treatment, only slight scleral swelling was observed, and there were no other adverse reactions. Theoretically, single-dose administration is riskier than two-dose administration. However, in the 2000 mg / kg dosing group after nanofiltration treatment, the single-dose toxic and side reactions were milder than those of the non-nanofiltration two-dose administration, indicating that the toxicity of the solution after nanofiltration was significantly reduced.

[0134] The above experimental results indicate that the osmotic pressure of the solution after nanofiltration decreased significantly, and the toxicity to animals also decreased.

[0135] Example 5 Prescription Screening (Screening of Cosolvents or Solubilizers)

[0136] The prescriptions are shown in Tables 13 and 14.

[0137] Table 13

[0138]

[0139] Table 14

[0140]

[0141] Note: HP-β-CD is hydroxypropyl-β-cyclodextrin, and SBECD is sulfobutyl-β-cyclodextrin.

[0142] Preparation process:

[0143] (1) Weigh the prescribed amount of the active ingredient and disperse it in purified water;

[0144] (2) Add an appropriate amount of sodium hydroxide and stir until the active ingredient is completely dissolved;

[0145] (3) Add the prescribed amount of cosolvent or solubilizer and stir until completely dissolved;

[0146] (4) Use hydrochloric acid to adjust the pH back to about 7.8, and observe whether the active ingredient precipitates during the adjustment process and the state of the solution after the adjustment is completed.

[0147] L-BPA is difficult to dissolve in water and requires a cosolvent or solubilizer to increase its solubility. According to the clinically commonly used dose, with a fixed concentration of 30 mg / ml, the effects of commonly used solubilizers or cosolvents on its dissolution were investigated. The results are shown in Table 15.

[0148] Table 15

[0149]

[0150] According to the above results, L-BPA can be solubilized well by fructose, sorbitol, mannitol, and meglumine, enabling its solubility to reach over 30 mg / ml. Among them, meglumine is a sugar alcohol derivative and a polyhydroxy strong base substance, but it has relatively high toxicity. With a large dosage of L-BPA, it is not suitable as a solubilizer for this drug. Therefore, ultimately, the suitable solubilizers for L-BPA are fructose, sorbitol, and mannitol, all of which are sugar alcohol substances.

[0151] Example 6 Prescription Screening (Screening of Solubilizer Dosage)

[0152] The prescriptions are shown in Tables 16 and 17.

[0153] Table 16

[0154]

[0155]

[0156] Table 17

[0157]

[0158] Preparation process:

[0159] (1) Weigh the prescribed amount of the active ingredient and disperse it in purified water;

[0160] (2) Add an appropriate amount of sodium hydroxide and stir until the active ingredient is completely dissolved;

[0161] (3) Add the prescribed amount of fructose and stir until completely dissolved;

[0162] (4) Use hydrochloric acid to adjust the pH back to 7.8, and observe whether there is precipitation of the active ingredient during the adjustment process and the solution state after the adjustment is completed.

[0163] The experimental results are shown in Table 18.

[0164] Table 18

[0165]

[0166]

[0167] According to the above results, the higher the proportion of fructose within a certain range, the better the solubilization effect and the better the physical stability. When the fructose concentration is 0.78 times that of L-BPA, a good solubilization effect can be achieved. When the fructose concentration is 0.78 times to 1.30 times that of L-BPA, its physical stability is good and no precipitation will occur. According to the prescription analysis, L-BPA and fructose are complexed in a mass ratio of 1:1. Slightly excessive fructose can improve the stability of the complex, and the fructose solution has a certain viscosity. Excessive fructose will make the viscosity of the injection solution too high, affecting the administration compliance. Therefore, the amount of fructose cannot be too large.

[0168] Example 7

[0169] The prescription is shown in Table 19.

[0170] Table 19

[0171]

[0172] Preparation process:

[0173] (1) Weigh the prescribed amount of API and disperse it in water for injection;

[0174] (2) Add an appropriate amount of sodium hydroxide and stir until the API is completely dissolved;

[0175] (3) Add the prescribed amount of fructose and stir until completely dissolved;

[0176] (4) Adjust the pH to 8.2 with hydrochloric acid, make up the volume, and filter through a 0.22 μm PES membrane;

[0177] (5) Conduct nanofiltration, control the nanofiltration process parameters: the pressure is 3.4 - 4.0 Mpa, the feed frequency is 50 Hz. During the nanofiltration process, add 28 L of water for injection for dilution;

[0178] (6) After nanofiltration, conduct pre-filtration and sterile filtration, fill, freeze-dry, and crimp the cap.

[0179] The experimental results are shown in Table 20.

[0180] Table 20

[0181]

[0182] The above results show that the appearance and properties of the L-BPA lyophilized powder produced by this process are good, the content and impurities meet the requirements, and the osmotic pressure meets the requirements.

[0183] The method of the present invention has been described through preferred embodiments. It is obvious that relevant personnel can make changes or appropriate alterations and combinations to the methods and applications described herein within the content, spirit, and scope of the present invention to implement and apply the technology of the present invention. Those skilled in the art can draw on the content of this article and appropriately improve the process parameters to achieve it. It should be particularly noted that all similar substitutions and alterations are obvious to those skilled in the art, and they are all considered to be included in the present invention.

[0184] In the description of this specification, the description with reference to terms such as "one embodiment", "some embodiments", "some implementation manners", "some implementation schemes", "examples", "specific examples", or "some examples", etc. means that the specific features, structures, materials, or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials, or characteristics described can be combined in a suitable manner in any one or more embodiments or examples. In addition, without contradiction, those skilled in the art can combine and combine the different embodiments or examples described in this specification and the features of different embodiments or examples.

[0185] Although the embodiments of the present invention have been shown and described above, it can be understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those of ordinary skill in the art can make changes, modifications, substitutions, and variations to the above embodiments within the scope of the present invention.

Claims

1. A method for reducing the osmotic pressure of L-BPA injection, comprising: using nanofiltration technology to obtain an L-BPA injection with an osmotic pressure of 280 mosm / kg - 330 mosm / kg.

2. The method according to claim 1, wherein the pressure used in the nanofiltration technology is 2.0 Mpa - 5.0 Mpa.

3. The method according to claim 1 or 2, wherein the rotation speed used in the nanofiltration technology is 10 rpm - 50 rpm.

4. The method according to claim 1 or 2, wherein the frequency used in the nanofiltration technology is 5 Hz - 50 Hz.

5. The method according to claim 1 or 2, according to the ratio of the weight of L-BPA to the total volume of the injection, before using the nanofiltration technology, the concentration range of L-BPA is 10.0 mg / mL - 50.0 mg / mL.

6. The method according to claim 1 or 2, the preparation method of the L-BPA injection comprising: adding L-BPA to an appropriate amount of water for dispersion, adding sodium hydroxide to completely dissolve it, then adding sugar alcohol substances, and after completely dissolving, adding hydrochloric acid to adjust the pH.

7. The method according to claim 6, wherein the sugar alcohol substances include at least one selected from fructose, sorbitol, and mannitol.

8. The method according to claim 6, wherein the concentration of the sugar alcohol substances is 0.80 times - 1.30 times the concentration of L-BPA.

9. The method according to claim 1 or 2, after using the nanofiltration technology, the L-BPA injection is further prepared into a freeze-dried powder injection.

10. The method according to claim 9, the preparation of the freeze-dried powder injection includes the following steps: (1) Pre-freezing at -50°C to -35°C and performing rewarming at -15°C to -5°C; (2) After the first sublimation at -20°C to -10°C, then cooling to -25°C to -15°C for sublimation; (3) Heating to 30°C to 50°C for drying.

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