An in vitro release method for aprepitant injection

Abstract

The application provides an in-vitro release method of an aprepitant injection. The in-vitro release method of the application uses cation exchange resin 50WX4 as an adsorption material and a phosphate buffer with an ethanol content of 15-25% (v / v) as a release medium, can be used for detecting the in-vitro release condition of the aprepitant injection, and is simple in operation, can completely separate free drugs and milk particles, and makes the aprepitant injection release completely.

Description

TECHNICAL FIELD

[0001] The application belongs to the field of drug detection, and particularly relates to an in-vitro release method of aprepitant injection. BACKGROUND

[0002] Aprepitant injection is a kind of intravenous administration fat emulsion injection (i.e. emulsion), and quality control is an important part in its production and development, and release is one of the key items in quality control. According to the investigation, the dissolution detection of aprepitant capsules mainly adopts USPI (basket method), USPII (paddle method) and USPIII (reciprocating cylinder method), but considering the complexity of the composition and structure of the milk particles in the aprepitant injection, the above methods cannot realize the separation of the free drug released in the release medium and the milk particles, and therefore, the above methods are not applicable to the aprepitant injection.

[0003] The prior art also discloses various in-vitro release detection methods for fat emulsion, such as flow cell method, forward dialysis method, reverse dialysis method, ultrafiltration centrifugation method and the like Document 1 : FDA published aprepitant injection review report; Document 2 : Zhang Penghua et al., preparation, properties and in-vitro release of silibinin fat emulsion, Chinese pharmacist, 2014.01, volume 17, No. 4, pages 568-571; Document 3 : Zhu Wenqi et al., determination of forsythoside ester compound liposome encapsulation efficiency by HPLC-ultrafiltration centrifugation method, Chinese modern applied pharmacy, 2013.09, volume 30, No. 9, pages 1001-1004) The above methods have problems such as lag caused by drug crossing dialysis membrane, incomplete ultrafiltration centrifugation separation and complex on-line detection method.

[0004] And CN110736813A provides a new method for measuring the in-vitro release of emulsion by using resin adsorption, and the known emulsion detected includes but is not limited to propofol fat emulsion, flurbiprofen axetil emulsion, clevidipine butyrate emulsion and the like, wherein the resin used in the release method is a macroporous resin and the release medium is a phosphate buffer, but the present inventors found through experimental research that when the method in the patent is used to measure the aprepitant injection, the aprepitant injection is not completely released.

[0005] The present application aims to develop a simple and effective in-vitro release method of aprepitant injection, which is used to simulate the in-vivo release of aprepitant injection and can evaluate the quality consistency of self-prepared and reference preparations. SUMMARY

[0006] Through a large number of researches, the present inventors find that the cation exchange resin 50WX4 is used as the adsorption material, and the phosphate buffer solution with the ethanol content v / v of 15-25% is used as the releasing medium, so that the in-vitro release of the aprepitant injection can be detected, and the operation is simple, the free drug and the milk particles can be completely separated, and the aprepitant injection is completely released.

[0007] The present application provides an in-vitro release method of aprepitant injection, characterized by comprising the following steps: adding the cation exchange resin 50WX4 into the releasing medium, preheating in the water bath oscillator, then adding the aprepitant injection, and continuing to oscillate, and then taking the releasing medium at different time points, and determining the content of the aprepitant; wherein the releasing medium is the phosphate buffer solution with the ethanol content v / v of 15-25%.

[0008] Further, in the in-vitro release method, the releasing medium is diluted with the anhydrous ethanol, and filtered through the microporous filter membrane.

[0009] Further, in the in-vitro release method, the amount of the cation exchange resin is w / v of 16%-24%.

[0010] Further, in the in-vitro release method, the phosphate is selected from the group consisting of potassium dihydrogen phosphate, disodium hydrogen phosphate, sodium dihydrogen phosphate, dipotassium phosphate, ammonium dihydrogen phosphate, and sodium monohydrogen phosphate, and the potassium dihydrogen phosphate is preferred.

[0011] Further, in the in-vitro release method, the releasing medium is the phosphate buffer solution with the ethanol content v / v of 20%.

[0012] Further, in the in-vitro release method, the temperature and the speed of the water bath oscillator are 37℃ and 150rpm. Further, in the in-vitro release method, the pH of the phosphate buffer solution is 6-8, and the pH of 7.4 is preferred.

[0013] Further, in the in-vitro release method, the preparation method of the phosphate buffer solution is as follows: taking the potassium dihydrogen phosphate, dissolving in water, then adding the sodium hydroxide solution, adding water, and adjusting the pH value to 7.4 by using the 0.2mol / L sodium hydroxide solution.

[0014] Further, in the in-vitro release method, the preparation method of the phosphate buffer solution containing ethanol is as follows: taking the potassium dihydrogen phosphate, dissolving in water, then adding the sodium hydroxide solution, adding water, adjusting the pH by using the sodium hydroxide solution, adding the anhydrous ethanol, mixing, and ultrasonic degassing.

[0015] The present application has the following beneficial effects:

[0016] The release method of the present application meets the requirements of simulating the in-vivo release of the aripipitant injection liquid, and can be used for evaluating the product quality of the generic enterprises, and comparing the consistency of the quality of the generic drug preparation and the reference preparation of the original research. DETAILED DESCRIPTION

[0017] The in-vitro release method of the aripipitant injection liquid provided by the present application is described in detail below through some embodiments, but the present application is not limited to the following embodiments. Any method and material similar or equivalent to the described content can be applied to the method of the present application. The preferred implementation method and material described herein are only for demonstration.

[0018] The specific conditions not specified in the examples are carried out according to the conventional conditions or the manufacturer's recommended conditions. The reagents or instruments used are all conventional products that can be purchased on the market. Unless otherwise defined, all professional and scientific terms used herein have the same meaning as understood by those skilled in the art.

[0019] Liquid chromatography: chromatographic column: octadecylsilane bonded silica gel column (RP18, 4.6x150mm, 5μm or equivalent performance chromatographic column), mobile phase: 0.1% phosphoric acid aqueous solution: acetonitrile = 55:45, flow rate: 1.0ml / min, column temperature: 35℃, detection wavelength: 210nm, injection volume: 20μl.

[0020] Method for calculating the release rate (note: ① used to characterize the release of aripipitant injection liquid in the release medium, also to evaluate the adsorption of the resin to the free drug of aripipitant injection liquid in the medium; ② when the detected sample is free drug, the release rate in the present application is called adsorption rate, which characterizes the adsorption of the resin to the free drug):

[0021]

[0022] Er: aripipitant adsorption rate; Cn: aripipitant content at t=n; Vo: volume of release medium; Ve: sampling volume; Ci: aripipitant concentration in the release medium at the i-th sampling; n: number of sampling at different time points; m0: total mass of aripipitant loaded.

[0023] Release medium (phosphate buffer solution containing 20% ethanol (v / v)): take 13.61g of potassium dihydrogen phosphate, dissolve in 1000ml of water, add 395ml of 0.2mol / L sodium hydroxide solution, add water to 2000ml, adjust the pH value to 7.4 with 0.2mol / L sodium hydroxide solution, add 500ml of anhydrous ethanol and mix well, and ultrasonic degassing.

[0024] Aripipitant: purchased from Jiangsu Huiju Pharmaceutical Co., Ltd.

[0025] Comparative Example: Using the release method of CN110736813A Example 2

[0026] Release medium: phosphate buffer (PBS, pH 7.4, containing sodium chloride 137 mmol / L, potassium chloride 2.7137 mmol / L, Na2HPO410 mmol / L, KH2PO42 mmol / L)

[0027] Release temperature: 37℃

[0028] Resin type: macroporous adsorption resin AB-8

[0029] 8g macroporous adsorption resin was added to 200ml release medium, then 1ml aripiprazole injection was added, and the solution was stirred at 37℃. At a certain time, 0.5ml release medium was taken and diluted to 1ml with methanol. The concentration of aripiprazole was determined by HPLC, and the release rate was calculated.

[0030]

[0031] The release rate of aripiprazole injection at 5min was 6.7%, and the release rate at 30min and 60min was about 14%. The CN110736813A specification book records that the release basically reaches the platform at 30min, that is, the resin adsorption reaches saturation, and only about 14% is released. Such an experimental method cannot meet the requirements of aripiprazole injection in vitro release, and cannot reflect the characteristics of rapid release of the preparation.

[0032] Example 1: Investigation of the influence of different ethanol concentrations of release medium (sample is free drug)

[0033] Investigation of the influence of different ethanol concentrations of release medium on resin adsorption (to exclude the interference of excipients, free drug was used for experimental research).

[0034] Aripiprazole free drug: Aripiprazole 160mg was weighed into a 20ml volumetric flask, dissolved in ethanol, diluted to volume, and shaken well.

[0035] In vitro release method: 8 g of macroporous exchange resin AB-8 was weighed into a conical flask, 100 ml of release medium was added, wherein the release medium contained different concentrations of ethanol (0%, 5%, 10%, 15%, 20%, 25% or 35%), and was preheated in a 37°C, 150 rpm water bath shaker. 0.2 ml of aripiprazole free drug was added to the flask, and the 37°C, 150 rpm water bath shaker was started. At different times, 1 ml of release medium was taken, the sample was diluted with 1 ml of anhydrous ethanol, and microfiltration membrane filtration was performed. The aripiprazole content of the sample taken at different time points was determined by liquid chromatography, and the effect of different ethanol concentrations on the adsorption of free drug was investigated (adsorption rate: the adsorption of resin to aripiprazole free drug was characterized, and the calculation method was consistent with that of release rate).

[0036] Table 1 Adsorption rate of release medium with different ethanol concentrations

[0037]

[0038] According to Table 1, when the ethanol concentration in the release medium is 0%, 5% and 10%, there are floating substances on the liquid surface, that is, the free drug precipitates from the release medium and floats on the liquid surface. Although the adsorption rate at 60 min is as high as 93%-98%, the adsorption rate of the resin further obtained by calculating the free drug content in the release medium cannot truly represent the adsorption situation. When the ethanol concentration in the release medium is 35%, the adsorption of the resin decreases, indicating that the ethanol concentration is too high, which increases the solubility of the release medium to the drug, but also affects the adsorption of the resin to the drug.

[0039] Only when the ethanol concentration in the release medium is 15-25%, the state of the release medium sample taken is normal, which can meet the adsorption of the resin to the drug.

[0040] Example 2: Investigation of the effect of different types of resins (sample is free drug)

[0041] (1) Resin type:

[0042] Different types of resins on the adsorption of free drug aripiprazole: ① macroporous adsorption resin D101, purchased from the national pharmaceutical group chemical reagent co., LTD; ② macroporous adsorption resin AB-8, purchased from tianjin guangfu fine chemical research institute; ③ cation exchange resin DOwex 50WX8, H+-form, 50-100 mesh (hereinafter referred to as 50WX8), purchased from aladdin; ④ cation exchange resin DOwex 50WX4, Na+-form, 200-400 mesh (hereinafter referred to as 50WX4), purchased from Sigma; ⑤ macroporous adsorption resin H-4, purchased from hecheng new materials; ⑥ macroporous adsorption resin NKA-II, purchased from hecheng new materials; ⑦ cation exchange resin 50WX2, Na+-form, 100-200 mesh (hereinafter referred to as 50WX2), purchased from Sigma; ⑧ macroporous adsorption resin S-8, purchased from hecheng new materials.

[0043] (2) preparation method: in vitro release method: 8g of different types of resin (D101, AB-8, 50WX8, 50WX4, H-40, NKA-II, 50WX2 and S-8) was weighed in a conical flask, 100ml release medium (20% ethanol in phosphate buffer solution) was added, and it was preheated in a 37℃, 150rpm water bath shaker. 0.2ml of aripiprazole free drug was added to the flask, and the 37℃, 150rpm water bath shaker was started. At different time, 1ml of release medium was taken, the sample was diluted with 1ml of anhydrous ethanol, and microfiltration membrane was used for filtration. The content of aripiprazole in the sample taken at different time points was determined by liquid chromatography, and the adsorption of different resins on free drug was investigated (calculation method was consistent with the calculation method of release rate).

[0044] Table 2 different types of resin on the adsorption rate of aripiprazole free drug

[0045]

[0046] According to the data in table 2, it can be seen that only using cation exchange resin 50WX4 (serial number 2-4), the adsorption rate at 15min is as high as 72.5%, which meets the requirement of rapid release, and the adsorption rate at 60min is not much different, which shows that the reason why the release at 15min is not complete may be that the amount of resin is too small, rather than the type of resin.

[0047] Other types of resin, 15min only release about 35-48%, up to 60min release about 75%, which shows that the adsorption speed of these resins is too slow, and even increasing the amount of resin cannot increase the adsorption speed, which cannot meet the fast release effect (FDA website public reference preparation review data shows that the reference preparation of this product uses USP4 release method, combined with the detection results of our company according to the release method of reference preparation, the release rate of aripiprazole injection at 15min is >70%).

[0048] Example 3: Investigation of the effect of the amount of resin added (sample is free drug)

[0049] Investigation of the effect of the amount of resin added on the adsorption of the resin

[0050] Respectively, 50WX4 cation exchange resin 8g, 16g and 24g were weighed into conical flasks, 100ml release medium (20% ethanol-containing phosphate buffer) was added, and preheated in a 37°C, 150rpm water bath. 0.2ml of aripiprazole free drug was added to the flask, and the 37°C, 150rpm water bath was started. 1ml of release medium was taken at different times, the sample was diluted with 1ml of anhydrous ethanol, and microfiltration membrane filtration was performed. The aripiprazole content of the sample taken at different time points was determined by liquid chromatography, and the adsorption of different amounts of resin on free drug was investigated (calculation method is consistent with the calculation method of release rate).

[0051] The calculation results of different amounts of resin are shown in the following table.

[0052] Table 3 Adsorption rate of different amounts of resin

[0053]

[0054] According to Table 3, increasing the amount of resin added significantly improves the adsorption effect of the resin. When the amount of resin added is 16%-24%, the 5min adsorption rate is as high as 99%, which can meet the detection requirements of immediate-release drugs.

[0055] Therefore, according to the investigation of the release method in Examples 1-3, the in vitro release of aripiprazole injection was detected using 16%-24% w / v of cation exchange resin 50WX4 and 15-25% ethanol v / v of phosphate buffer solution as the dilution medium.

[0056] Example 4: In vitro release detection (sample is aripiprazole injection)

[0057] Aripiprazole injection (generic): self-made by Shizhu Pharmaceutical Group Zhongqi Pharmaceutical Technology (Shijiazhuang) Co., Ltd.

[0058] Aripiprazole injection (reference): purchased from Heron Therapeutics

[0059] Respectively, 50WX4 cation exchange resin 16g was weighed into different conical flasks, 100ml release medium was added, and preheated in a 37°C, 150rpm water bath. 0.2ml of reference aripiprazole injection was added to the flask, and the water bath was continued. 1ml of release medium was taken at different times, the sample was diluted with 1ml of anhydrous ethanol, and microfiltration membrane filtration was performed. The aripiprazole content of the sample taken at different time points was determined by liquid chromatography.

[0060] The results of the in vitro release rate calculation of the aprepitant injection liquid are shown in the following table.

[0061] Table 4 In vitro release rate of aprepitant injection liquid

[0062]

[0063]

[0064] According to Table 4, the release rate of the self-developed aprepitant injection liquid at 15 min is only 10.9%, compared with the comparative example, while the release rate of the self-developed aprepitant injection liquid of the present application at 15 min reaches 94%. It can be seen that the release method of the present application meets the requirements of simulating in vivo release. Moreover, the in vitro release method of the present application can be used to evaluate the quality of the self-developed aprepitant injection liquid, and to compare and investigate the consistency of the quality of the self-developed generic drug preparation and the reference preparation of the original research.

Claims

1. An in vitro release method for a fosaprepitant fat emulsion injection, characterized by, The method comprises the following steps: adding cation exchange resin 50WX4 into a release medium, preheating in a water bath oscillator, then adding the fat emulsion injection of alarepride, taking the release medium at different time points after continuous oscillation, and determining the content of alarepride by using liquid chromatography; wherein the release medium is phosphate buffer solution with the content of ethanol v / v being 15-25%; The content of the cation exchange resin is 16%-24% w / v.

2. The in vitro release method of claim 1, wherein, After the release medium is taken, it is diluted with anhydrous ethanol and filtered by using a microporous filter membrane.

3. The in vitro release method of claim 1, wherein, The phosphate is selected from potassium dihydrogen phosphate, disodium hydrogen phosphate, sodium dihydrogen phosphate, dipotassium hydrogen phosphate or ammonium dihydrogen phosphate.

4. The in vitro release method of claim 1, wherein, The phosphate is potassium dihydrogen phosphate.

5. The in vitro release method of claim 1, wherein, The content of ethanol in the release medium is 20% v / v.

6. The in vitro release method of claim 1, wherein, The temperature and speed of the water bath oscillator are 37 DEG C and 150 rpm.

7. The in vitro release method of claim 1, wherein, The pH of the phosphate buffer solution is 6-8.

8. The in vitro release method of claim 1, wherein, The pH of the phosphate buffer solution is 7.

4.

9. The in vitro release method of claim 1, wherein, The preparation method of the phosphate buffer solution containing ethanol is as follows: taking potassium dihydrogen phosphate, dissolving in water, then adding sodium hydroxide solution, adding water, adjusting the pH by using sodium hydroxide solution, adding anhydrous ethanol and mixing uniformly, and ultrasonic degassing.

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