Method for detecting related substances in memantine hydrochloride donepezil preparation

Abstract

The invention discloses a method for detecting related substances in a memantine hydrochloride donepezil preparation. The detection method comprises the following steps: preparing and measuring a test solution and a reference solution by adopting a GC (Gas Chromatography) method, measuring a solvent acetonitrile, and respectively injecting into a gas chromatograph; and recording a chromatogram and calculating by peak area according to an external standard method. The detection method can be used for simultaneously detecting memantine impurity E, memantine impurity F, memantine impurity G, memantine impurity H and memantine impurity I in the memantine hydrochloride donepezil preparation. The method has the advantages of good sensitivity, durability, system applicability, specificity and accuracy, and qualified recovery rate.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical technology, specifically relating to a method for detecting related substances in memantine-dopezil hydrochloride preparations. Background Technology

[0002] Memantine-Donepizil Hydrochloride Extended-Release Capsules are a combination of the NMDA receptor antagonist memantine hydrochloride and the acetylcholinesterase inhibitor donepezil hydrochloride, and are indicated for patients with moderate to severe Alzheimer's disease.

[0003] USP PF 44(6):Nov.-Dec.2018 discloses that the memantine donepezil hydrochloride sustained-release capsules may introduce degradation impurities: memantine impurity E (N-formyl-3,5-dimethyladamantane), memantine impurity F (1-hydroxy-3-amino-5,7-dimethyladamantane hydrochloride), and process impurities: memantine impurity G (3-methyl-1-adamantyl)amine hydrochloride), memantine impurity H (3,5,7-trimethyladamantaneamine hydrochloride), and memantine impurity I (1-amino-3-ethyladamantane).

[0004] During pharmaceutical manufacturing, residual impurities in active pharmaceutical ingredients (APIs) or pharmaceutical intermediates can pose potential risks to the final product. These impurities can not only affect drug efficacy but also cause production and storage problems, and some impurities can even cause toxic side effects. Therefore, studying potential drug impurities can ensure the safety, efficacy, and stability of medication, while also providing a basis for quality assurance in the production and distribution processes.

[0005] For memantine donepezil hydrochloride sustained-release capsules, a systematic study of its process impurities is required to ensure medication safety. Currently, the research methods for these process impurities disclosed in USP PF 44(6):Nov.-Dec.2018 for memantine hydrochloride sustained-release capsules are not applicable to the study of process impurities in memantine donepezil hydrochloride sustained-release capsules. The recovery rate of this research method cannot meet the pharmacopoeia requirements, and the accuracy of the results obtained by the research method is poor, which cannot guarantee the safety and stability of medication.

[0006] In existing technologies, there are numerous reports on the detection of impurities in memantine hydrochloride, but few methods are disclosed for the detection of related substances in memantine-dopedizine hydrochloride formulations. Current detection methods cannot be applied to the detection of degradation impurities memantine E and F in memantine-dopedizine hydrochloride formulations with complex matrices, nor can they simultaneously and effectively locate process impurities memantine G, H, and I. Therefore, existing methods cannot be applied to the detection of related substances in memantine-dopedizine hydrochloride formulations with complex matrices, and thus cannot guarantee medication safety. Summary of the Invention

[0007] To address the above-mentioned problems in the prior art, this invention provides a method for detecting related substances in memantine donepezil hydrochloride preparations and its application.

[0008] A method for detecting related substances in memantine donepezil hydrochloride preparations, wherein the related substances include at least one of memantine impurity E, memantine impurity F, memantine impurity G, memantine impurity H and memantine impurity I;

[0009] The detection method employs GC chromatography, and the detection conditions include: a capillary column using 4-6% (e.g., 5%) phenyl-methylpolysiloxane as the stationary phase; a flame ionization detector; a split ratio of (4-6):1 (e.g., 4:1, 5:1, 6:1); an initial temperature of 113-117℃ (e.g., 114℃, 115℃, 116℃); a programmed temperature increase to 250-300℃ (e.g., 255℃, 260℃, 265℃, 270℃, 275℃, 280℃, 285℃, 290℃, 295℃); and a maintenance temperature of 20-35 minutes; an injection port temperature of 240-260℃ (e.g., 245℃, 250℃, 255℃); and a detector temperature of 295-305℃ (e.g., 300℃).

[0010] Prepare and measure the test solution and reference solution, measure the solvent acetonitrile, and inject them into the gas chromatograph separately; record the chromatograms and calculate the peak area according to the external standard method.

[0011] According to an embodiment of the present invention, the duration of maintenance is 20-30 minutes.

[0012] According to an embodiment of the present invention, the preparation of the test solution includes: first, mixing the test sample with an alkaline solution until dissolved, then adding acetonitrile and mixing; after standing and separating into layers, adding a drying agent to the organic layer to dry it, and taking the supernatant to obtain the test solution;

[0013] Preferably, the mixing method is shaking or vortexing; for example, the test sample is placed in an alkaline solution and shaken or vortexed for 3-5 minutes to dissolve the test sample; or, after adding acetonitrile, vortexing for at least 30 seconds until extraction is completely stopped.

[0014] The preparation of the test solution of the present invention adopts the opposite method to that of USP PF 44(6):Nov.-Dec.2018 (the test sample is first added to acetonitrile, and then mixed with alkaline solution). That is, the test sample is first mixed with alkaline solution, and then mixed with acetonitrile. Combined with the chromatographic column detection conditions, the technical problem of unqualified recovery rate of related substances is solved.

[0015] According to an embodiment of the present invention, the preparation of the reference solution includes: dissolving the reference standard in acetonitrile solution, quantitatively diluting it with acetonitrile to prepare a reference standard stock solution; measuring the reference standard stock solution and mixing it with an alkaline solution, allowing it to stand and separate into layers, adding a drying agent to the organic layer to dry it, and taking the supernatant to obtain the reference standard solution;

[0016] Preferably, the mixing method is shaking or vortexing; for example, the reference standard stock solution is mixed with an alkaline solution and then shaken or vortexed for 1-5 minutes.

[0017] For example, the concentration of the acetonitrile solution is 40-60%, such as 50%.

[0018] According to an embodiment of the present invention, the memantine donepezil hydrochloride formulation used as a test sample includes, but is not limited to, memantine donepezil hydrochloride capsules, such as sustained-release capsules.

[0019] According to an embodiment of the present invention, the chromatographic column is an HP Ultra2, 0.32 mm × 50 m, 0.52 μm; or a chromatographic column with similar polarity.

[0020] According to an embodiment of the present invention, the programmed temperature rise is as follows: the temperature is increased to 145°C at a rate of 2°C / min, and then increased to 250-300°C at a rate of 30°C / min.

[0021] According to an embodiment of the present invention, the carrier gas used in the hydrogen flame ionization detector is helium, the flow rate is 0.5-5 ml / min, for example 1.5-2.0 ml / min, and the injection volume is 1 ml.

[0022] According to an embodiment of the present invention, the alkaline solution is an aqueous solution of sodium hydroxide or an aqueous solution of potassium hydroxide.

[0023] According to an embodiment of the present invention, the molar concentration of the alkaline solution is 1-5 mol / L, for example, 1 mol / L, 2 mol / L, 3 mol / L, 4 mol / L, or 5 mol / L.

[0024] According to an embodiment of the present invention, the volume ratio of alkaline solution to acetonitrile in the test solution is 1:2.

[0025] According to an embodiment of the present invention, the desiccant is at least one selected from anhydrous sodium sulfate, anhydrous calcium chloride, anhydrous magnesium sulfate, and anhydrous magnesium perchlorate.

[0026] The term "at least one" means that it can be one, two, or more of the above-defined ranges, covering all of them.

[0027] Beneficial effects

[0028] The detection method of this invention can simultaneously detect memantine impurity E, memantine impurity F, memantine impurity G, memantine impurity H, and memantine impurity I in memantine-donepezil hydrochloride formulations. The method exhibits good sensitivity, robustness, system suitability, specificity, and accuracy, with satisfactory recovery. Attached Figure Description

[0029] Figure 1 The test chromatogram of the test solution in Example 1;

[0030] Figure 2 The test chromatogram is shown for the spiked test solution in Example 1;

[0031] Figure 3 The spectrum is that of the test solution in Example 12;

[0032] Figure 4 The spectrum is shown for the test solution in Comparative Example 1;

[0033] Figure 5 The spectrum is for the test solution in Comparative Example 2;

[0034] Figure 6 The spectrum is for solutions suitable for system use. Detailed Implementation

[0035] The technical solution of the present invention will be further described in detail below with reference to specific embodiments. It should be understood that the following embodiments are merely illustrative and explanatory of the present invention, and should not be construed as limiting the scope of protection of the present invention. All technologies implemented based on the above content of the present invention are covered within the scope of protection intended by the present invention.

[0036] Unless otherwise stated, the raw materials and reagents used in the following examples are commercially available products or can be prepared by known methods.

[0037] Related substances: Memantine impurity E sourced from TRC; Memantine impurity F sourced from QCC; Memantine impurity G sourced from SINCO; Memantine impurity H sourced from SINCO; Memantine impurity I sourced from Shanghai Kewei Chemical Technology Co., Ltd.; Memantine hydrochloride reference standard sourced from the China National Institutes for Food and Drug Control; Memantine hydrochloride donepezil sustained-release capsules (main ingredients: memantine hydrochloride, donepezil hydrochloride, and pharmaceutical excipients) sourced from Sichuan Keruide Pharmaceutical Co., Ltd., batch number: 240201; Memantine hydrochloride sustained-release microcapsules blank excipient sourced from Sichuan Keruide Pharmaceutical Co., Ltd., batch number: M20240408-K.

[0038] The contents of memantine-dopezil hydrochloride sustained-release capsules consist of memantine-dopezil hydrochloride sustained-release microspheres and a mixture of donepezil hydrochloride powder. The donepezil hydrochloride powder contains pharmaceutical excipients and donepezil hydrochloride. The "blank excipient in mixed powder" in the following test examples refers to the pharmaceutical excipients in the donepezil hydrochloride powder.

[0039] Memantine impurity E stock solution: Take an appropriate amount of memantine impurity E hydrochloride, accurately weigh it, place it in a 20ml volumetric flask, add 2ml of 50% acetonitrile to dissolve it, and quantitatively dilute it with acetonitrile to obtain a memantine impurity E stock solution containing approximately 1.12mg of memantine impurity E per 1ml.

[0040] The same method was used to prepare stock solutions of the same concentrations of micranthae impurity F, micranthae impurity G, micranthae impurity H, micranthae impurity I, micranthae hydrochloride, and donepezil hydrochloride.

[0041] In all embodiments of the present invention, the solution preparation system suitability solution is as described in Example 2:

[0042] System suitability solutions: Measure 1 ml each of the stock solutions of memantine impurity E, memantine impurity F, memantine impurity G, memantine impurity H, and memantine impurity I, and 10 ml each of memantine hydrochloride stock solution and donepezil hydrochloride stock solution. Place them in the same 200 ml volumetric flask, dilute to the mark with acetonitrile, and shake well. Accurately measure 10 ml of the mixed solution and mix with 5 ml of 3 mol / L sodium hydroxide solution. Vortex for 1 minute, allow to stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and collect the supernatant.

[0043] If impurity peaks are present in the chromatogram of the test solution, excluding solvent peaks, excipient peaks, memantine impurity G peak, memantine impurity H peak, memantine impurity I peak, and donepezil peaks, if there is a chromatographic peak with the same retention time as memantine impurity E in the reference solution, calculated using the external standard method of principal component, it shall not exceed 0.2%; if there is a chromatographic peak with the same retention time as memantine impurity F in the reference solution, calculated using the external standard method of principal component with the corrected peak area (multiplied by a correction factor of 1.3), it shall not exceed 0.2%; other individual impurities shall be calculated using the external standard method of principal component with the corrected peak area (multiplied by a correction factor of 1.3), and shall not exceed 0.2%. The total amount of impurities shall not exceed 0.5%, and peaks with an area smaller than the main peak area of ​​the sensitivity solution shall be ignored.

[0044] Impurity calculation formula:

[0045] In the formula:

[0046] A -- The peak area of ​​impurities in the test solution;

[0047] V -- Volume of the test solution diluted, in ml;

[0048] F--Response factor of donepezil hydrochloride reference solution;

[0049] L--Memantine-Donepizil Extended-Release Capsules, Memantine Hydrochloride Label (mg);

[0050] B -- Theoretical fill weight (mg);

[0051] m -- Sample weight (mg);

[0052] Note: The sum of all impurities is the sum of all known impurities and other individual impurities.

[0053] Example 1

[0054] The method for detecting related substances in memantine donepezil hydrochloride preparations, the related substances to be detected include: memantine impurity E and memantine impurity F.

[0055] GC chromatography was used with a capillary column containing 5% phenyl-methylpolysiloxane (HP Ultra2, 0.32 mm × 50 m, 0.52 μm; or similar polarity) as the stationary phase. The initial temperature was 115 °C, increased to 145 °C at a rate of 2 °C / min, and then increased to 300 °C at a rate of 30 °C / min, and held for 29.5 min. The injection port temperature was 250 °C, the split ratio was 5:1, the detector temperature was 300 °C, the carrier gas was helium at a flow rate of 1.8 ml / min, and the injection volume was 1 ml.

[0056] For the test solution, take 20 capsules of memantine-dopezil hydrochloride, combine the contents, grind the separated microparticles into a fine powder, take an appropriate amount of the fine powder (approximately equivalent to 28 mg of memantine hydrochloride), accurately weigh it, place it in a 25 ml volumetric flask, add 5 ml of 3 mol / L sodium hydroxide solution, shake or vortex for 3 minutes, then add 10 ml of acetonitrile and mix, vortex for 1 minute, let stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and take the supernatant.

[0057] For the reference solution, accurately weigh an appropriate amount of memantine hydrochloride, add 2 ml of 50% acetonitrile solution to dissolve the memantine hydrochloride, and then quantitatively dilute with acetonitrile to prepare a solution containing approximately 5.6 μg of memantine hydrochloride per ml. Accurately measure 10 ml of this solution and mix it with 5 ml of 3 mol / L sodium hydroxide solution, vortex for 1 minute, allow to stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and collect the supernatant.

[0058] Spiked test solution: Take 20 capsules of memantine donepezil hydrochloride, combine the contents, grind the separated microcapsules into fine powder, take about 178 mg of the fine powder, and accurately add 10 ml of impurity control mixed solution (accurately measure 1 ml each of memantine impurity E stock solution and memantine impurity F stock solution, place them in the same 200 ml volumetric flask, dilute to the mark with acetonitrile, and mix well), place them in the same 25 ml volumetric flask, shake or vortex for 3 minutes, add 5 ml of 3 mol / L sodium hydroxide solution, vortex for 1 minute, let stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and take the supernatant.

[0059] For the determination, inject the test solution, reference solution, and acetonitrile into the gas chromatograph separately; record the chromatograms and calculate the peak area using the external standard method.

[0060] Figure 1 The chromatogram of the test solution is shown. Figure 2 The figure shows the test chromatogram of the spiked test solution. As can be seen from the figure, the method of the present invention can effectively detect and separate memantine hydrochloride and donepezil in memantine-dopedizine hydrochloride capsules; it also indicates that the memantine-dopedizine hydrochloride capsules do not contain impurities E and F. That is, the method of the present invention can effectively detect and separate impurities E / F, memantine hydrochloride and donepezil in memantine-dopedizine hydrochloride capsules, and is applicable to the detection of related substances in memantine-dopedizine hydrochloride capsules.

[0061] Example 2

[0062] The detection conditions are basically the same as in Example 1, except that:

[0063] For the reference solution, accurately weigh an appropriate amount of methimazole impurity E, place it in a 20 ml volumetric flask, add 2 ml of 50% acetonitrile to dissolve it, and quantitatively dilute with acetonitrile to obtain a methimazole impurity E stock solution containing approximately 1.12 mg of methimazole impurity E per ml. Prepare methimazole impurity F, G, H, and I stock solutions of the same concentration using the same method. Accurately measure 1 ml of each of the above stock solutions, place them in a 200 ml volumetric flask, dilute with acetonitrile to the mark, and shake well. Take 10 ml of the shaken solution and mix it with 5 ml of 3 mol / L sodium hydroxide solution, vortex for 1 minute, allow to stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and collect the supernatant.

[0064] The injection port temperature is 245℃.

[0065] Example 3

[0066] The detection conditions are basically the same as in Example 2, except that:

[0067] The injection port temperature is 255℃.

[0068] Example 4

[0069] The detection conditions are basically the same as in Example 2, except that:

[0070] The split ratio is 4:1.

[0071] Example 5

[0072] The detection conditions are basically the same as in Example 2, except that:

[0073] The split ratio is 6:1.

[0074] Example 6

[0075] The detection conditions are basically the same as in Example 2, except that:

[0076] Flow rate 1.7 ml / min.

[0077] Example 7

[0078] The detection conditions are basically the same as in Example 2, except that:

[0079] Flow rate 1.9 ml / min.

[0080] Example 8

[0081] The detection conditions are basically the same as in Example 2, except that:

[0082] Starting temperature: 113℃.

[0083] Example 9

[0084] The detection conditions are basically the same as in Example 2, except that:

[0085] Starting temperature: 117℃.

[0086] Example 10

[0087] The detection conditions are basically the same as in Example 2, except that:

[0088] Detector temperature: 295℃.

[0089] Example 11

[0090] The detection conditions are basically the same as in Example 2, except that:

[0091] Detector temperature: 305℃.

[0092] Table 1 Test Results of Example 2-11

[0093]

[0094] As shown in Examples 2-11, under various chromatographic conditions, the peak area RSD value of the reference solution was not greater than 8.0% when injected five times consecutively. The minimum resolution between each component in the system suitability solution was not less than 1.9. The RSD of the content detection results of methimazole impurity E and methimazole impurity F was small, and the results all met the requirements, indicating that the detection method has good robustness.

[0095] Example 12

[0096] Chromatographic conditions

[0097] Chromatographic column: HP Ultra2, 0.32mm × 50m, 0.52μm

[0098] Heating program: The starting temperature is 115℃, the temperature is increased to 145℃ at a rate of 2℃ per minute, and then increased to 300℃ at a rate of 30℃ per minute, and maintained for 29.5 minutes.

[0099] Inlet temperature: 250℃;

[0100] Hydrogen flame ionization detector;

[0101] Flow split ratio: 5:1;

[0102] Detector temperature: 300℃;

[0103] Carrier gas: Helium

[0104] Flow rate: 1.8 ml per minute;

[0105] Injection volume: 1 μl.

[0106] Test solution: Take appropriate amounts of memantine hydrochloride reference standard, donepezil hydrochloride reference standard, memantine impurity E, memantine impurity F, memantine impurity G, memantine impurity H, and memantine impurity I, add 2 ml of 50% acetonitrile to dissolve, and quantitatively dilute with acetonitrile to prepare a solution containing approximately 2.8 mg of memantine hydrochloride and 5.6 μg each of donepezil hydrochloride, memantine impurity E, memantine impurity F, memantine impurity G, memantine impurity H, and memantine impurity I per ml. Accurately measure 10 ml of this solution and mix with 5 ml of 3 mol / L sodium hydroxide solution, vortex for 1 minute, allow to stand to separate into layers, dry the organic layer with anhydrous sodium sulfate, and collect the supernatant.

[0107] The results are as follows Figure 3 As shown, under these conditions, the peak separation is good, all components can precipitate peaks, and the sensitivity is relatively high.

[0108] Screening of analyte solution treatment conditions:

[0109] The chromatographic conditions were as described in Comparative Example 1. The test solution and spiked test solution were prepared according to the method of Example 1. The molar concentration and amount of sodium hydroxide and the amount of acetonitrile used in the preparation of the test solution are shown in the corresponding group in the table below. The sodium hydroxide solution used in the preparation of the spiked test solution is shown in the corresponding group in the table below. The recovery rates of E and F in the spiked test solution are shown in Table 2 below.

[0110] Table 2. Screening results of analyte solution treatment conditions

[0111]

[0112] Conclusion: Through comparison, it was found that when the ratio of aqueous phase to organic phase was 5:10 and the sodium hydroxide concentration was 1-5 mol / L, the recovery rates of impurities E and F in memantine were relatively stable and met the requirements.

[0113] Comparative Example 1

[0114] Referencing USP PF 44(6):Nov.-Dec.2018, the following methods for the detection of related substances were proposed:

[0115] Chromatographic conditions

[0116] Chromatographic column: HP-5ms, 0.32mm × 30m, 0.25μm

[0117] Heating program: Start at 115℃ and hold for 6 minutes, then increase to 200℃ at a rate of 20℃ per minute and hold for 1.5 minutes, then increase to 300℃ at a rate of 20℃ per minute and hold for 8.25 minutes.

[0118] Inlet temperature: 250℃;

[0119] Flow split ratio: 16:1;

[0120] Detector: Flame Ionization Detector (FID)

[0121] Detector temperature: 300℃;

[0122] Carrier gas: Helium

[0123] Flow rate: 1.8 ml per minute;

[0124] Injection volume: 1 μl.

[0125] Test solution: Take appropriate amounts of memantine hydrochloride reference standard, donepezil hydrochloride reference standard, memantine impurity E, memantine impurity F, memantine impurity G, memantine impurity H, and memantine impurity I, add 2 ml of 50% acetonitrile to dissolve them, and quantitatively dilute with acetonitrile to prepare a solution containing approximately 20 μg of memantine hydrochloride and 5 μg each of donepezil hydrochloride, memantine impurity E, memantine impurity F, memantine impurity G, memantine impurity H, and memantine impurity I per ml. Accurately measure 10 ml of this solution and mix it with 5 ml of 3 mol / L sodium hydroxide solution, vortex for 1 minute, allow to stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and take the supernatant.

[0126] The results are as follows Figure 4 As shown, from Figure 4 It is known that the separation degree between the methyl methacrylate impurity H peak and the methyl methacrylate peak in the test solution chromatogram is only 1.9, and the concentration of methyl methacrylate is low. If the concentration of methyl methacrylate increases, the methyl methacrylate impurity H peak and the methyl methacrylate peak may not reach baseline separation. At the same time, the recovery rate of methyl methacrylate impurity E in the spiked test solution (prepared by the same method as in Example 1) is 121.8%, and the recovery rate of methyl methacrylate impurity F is 77.1%, both of which do not meet the requirements.

[0127] Table 3. Recovery results of impurity E (Mexican) in the spiked test solution.

[0128]

[0129] Table 4. Recovery results of the impurity F, known as the "Medaling King Kong," in the spiked test solution.

[0130]

[0131] Comparative Example 2

[0132] Chromatographic conditions

[0133] Chromatographic column: HP Ultra2, 0.32mm × 50m, 0.52μm

[0134] Heating program: The starting temperature is 50℃, and the temperature is increased to 100℃ at a rate of 5℃ per minute, then increased to 110℃ at a rate of 0.5℃ per minute, and then increased to 250℃ at a rate of 10℃ per minute, and maintained for 20 minutes.

[0135] Carrier gas: Helium

[0136] Flow rate: 4 ml per minute

[0137] Detector: Flame Ionization Detector (FID)

[0138] Detector temperature: 300℃

[0139] Inlet temperature: 220℃

[0140] Flow split ratio: 3:1

[0141] Injection volume: lμl

[0142] Test solutions: Take appropriate amounts of reference standards for impurities A, B, C, D, E, J, G, and H of methimazole (take appropriate amounts of reference standards for impurities G and H of methimazole, dissolve them in 5 mol / L sodium hydroxide aqueous solution, and shake for 5 minutes). Add appropriate amounts of internal standard solution to make the concentrations of impurities A, B, C, D, J, G, and H of methimazole 2.5 mg / ml, and the concentration of impurity J of methimazole 1.0 mg / ml. Vortex and shake for 10 minutes. After separation, take the n-hexane layer and dry it with anhydrous sodium sulfate. These are used as stock solutions for impurities A, B, C, D, E, J, G, and H of methimazole, respectively. Accurately weigh approximately 120 mg of memantine hydrochloride reference standard and place it in a 10 ml centrifuge tube. Add 2 ml of 5 mol / L sodium hydroxide solution and vortex for 5 minutes. Accurately add stock solutions of memantine impurities A, B, C, D, E, J, G, and H, and an appropriate amount of n-hexane, so that the concentration of each of the memantine impurities A, B, C, D, and E is 90 μg / ml, and the concentration of each of the memantine impurities J, G, and H is 60 μg / ml. Vortex for 10 minutes and allow the layers to separate. Dry the n-hexane layer with anhydrous sodium sulfate to obtain the final product.

[0143] The results are as follows Figure 5 As shown, the separation degree between memantine and memantine impurity H is 1.5, which is a good separation degree. However, memantine impurity F is insoluble in n-hexane, so this method is not applicable to the detection of memantine impurity F and has poor sensitivity.

[0144] Comparative Example 3

[0145] Chromatographic conditions

[0146] Chromatographic column: HP Ultra2, 0.32mm × 50m, 0.52μm

[0147] Heating program: The starting temperature is 115℃, and the temperature is increased to 145℃ at a rate of 2℃ per minute, then increased to 300℃ at a rate of 30℃ per minute, and maintained for 9.5 minutes.

[0148] Inlet temperature: 250℃;

[0149] Flow split ratio: 5:1;

[0150] Detector: Flame Ionization Detector (FID)

[0151] Detector temperature: 300℃;

[0152] Carrier gas: Helium

[0153] Flow rate: 1.8 ml per minute;

[0154] Injection volume: 1 μl.

[0155] Test solution: Take 20 capsules of memantine donepezil hydrochloride sustained-release capsules, combine the contents, grind the sieved microparticles into fine powder, take an appropriate amount of fine powder (approximately equivalent to 28 mg of memantine hydrochloride), accurately weigh it, place it in a 25 ml volumetric flask, add 5 ml of 3 mol / L sodium hydroxide solution, vortex for 3 minutes, then add 10 ml of acetonitrile and mix, vortex for 1 minute, let stand to allow the layers to separate, dry the organic layer with anhydrous sodium sulfate, and take the supernatant.

[0156] Results: The peaks were well separated, but when the samples were tested, donepezil attached to the microspheres also showed peaks. Because donepezil has a high boiling point, it would remain in the system and affect the detection of the next sample.

[0157] Test case

[0158] The following example, Example 1, is used to verify the methodology and further validate the technical solution of the present invention.

[0159] 1. System Applicability

[0160] Solution preparation:

[0161] Reference solution: Accurately measure 1 ml of memantine hydrochloride stock solution (as described above), place it in a 200 ml volumetric flask, dilute to the mark with acetonitrile, shake well to obtain the reference stock solution; take 10 ml of the reference stock solution and mix with 5 ml of 3 mol / L sodium hydroxide solution, vortex for 1 min, let stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and take the supernatant to obtain the reference solution.

[0162] System suitability solution: as described above.

[0163] Sensitivity solution: Accurately measure 10 ml of the reference stock solution and place it in a 20 ml volumetric flask. Dilute to the mark with acetonitrile and shake well. Accurately measure 10 ml of this solution and mix it with 5 ml of 3 mol / L sodium hydroxide solution. Vortex for 1 min, allow to stand for separation, dry the organic layer with anhydrous sodium sulfate, and take the supernatant.

[0164] Accurately measure 1 μL each of the reference solution and the system suitability solution, and inject them into the gas chromatograph. The chromatogram of the system suitability solution is as follows: Figure 6 The results are shown in Table 5 below.

[0165] Table 5. System Suitability Solution Spectrum Data Results

[0166] name Retention time Resolution Impurities G in Mamanite 12.307 / Megatron 12.653 3.2 Impurities H in Mamanite 12.880 2.1 Impurities in Magnesium I 16.606 46.5 Impurities in Magnesium Oxide F 17.216 13.1 Impurities E in Mamanite 19.792 72.2 Donepezil 41.374 167.1

[0167] Five consecutive injections of the reference solution resulted in a peak area RSD of 5.5% for memantine. In the system suitability solution, the resolution of each component was greater than 1.5, with a minimum resolution of 2.1. In the sensitivity solution, the high signal-to-noise ratio of the memantine peak was 42.1, all of which met the detection requirements, indicating good system suitability.

[0168] 2. Exclusivity

[0169] Solution preparation:

[0170] Blank solution: Accurately measure 10 ml of acetonitrile and mix with 5 ml of 3 mol / L sodium hydroxide solution. Vortex for 1 min, allow to stand for separation, dry the organic layer with anhydrous sodium sulfate, and take the supernatant.

[0171] Blank excipient solution: Weigh 150 mg of blank excipient for memantine hydrochloride sustained-release microcapsules and 90 mg of blank excipient for mixed powder, place them in a 25 ml volumetric flask, add 5 ml of 3 mol / L sodium hydroxide solution, vortex for 3 min, add 10 ml of acetonitrile, mix, vortex for 1 min, let stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and take the supernatant.

[0172] Mimantan impurity E positioning solution: Take 1 ml of mimantan impurity E stock solution and place it in a 10 ml volumetric flask. Dilute to the mark with acetonitrile and shake well. Accurately take 1 ml of this solution and place it in a 20 ml volumetric flask. Shake well. Accurately take 10 ml of this solution and mix it with 5 ml of 3 mol / L sodium hydroxide solution. Vortex for 1 min, allow to stand to separate the layers, dry the organic layer with anhydrous sodium sulfate, and take the supernatant. Prepare mimantan impurity F, mimantan impurity G, mimantan impurity H, and mimantan impurity I positioning solutions using the same method.

[0173] Donepezil positioning solution: Measure 1 ml of donepezil stock solution into a 20 ml volumetric flask, dilute to the mark with acetonitrile, and shake well. Accurately measure 10 ml of this solution and mix with 5 ml of 3 mol / L sodium hydroxide solution, vortex for 1 minute, allow to stand to separate into layers, dry the organic layer with anhydrous sodium sulfate, and collect the supernatant. Prepare memantine positioning solution using the same method.

[0174] The preparation methods for the following stock solutions are as described above: Memantine Impurity E Stock Solution, Memantine Impurity F Stock Solution, Memantine Impurity G Stock Solution, Memantine Impurity H Stock Solution, Memantine Impurity I Stock Solution, Memantine Hydrochloride Stock Solution, and Donepezil Hydrochloride Stock Solution.

[0175] The system suitability solution is as described above.

[0176] Test solution: Same as in Example 1.

[0177] Take 1 μL of each of the above solutions and inject them into the gas chromatograph. The test results are shown in Table 6 below.

[0178] Table 6 Specificity Test Results

[0179]

[0180] Table 7. Results of the positioning solution test

[0181]

[0182] Specificity tests showed that the blank solvent, blank excipient, and donepezil did not interfere with the detection of memantine hydrochloride impurities. In the test sample and system suitability solution, the peak resolution was not lower than 2.1. This method has high specificity.

[0183] 3. Accuracy

[0184] Solution preparation:

[0185] The test solution is the same as in Example 1.

[0186] The same applies to the stock solution for impurity E in jamanite and the same applies to the stock solution for impurity F in jamanite.

[0187] 50% Impurity Control Mixed Solution: Accurately measure 0.5 ml of micranis impurity E stock solution and 0.5 ml of micranis impurity F stock solution, place them in the same 200 ml volumetric flask, dilute to the mark with acetonitrile, and mix well.

[0188] 100% Impurity Control Mixed Solution: Accurately measure 1 ml of micranthin impurity E stock solution and 1 ml of micranthin impurity F stock solution, place them in a 200 ml volumetric flask, dilute to the mark with acetonitrile, and mix to obtain the solution.

[0189] 150% Impurity Control Mixed Solution: Accurately measure 1.5 ml of micranis impurity E stock solution and 1.5 ml of micranis impurity F stock solution, place them in the same 200 ml volumetric flask, dilute to the mark with acetonitrile, and mix well.

[0190] 50% spiked test solution: Weigh about 178 mg of memantine donepezil hydrochloride microparticles (the same fine powder as in Example 1), and accurately measure 10 ml of 50% impurity control mixed solution. Place them in the same 25 ml volumetric flask, shake or vortex for 3 minutes, add 5 ml of 3 mol / L sodium hydroxide solution, vortex for 1 minute, let stand to allow the layers to separate, dry the organic layer with anhydrous sodium sulfate, and take the supernatant.

[0191] (Prepare three samples in parallel)

[0192] 100% Spiked Test Solution: Weigh 178 mg of memantine-dopezil hydrochloride microgranules (same as the fine powder in Example 1), and accurately add 10 ml of 100% impurity control mixed solution. Place the solutions in the same 25 ml volumetric flask, shake or vortex for 3 minutes, add 5 ml of 3 mol sodium hydroxide solution, vortex for 1 minute, and allow to stand to separate the layers. Dry the organic layer with anhydrous sodium sulfate and collect the supernatant. (Prepare three parallel aliquots)

[0193] 150% Spiked Test Solution: Weigh 178 mg of memantine-dopezil hydrochloride microgranules (same as the fine powder in Example 1), and accurately add 10 ml of 150% impurity control mixed solution. Place in the same 25 ml volumetric flask, shake or vortex for 3 minutes, add 5 ml of 3 mol sodium hydroxide solution, vortex for 1 minute, and allow to stand to separate the layers. Dry the organic layer with anhydrous sodium sulfate and collect the supernatant. (Prepare three parallel aliquots)

[0194] Reference solution: Accurately measure 10 ml of 100% impurity reference mixed solution and place it in a 25 ml volumetric flask. Add 5 ml of 3 mol / L sodium oxide solution, vortex for 1 minute, and let stand to allow the layers to separate. Dry the organic layer with anhydrous sodium sulfate and take the supernatant.

[0195] Test: Take the above reference solution, test solution and spiked test solution, inject them sequentially, and collect chromatograms.

[0196] Acceptable criteria: At three concentration levels of 50%, 100%, and 150%, the recoveries of the nine samples should be in the range of 80%-120%, and the RSD of the recovery data for the nine samples should not exceed 20%.

[0197] Recovery rate = (Measured amount - Background amount) × Added amount × 100%.

[0198] Table 8. Accuracy data for impurity E in Amanita muscaria.

[0199]

[0200] Table 9. Accuracy data for impurity F in Amanita muscaria.

[0201]

[0202]

[0203] Conclusion: At three concentration levels (50%, 100%, and 150%), the recoveries of impurity E in memantine ranged from 105.75% to 115.56% across the nine samples; the RSD for all nine recoveries was 2.7%. The recoveries of impurity F in memantine ranged from 110.30% to 115.75%; the RSD for all nine recoveries was 1.5%. The results met the requirements, and the accuracy was good.

[0204] The detection method of the present invention can simultaneously detect impurities E, F, G, H and I of methimazole hydrochloride donepezil preparations, with qualified recovery rate and accurate detection results.

[0205] The embodiments of the present invention have been described above. However, the present invention is not limited to the above embodiments. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A method for detecting related substances in memantine-donepizil hydrochloride preparations, characterized in that, The relevant substances include at least one of the following: methyl methacrylate impurity E, methyl methacrylate impurity F, methyl methacrylate impurity G, methyl methacrylate impurity H, and methyl methacrylate impurity I; The detection method employs GC chromatography, with the following detection conditions: a capillary column using 4-6% phenyl-methylpolysiloxane as the stationary phase, a flame ionization detector, a split ratio of (4-6):1, an initial temperature of 113-117℃, a programmed temperature increase to 250-300℃, and a holding time of 20-35 minutes; an injection port temperature of 240-260℃; and a detector temperature of 295-305℃. Prepare and measure the test solution and reference solution, measure the solvent acetonitrile, and inject them into the gas chromatograph separately; record the chromatograms and calculate the peak area according to the external standard method.

2. The detection method according to claim 1, characterized in that, The duration of maintenance is 20-30 minutes.

3. The detection method according to claim 1, characterized in that, The preparation of the test solution includes: first, mixing the test sample with an alkaline solution until dissolved, then adding acetonitrile and mixing; after standing and separating the layers, adding a drying agent to the organic layer to dry it, and taking the supernatant to obtain the test solution; Preferably, the mixing method is shaking or vortexing; for example, the test sample is placed in an alkaline solution and shaken or vortexed for 3-5 minutes to dissolve the test sample; or, after adding acetonitrile, it is shaken or vortexed for at least 30 seconds until the extraction is completely stopped.

4. The detection method according to claim 1, characterized in that, The preparation of the reference solution includes: dissolving the reference standard in acetonitrile solution, quantitatively diluting it with acetonitrile to prepare a reference standard stock solution; measuring the reference standard stock solution and mixing it with an alkaline solution, allowing it to stand and separate into layers, adding a drying agent to the organic layer to dry it, and taking the supernatant to obtain the reference standard solution; Preferably, the mixing method is shaking or vortexing; for example, the reference standard stock solution is mixed with the alkaline solution and vortexed for 1-5 minutes. For example, the concentration of the acetonitrile solution is 40-60%.

5. The detection method according to any one of claims 1-4, characterized in that, The memantine donepezil hydrochloride formulation used as the test sample includes, but is not limited to, memantine donepezil hydrochloride capsules, such as sustained-release capsules.

6. The detection method according to claim 1, characterized in that, The chromatographic column is an HP Ultra2, 0.32 mm × 50 m, 0.52 μm; or a chromatographic column with similar polarity; And / or, the programmed temperature rise is as follows: heat to 145°C at a rate of 2°C / min, and then heat to 250-300°C at a rate of 30°C / min; And / or, the carrier gas used in the hydrogen flame ionization detector is helium, with a flow rate of 0.5-5 ml / min and an injection volume of 1 ml.

7. The detection method according to claim 3 or 4, characterized in that, The alkaline solution is an aqueous solution of sodium hydroxide or potassium hydroxide.

8. The detection method according to claim 7, characterized in that, The molar concentration of the alkaline solution is 1-5 mol / L.

9. The detection method according to claim 3, characterized in that, In the test solution, the volume ratio of alkaline solution to acetonitrile is 1:

2.

10. The detection method according to claim 3 or 4, characterized in that, The desiccant is at least one of anhydrous sodium sulfate, anhydrous calcium chloride, anhydrous magnesium sulfate, and anhydrous magnesium perchlorate.

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