A quality control method for melogabalin benzenesulfonate intermediate
Through high performance liquid chromatography and gradient elution procedures, the problem of insufficient detection methods for specific impurities in melogabarin parabenzenesulfonate intermediates in the prior art was solved, and effective elution, separation and quantitative detection of various impurities were achieved, and the quality control ability of the intermediate and its subsequent preparation products was improved.
Patent Information
- Application Number
- CN202411860497.4
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-12-17
- Publication Date
- 2025-04-04
AI Technical Summary
The prior art lacks effective detection methods to control the presence of specific impurities in the melogabarin benzenesulfonate intermediate, affecting the quality of the intermediate and its subsequent preparation products.
Using high-performance liquid chromatography, a gradient elution procedure was used, using an octadecylsilane-bonded silica gel chromatography column, combined with a phosphate buffered solution-acetonitrile mixed solution and a methanol-acetonitrile mixed solvent as the mobile phase, to achieve the elution, separation and quantitative detection of a variety of specific impurities in the melogabarin terebenzene sulfonate intermediate.
This method can effectively elute, separate and quantitatively detect a variety of specific impurities in the melogabarin benzenesulfonate intermediate. It has strong specificity, wide linear range, high sensitivity, good accuracy, and good solution stability and durability, meeting quality control requirements.
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Abstract
Description
Technical Field
[0001] The invention relates to a quality control method for a melogabalin benzenesulfonate intermediate, and in particular to a quality control method for impurities in a melogabalin benzenesulfonate intermediate. Background Art
[0002] Melogabaline benzenesulfonate (2-((1R,5S,6S)-6-(aminomethyl)-3-ethylbicyclo[3.2.0]hept-3-en-6-yl)acetate benzenesulfonate, C 18 H 25 NO5S) is a GABA analogue and a new type of preferentially selective α2δ-1 ligand that acts on the α2-δ1 subunit of the voltage-sensitive calcium channel complex (widely present in the nervous system that mediates pain transmission and processing throughout the body) and is used to treat pain associated with diabetic peripheral neuropathy, fibromyalgia and postherpetic neuralgia.
[0003]
[0004] Tert-butyl 2-((1R,5S,6S)-6-(aminomethyl)-3-ethylbicyclo[3.2.0]hept-3-en-6-yl)acetate hydrochloride is an important intermediate for the preparation of melogabalin benzenesulfonate. The following 6 specific impurities are generated during the synthesis of the above intermediates and can be transferred to the raw materials and preparations along the process. At present, there is no effective detection method for these specific impurities, which is not conducive to the quality control of the intermediate and the raw materials and preparations prepared subsequently.
[0005] Summary of the invention
[0006] Purpose of the invention: The present invention aims to provide a quality control method capable of effectively eluting, separating and quantifying specific impurities in milogabalin besylate intermediates.
[0007] Technical solution: The quality control method of the melogabalin besylate intermediate of the present invention,
[0008]
[0009] The following steps are involved:
[0010] (1) preparing sample solution;
[0011] (2) Detection: High performance liquid chromatography was used, using an octadecylsilane bonded silica gel column, a phosphate buffer solution-acetonitrile mixed solution as mobile phase A, and a methanol-acetonitrile mixed solvent as mobile phase B, and gradient elution was performed according to the following procedure:
[0012] The volume fraction of mobile phase A was reduced from 50% to 25% from 0 to 30 min, and maintained at 25% from 30 to 45 min.
[0013] (3)Analysis results.
[0014] Preferably, in the melogabalin besylate intermediate, the quality control impurities are as follows:
[0015]
[0016] Preferably, the gradient elution procedure described in step (2) is as follows:
[0017]
[0018] Preferably, the phosphate buffer solution described in step (2) is a dipotassium hydrogen phosphate buffer solution with a concentration of 10 to 50 mmol / L and a pH of 6 to 8.
[0019] More preferably, the concentration of the dipotassium hydrogen phosphate buffer solution is 30 mmol / L and the pH is 7.2.
[0020] More preferably, the pH of the dipotassium hydrogen phosphate buffer solution is adjusted to 7.2 using phosphoric acid.
[0021] Preferably, in the mobile phase A described in step (2), the volume ratio of phosphate buffer solution to acetonitrile is 95:5; and in the mobile phase B, the volume ratio of methanol to acetonitrile is 40:60.
[0022] Preferably, the chromatographic column in step (2) has a length of 150 to 250 mm, a diameter of 4.6 mm, and a filler particle size of 3.5 to 5 μm.
[0023] More preferably, the chromatographic column has a length of 250 mm, a diameter of 4.6 mm, and a filler particle size of 5 μm.
[0024] More preferably, the chromatographic column is a Thermo Hypersil GOLD chromatographic column or an equivalent chromatographic column, with a column length of 250 mm, a diameter of 4.6 mm, and a filler particle size of 5 μm.
[0025] Preferably, the detection wavelength of the detection in step (2) is 195-220 nm, the column temperature of the chromatographic column is 20-40° C., and the flow rate of the mobile phase is 0.5-1.5 ml / min.
[0026] More preferably, the detection wavelength is 215 nm, the column temperature of the chromatographic column is 30° C., and the flow rate of the mobile phase is 1 ml / min.
[0027] Preferably, the temperature of the sample plate detected in step (2) is 2-8°C.
[0028] More preferably, the sample tray temperature is 5°C.
[0029] Preferably, the injection volume for detection in step (2) is 5 μl.
[0030] Preferably, the detection linear range of the melogabalin besylate intermediate is 498.0015 to 7506.4860 μg / ml.
[0031] More preferably, the detection linear equation of the melogabalin besylate intermediate is y=0.1148x+17.3458.
[0032] Preferably, in the melogabalin besylate intermediate, the detection limit and quantification limit of each quality control component are as follows:
[0033]
[0034] Preferably, the sample described in step (1) is prepared using an acetonitrile-water solution with a volume ratio of 95:5.
[0035] Further preferably, the sample solution comprises an impurity localization solution and a system suitability solution, and the specific preparation method is as follows:
[0036] Preparation of impurity localization solution: Take appropriate amount of impurity 1, impurity 2, impurity 3, impurity 4, impurity 5 and impurity 6 reference substances, weigh them accurately, add diluent to dissolve and dilute them respectively, to make solutions with the contents of impurities 1 to impurity 6 of 15μg, 15μg, 15μg, 25μg, 15μg and 100μg per 1ml, which are used as localization solutions of impurities 1 to impurity 6 respectively.
[0037] Preparation of system suitability solution: Take appropriate amounts of melogabalin besylate intermediate, impurity 1, impurity 2, impurity 3, impurity 4, impurity 5 and impurity 6, dissolve in diluent and quantitatively dilute to make a solution containing approximately 5 mg of melogabalin besylate intermediate, 15 μg each of impurities 1 to 3 and impurity 5, 25 μg of impurity 4 and 100 μg of impurity 6 per 1 ml.
[0038] Preparation of test solution: Take an appropriate amount of melogabalin benzenesulfonate intermediate, dissolve it in a diluent and quantitatively dilute it to make a solution containing about 5 mg of melogabalin benzenesulfonate intermediate per 1 ml.
[0039] Preferably, in step (3), the content of each component is calculated using an area normalization method.
[0040] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages:
[0041] The quality control method designed by the present invention can effectively elute, separate and quantify various specific impurities in the melogabalin besylate intermediate by optimizing the chromatographic conditions and adopting gradient elution. The method has strong specificity, wide linear range, high sensitivity, good accuracy, and good solution stability and durability. The effective detection of the melogabalin besylate intermediate can be achieved, which is beneficial to the quality control of the subsequently prepared raw materials and preparation products. BRIEF DESCRIPTION OF THE DRAWINGS
[0042] Figure 1 The HPLC chromatogram of the system suitability solution of Example 1;
[0043] Figure 2 The HPLC chromatogram of the test solution of Example 1;
[0044] Figure 3 This is the HPLC chromatogram of the system suitability solution of Comparative Example 1;
[0045] Figure 4 The HPLC chromatogram of the system suitability solution of Comparative Example 2;
[0046] Figure 5 This is the HPLC chromatogram of the system suitability solution of Comparative Example 3. DETAILED DESCRIPTION
[0047] The technical solution of the present invention is further described below in conjunction with embodiments.
[0048] Example 1
[0049] 1. Experimental methods
[0050] Instrument: Thermo Ultimate 3000;
[0051] Column: Thermo Hypersil GOLD 4.6×250 mm, 5 μm;
[0052] Mobile phase: Phase A: 30 mmol / L potassium dihydrogen phosphate solution (pH adjusted to 7.2 with phosphoric acid)-acetonitrile (95:5), Phase B: methanol-acetonitrile (40:60);
[0053] Detection wavelength: 215nm;
[0054] Flow rate: 1.0ml / min;
[0055] Column temperature: 30°C;
[0056] Injector temperature: 5°C;
[0057] Injection volume: 5μl.
[0058] Table 1 Gradient elution program of Example 1
[0059]
[0060]
[0061] Dilution solution (blank solution): acetonitrile-water (95:5).
[0062] Preparation of impurity localization solution: Take appropriate amount of impurity 1, impurity 2, impurity 3, impurity 4, impurity 5 and impurity 6 reference substances, accurately weigh them, add diluent to dissolve and dilute them respectively to make solutions with the contents of impurities 1 to 6 of 15μg, 15μg, 15μg, 25μg, 15μg and 100μg per 1ml, which are used as localization solutions for impurities 1 to 6 respectively.
[0063] Preparation of system suitability solution: Take appropriate amounts of melogabalin besylate intermediate, impurity 1, impurity 2, impurity 3, impurity 4, impurity 5 and impurity 6, add diluent to dissolve and quantitatively dilute to make a solution containing approximately 5 mg of melogabalin besylate intermediate, 15 μg of impurities 1 to 3 and impurity 5, 25 μg of impurity 4, and 100 μg of impurity 6 per 1 ml, shake well and set aside.
[0064] Preparation of test solution: Take an appropriate amount of melogabalin benzenesulfonate intermediate, add diluent to dissolve and quantitatively dilute to make a solution containing about 5 mg of melogabalin benzenesulfonate intermediate per 1 ml, shake well and set aside.
[0065] Sample determination: 5 μl of blank solution, impurity location solution, system suitability solution and test solution were measured and injected into the liquid chromatograph. The amount of intermediates and impurities and their measured retention times are shown in Table 2.
[0066] Table 2 Positioning detection results of Example 1
[0067] name Impurity localization solution content (μg / ml) System suitability solution content (μg / ml) Retention time (min) Intermediates N / A 5000 17.470 Impurity 1 15 15 9.350 Impurity 2 15 15 33.660 Impurity 3 15 15 32.323 Impurity 4 25 25 23.917 Impurity 5 15 15 21.173 Impurity 6 100 100 16.443
[0068] 2. Experimental results
[0069] Depend on Figure 1It can be seen that RT = 17.470min is the chromatographic peak of the melogabalin benzenesulfonate intermediate, RT = 9.350min is the chromatographic peak of impurity 1, RT = 16.443min is the chromatographic peak of impurity 6, RT = 21.173min is the chromatographic peak of impurity 5, RT = 23.917min is the chromatographic peak of impurity 4, RT = 32.323min is the chromatographic peak of impurity 3, and RT = 33.660min is the chromatographic peak of impurity 2. The main peak of the melogabalin benzenesulfonate intermediate detected by this method can achieve baseline separation with the chromatographic peaks of adjacent impurities and known impurities, and the minimum resolution is 2.12.
[0070] 3. Methodological validation
[0071] With reference to the "Guidelines for Validation of Analytical Methods", the quality control method of Example 1 was methodologically validated.
[0072] Table 3 Methodology validation results
[0073]
[0074]
[0075] It can be seen from the above methodological verification results that the quality control method designed in the present invention has strong specificity, wide linear range, high sensitivity, high accuracy, good solution stability and durability, and can achieve effective detection of melogabalin besylate intermediates and their impurities, thus meeting the quality control requirements.
[0076] Comparative Example 1
[0077] 1. Experimental methods
[0078] Instrument: Thermo Ultimate 3000;
[0079] Chromatographic column: Welch Ultimate XB-C18 4.6×250mm, 5μm;
[0080] Mobile phase: Phase A: 10 mmol / L potassium dihydrogen phosphate solution (pH adjusted to 7.2 with phosphoric acid)-acetonitrile (95:5), Phase B: methanol-acetonitrile (50:50);
[0081] Detection wavelength: 215nm;
[0082] Flow rate: 1.0ml / min;
[0083] Column temperature: 30°C;
[0084] Injector temperature: 5°C;
[0085] Injection volume: 5μl.
[0086] Table 4 Gradient elution program of Comparative Example 1
[0087]
[0088] 2. Experimental results
[0089] Depend on Figure 3 It can be seen that RT = 26.600min is the peak of the melogabalin benzenesulfonate intermediate, RT = 13.763min is the peak of impurity 1, RT = 26.057min is the chromatographic peak of impurity 6, RT = 28.893min is the chromatographic peak of impurity 5, RT = 30.933min is the chromatographic peak of impurity 4, RT = 38.903min is the chromatographic peak of impurity 3, RT = 42.207min is the chromatographic peak of impurity 2, and the others are all chromatographic peaks of unknown impurities. The chromatographic peak of the melogabalin benzenesulfonate intermediate detected by this method cannot achieve baseline separation with the chromatographic peak of impurity 6, and therefore cannot meet the quality control requirements.
[0090] Comparative Example 2
[0091] 1. Experimental methods
[0092] Instrument: Thermo Ultimate 3000;
[0093] Column: Agilent Poroshell EC-C18, 4.6 mm × 250 mm, 4 μm;
[0094] Mobile phase: Phase A: 10 mmol / L potassium dihydrogen phosphate solution (pH adjusted to 7.2 with phosphoric acid)-acetonitrile (95:5), Phase B: methanol-acetonitrile (50:50);
[0095] Detection wavelength: 215nm;
[0096] Flow rate: 1.0ml / min;
[0097] Column temperature: 30°C;
[0098] Injector temperature: 5°C;
[0099] Injection volume: 10 μl.
[0100] Table 5 Gradient elution program of Comparative Example 2
[0101]
[0102] The impurity localization solution and system suitability solution were prepared in the same manner as in Example 1, wherein the amounts of the intermediates and impurities used and their measured retention times are shown in Table 6.
[0103] Table 6 Positioning detection results of comparative example 2
[0104] name Impurity localization solution content (μg / ml) System suitability solution content (μg / ml) Retention time (min) Intermediates N / A 5000 22.070 Impurity 1 15 15 10.900 Impurity 2 15 15 36.650 Impurity 3 15 15 34.637 Impurity 4 25 25 27.353 Impurity 5 15 15 25.173 Impurity 6 100 100 N / A
[0105] 2. Experimental results
[0106] Depend on Figure 4 It can be seen that RT=22.070min is the chromatographic peak of the melogabalin benzenesulfonate intermediate, RT=10.900min is the chromatographic peak of impurity 1, RT=25.173min is the chromatographic peak of impurity 5, RT=27.353min is the chromatographic peak of impurity 4, RT=34.637min is the chromatographic peak of impurity 3, RT=36.650min is the chromatographic peak of impurity 2, impurity 6 cannot be detected in this detection method, so it cannot meet the quality control requirements, and the others are chromatographic peaks of unknown impurities.
[0107] Comparative Example 3
[0108] 1. Experimental methods
[0109] Instrument: Thermo Ultimate 3000;
[0110] Column: Thermo Hypersil GOLD 4.6×250 mm, 5 μm;
[0111] Mobile phase: Phase A: 30 mmol / L potassium dihydrogen phosphate solution (pH adjusted to 7.2 with phosphoric acid)-acetonitrile (95:5), Phase B: acetonitrile;
[0112] Detection wavelength: 215nm;
[0113] Flow rate: 1.0ml / min;
[0114] Column temperature: 30°C;
[0115] Injector temperature: 5°C
[0116] Injection volume: 10 μl.
[0117] Table 7 Gradient elution program of Comparative Example 3
[0118]
[0119] The impurity localization solution and the system suitability solution were prepared in the same manner as in Example 1. The amounts of the intermediates and impurities used and their measured retention times are shown in Table 8.
[0120] Table 8 Positioning detection results of comparative example 3
[0121] name Impurity localization solution content (μg / ml) System suitability solution content (μg / ml) Retention time (min) Intermediates N / A 5000 9.337 Impurity 1 15 15 8.067 Impurity 2 15 15 26.620 Impurity 3 15 15 25.473 Impurity 4 25 25 16.023 Impurity 5 15 15 12.783 Impurity 6 100 100 8.730
[0122] 2. Experimental results
[0123] Depend on Figure 5 It can be seen that RT = 9.337min is the chromatographic peak of the melogabalin benzenesulfonate intermediate, RT = 8.067min is the chromatographic peak of impurity 1, RT = 8.730min is the chromatographic peak of impurity 6, RT = 12.783min is the chromatographic peak of impurity 5, RT = 16.023min is the chromatographic peak of impurity 4, RT = 25.473min is the chromatographic peak of impurity 3, RT = 26.620min is the chromatographic peak of impurity 2, and the others are all chromatographic peaks of unknown impurities. The chromatographic peak of impurity 2 detected by this method cannot achieve baseline separation with the chromatographic peak on the left, so it cannot meet the quality control requirements.
Claims
1. A quality control method for a melogabalin benzenesulfonate intermediate, It is characterized in that The following steps are involved: (1) preparing sample solution; (2) Detection: High performance liquid chromatography was used, using an octadecylsilane bonded silica gel column, a phosphate buffer solution-acetonitrile mixed solution as mobile phase A, and a methanol-acetonitrile mixed solvent as mobile phase B, and gradient elution was performed according to the following procedure: The volume fraction of mobile phase A was reduced from 50% to 25% from 0 to 30 min, and maintained at 25% from 30 to 45 min. (3)Analysis results.
2. The quality control method according to claim 1, characterized in that: In the melogabalin besylate intermediate, the quality control impurities are as follows:
3. The quality control method according to claim 1, characterized in that: The gradient elution procedure described in step (2) is as follows:
4. The quality control method according to claim 1, characterized in that: The phosphate buffer solution described in step (2) is a dipotassium hydrogen phosphate buffer solution with a concentration of 10 to 50 mmol / L and a pH of 6 to 8.
5. The quality control method according to claim 1, characterized in that: In the mobile phase A described in step (2), the volume ratio of phosphate buffer solution to acetonitrile is 95:5; in the mobile phase B, the volume ratio of methanol to acetonitrile is 40:
60.
6. The quality control method according to claim 1, characterized in that: The column length of the chromatographic column in step (2) is 150-250 mm, the diameter is 4.6 mm, and the filler particle size is 3.5-5 μm.
7. The quality control method according to claim 6, characterized in that: The chromatographic column has a length of 250 mm, a diameter of 4.6 mm, and a filler particle size of 5 μm.
8. The quality control method according to claim 1, characterized in that: The detection wavelength of the detection in step (2) is 195-220 nm, the column temperature of the chromatographic column is 20-40° C., and the flow rate of the mobile phase is 0.5-1.5 ml / min.
9. The quality control method according to claim 1, characterized in that: The temperature of the sample plate detected in step (2) is 2 to 8°C.
10. The quality control method according to claim 1, characterized in that: The sample solution described in step (1) is prepared using an acetonitrile-water solution with a volume ratio of 95:5.