A method for determining the dissolution rate of solid preparations of thienopyridine compounds

Abstract

The invention discloses a dissolution determination method for a solid preparation of a thienopyridine compound. The method comprises the following steps: using a pH 4.5 acetate solution containing 0.1% CTAB as a dissolution medium, placing the solid preparation of the compound in the dissolution medium, and performing a dissolution test by a paddle method, wherein the rotation speed is 65 to 85 revolutions per minute, filtering the dissolution solution at different time points, and taking each subsequent filtrate as a test solution; taking an appropriate amount of a reference substance of the compound, adding acetonitrile-water with a volume ratio of 70:30 to dissolve and dilute, obtaining a reference substance stock solution, and then diluting the reference substance stock solution with the dissolution medium to obtain a reference substance solution containing about 27.8 μg / ml of the compound; detecting the test solution and the reference substance solution by high performance liquid chromatography, comparing the solution with the labeled amount of the solid preparation of the compound, and calculating the dissolution by peak area according to an external standard method.

Description

Technical Field

[0001] The present invention relates to the field of drug analysis and detection, and in particular to a method for determining the dissolution rate of a solid preparation of a thienopyridine compound. Background Art

[0002] As a Class 1 new drug, thienopyridine compounds have not been included in the relevant quality standard control methods for thienopyridine compound tablets in the pharmacopoeias and patents of various countries. Dissolution is a key indicator for the quality control of solid preparations. Therefore, for thienopyridine compound tablets, it is necessary to establish a dissolution curve determination method with good repeatability and discrimination to provide a basis for the quality control of thienopyridine compound tablets. Summary of the invention

[0003] The chemical name of the thienopyridine compound in the present invention is 2-(4-(7-(2-fluoro-4-(1-((4-fluorophenyl)carbamoyl)cyclopropane-1-carboxamide)phenoxy)thieno[3,2-b]pyridin-2-yl)-1H-pyrazol-1-yl)ethyl valine ester dihydrochloride monohydrate, and its chemical formula is C 34 H 32 F2N6O5S·2HCl·H2O, the chemical structure is shown in (I), and the molecular weight is 765.65.

[0004]

[0005] The purpose of the present invention is to provide a dissolution determination method for the solid preparation of the above-mentioned compound, which is used to evaluate the consistency of the quality of the preparation batches and provide an indicator for the research and development of innovative drug preparations. It can be used as an indicator to confirm the consistency of drug quality and efficacy after product stability inspection and some changes (such as prescription, production process, production site changes and production process scale-up).

[0006] The dissolution determination method of the present invention comprises the following steps: using a pH 4.5 acetate solution containing 0.1% hexadecyltrimethylammonium bromide (CTAB) as a dissolution medium, placing the solid preparation of the compound in the above dissolution medium, performing a dissolution test by a paddle method, with a rotation speed of 65 to 85 revolutions per minute, filtering the dissolution solution at different time points, and taking each subsequent filtrate as a test solution; taking an appropriate amount of the compound reference substance, adding acetonitrile-water with a volume ratio of 70:30 to dissolve and dilute to obtain a reference substance stock solution, and then diluting the reference substance stock solution with the above dissolution medium to obtain a reference substance solution containing about 27.8 μg / ml of the compound; using a high performance liquid chromatography method to detect the above test solution and the reference substance solution, and comparing them with the labeled amount of the solid preparation of the compound, and calculating the dissolution by peak area according to the external standard method.

[0007] Preferably, the dissolution medium used in the present invention is prepared according to the following steps: take 2.99g of sodium acetate and 14.0ml of 2mol / L acetic acid solution, add water to dissolve and dilute to 1000ml, then add 1.0g of CTAB, dissolve it, and shake well to obtain the dissolution medium.

[0008] Preferably, the rotation speed adopted in the present invention is 75 revolutions per minute.

[0009] Preferably, the time for taking the dissolution solution in the present invention is 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 45 minutes and 60 minutes respectively.

[0010] Preferably, the present invention uses a microporous filter membrane to filter the dissolution solution to obtain each subsequent filtrate.

[0011] Preferably, the high performance liquid chromatography conditions used in the present invention are: the chromatographic column is an octadecylsilane bonded silica gel column, the mobile phase is a potassium dihydrogen phosphate solution-acetonitrile with a volume ratio of 30:70 and a pH value of 6.0, isocratic elution, the concentration of the potassium dihydrogen phosphate solution is 0.01 mol / L, the column temperature of the chromatographic column is 40°C, the flow rate is 1.5 ml / min, and the detection wavelength is 252 nm.

[0012] Preferably, the chromatographic column is Agilent Zorbax SB-C18.

[0013] Preferably, the solid preparation of the present invention is a tablet.

[0014] Preferably, the solid preparation of the present invention is a film-coated tablet.

[0015] The high performance liquid chromatography method adopted in the present invention realizes an extremely short testing time (the time for testing one injection) and can separate the main component (i.e. the raw material drug thienopyridine compound) from all other impurities, and quickly and accurately quantitatively detect the content of the main component.

[0016] The present invention adopts the above technical solution and has the following technical effects:

[0017] The thienopyridine compound of the present invention belongs to Class 1 new drugs, and its relevant dissolution determination method is not currently included in the pharmacopoeias and literature patents of various countries. The present invention provides a dissolution determination method for a solid preparation of a thienopyridine compound with discrimination, which is of great significance for the quality control and batch difference comparison of thienopyridine compound tablets. The determination method of the present invention can quickly and accurately determine the dissolution of a solid preparation of a thienopyridine compound. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] The technical scheme of the present invention will be described in detail below through examples and drawings, so as to better understand the advantages of the present invention. The provided examples should be interpreted as an illustration of the method of the present invention, and do not limit the technical scheme and protection scope disclosed by the present invention in any way.

[0019] Figure 1 is a high performance liquid chromatogram of a thienopyridine compound reference solution;

[0020] Figure 2 is the HPLC chromatogram of the test solution;

[0021] Figure 3 is the HPLC chromatogram of the blank dissolution medium of Example 3;

[0022] Figure 4 It is the HPLC chromatogram of the blank auxiliary material solution of Example 3;

[0023] Figure 5 A schematic diagram of the dissolution curves of the thienopyridine compound tablets obtained when the three batches of self-developed preparations provided in Example 2 were dissolved in an acetate solution containing 0.1% CTAB at pH 4.5 as the dissolution medium;

[0024] Figure 6 This is the linear graph of Example 6. DETAILED DESCRIPTION

[0025] The following will be combined with the accompanying drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. This detailed description should not be considered as a limitation of the present invention, but should be understood as a more detailed description of the characteristics and implementation schemes of a part of the embodiments of the present invention, rather than all the embodiments. Therefore, the following detailed description of the embodiments of the present invention provided in the accompanying drawings is not intended to limit the scope of the claimed invention, but merely represents selected embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without making creative work belong to the scope of protection of the present invention.

[0026] In the following specific embodiments, the reference substances, samples, reagents and instruments used are:

[0027] Instruments: SOTAX XTend series intelligent dissolution apparatus; Agilent 1260 series high performance liquid chromatograph;

[0028] Reagents: sodium acetate (analytical grade), acetic acid (analytical grade), hydrochloric acid (analytical grade), sodium dodecyl sulfate (analytical grade), hexadecyltrimethylammonium bromide (analytical grade), acetonitrile (chromatographic grade), potassium dihydrogen phosphate (analytical grade).

[0029] Chromatographic column:

[0030] Agilent Zorbax SB-C18, 4.6mm×150mm, 5μm.

[0031] The solvent mentioned in the present invention, unless otherwise specified, is ultrapure water.

[0032] The thienopyridine compound tablets of the present invention are prepared by the preparation room of Shenzhen Neptuno Pharmaceutical Technology Research Institute Co., Ltd., the main component: the thienopyridine compound of the present invention (code name td32-4P10), specification: 25 mg, batch number: 20230301, containing td32-4P10 (C 34 H 32 F2N6O5S·2HCl) is 102.8% of the labeled amount. The preparation method is to mix the crystalline form A of the compound prepared according to the method disclosed in Chinese patent application ZL202210358212.1 with the auxiliary materials, and then compress the tablets and coat them; wherein the raw material of compound td32-4P10 is commercially available or prepared according to the method of Chinese patent application ZL202010099747.2, batch number: J20220802, content is 99.1%.

[0033] The thienopyridine compound reference substance of the present invention is commercially available or prepared according to the method of Chinese patent application ZL202010099747.2 (compound code (td32-4P10)), batch number: 20221201, and the content is 96.5%.

[0034] [Example 1] Determination of Thienopyridine Compounds by High Performance Liquid Chromatography

[0035] 1.1 Solution preparation

[0036] Intermediate stock solution: Take about 35 mg of each of thienopyridine compound intermediate 1, thienopyridine compound intermediate 2 and thienopyridine compound intermediate 3 (for detailed information on the intermediates, see Patent No.: ZL), accurately weigh, place in a 25 ml volumetric flask, add about 4 ml of tetrahydrofuran to dissolve, dilute to the scale with acetonitrile-water (70:30), shake well, and obtain.

[0037]

[0038] Preparation of mixed solution: Take about 35 mg of the thienopyridine compound reference substance, accurately weigh it, place it in a 25 ml volumetric flask, add acetonitrile-water (70:30) to dissolve and dilute to the scale as the reference substance stock solution; accurately measure 1 ml of the reference substance stock solution and 10 μl of the intermediate stock solution, place them in a 50 ml volumetric flask, dilute to the scale with water, shake well, and obtain.

[0039] 1.2 Selection of chromatographic columns

[0040] Taking the detection method of Chinese patent application ZL202111625993.8 as a reference, 0.02 mol / L potassium dihydrogen phosphate buffer (pH 6.0)-acetonitrile (30:70) was used as the mobile phase, the detection wavelength was 252 nm, the column temperature was 30°C, the flow rate was 1.0 ml / min, and different chromatographic columns were selected. The chromatographic column information is shown in Table 1. Take 10 μl of the mixed solution under item 1.1 and inject it into the liquid chromatograph. Record the chromatogram. The main peak retention time, theoretical plate number, tailing factor, and separation between the main peak and adjacent peaks were used as evaluation indicators to screen and optimize the chromatographic column. The results are summarized in Table 2.

[0041] Table 1 Dissolution test method screening - chromatographic column information table

[0042] Serial number Chromatographic columns ① Waters XBridge Shield RP18 (4.6mm×150mm, 3.5μm) ② Waters XBridge C18 (4.6mm×150mm, 3.5μm) ③ Agilent Zorbax SB-C18 (4.6mm×150mm, 5μm) ④ Waters Symmetry C18 (3.9mm×150mm, 5μm)

[0043] Table 2 Dissolution test method screening - chromatographic column selection test results

[0044]

[0045] Under the conditions of chromatographic columns ① and ②, the impurities before the main peak did not reach baseline separation with the main peak; under the conditions of chromatographic column ④, the tailing factor of the main peak was large and the number of theoretical plates was too low; under the conditions of chromatographic column ③, the main peak was well separated from the impurities before and after it, and the tailing factor and theoretical plate number were better, so the chromatographic column was tentatively determined to be Agilent Zorbax SB-C18 (4.6mm×150mm, 5μm).

[0046] 1.3 Selection of mobile phase

[0047] Under the premise that other chromatographic conditions remain unchanged, the proportion of acetonitrile in the mobile phase was adjusted, 10 μl of the mixed solution prepared in Example 1.1 was accurately measured and injected into the liquid chromatograph respectively. The main peak elution time, tailing factor, theoretical plate number, separation degree, and maximum retention time of impurities were used as the evaluation indicators for comparison. The results are shown in Table 3.

[0048] Table 3 Dissolution test method screening - selection test results of mobile phase ratio

[0049]

[0050] The results showed that when the mobile phase A-mobile phase B, i.e. 0.02 mol / L potassium dihydrogen phosphate buffer (pH 6.0)-acetonitrile, was adjusted to 25:75, the maximum retention time of the impurity was reduced to 5.098 min, but the separation between the impurity and the main peak was reduced. At the same time, considering that the solubility of potassium dihydrogen phosphate is lower in a high-proportion organic phase, the risk of salting out in a high-proportion organic phase is higher, so the mobile phase A-mobile phase B ratio is tentatively set to (30:70).

[0051] 1.4 Selection of buffer salt concentration

[0052] Under the premise that other chromatographic conditions remain unchanged, different concentrations of buffer salts were used to accurately measure 10 μl of the mixed solution prepared in Example 1.1 and injected into the liquid chromatograph respectively. The main peak elution time, tailing factor, theoretical plate number and separation degree were used as the evaluation indicators for comparison. The results are shown in Table 4.

[0053] Table 4 Dissolution test method screening - buffer salt concentration selection test results

[0054]

[0055] The results showed that reducing the buffer salt concentration from 0.02 mol / L to 0.01 mol / L had no significant effect on the retention time, peak area, theoretical plate number and tailing factor of NEP018, and reduced the risk of salting out of the buffer salt in a high proportion of organic phase. Therefore, the buffer salt concentration of the mobile phase was tentatively set at 0.01 mol / L.

[0056] 1.5 Selection of column temperature

[0057] Under the premise that other chromatographic conditions remain unchanged, different column temperatures were used, and 10 μl of the mixed solution prepared in Example 1.1 was accurately measured and injected into the liquid chromatograph respectively. The main peak elution time, tailing factor, theoretical plate number and separation degree were used as the evaluation indicators for comparison. The results are shown in Table 5.

[0058] Table 5 Dissolution test method screening - chromatographic column temperature selection test results

[0059]

[0060] The results showed that when the column temperature increased from 30℃ to 40℃, the number of theoretical plates increased and the tailing factor decreased, so the column temperature was temporarily set at 40℃.

[0061] 1.6 Selection of mobile phase flow rate

[0062] Under the premise that other chromatographic conditions remain unchanged, different flow rates were used to accurately measure 10 μl of the mixed solution in item 1.1 and inject them into the liquid chromatograph respectively. The main peak elution time, tailing factor, theoretical plate number and maximum retention time of impurities were used as evaluation indicators for comparison. The results are shown in Table 6.

[0063] Table 6 Dissolution test method screening - mobile phase flow rate selection test results

[0064]

[0065] The results showed that when the flow rate was increased to 1.5 ml / min, the main peak elution time and the maximum retention time of impurities decreased to 4.124 min, which could shorten the running time and minimize the main peak tailing factor. Therefore, the flow rate was temporarily set to 1.5 ml / min.

[0066] Therefore, the chromatographic conditions adopted by the present invention are as follows: the chromatographic column is an octadecylsilane bonded silica gel column, the length of the chromatographic column is 150 mm, the inner diameter is 4.6 mm, and the size of the filler particles is 5 μm; the mobile phase is 0.01 mol / L potassium dihydrogen phosphate solution (pH value is adjusted to 6.0 with potassium hydroxide)-acetonitrile (30:70) as the mobile phase, isocratic elution; column temperature: 40°C; flow rate is 1.5 ml / min; detection wavelength is 252 nm; solvent: acetonitrile-water (70:30). The high performance liquid chromatography method of the present invention achieves an extremely short test time (the time for testing one injection) and can separate the main component (i.e., the raw material drug thienopyridine compound) from all other impurities, and quickly and accurately quantitatively detect the content of the main component.

[0067] [Example 2] Determination of the dissolution rate of thienopyridine compound tablets

[0068] 2.1 Selection of dissolution curve

[0069] 2.1.1 Selection of dissolution medium

[0070] According to the "Guidelines for Determination and Comparison of Dissolution Curves of Ordinary Oral Solid Preparations" and "Technical Guidelines for Dissolution Tests of Ordinary Oral Solid Preparations", the dissolution curves of thienopyridine compound tablets were measured using the paddle method at 50 rpm, using conventional media: 900 ml each of water, pH 1.0 hydrochloric acid solution, pH 4.5 acetate buffer, and pH 6.8 phosphate buffer without surfactant as the dissolution medium. The results are shown in Table 7.

[0071] Table 7 Dissolution curves of thienopyridine compounds in different dissolution media

[0072]

[0073]

[0074] The results showed that the main component dissolution amount in pH 1.0 medium was greater than 85% in 20 minutes, and the RSD values ​​of the dissolution amounts at each time point were less than 15%, indicating a faster dissolution; except for 15 minutes, the dissolution amounts at each time point in pH 6.8 medium were less than 2%, and the calculable RSDs were greater than 30%, so this medium was not used for subsequent dissolution curve determination; within the cutoff time, the average dissolution amount of NEP018 in water medium and pH 4.5 medium did not reach 85%, so subsequent attempts were made to increase the rotation speed or add an appropriate amount of surfactant to develop the dissolution curves in water medium and pH 4.5 medium.

[0075] 2.1.2 Investigation of surfactant types

[0076] The solubility of this product in water medium and pH4.5 medium at each time point is low, and the RSD of dissolution amount is large. Increasing the rotation speed alone cannot improve the above problems. Therefore, different types and concentrations of surfactants (such as SDS: sodium dodecyl sulfate, CTAB: hexadecyltrimethylammonium bromide) were added to water medium and pH4.5 medium as dissolution medium, paddle method 50rpm, medium volume 900ml, and dissolution test on tablets containing thienopyridine compounds was carried out to investigate the types of surfactants added, so as to develop the types of surfactants added to the water medium and pH4.5 medium of this product. The results are shown in Table 8.

[0077] Table 8 Surfactant type selection results

[0078]

[0079]

[0080] The results showed that: ① In the aqueous medium with 0.1% SDS, the dissolution amount of the main component was 58.3% at 90 minutes, and in the aqueous medium with 0.1% CTAB, the dissolution amount of the main component increased to 81.0% at 90 minutes, so CTAB was selected as the surfactant of the aqueous medium; ② In the pH4.5 medium, the dissolution curve of 0.1% SDS added did not change from 5 minutes to 90 minutes (75.1% to 79.3%), which was not discriminatory. The dissolution curve measured in the pH4.5 medium with 0.1% CTAB was more discriminatory, so CTAB was selected as the surfactant of the pH4.5 medium. On this basis, the speed was increased after adding CTAB in the two media to investigate whether it could effectively increase the end point dissolution.

[0081] 2.1.3 Investigation of rotation speed

[0082] Using water (containing 0.1% CTAB) medium and pH4.5 (containing 0.1% CTAB) medium as dissolution medium, paddle method at 50rpm and 75rpm, medium volume of 900ml, dissolution paddle method speed investigation test was carried out on tablets containing thienopyridine compound to determine the dissolution speed of this product. The test results are shown in Table 9.

[0083] Table 9 Results of speed selection

[0084]

[0085] The results showed that with the increase of rotation speed, the interference of excipient stacking after tablet disintegration, which led to a large RSD of dissolution rate, was improved in water (containing 0.1% CTAB) medium and pH 4.5 (containing 0.1% CTAB) medium. The dissolution amount at the same time point increased, the RSD value of the dissolution amount decreased, and the repeatability between parallel samples was higher. In addition, the endpoint dissolution of pH 4.5 (containing 0.1% CTAB) medium was greater than that of water (containing 0.1% CTAB) medium. Therefore, the paddle method of 75 rpm was used as the dissolution speed of pH 4.5 (containing 0.1% CTAB) medium.

[0086] 2.2 Finalization of the dissolution test method#

[0087] 2.2.1 Preparation of reference solution

[0088] Accurately weigh an appropriate amount of thienopyridine compound reference substance and place it in a volumetric bottle, add acetonitrile-water (70:30) to dissolve and dilute to the scale, shake well, accurately measure an appropriate amount, dilute with acetonitrile-water (70:30) to make a solution containing about 27.8μg of thienopyridine compound per 1ml, shake well, and use it as the reference substance solution.

[0089] 2.2.2 Test solution

[0090] 900 ml of pH 4.5 acetate solution (containing 0.1% CTAB) (take 2.99 g of sodium acetate and 14.0 ml of 2 mol / L acetic acid solution, add water to dissolve and dilute to 1000 ml, add 1.0 g of CTAB to dissolve, and shake well) is used as the dissolution medium; use the paddle method at a speed of 75 revolutions per minute. After 5, 10, 15, 20, 30, 45, and 60 minutes, take the dissolution solution and filter it through a microporous filter membrane, and the filtrate is used as the test solution.

[0091] 2.2.3 Chromatographic conditions

[0092] The dissolution test method for tablets containing thienopyridine compounds adopts high performance liquid chromatography, and the chromatographic conditions are as follows:

[0093] Chromatographic column: octadecylsilane bonded silica gel column, the length of the chromatographic column is 150 mm, the inner diameter is 4.6 mm, and the size of the filler particles is 5 μm;

[0094] Mobile phase: 0.01 mol / L potassium dihydrogen phosphate solution (pH adjusted to 6.0 with potassium hydroxide)-acetonitrile (30:70) as mobile phase, isocratic elution;

[0095] Column temperature: 40°C;

[0096] Flow rate: 1.5ml / min;

[0097] Detection wavelength: 252nm;

[0098] Solvent: acetonitrile-water (70:30)

[0099] 2.2.4 Determination

[0100] Accurately measure 10 μl of the reference solution and test solution in Examples 2.2.1 and 2.2.2, respectively, inject into a high performance liquid chromatograph, record the chromatogram, and calculate the amount of thienopyridine compound dissolved from each tablet at each time point by the peak area using the external standard method.

[0101] Calculation formula

[0102]

[0103] Where C is the dissolution amount of the test sample, %;

[0104] A X is the main peak area of ​​the test solution;

[0105] f is the average value of the correction factor f of the reference solution, f = G' R / A R ;

[0106] G' R Weigh the sample amount for the reference substance (pure);

[0107] V R is the dilution multiple of the reference solution;

[0108] A R is the main peak area in the reference solution;

[0109] 900 is the volume of dissolution medium, ml;

[0110] 25 represents the labeled weight of the thienopyridine compound tablet: 25 mg.

[0111] [Example 3] Specificity test

[0112] Blank excipient solution: weigh 26.4 mg of blank excipient according to the prescription ratio (equivalent to the amount of excipient corresponding to 2.8 mg of the main ingredient), place in a 100 ml volumetric flask, add an appropriate amount of dissolution medium, shake to disperse, sonicate for 5 minutes, cool, dilute to the scale with dissolution medium, shake well, and filter.

[0113] The blank dissolution medium and blank auxiliary material solution were tested respectively using the chromatographic conditions summarized in Example 2, and the chromatograms were recorded, which were respectively Figure 3-4 shown.

[0114] Results: No interfering peaks were detected in the blank dissolution medium and blank excipient solution at the corresponding positions of the main peak, indicating that this method has good specificity for the dissolution determination of this product.

[0115] [Example 4] System suitability test

[0116] Take 10 μl of the reference solution in Example 2.2.1 and inject it into a high performance liquid chromatograph. Perform the test according to the chromatographic conditions summarized in Example 2.2.3 and record the chromatogram. The chromatographic test results are shown in Figure 1 And Table 10.

[0117] Table 10 System suitability test results

[0118]

[0119] In the chromatogram of the reference solution, the theoretical plate number of the main peak was greater than 2000, and the RSD of the main peak area after 5 consecutive injections was less than 2.0%, meeting the system suitability requirements.

[0120] [Example 5] Filter membrane adsorption test

[0121] Test solution: Take the product and operate according to the dissolution curve determination method. After 30 minutes, take a sample and divide it into two parts. One part uses a microporous filter membrane (PES water system), discards 3ml, 5ml, and 10ml of the initial filtrate, and takes the subsequent filtrate as the test solution; the other part is centrifuged for 5min (10000 rpm), and the supernatant is taken as the test solution.

[0122] At least one injection of blank solvent was made, and one injection of each of the test solution obtained by centrifugation and filtration was made, and the chromatogram was recorded, and the change in the peak area of ​​the main peak was calculated. The results are shown in Table 11.

[0123] Table 11 Filter membrane adsorption test results

[0124]

[0125]

[0126] Results: After testing and calculation, the change in the peak area of ​​the main peak under different treatment methods was no more than 2.0%. The filtration should have no interference with the dissolution determination and meet the test requirements.

[0127] [Example 6] Linearity and range

[0128] 6.1 Preparation of linear solution

[0129] Linear stock solution (0.694 mg / ml): Take about 36.0 mg of the thienopyridine compound reference substance, accurately weigh it, place it in a 50 ml volumetric flask, add an appropriate amount of solvent to dissolve and dilute to the scale, and shake well.

[0130] Linear series solutions: Accurately measure an appropriate amount of linear stock solution or linear solution, and dilute with dissolution medium according to Table 12 to make solutions containing approximately 2.78 μg, 6.94 μg, 13.89 μg, 27.78 μg, and 41.67 μg of thienopyridine compound per 1 ml (equivalent to 10%, 25%, 50%, 100%, and 150% of the concentration of the test solution, respectively).

[0131] Table 12 Linear solution preparation list

[0132] Solution name Theoretical concentration (μg / ml) Equivalent to the concentration of the test solution (%) Aspirate solution / volume Dilute to Linear solution① 2.78 10％ Linear solution④ / 5ml 50ml Linear solution ② 6.94 25％ Linear solution④ / 5ml 20ml Linear solution ③ 13.89 50％ Reference substance stock solution / 1ml 50ml Linear solution ④ 27.78 100％ Reference stock solution / 2ml 50ml Linear solution⑤ 41.67 150％ Reference stock solution / 3ml 50ml

[0133] 6.2 Determination

[0134] Accurately measure 10 μl of each of the above linear series solutions, inject them into the liquid chromatograph, record the chromatogram, plot with the peak area (A) as the ordinate and the main component concentration (C) as the abscissa, and calculate the regression equation and correlation coefficient. Figure 6 And Table 13.

[0135] Table 13 Linearity test results

[0136]

[0137]

[0138] Results: The linear equation correlation coefficient R of thienopyridine compounds was greater than 0.990, the intercept was less than 25%, and the residual standard deviation was less than 10%, indicating that thienopyridine compounds had a good linear relationship in the concentration range of 2.7564μg / ml to 41.3464μg / ml (equivalent to 9.9181% to 148.7708% of the test substances).

[0139] [Example 7] Precision test

[0140] The reference solution and the 30-min test solution were prepared according to the preparation method of Example 2.2, 10 μl of the above solution was accurately measured and injected into the liquid chromatograph, and the solubility of the thienopyridine compound was calculated as the repeatability test result. The results are shown in Table 14.

[0141] Table 14 Repeatability test results

[0142] Repeatability 1 2 3 4 5 6 RSD% Dissolution (%) 97.37 88.10 88.02 91.96 89.24 86.40 2.2

[0143] Results: In the 6 test solutions, the RSD of the solubility of thienopyridine compounds measured was less than 10.0%, indicating that the method had good repeatability.

[0144] The product was tested by different people, different equipment and different dates according to the repeatability method, and the RSD of the dissolution of the thienopyridine compound was calculated as the intermediate precision test result. The results of the repeatability and intermediate precision tests were combined to calculate the RSD of the dissolution of the twelve samples. The results are shown in Table 15.

[0145] Table 15 Intermediate precision results

[0146]

[0147] Results: A and B were the operators. The RSD of the dissolution rates of the 12 test solutions was less than 10.0%, indicating that the intermediate precision of this method was good.

[0148] [Example 8] Solution stability

[0149] Accurately measure 10 μl of the reference solution and the test solution in Example 2.2, and inject them into the liquid chromatograph at 0h, 2h, 4h, 6h, 8h, 10h, 12h, 16h, 20h, and 24h, respectively. Record the chromatogram and calculate the change in the peak area of ​​the main peak. The results are shown in Table 17.

[0150] Table 17 Solution stability results

[0151]

[0152] Compared with the stability solution at 0h, the changes in the main peak area of ​​the reference solution within 24h and the test solution within 12h were both less than 2.0%. The solution had good stability and met the detection requirements.

[0153] [Example 9] Durability test

[0154] The durability test was carried out on different types of chromatographic columns with column temperature ±2°C, flow rate ±0.1ml / min, mobile phase pH ±0.5.

[0155] The reference solution was prepared according to the preparation method of Example 2, and 10 μl was accurately measured and injected into the liquid chromatograph. The test was performed after appropriate adjustments based on the chromatographic conditions summarized in Example 2.2.3. The test results are shown in Table 18.

[0156] Table 18 Durability test results

[0157]

[0158] The results showed that when all parameters changed slightly, the theoretical plate number of the main peak and the RSD of the main peak area of ​​5 consecutive injections met the system suitability requirements.

Claims

1. A method for determining the dissolution rate of a solid preparation of a compound having a structure represented by formula (I), characterized in that: The following steps are involved: The method comprises the following steps: using a pH 4.5 acetate solution containing 0.1% CTAB as a dissolution medium, placing the solid preparation of the compound in the dissolution medium, and performing a dissolution test by a paddle method, wherein the rotation speed is 65 to 85 revolutions per minute, filtering the dissolution solution at different time points, and taking each subsequent filtrate as a test solution; taking an appropriate amount of the reference compound, dissolving and diluting the reference compound stock solution in acetonitrile-water at a volume ratio of 70:30 to obtain a reference compound stock solution, and then diluting the reference compound stock solution with the dissolution medium to obtain a reference compound solution containing about 27.8 μg / ml of the compound; detecting the test solution and the reference compound solution by high performance liquid chromatography, and comparing them with the labeled amount of the solid preparation of the compound, and calculating the dissolution by peak area according to the external standard method; 2. The dissolution determination method according to claim 1, characterized in that The dissolution medium was prepared according to the following steps: 2.99 g of sodium acetate and 14.0 ml of 2 mol / L acetic acid solution were dissolved in water and diluted to 1000 ml, and then 1.0 g of CTAB was added to dissolve the mixture and shaken to obtain the dissolution medium.

3. The dissolution determination method according to claim 1, characterized in that: The rotation speed is 75 revolutions per minute.

4. The dissolution determination method according to claim 1, characterized in that: The times for taking the dissolution solution are 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 45 minutes and 60 minutes respectively.

5. The dissolution determination method according to claim 4, characterized in that: The dissolution solution is filtered through a microporous filter membrane to obtain the subsequent filtrate.

6. The dissolution determination method according to claim 1, characterized in that: The high performance liquid chromatography conditions are as follows: the chromatographic column is an octadecylsilane bonded silica gel column, the mobile phase is a potassium dihydrogen phosphate solution-acetonitrile with a volume ratio of 30:70 and a pH value of 6.0, isocratic elution, the concentration of the potassium dihydrogen phosphate solution is 0.01 mol / L, the column temperature of the chromatographic column is 40°C, the flow rate is 1.5 ml / min, and the detection wavelength is 252 nm.

7. The dissolution determination method according to claim 6, characterized in that: The chromatographic column is Agilent ZorbaxSB-C18.

8. The dissolution determination method according to claim 1, characterized in that: The solid preparation of the compound is a tablet.

9. The dissolution determination method according to claim 8, characterized in that: The tablet is a film-coated tablet.

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