A quality control method for pazufloxacin mesylate injection

Abstract

The present invention relates to the technical field of drug analysis, and is used to solve the problem that the existing quality method of pazufloxacin mesylate cannot separate photodegradable impurities, and provides a quality control method for pazufloxacin mesylate injection, comprising the following steps: (1) preparing a test solution; preparing a control solution; preparing a reference solution; (2) recording a chromatogram of the reference solution and the reference solution; (3) recording the chromatograms of the test solution and the reference solution; (4) calculating the content of pazufloxacin mesylate by peak area according to the external standard method. The present invention adjusts the ratio of two mobile phases in the gradient according to the polarity of each impurity, and changes isocratic elution to gradient elution, so that each impurity with different polarities can be well separated.

Description

Technical Field

[0001] The invention relates to the technical field of drug analysis, in particular to a quality control method for pazufloxacin mesylate injection. Background Art

[0002] Pazufloxacin mesylate sodium chloride injection is widely used in clinical practice. It is a large-volume injection and belongs to the quinolone antibiotic class. Its main ingredient is pazufloxacin mesylate, which has the characteristics of a broad antibacterial spectrum and strong antibacterial effect.

[0003] Pazufloxacin will degrade under light conditions and needs to be kept away from light during storage. The document "Determination of the Content and Related Substances of Pazufloxacin Mesylate Sodium Chloride Injection by High Performance Liquid Chromatography" Pan Xigui, Wang Yang, Zhang Xianzhou and others reported: Pazufloxacin Mesylate Sodium Chloride Injection is easily affected by light. The injection will change color to varying degrees during storage under different conditions and its content will decrease. The color change of pazufloxacin is mainly caused by the change of the side chain at position 7, which produces a small amount of new related substances. Referring to the reference preparation IF file published by PMDA, impurities D-1 and D-9 are photodegradable impurities. The structural formulas of D-1 and D-9 are as follows:

[0004]

[0005] However, the isocratic elution chromatographic conditions of the related substance detection methods in the existing quality standards for raw materials of pazufloxacin mesylate cannot separate the main degradation impurities (especially photodegradation impurities) before the main peak of pazufloxacin mesylate injection. Pazufloxacin mesylate injection is exposed to light during production and use, which may produce photodegradation impurities. Therefore, being able to accurately detect photodegradation impurities is of great significance for the quality control of this product. Summary of the invention

[0006] <Technical Problems Solved by the Invention>

[0007] The current quality control method for raw materials of pazufloxacin mesylate cannot separate the photodegradation impurities.

[0008] <Technical solution adopted by the present invention>

[0009] In view of the above technical problems, the object of the present invention is to provide a quality control method for pazufloxacin mesylate injection.

[0010] The specific contents are as follows:

[0011] A quality control method for pazufloxacin mesylate injection, wherein mobile phase A and mobile phase B are blended to obtain a mixed reagent;

[0012] The method for detecting related substances of pazufloxacin mesylate comprises the following steps:

[0013] (1-1) diluting pazufloxacin mesylate injection with a mixed reagent to obtain a test solution; accurately measuring the test solution and diluting it with the mixed reagent to obtain a control solution;

[0014] (1-2) Analyze the test solution and the control solution using high performance liquid chromatography:

[0015] Chromatographic conditions and system suitability experiments: Octadecylsilane bonded silica gel was used as filler, mobile phase A was phosphoric acid-sodium octane sulfonate solution, mobile phase B was acetonitrile and / or methanol, linear gradient elution was performed, and the detection wavelengths were 245nm and 250nm;

[0016] (1-3) Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram;

[0017] The method for detecting the content of pazufloxacin mesylate comprises the following steps:

[0018] (2-1) diluting the pazufloxacin mesylate injection with the mixed reagent to obtain a test solution; accurately measuring the pazufloxacin reference substance and diluting it with the mixed reagent to obtain a reference substance solution;

[0019] (2-2) Analyze the test solution and the reference solution using high performance liquid chromatography:

[0020] Chromatographic conditions and system suitability experiments: Octadecylsilane bonded silica gel was used as filler, mobile phase A was phosphoric acid-sodium octane sulfonate solution, mobile phase B was acetonitrile and / or methanol, linear gradient elution was performed, and the detection wavelength was 245 nm;

[0021] (2-3) Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, record the chromatogram, and calculate the content of pazufloxacin mesylate based on the peak area using the external standard method.

[0022] <Beneficial Effects of the Invention>

[0023] The chromatographic conditions were validated by methodology, and the specificity test, linearity and range, detection limit, quantification limit, precision, stability, accuracy and method robustness all met the requirements of the Chinese Pharmacopoeia, proving that the method is sensitive, reliable, reproducible and has the advantages of accurate results. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 is a chromatogram of a pazufloxacin mesylate reference substance solution (measured using the method of Example 1);

[0025] Figure 2 : is the chromatogram of pazufloxacin mesylate injection (without destruction) (measured by the method of Example 1);

[0026] Figure 3 is a chromatogram of a sample obtained by strong light irradiation destruction treatment (measured using the method of Comparative Example 1); wherein 3-1 is a complete chromatogram with a retention time of 0 to 42 min, and 3-2 is a chromatogram with a retention time of 0 to 42 min;

[0027] Figure 4 The chromatogram of the sample obtained by the strong light irradiation destruction treatment (measured by the method of Example 1);

[0028] Figure 5 is a chromatogram of a sample obtained after high temperature oxidation destruction treatment (measured using the method of Example 1);

[0029] Figure 6 is a chromatogram of a sample obtained after high temperature destruction treatment (measured using the method of Example 1);

[0030] Figure 7 is a chromatogram of a sample obtained after strong acid destruction (measured using the method of Example 1);

[0031] Figure 8 This is the chromatogram of the sample obtained after strong alkali destruction treatment (measured using the method of Example 1). DETAILED DESCRIPTION

[0032] In order to make the purpose, technical scheme and advantages of the embodiments of the present invention clearer, the technical scheme in the embodiments of the present invention will be described clearly and completely below. If the specific conditions are not specified in the embodiments, they are carried out according to conventional conditions or conditions recommended by the manufacturer. If the manufacturer of the reagents or instruments used is not specified, they are all conventional products that can be purchased commercially.

[0033] The invention provides a quality control method for pazufloxacin mesylate injection, which mainly comprises adjusting the ratio of two mobile phases in a gradient according to the polarity of each impurity, changing isocratic elution into gradient elution, and achieving good separation of impurities with different polarities.

[0034] <Related substances> are determined according to high performance liquid chromatography (Chinese Pharmacopoeia 2020 Edition, Part IV, General Rules 0512).

[0035] 1. Mixed solvent: Take mobile phase A and mobile phase B, the mixing volume ratio is 70-85:15-30, mix well, and you have it.

[0036] 2. Test solution: Take an appropriate amount of this product (pazufloxacin mesylate injection) and dilute it with a mixed solvent to make a solution containing approximately 0.24 mg of pazufloxacin per 1 ml.

[0037] 3. Control solution: Accurately measure an appropriate amount of the test solution and quantitatively dilute it with a mixed solvent to make a solution containing approximately 0.48 μg of pazufloxacin mesylate per 1 ml.

[0038] 4. Sensitivity solution: Accurately measure an appropriate amount of the pazufloxacin mesylate reference solution under the content determination item and quantitatively dilute it with a mixed solvent to make a solution containing approximately 0.06 μg per 1 ml.

[0039] 5. Chromatographic conditions: Octadecylsilane bonded silica gel is used as filler; mobile phase A is phosphoric acid-sodium octane sulfonate solution (add 0.8-1.8 g of 85% concentration phosphoric acid and 1-2 g of sodium octane sulfonate to 1000 ml and stir until dissolved); mobile phase B is acetonitrile and / or methanol.

[0040] Linear gradient elution was performed according to Table 1, with a flow rate of 1.5 ml per minute, a column temperature of 40°C, detection wavelengths of 245 nm and 250 nm, and an injection volume of 20 μl.

[0041] Table 1

[0042] Time (minutes) Mobile phase A (%) Mobile phase B (%) 0 90 10 25 78 22 60 78 22 60.1 90 10 70 90 10

[0043] System suitability requires that the signal-to-noise ratio of the main component peak height in the system sensitivity solution chromatogram (245nm) should be greater than 10.

[0044] 6. Determination method: Accurately measure the test solution and reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0045] 7. If there are impurity peaks with the same retention time as impurities D-1 and D-9 (retention time relative to the main peak of pazufloxacin is approximately 0.28 and 0.33, respectively) in the chromatogram of the limit test solution (245nm), the peak areas of impurities D-1 and D-9 (multiplied by correction factors 1.22 and 1.43, respectively) shall not be greater than 0.5 times (0.1%) the main peak area of ​​the reference solution (245nm); excluding the blank excipient peak, the peak area of ​​other single impurities (250nm) shall not be greater than 0.5 times (0.1%) the main peak area of ​​the reference solution (250nm), the sum of the known impurity content and the unknown impurity content shall not be greater than 0.2%, and the peaks less than 0.5 times the main peak area of ​​the sensitivity solution shall be ignored.

[0046] <Determination of the content of pazufloxacin mesylate> Determined according to high performance liquid chromatography (General Rules 0512 of Part IV of the Chinese Pharmacopoeia 2020 Edition).

[0047] 1. Mixed solvent: Take mobile phase A and mobile phase B, the mixing volume ratio is 70-85:15-30, mix well, and you have it.

[0048] 2. Test solution: Take an appropriate amount of this product (pazufloxacin mesylate injection) and dilute it with a mixed solvent to make a solution containing approximately 0.24 mg of pazufloxacin per 1 ml.

[0049] 3. Reference solution Take an appropriate amount of pazufloxacin reference substance (standard substance), weigh it accurately, add mixed solvent to dissolve and quantitatively dilute to make a solution containing about 0.24 mg per 1 ml.

[0050] 4. Chromatographic conditions: Use octadecylsilane bonded silica gel as filler (Medium Spectrum Red RD-C18, 4.6mm×250mm, 5μm or chromatographic column with equivalent performance); mobile phase A is phosphoric acid-sodium octane sulfonate solution (add 0.8-1.8g of 85% concentration phosphoric acid to 1000ml, add 1-2g of sodium octane sulfonate, and stir until dissolved); mobile phase B is acetonitrile and / or methanol.

[0051] Linear gradient elution was performed according to Table 1, with a flow rate of 1.5 ml per minute, a column temperature of 40°C, a detection wavelength of 245 nm, and an injection volume of 20 μl.

[0052] 5. Determination method: Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram. Calculate the peak area of ​​pazufloxacin mesylate (C 16 H 15 FN2O4) content.

[0053] <Example>

[0054] Example 1

[0055] The relevant substances were determined according to high performance liquid chromatography (General Rules 0512 of Part IV of the Chinese Pharmacopoeia 2020 Edition).

[0056] 1. Mixed solvent: Take 85 ml of mobile phase A and 15 ml of mobile phase B, mix well.

[0057] 2. Test solution: Take an appropriate amount of this product (pazufloxacin mesylate injection) and dilute it with a mixed solvent to make a solution containing approximately 0.24 mg of pazufloxacin per 1 ml.

[0058] 3. Control solution: Accurately measure an appropriate amount of the test solution and quantitatively dilute it with a mixed solvent to make a solution containing approximately 0.48 μg of pazufloxacin mesylate per 1 ml.

[0059] 4. Sensitivity solution: Accurately measure an appropriate amount of the pazufloxacin mesylate reference solution under the content determination item and quantitatively dilute it with a mixed solvent to make a solution containing approximately 0.06 μg per 1 ml.

[0060] 5. Chromatographic conditions: Use octadecylsilane bonded silica gel as filler (Medium Spectrum Red RD-C18, 4.6 mm × 250 mm, 5 μm or chromatographic column with equivalent performance); mobile phase A is phosphoric acid-sodium octane sulfonate solution (add 1.18 g of 85% concentration phosphoric acid and 1.38 g of sodium octane sulfonate to 1000 ml and stir until dissolved); mobile phase B is acetonitrile.

[0061] Linear gradient elution was performed according to Table 1, with a flow rate of 1.5 ml per minute, a column temperature of 40°C, detection wavelengths of 245 nm and 250 nm, and an injection volume of 20 μl.

[0062] System suitability requires that the signal-to-noise ratio of the main component peak height in the system sensitivity solution chromatogram (245nm) should be greater than 10.

[0063] 6. Determination method: Accurately measure the test solution and reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0064] 7. If there are impurity peaks with the same retention time as impurities D-1 and D-9 (retention time relative to the main peak of pazufloxacin is approximately 0.28 and 0.33, respectively) in the chromatogram of the limit test solution (245nm), the peak areas of impurities D-1 and D-9 (multiplied by correction factors 1.22 and 1.43, respectively) shall not be greater than 0.5 times (0.1%) the main peak area of ​​the reference solution (245nm); excluding the blank excipient peak, the peak area of ​​other single impurities (250nm) shall not be greater than 0.5 times (0.1%) the main peak area of ​​the reference solution (250nm), the sum of the known impurity content and the unknown impurity content shall not be greater than 0.2%, and the peaks less than 0.5 times the main peak area of ​​the sensitivity solution shall be ignored.

[0065] The content of pazufloxacin mesylate was determined according to the high performance liquid chromatography method (General Rules 0512 of Part IV of the Chinese Pharmacopoeia 2020 Edition).

[0066] 1. Mixed solvent: Take 85 ml of mobile phase A and 15 ml of mobile phase B, mix well.

[0067] 2. Test solution: Take an appropriate amount of this product (pazufloxacin mesylate injection) and dilute it with a mixed solvent to make a solution containing approximately 0.24 mg of pazufloxacin per 1 ml.

[0068] 3. Reference solution Take an appropriate amount of pazufloxacin reference substance (standard substance), weigh it accurately, add mixed solvent to dissolve and quantitatively dilute to make a solution containing about 0.24 mg per 1 ml.

[0069] 4. Chromatographic conditions: Use octadecylsilane bonded silica gel as filler (Medium Spectrum Red RD-C18, 4.6 mm × 250 mm, 5 μm or chromatographic column with equivalent performance); mobile phase A is phosphoric acid-sodium octane sulfonate solution (add 1.18 g of 85% concentration phosphoric acid and 1.38 g of sodium octane sulfonate to 1000 ml and stir until dissolved); mobile phase B is acetonitrile.

[0070] Linear gradient elution was performed according to Table 1, with a flow rate of 1.5 ml per minute, a column temperature of 40°C, a detection wavelength of 250 nm, and an injection volume of 20 μl.

[0071] 5. Determination method: Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram. Calculate the peak area of ​​pazufloxacin mesylate (C 16 H 15 FN2O4) content.

[0072] <Comparative Example>

[0073] Comparative Example 1

[0074] The chromatographic conditions for the inspection of related substances in Japanese Pharmacopoeia JP17 are as follows:

[0075] Chromatographic column: stainless steel column with an inner diameter of 4.6 mm and a length of 15 cm, using octadecylsilane bonded silica gel as filler (particle size of 5 μm); column temperature: 40°C; mobile phase: 1.08 g of sodium 1-octane sulfonate dissolved in 1000 mL of a mixed solution of dilute phosphoric acid (1 / 1000) and acetonitrile (39:11). Flow rate: adjust the flow rate so that the retention time of pazufloxacin is about 8 minutes. Determination time: starting from the appearance of the solvent peak, the total determination time is about 6 times the retention time of pazufloxacin.

[0076] <Test example>

[0077] The separation of impurities in samples subjected to strong light (ultraviolet light) irradiation, high temperature oxidation, high temperature destruction, strong acid, and strong base destruction under the selected chromatographic conditions was compared using the methods described in Example 1 and Comparative Example 1. The measurement wavelength was 245 nm.

[0078] The chromatogram obtained by measuring the pazufloxacin mesylate reference substance solution using the method of Example 1 is as follows: Figure 1 shown.

[0079] The chromatogram of the pazufloxacin mesylate injection reagent (not destroyed) obtained by the method of Example 1 is as follows Figure 2 shown.

[0080] Strong light exposure: 366nm destruction for 5 days; the chromatogram obtained by the method of Comparative Example 1 is shown in Figure 3, and the chromatogram obtained by the method of Example 1 is shown in Figure 4 shown.

[0081] High temperature oxidation : Add 30% hydrogen peroxide at a volume ratio of 5% and destroy at 121°C for 10 min; the chromatogram obtained by the method of Example 1 is as follows Figure 5 shown.

[0082] High temperature damage : 121°C destruction for 30 min; the chromatogram obtained by the method of Example 1 is as follows Figure 6 shown.

[0083] strong acid : 1mol / L HCl at 90°C for 3h; the chromatogram obtained by the method of Example 1 is as follows Figure 7 shown.

[0084] strong base : 1mol / L NaOH 90℃ destruction 3h; the chromatogram obtained by the method of Example 1 is as follows Figure 8 shown.

[0085] Depend on Figures 1 to 8 It can be seen that the optimized method can achieve good separation of the main peak of pazufloxacin and adjacent impurities, as well as the main photodegradation impurities before the main peak.

[0086] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A quality control method for pazufloxacin mesylate injection, characterized in that: The quality control method includes a method for detecting related substances of pazufloxacin mesylate and a method for detecting the content of pazufloxacin mesylate; mobile phase A and mobile phase B are blended to obtain a mixed reagent; The method for detecting related substances of pazufloxacin mesylate comprises the following steps: (1-1) diluting pazufloxacin mesylate injection with the mixed reagent to obtain a test solution; accurately measuring the test solution and diluting it with the mixed reagent to obtain a control solution; (1-2) Analyze the test solution and the control solution using high performance liquid chromatography: Chromatographic conditions and system suitability experiments: octadecylsilane bonded silica gel was used as filler, mobile phase A was phosphoric acid-sodium octane sulfonate solution, mobile phase B was acetonitrile and / or methanol, linear gradient elution was performed, and the detection wavelengths were 245nm and 250nm; in the mixed reagent, the volume ratio of mobile phase A to mobile phase B was 70~85:15~30; (1-3) Accurately measure the test solution and reference solution, inject them into the liquid chromatograph respectively, and record the chromatogram; The method for detecting the content of pazufloxacin mesylate comprises the following steps: (2-1) diluting pazufloxacin mesylate injection with the mixed reagent to obtain a test solution; accurately measuring pazufloxacin reference substance and diluting it with the mixed reagent to obtain a reference substance solution; (2-2) Analyze the test solution and reference solution using high performance liquid chromatography: Chromatographic conditions and system suitability experiment: octadecylsilane bonded silica gel was used as filler, mobile phase A was phosphoric acid-sodium octane sulfonate solution, mobile phase B was acetonitrile and / or methanol, linear gradient elution was performed, and the detection wavelength was 245nm; the preparation method of phosphoric acid-sodium octane sulfonate solution was as follows: 0.8-1.8g of 85% concentration phosphoric acid was added to 1000ml, and 1-2g of sodium octane sulfonate was added, and stirred until dissolved; (2-3) Accurately measure the test solution and the reference solution, inject them into the liquid chromatograph respectively, record the chromatogram, and calculate the content of pazufloxacin mesylate by the peak area according to the external standard method; (1-1) also includes a sensitivity solution; the sensitivity solution is prepared by accurately measuring an appropriate amount of the reference solution in (2-1), and quantitatively diluting it with a mixed solvent to prepare a solution containing about 0.06 μg per 1 ml; In gradient elution, the mobile phase ratios are as follows, where the mobile phase ratios are in volume percentages: 0-25min, mobile phase A is 90%-78%, mobile phase B is 10%-22%; 25-60min, mobile phase A is 78%-78%, mobile phase B is 22%-22%; 60-60.1min, mobile phase A is 78%-90%, mobile phase B is 22%-10%; 60.1-70min, mobile phase A is 90%-90%, mobile phase B is 10%-10%.

2. The quality control method of pazufloxacin mesylate injection according to claim 1, characterized in that: In (1-2) and (2-2), the flow rate was 1.5 ml per minute and the column temperature was 40 °C.

3. The quality control method of pazufloxacin mesylate injection according to claim 1, characterized in that: In (1-3) and (2-3), the injection volume was 20 μL.

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