Method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection

Abstract

The invention provides a method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection, which adopts a liquid chromatography detection method, and the chromatographic conditions are as follows: Thermo Hypersil GOLDTM 4.6 * 250mm, 5 [mu] m chromatographic column; octadecyl silane bonded silica gel is used as a filling agent; a mobile phase A is water, the pH value is adjusted to 3.5 by using phosphoric acid, a mobile phase B is acetonitrile, and gradient elution is performed; the flow rate is 0.9 to 1.1 ml / min; the column temperature is 35-45 DEG C; the detection wavelength is 240 nm; the sample injection volume is 10 microliters. According to the method, the photodegradable impurities in the pazufloxacin mesilate sodium chloride injection can be rapidly detected by selecting proper liquid chromatography conditions, the separation degree of pazufloxacin mesilate and each impurity is good, and the detection result is accurate and credible; based on the method, the quality of the pazufloxacin mesylate sodium chloride injection can be quickly and effectively controlled, and the safety of the medicine is ensured.

Description

Technical Field

[0001] The invention relates to the technical field of drug analysis and detection, and in particular to a method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection. Background Art

[0002] Pazufloxacin mesylate sodium chloride injection is an antibiotic drug used to treat a variety of infections caused by sensitive bacteria. Its main ingredients include pazufloxacin mesylate and sodium chloride. Among them, pazufloxacin mesylate is sensitive to light and is prone to photodegradation impurities. However, the detection method for photodegradation impurities in the domestic and international pharmacopoeia standards for pazufloxacin mesylate sodium chloride injection is not included, and its risks cannot be controlled.

[0003] In this regard, those skilled in the art began to study a method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection. For example, patent application number 202111645340.6 discloses a quality control method for pazufloxacin mesylate injection. However, the detection time of the method disclosed in the above patent is relatively long, and the optical gradient elution requires 70 minutes, which is inefficient.

[0004] Therefore, a new method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection with higher detection efficiency is urgently needed. Summary of the invention

[0005] The technical problem to be solved by the present invention is to provide a method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection, the method has short detection time and high efficiency, and can achieve good separation between pazufloxacin mesylate and various impurities; based on the method, the present invention can effectively control the quality of pazufloxacin mesylate sodium chloride injection and ensure the safety of the drug.

[0006] The present invention adopts the following technical solutions to solve the above technical problems:

[0007] A method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection, using liquid chromatography detection method, the chromatographic conditions are: using Thermo Hypersil GOLD TM 4.6×250mm, 5μm chromatographic column; octadecylsilane bonded silica gel as filler; mobile phase A is water, pH value is adjusted to 3.5 with phosphoric acid, mobile phase B is acetonitrile, gradient elution; flow rate is 0.9~1.1ml / min; column temperature is 35~45℃; detection wavelength is 240nm; injection volume is 10μl.

[0008] As one of the preferred embodiments of the present invention, in the gradient elution, the mobile phase ratio is as follows:

[0009] 0 min, mobile phase A was 60%, mobile phase B was 40%;

[0010] 5 min, mobile phase A: 70%, mobile phase B: 30%;

[0011] 10 min, mobile phase A: 50%, mobile phase B: 50%;

[0012] 15 min, mobile phase A: 60%, mobile phase B: 40%;

[0013] 20min, mobile phase A was 60%, mobile phase B was 40%.

[0014] As one of the preferred embodiments of the present invention, the method comprises the following specific steps:

[0015] (1) Solution preparation

[0016] Test solution: Take pazufloxacin mesylate sodium chloride injection and dilute it with diluent to obtain the test solution;

[0017] Reference solution: Take the impurity raw material and dilute it with diluent to obtain the corresponding reference solution;

[0018] (2) Liquid chromatography analysis

[0019] The liquid chromatography was used for detection. The chromatographic conditions were: Thermo Hypersil GOLD TM 4.6×250mm, 5μm chromatographic column; octadecylsilane bonded silica gel as filler; mobile phase A is water, pH value adjusted to 3.5 with phosphoric acid, mobile phase B is acetonitrile, gradient elution; flow rate 0.9~1.1ml / min; column temperature 35~45℃; detection wavelength 240nm; injection volume 10μl;

[0020] Take the reference solution and the test solution, inject them separately, record the chromatograms, and calculate the content of each component.

[0021] As one of the preferred embodiments of the present invention, in step (1), the diluent is formulated as follows: adjust the pH of water to 3.5 with phosphoric acid, and then mix with acetonitrile in a volume ratio of 60:40.

[0022] As one of the preferred embodiments of the present invention, in step (1), the concentration of the reference solution is 0.2 mg / ml.

[0023] As one of the preferred embodiments of the present invention, in the reference solution, the impurity raw materials include impurity 10, impurity 11, pazufloxacin ethyl ester, and levofloxacin ester; wherein impurities 10 and 11 are known photodegradable impurities in pazufloxacin mesylate sodium chloride injection, and the structural formula is as follows:

[0024]

[0025] As one of the preferred embodiments of the present invention, in step (1), the concentration of the test solution is 1 mg / ml.

[0026] As one of the preferred embodiments of the present invention, in step (2), the content of each component is calculated according to the external standard method.

[0027] The advantages of the present invention compared to the prior art are:

[0028] (1) The present invention can quickly detect the photodegradable impurities in pazufloxacin mesylate sodium chloride injection by selecting appropriate liquid chromatography conditions (chromatographic column, filler, mobile phase composition, mobile phase pH, 20 min elution program, etc.), and the separation degree between pazufloxacin mesylate and various impurities is good, and the detection result is accurate and reliable; based on this method, the present invention can quickly and effectively control the quality of pazufloxacin mesylate sodium chloride injection and ensure the safety of the drug;

[0029] (2) The detection method provided by the present invention has strong specificity and good durability. Changing the chromatographic conditions such as column flow rate and column temperature within a small range will not affect the detection of impurities. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 It is the location diagram of each impurity in the separation test in Experimental Example 1 (in the figure, the main peak of the test product is the peak of pazufloxacin mesylate);

[0031] Figure 2 is a comparative chromatogram of the blank solvent and the separation test solution in the separation test in Experimental Example 1 (in the figure, the illuminated sample refers to the separation test solution);

[0032] Figure 3 This is a comparative chromatogram of the comparative test of the existing method in Experimental Example 1 (in the figure, the main peak of the test product is the pazufloxacin mesylate peak). DETAILED DESCRIPTION

[0033] The embodiments of the present invention are described in detail below. The embodiments are implemented on the premise of the technical solution of the present invention, and detailed implementation methods and specific operation processes are given, but the protection scope of the present invention is not limited to the following embodiments. The raw materials, reagents, equipment and methods used in the following embodiments are conventional raw materials, reagents, equipment and methods in the art unless otherwise specified, and will not be repeated.

[0034] Meanwhile, the structural information of pazufloxacin mesylate and various common impurities of the present invention is shown in Table 1.

[0035] Table 1 Structural information of pazufloxacin mesylate and common impurities

[0036]

[0037]

[0038] Example 1

[0039] A method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection in this embodiment comprises the following steps:

[0040] (1) Solution preparation

[0041] Diluent: Adjust the pH of water to 3.5 with phosphoric acid and mix with acetonitrile in a volume ratio of 60:40.

[0042] Test solution: Take a sample of pazufloxacin mesylate sodium chloride injection and dilute it to 1 mg / ml (pazufloxacin mesylate sodium chloride concentration) with a diluent to obtain the test solution.

[0043] Reference solution: Take impurity 10, impurity 11, pazufloxacin ethyl ester and levofloxacin ester raw materials respectively, dilute them to 0.2 mg / ml with diluent, and obtain reference solution.

[0044] (2) Liquid chromatography analysis

[0045] Agilent 1260infinity II high performance liquid chromatograph was used for detection. The chromatographic conditions were: Liquid chromatography detection method was used. The chromatographic conditions were: Thermo Hypersil GOLD TM 4.6×250mm, 5μm chromatographic column; octadecylsilane bonded silica gel as filler; mobile phase A is water, pH value is adjusted to 3.5 with phosphoric acid, mobile phase B is acetonitrile, gradient elution is performed according to Table 2; flow rate is 0.9~1.1ml / min; column temperature is 35~45℃; detection wavelength is 240nm; injection volume is 10μl.

[0046] Table 2 Gradient elution program (mobile phase ratio, volume percentage)

[0047] Time (min) Mobile phase A (%) Mobile phase B (%) 0 60 40 5 70 30 10 50 50 15 60 40 20 60 40

[0048] Take the reference solution and the test solution, inject them separately under the above-mentioned chromatographic conditions, and record the chromatograms. If there are chromatographic peaks in the chromatogram of the test solution that are consistent with the retention time of each impurity peak, calculate the peak area according to the external standard method.

[0049] Example 2

[0050] The method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection of this embodiment is basically the same as that of Example 1, with the main difference being that the flow rate is 0.9 ml / min.

[0051] Example 3

[0052] The method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection of this embodiment is basically the same as that of Example 1, with the main difference being that the flow rate is 1.1 ml / min.

[0053] Example 4

[0054] The method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection of this embodiment is basically the same as that of Example 1, with the main difference being that the column temperature is 35°C.

[0055] Example 5

[0056] The method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection of this embodiment is basically the same as that of Example 1, with the main difference being that the column temperature is 45°C.

[0057] Comparative Example 1

[0058] This comparative example provides a method for detecting photodegradation impurities in an existing pazufloxacin mesylate sodium chloride injection (i.e., the method disclosed in patent application number 202111645340.6).

[0059] Experimental Example 1

[0060] This experimental example is used to verify the feasibility and application advantages of the method of the present invention.

[0061] 1. Separation test

[0062] The instrument and chromatographic conditions were the same as those in Example 1.

[0063] Blank solvent (ie, diluent): The pH of water was adjusted to 3.5 with phosphoric acid, and then mixed with acetonitrile in a volume ratio of 60:40.

[0064] Separation test solution: Take a sample of pazufloxacin mesylate sodium chloride injection for illumination and dilute it to 1 mg / ml (pazufloxacin mesylate sodium chloride concentration) with a diluent.

[0065] Impurity 10 localization solution: Take impurity 10 and dissolve it in a solvent to dilute it to 0.2 mg / ml.

[0066] Impurity 11 localization solution: Take impurity 11 and dissolve and dilute it to 0.2 mg / ml with solvent.

[0067] Pazufloxacin ethyl ester localization solution: Take pazufloxacin ethyl ester and dissolve it in solvent to dilute it to 0.2 mg / ml.

[0068] Levoflurane localization solution: Take levoflurane and dissolve it in a solvent to dilute it to 0.2 mg / ml.

[0069] Take the blank solvent, separation test solution and each location solution and inject them respectively, and record the chromatogram. Among them, the location map of each impurity is as follows: Figure 1 As shown, the comparison between blank solvent and separation test solution is shown in Figure 2 The separation result data are shown in Table 3.

[0070] Table 3 Separation test results

[0071]

[0072]

[0073] The above results show that the blank solvent has no interference with the components to be tested; the impurities 10, 11, levofloxacin ester and ethyl ester of the tested solvent have no interference with the detection.

[0074] 2. Durability test

[0075] Referring to the method of Example 1, the flow rate, column temperature, mobile phase pH and chromatographic column were changed within a certain range to test the durability of the chromatographic conditions and provide a basis for determining the chromatographic conditions. The impurity peaks and other peaks must be baseline separated.

[0076] ① Different flow rates

[0077] The flow rate was changed to 0.9ml / min, 1.0ml / min, and 1.1ml / min to examine the tolerance of this method to flow rate changes.

[0078] ② Different column temperatures

[0079] The column temperature was changed to 35℃, 40℃, and 45℃ to examine the tolerance of this method to column temperature changes.

[0080] ③ Different pH of mobile phase

[0081] The pH of the mobile phase was changed to 3.3, 3.5, and 3.7 to investigate the tolerance of this method to changes in the mobile phase pH.

[0082] ④Different chromatographic columns

[0083] Another column of the same brand and model (Thermo Hypersil GOLD TM 4.6×250mm) to investigate the tolerance of this method to chromatographic column changes.

[0084] Take the above-mentioned pazufloxacin mesylate sodium chloride injection illumination sample solution, change the flow rate, column temperature, different mobile phase pH and chromatographic column, accurately measure 10 μl and inject it into the liquid chromatograph, record the chromatogram, and the results are shown in Tables 4 to 6.

[0085] Table 3 Test results at different flow rates

[0086]

[0087] Table 4 Test results at different column temperatures

[0088]

[0089] Table 5 Test results of different mobile phase pH

[0090]

[0091] Table 6 Test results of different chromatographic columns

[0092]

[0093] From the above results, it can be seen that by changing the flow rate, column temperature and chromatographic column, the separation of pazufloxacin mesylate, impurity 10 and various unknown impurities meets the requirements, indicating that different flow rates, different column temperatures and different chromatographic columns of the same brand and model have little effect on the chromatographic conditions; but different pH values ​​of the mobile phase will affect the separation of the photodegradable impurity 1 and adjacent impurities, so the pH of the mobile phase has a greater impact on the chromatographic conditions, and the optimal pH is 3.5.

[0094] It should be noted that in this durability test, because impurity 11, pazufloxacin ethyl ester and levofloxacin ester were not detected in the samples, the specific data were not counted.

[0095] 3. Comparative Test of Existing Methods

[0096] According to the method of the present invention (Example 1 as an example) and the comparative example 1, the same sample was tested, and the results are as follows Figure 3 shown.

[0097] Depend on Figure 3 The results show that the comparative method has a longer detection time than the present invention, and the pazufloxacin ethyl ester impurity cannot be effectively separated from the pazufloxacin main peak, which is not conducive to the detection of impurities. Therefore, the present invention has a better application effect than the comparative method.

[0098] In summary, the method of the present invention has short detection time and high efficiency, can achieve good separation between pazufloxacin mesylate and various impurities, effectively control the quality of pazufloxacin mesylate sodium chloride injection, and ensure the safety of the drug; at the same time, the detection method provided by the present invention has strong specificity and good durability, and changing chromatographic conditions such as flow rate and column temperature within a small range will not affect the detection of impurities.

[0099] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection, characterized in that: Liquid chromatography was used for detection. The chromatographic conditions were as follows: Thermo Hypersil GOLD TM 4.6×250mm, 5μm chromatographic column; octadecylsilane bonded silica gel as filler; mobile phase A is water, pH value is adjusted to 3.5 with phosphoric acid, mobile phase B is acetonitrile, gradient elution; flow rate is 0.9~1.1ml / min; column temperature is 35~45℃; detection wavelength is 240nm; injection volume is 10μl.

2. The method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection according to claim 1, characterized in that: In the gradient elution, the mobile phase examples are as follows: 0 min, mobile phase A was 60%, mobile phase B was 40%; 5 min, mobile phase A: 70%, mobile phase B: 30%; 10 min, mobile phase A: 50%, mobile phase B: 50%; 15 min, mobile phase A: 60%, mobile phase B: 40%; 20min, mobile phase A was 60%, mobile phase B was 40%.

3. The method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection according to claim 1 or 2, characterized in that: The specific steps include: (1) Solution preparation Test solution: Take pazufloxacin mesylate sodium chloride injection and dilute it with diluent to obtain the test solution; Reference solution: Take the impurity raw material and dilute it with diluent to obtain the corresponding reference solution; (2) Liquid chromatography analysis The liquid chromatography was used for detection. The chromatographic conditions were: ThermoHypersil GOLD TM 4.6×250mm, 5μm chromatographic column; octadecylsilane bonded silica gel as filler; mobile phase A is water, pH value is adjusted to 3.5 with phosphoric acid, mobile phase B is acetonitrile, gradient elution; flow rate is 0.9~1.1ml / min; column temperature is 35~45℃; detection wavelength is 240nm; injection volume is 10μl; Take the reference solution and the test solution, inject them separately, record the chromatograms, and calculate the content of each component.

4. The method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection according to claim 3, characterized in that: In the step (1), the diluent is formulated as follows: adjust the pH of water to 3.5 with phosphoric acid, and then mix with acetonitrile in a volume ratio of 60:

40.

5. The method for detecting photodegradation impurities in pazufloxacin mesylate sodium chloride injection according to claim 3, characterized in that: In the step (1), the concentration of the reference solution is 0.2 mg / ml.

6. The method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection according to claim 3, characterized in that: In the reference solution, the impurity raw materials include impurity 10, impurity 11, pazufloxacin ethyl ester, and levofloxacin ester; wherein impurities 10 and 11 are known photodegradable impurities in pazufloxacin mesylate sodium chloride injection, and the structural formula is as follows:

7. The method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection according to claim 3, characterized in that: In the step (1), the concentration of the test solution is 1 mg / ml.

8. The method for detecting photodegradable impurities in pazufloxacin mesylate sodium chloride injection according to claim 3, characterized in that: In the step (2), the content of each component is calculated according to the external standard method.

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