WO2023272962A1

WO2023272962A1 - Method for detecting small polar impurities in pazufloxacin mesylate bulk drug

Info

Publication number: WO2023272962A1
Application number: PCT/CN2021/119262
Authority: WO
Inventors: 刘玉; 韦家华; 王汕桃
Original assignee: 海南海神同洲制药有限公司
Priority date: 2021-06-30
Filing date: 2021-09-18
Publication date: 2023-01-05
Also published as: CN113702517A

Abstract

Provided is a method for detecting small polar impurities in a pazufloxacin mesylate bulk drug, wherein detection is performed using a high-performance liquid chromatography method; and a mobile phase comprises acetonitrile, a 10% triethylamine mesylate solution, a 1.0 mol/L dipotassium hydrogen phosphate, and water at a volume ratio of 45 : 10 : 7 : 138. The method comprises the following steps: respectively injecting a test sample solution and a control solution into a high-performance liquid chromatograph, and recording a chromatogram until 7 times the retention time of a pazufloxacin mesylate characteristic peak. According to the method, a pazufloxacin mesylate bulk drug is subjected to detection by using HPLC, and small polar impurities with a retention time longer than that of a pazufloxacin mesylate characteristic peak can be effectively detected within 7 times the retention time of pazufloxacin mesylate by means of a mobile phase with a specific ratio. Therefore, the quality control of a pazufloxacin mesylate bulk drug can be more accurately and scientifically realized.

Description

Detection method of small polar impurities in Pazufloxacin mesylate API

technical field

The application relates to the field of quality control of the raw materials of pazufloxacin mesylate, in particular to a detection method for small polar impurities in the raw materials of pazufloxacin mesylate.

Background technique

Pazufloxacin mesylate is a new type of fluoroquinolone antibacterial drug developed by Japan. It is the mesylate salt of Pazufloxacin. It has the characteristics of broad antibacterial spectrum, high antibacterial activity, small side effects and good tolerance. It is clinically used to treat Gram-positive and negative bacterial infections, such as bronchial and lung infections, bacillary dysentery, urinary system, skin and soft tissue infections.

Pazufloxacin mesylate API may produce some degradation products after exposure to light, oxidation, high temperature, and acid and alkali during storage, transportation and use. These degradation products may affect the efficacy and even safety of the drug. will have an impact, therefore, it is necessary to detect these degradation products.

In the prior art, high performance liquid chromatography (HPLC) is usually used to detect related substances in the bulk drug of pazufloxacin sulfonate. The inventor finds after studying a large amount of HPLC detection methods, the flow that these HPLC detection methods adopts is relative to the impurity of small polarity, especially the impurity that retention time is greater than main drug pazufloxacin mesylate characteristic peak, and cannot be detected effectively. These small polar impurities may be derived from some degradation products of Pazufloxacin Mesylate bulk drug due to light, oxidation, high temperature, and acid and alkali during storage, transportation and use. The curative effect and even the safety of the drug may be affected. Therefore, a comprehensive and accurate detection of these small polar impurities is very necessary for the quality control of the drug.

Contents of the invention

The invention provides a method for detecting small polar impurities in the raw drug of pazufloxacin mesylate, so as to control the quality of the raw drug of pazufloxacin mesylate more comprehensively and accurately.

The technical solution is as follows:

A method for detecting small polar impurities in the bulk drug of pazufloxacin mesylate, which is detected by high performance liquid chromatography, and the mobile phase is acetonitrile-10% trimethanesulfonate with a volume ratio of 45:10:7:138 Ethylamine solution-1.0mol/L dipotassium hydrogen phosphate-water;

The method comprises the steps of:

With mobile phase as solvent, prepare the test solution of Pazufloxacin mesylate bulk drug;

Get described need testing solution and dilute with mobile phase, obtain contrast solution;

Get need testing solution and contrast solution inject high performance liquid chromatography respectively, record chromatogram to the retention time of 7 times of pazufloxacin mesylate characteristic peak;

The peak area of the impurity whose retention time is greater than the characteristic peak of pazufloxacin mesylate in the spectrum of the test solution obtained is compared with the characteristic peak area of pazufloxacin mesylate in the spectrum of the test solution to determine The content of this impurity relative to pazufloxacin mesylate.

In some embodiments of the present application, the test solution is diluted 100 times to obtain the control solution.

In some embodiments of the present application, the concentration of the bulk drug of pazufloxacin mesylate in the test solution is 0.3 mg/ml.

In some embodiments of the present application, the injection volume of the high performance liquid chromatograph is 20 μl.

In some embodiments of the present application, the chromatographic conditions of high performance liquid chromatography also include:

Stationary phase: Octadecylsilane bonded silica gel,

Flow rate: 1.0ml/min,

Detection wavelength: 254nm.

In some embodiments of the present application, the resolution of high performance liquid chromatography is 1.6.

In some embodiments of the present application, the number of theoretical plates of high-performance liquid chromatography calculated based on the characteristic peak of pazufloxacin mesylate should not be less than 2000.

In some embodiments of the present application, before taking need testing solution and control solution respectively and injecting high-performance liquid chromatograph, described method also comprises:

Take 20 μl of the control solution and inject it into the liquid chromatograph, and adjust the detection sensitivity so that the peak height of the main component chromatographic peak is 10-25% of the full scale.

Beneficial effect

The detection method of the present application adopts HPLC to detect the raw material drug of pazufloxacin mesylate, and through the mobile phase of specific proportioning, the retention time can be effectively reduced to 7 times of that of pazufloxacin mesylate. Small polar impurities larger than the characteristic peaks of pazufloxacin mesylate were detected. Therefore, the quality control of the Pazufloxacin Mesylate API can be realized more accurately and scientifically.

Description of drawings

Fig. 1A is the HPLC chromatogram that adopts mobile phase 1 to detect the bulk drug of pazufloxacin mesylate in embodiment 1;

Fig. 1B is the HPLC chromatogram that adopts mobile phase 2 to detect the bulk drug of pazufloxacin mesylate in Example 1;

Fig. 1 C is the HPLC chromatogram that adopts mobile phase 3 to detect the bulk drug of pazufloxacin mesylate in Example 1;

Fig. 1D is the HPLC chromatogram that adopts mobile phase 4 to detect the bulk drug of pazufloxacin mesylate in Example 1;

Fig. 2 is the HPLC chromatogram of the pazufloxacin mesylate crude drug after photodegradation in embodiment 2;

Fig. 3 is the HPLC chromatogram of the pazufloxacin mesylate crude drug after acid degradation in embodiment 3;

Fig. 4 is the HPLC chromatogram of the bulk drug of pazufloxacin mesylate after alkaline degradation in Example 4.

detailed description

In order to make the purpose, technical solution and advantages of the present application clearer, the technical solution of the present application will be clearly and completely described through specific examples below.

It should be noted that, in the following examples, those without specific conditions were carried out according to conventional conditions or conditions suggested by the manufacturer. The reagents or instruments used were not indicated by the manufacturer, and they were all conventional products that could be purchased from the market.

Embodiment 1 is used to detect the screening of the mobile phase of small polar impurity

The mobile phase pazufloxacin mesylate raw materials in Table 1 below were used for detection.

Table 1

流动相1mobile phase 1	乙腈-10％甲磺酸三乙胺-1M磷酸氢二钾-水(30:10:7:170)Acetonitrile-10% triethylamine methanesulfonate-1M dipotassium hydrogen phosphate-water (30:10:7:170)
流动相2mobile phase 2	乙腈-10％甲磺酸三乙胺-1M磷酸氢二钾-水(30:10:7:153)Acetonitrile-10% triethylamine methanesulfonate-1M dipotassium hydrogen phosphate-water (30:10:7:153)
流动相3mobile phase 3	乙腈-10％甲磺酸三乙胺-1M磷酸氢二钾-水(45:10:7:138)Acetonitrile-10% triethylamine methanesulfonate-1M dipotassium hydrogen phosphate-water (45:10:7:138)
流动相4mobile phase 4	乙腈-10％甲磺酸三乙胺-1M磷酸氢二钾-水(70:10:7:113)Acetonitrile-10% triethylamine methanesulfonate-1M dipotassium hydrogen phosphate-water (70:10:7:113)

Wherein, the preparation of 10% triethylamine methanesulfonate solution is as follows:

Under the condition of ice bath, slowly add 30ml of methanesulfonic acid and 30ml of triethylamine into 200ml of water, after the dissolution is complete, add water to 300ml.

The chromatographic conditions are as follows:

Stationary phase: Octadecylsilane bonded silica gel,

Mobile phase: see Table 1

Flow rate: 1.0ml/min,

Detection wavelength: 254nm,

Injection volume: 20 μl.

The detection steps are as follows:

(1) Prepare the test solution: take 30 mg of pazufloxacin mesylate crude drug, accurately weigh it, put it in a 100ml measuring bottle, add mobile phase to dissolve and dilute to the mark, shake well, and use it as the test solution.

(2) according to above-mentioned chromatographic condition, get need testing solution 20 μ l injection liquid chromatograph respectively, under the condition of mobile phase 1 and mobile phase 2, record chromatogram to pazufloxacin mesylate characteristic peak retention time 2 times, under the conditions of mobile phase 3 and mobile phase 4, record the chromatogram to 7 times of the retention time of the characteristic peak of pazufloxacin mesylate, and the corresponding HPLC chromatograms of different flows are shown in Figure 1A-Figure 1D respectively Show.

It can be seen from Figure 1A-Figure 1D that when mobile phase 1 and mobile phase 2 are used to detect the raw material drug of pazufloxacin mesylate, impurities whose retention time is greater than the characteristic peak of pazufloxacin mesylate cannot be detected . For mobile phase 3, the less polar related substances located after the retention time of pazufloxacin mesylate can be effectively detected within 7 times of the retention time of the main drug pazufloxacin mesylate, and the same as the main drug. Drugs are well separated. Compared with mobile phase 3, the detection effect of mobile phase 4 is not ideal.

It can be seen that the application adopts the mobile phase of acetonitrile-10% triethylamine methanesulfonate solution-1.0mol/L dipotassium hydrogen phosphate-water with a volume ratio of 45:10:7:138. Within the retention time of the characteristic peak of pazufloxacin acid, small polar impurities can be detected comprehensively and accurately.

Example 2 HPLC qualitative detection of Pazufloxacin mesylate raw material after photodegradation

Using mobile phase 3, other chromatographic conditions are the same as in Example 1.

The detection steps are as follows:

(1) Preparation of photodegradation test solution: take 30 mg of this product, put it in a 100 ml measuring bottle, use mobile phase to make a solution containing 300 μg of pazufloxacin mesylate per 1 ml, put it under ultraviolet light for 7 days, and use it as a Test solution.

(2) Perform HPLC detection according to the same method as in Example 1. The chromatogram is shown in Figure 3.

As can be seen from Figure 3, for the crude drug after photodegradation, the small polarity produced by the degradation can be reduced by using the mobile phase provided by the application within the retention time of the characteristic peak of pazufloxacin mesylate 7 times. Impurities are detected comprehensively and accurately.

Example 3 HPLC qualitative detection of pazufloxacin mesylate bulk drug after acid degradation

The detection steps are as follows:

(1) Prepare the test solution for acid degradation: take 30 mg of pazufloxacin mesylate crude drug, place it in a 50 ml measuring bottle, add 25 ml of 1.0 mol/L hydrochloric acid and heat it in a boiling water bath for 12 hours in the dark, and add 1.0 mol/L sodium hydroxide was diluted and neutralized to the mark as the test solution.

(2) Perform HPLC detection according to the same method as in Example 1. The chromatogram is shown in Figure 4. As can be seen from Figure 4, for the crude drug after acid degradation, the small polarity generated by the degradation can be reduced by using the mobile phase provided by the application within the retention time of the characteristic peak of pazufloxacin mesylate 7 times. Impurities are detected comprehensively and accurately.

Example 4 HPLC qualitative detection of Pazufloxacin mesylate raw material after alkali degradation

The detection steps are as follows:

(1) prepare alkali degradation need testing solution: get pazufloxacin mesylate crude drug 30mg, place in 50ml measuring bottle, add 25ml 1.0mol/L sodium hydroxide and heat in boiling water bath for 12 hours in the dark, another Add 1.0mol/L hydrochloric acid to dilute and neutralize to the mark as the test solution.

(2) Perform HPLC detection according to the same method as in Example 1. The chromatogram is shown in Figure 5.

As can be seen from Figure 5, for the bulk drug after alkali degradation, the low polarity produced by the degradation can be reduced by using the mobile phase provided by the application within the retention time of 7 times the characteristic peak of pazufloxacin mesylate. Impurities are detected comprehensively and accurately.

Example 5 Carry out HPLC quantitative detection of Pazufloxacin mesylate crude drug

The detection steps are as follows:

Get 30mg of pazufloxacin mesylate crude drug, accurately weighed, put in a 100ml measuring bottle, add mobile phase to dissolve and dilute to the scale, shake up, as the test solution;

Accurately measure 1ml of the test solution, put it in a 100ml measuring bottle, dilute to the mark with mobile phase, shake well, and use it as a contrast solution.

Take respectively 20 μl of the test solution and the control solution and inject into the liquid chromatograph, and record the chromatogram to 7 times of the retention time of the main component peak.

Measure the sum of the peak areas of each impurity whose retention time is greater than the characteristic peak of the main drug Pazufloxacin sulfonate in the chromatogram of the test product solution, and compare it with the peak area of the main drug Pazufloxacin mesilate in the chromatogram of the contrast solution , to determine the content of degradation products.

The above descriptions are only preferred embodiments of the present application, and are not intended to limit the present application. For those skilled in the art, various modifications and changes may be made to the present application. Any modifications, equivalent replacements, improvements, etc. made within the gist and principles of this application shall be included within the protection scope of this application.

Claims

A method for detecting small polar impurities in the bulk drug of pazufloxacin mesylate, characterized in that, it is detected by high performance liquid chromatography, and the mobile phase is acetonitrile-10% with a volume ratio of 45:10:7:138 Triethylamine methanesulfonate solution-1.0mol/L dipotassium hydrogen phosphate-water;

The method comprises the steps of:

With mobile phase as solvent, prepare the test solution of Pazufloxacin mesylate bulk drug;

Get described need testing solution and dilute with mobile phase, obtain contrast solution;

Get need testing solution and contrast solution inject high performance liquid chromatography respectively, record chromatogram to the retention time of 7 times of pazufloxacin mesylate characteristic peak;

The peak area of the impurity whose retention time is greater than the characteristic peak of pazufloxacin mesylate in the spectrum of the test solution obtained is compared with the characteristic peak area of pazufloxacin mesylate in the spectrum of the test solution to determine The impurity is relative to the content of pazufloxacin mesylate.
The method according to claim 1, wherein the test solution is diluted 100 times to obtain the control solution.
The method according to claim 2, wherein the concentration of pazufloxacin mesylate bulk drug in the test solution is 0.3 mg/ml.
The method according to claim 1, characterized in that the injection volume of the high performance liquid chromatograph is 20 μl.
The method according to any one of claims 1-4, wherein the chromatographic conditions of high performance liquid chromatography also include:

Stationary phase: Octadecylsilane bonded silica gel,

Flow rate: 1.0ml/min,

Detection wavelength: 254nm.
The method according to claim 1, characterized in that the high performance liquid chromatography has a resolution of 1.6.
The method according to claim 1, characterized in that the number of theoretical plates of high performance liquid chromatography should be no less than 2000 based on the characteristic peak of pazufloxacin mesylate.
The method according to any one of claims 1-6, wherein, before getting need testing solution and contrast solution to inject high performance liquid chromatograph respectively, said method also comprises:

Take 20 μl of the control solution and inject it into the liquid chromatograph, and adjust the detection sensitivity so that the peak height of the main component chromatographic peak is 10-25% of the full scale.