A method for separating and detecting moxifloxacin and its impurities

Abstract

The invention discloses a separation and detection method for moxifloxacin and its impurities, and belongs to the technical field of drug analysis and detection. The separation and detection method comprises the following steps: passing the sample to be tested into high performance liquid chromatography for separation and detection of moxifloxacin and its impurities; the high performance liquid chromatography detection conditions include: the chromatographic column is a chromatographic column with phenylsilane bonded silica gel as filler; the mobile phase includes mobile phase A and mobile phase B, mobile phase A is a buffer solution, and mobile phase B is methanol; the pH of the buffer solution is 2-3; the column temperature is 40-45°C; the mobile phase flow rate is 1.0-1.8mL / min; the injection volume is 5-15μL; the elution method is gradient elution; and the detection wavelength is 277nm. The separation and detection method of the invention has good specificity, accuracy and precision, high sensitivity, and can separate, detect and qualitatively analyze various impurities in moxifloxacin.

Description

Technical Field

[0001] The invention belongs to the technical field of drug analysis and detection, and particularly relates to a method for separating and detecting moxifloxacin and its impurities. Background Art

[0002] Moxifloxacin is a new fourth-generation serotonin drug with a broad antibacterial spectrum and good pharmacokinetic properties. In clinical applications, moxifloxacin has shown good safety and oral absorption, with an average absolute bioavailability of up to 90%. However, a variety of impurities will be generated during the production, preparation and storage of moxifloxacin hydrochloride injection; the presence of these impurities may not only affect the efficacy of the drug, but also increase the risk of adverse reactions. Therefore, it is necessary to separate and test moxifloxacin and its impurities. By testing the impurity content, it is possible to evaluate whether the quality of the drug meets the standards, thereby avoiding the use of unqualified products; secondly, impurities may cause adverse reactions to the human body. Through separation and testing, these impurities can be discovered and controlled in time to ensure the safety of patients' medication; at the same time, separation and testing can understand the source and conditions of impurity generation, which helps to optimize the production process, reduce the generation of impurities, and improve the purity and stability of the product.

[0003] High performance liquid chromatography (HPLC) is the main method for detecting and separating moxifloxacin and its impurities, but in practical applications, there are still problems such as poor specificity, poor separation effect and poor sensitivity. For some impurities with similar structures and similar properties, their molecular structures contain multiple functional groups, such as fluorine, hydroxyl, methoxy, carboxyl and complex heterocyclic structures. The interaction between these functional groups may lead to the formation of hydrogen bonds and π-π stacking between molecules. The similarity of these structures will also cause their physical and chemical properties to be similar. When HPLC is used for detection and separation, the separation efficiency is not high enough, resulting in the impurity peak overlapping with the main peak or being difficult to distinguish, which will affect the accuracy and reliability of the detection; and for impurities with extremely low content, the detection sensitivity of HPLC may be insufficient, and it is difficult to accurately determine their content, which may lead to missed detection or misjudgment. Summary of the invention

[0004] The object of the present invention is to provide a separation and detection method for moxifloxacin and its impurities, which has good specificity, accuracy and precision, high sensitivity, good linear relationship within the impurity limit concentration range of 0.05% to 0.4%, can realize single detection of moxifloxacin and its impurities, has low cost and high efficiency.

[0005] The technical solution adopted to achieve the above-mentioned purpose is to provide a method for separating and detecting moxifloxacin and its impurities, comprising the following steps:

[0006] Passing the sample to be tested into high performance liquid chromatography to separate and detect moxifloxacin and its impurities;

[0007] The high performance liquid chromatography detection conditions include: the chromatographic column is a chromatographic column filled with phenylsilane bonded silica gel; the mobile phase includes mobile phase A and mobile phase B, mobile phase A is a buffer solution, and mobile phase B is methanol; the pH of the buffer solution is 2-3; the column temperature is 40-45°C; the mobile phase flow rate is 1.0-1.8 mL / min; the injection volume is 5-15 μL; the elution method is gradient elution; the detection wavelength is 277 nm;

[0008] The impurities are impurity A, impurity B, impurity C, impurity D, impurity E, impurity F, impurity ACB-8, impurity ACB-9, impurity ACB-10, impurity I, impurity J, impurity K and impurity L. The specific structural formula is as follows:

[0009]

[0010]

[0011]

[0012] ;

[0013] The gradient elution program was:

[0014] .

[0015] Preferably, the pH of the buffer solution is 2.45-2.55; the column temperature is 45° C.; the mobile phase flow rate is 1.3 mL / min; and the injection volume is 10 μL.

[0016] Preferably, the buffer solution contains tetrabutylammonium hydrogen sulfate, potassium dihydrogen phosphate, phosphoric acid and water.

[0017] More preferably, the method for preparing the buffer solution comprises the following steps: dissolving tetrabutylammonium hydrogen sulfate and potassium dihydrogen phosphate in water, and adjusting the pH with phosphoric acid.

[0018] More preferably, the solid-liquid ratio of tetrabutylammonium hydrogen sulfate, potassium dihydrogen phosphate and water is 0.5g:1g:1L.

[0019] Preferably, the chromatographic column model is GL Inertsil Phenyl, with a size of 250 mm×4.6 mm and 5 μm.

[0020] The present invention has the following beneficial effects:

[0021] The method for separating and detecting moxifloxacin and its impurities provided by the present invention can achieve effective separation between the main component and its impurities, has good specificity, accuracy and precision, high sensitivity, good linear relationship within the impurity limit concentration range of 0.05% to 0.4%, can achieve single detection of moxifloxacin and its impurities, has low cost and high efficiency. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 is the HPLC separation chromatogram of Example 1;

[0023] Figure 2 It is the HPLC separation chromatogram of Comparative Example 1. DETAILED DESCRIPTION

[0024] The features and performance of the present invention are further described in detail below in conjunction with embodiments.

[0025] Example 1

[0026] A method for separating and detecting moxifloxacin and its impurities comprises the following steps:

[0027] (1) Weigh 0.5 g of tetrabutylammonium hydrogen sulfate and 1.0 g of potassium dihydrogen phosphate, add them into 1000 mL of water to dissolve, and adjust the pH to 2.5±0.05 with phosphoric acid to obtain a diluent;

[0028] (2) Take moxifloxacin hydrochloride eye drops as the sample to be tested, and quantitatively dilute it with a diluent to a solution containing 1 mg of moxifloxacin per 1 mL, which is the test solution;

[0029] (3) Take an appropriate amount of moxifloxacin hydrochloride reference substance, weigh it accurately, add diluent to dissolve it and quantitatively dilute it to make a solution containing 2 μg of moxifloxacin per 1 mL, which is the reference substance solution;

[0030] (4) Accurately measure an appropriate amount of the reference solution and quantitatively dilute it with a diluent to produce a solution containing 0.5 μg of moxifloxacin per 1 mL, which is the sensitivity solution;

[0031] (5) Take appropriate amounts of moxifloxacin hydrochloride and its impurities (the name, structural formula and source information of the impurities are shown in Table 1; the information of the moxifloxacin hydrochloride reference substance and each impurity reference substance is shown in Table 2), add diluent to dissolve and dilute to a solution containing approximately 1.0 mg of moxifloxacin and 1.0 μg of each impurity per 1 mL, as the separation solution;

[0032] Table 1 Moxifloxacin impurity information

[0033]

[0034]

[0035] Table 2 Summary of reference substance information

[0036]

[0037] Note: TRC is the abbreviation of Toronto Research Chemicals of Canada.

[0038] (6) The HPLC detection conditions include: the chromatographic column is a chromatographic column with phenylsilane bonded silica gel as the filler; the chromatographic column model is GL Inertsil Phenyl, the size is 250 mm × 4.6 mm, 5 μm; the mobile phase includes mobile phase A and mobile phase B, mobile phase A is a buffer solution, and mobile phase B is methanol; the column temperature is 45°C; the mobile phase flow rate is 1.3 mL / min; the injection volume is 10 μL; the elution method is gradient elution, and the gradient elution program is shown in Table 3; the detection wavelength is 277 nm;

[0039] The buffer solution contains tetrabutylammonium hydrogen sulfate, potassium dihydrogen phosphate, phosphoric acid and water. The preparation method comprises the following steps: weighing 0.5 g of tetrabutylammonium hydrogen sulfate and 1.0 g of potassium dihydrogen phosphate, adding them into 1000 mL of water to dissolve, adjusting the pH to 2.5±0.05 with phosphoric acid, and setting aside;

[0040] Table 3 Gradient elution program

[0041]

[0042] (7) Accurately measure 10 μL of the resolution solution, inject it into the HPLC and record the resolution chromatogram; accurately measure 10 μL of the sensitivity solution, inject it into the HPLC and adjust the detection sensitivity; accurately measure 10 μL of the test solution and the control solution respectively, inject them into the HPLC and record the chromatogram;

[0043] In the separation solution chromatogram, impurity K, impurity J, impurity ACB-9, impurity I, impurity F, main peak of moxifloxacin, impurity A, impurity B, impurity C, impurity D, impurity E, impurity L, impurity ACB-10, and impurity ACB-8 appear in sequence. The results are as follows Figure 1 As shown; the separation degree of impurity I and moxifloxacin is not less than 1.5; in the sensitivity solution chromatogram, the signal-to-noise ratio of the main component chromatographic peak height is not less than 8;

[0044] (8) If there are chromatographic peaks in the chromatogram of the test solution that coincide with the retention times of impurities K and L, the correction factor shall be calculated by the principal component external standard method and shall not exceed 0.2%. The correction factors and limits are shown in Table 4.

[0045] Table 4 Component correction factors and limits

[0046]

[0047] In summary, Figure 1 It can be seen that the separation and detection method of moxifloxacin and its impurities of the present invention has good specificity, accuracy and precision, high sensitivity, good linear relationship within the impurity limit concentration range of 0.05% to 0.4%, and is applicable to the separation and detection of moxifloxacin and all its impurities.

[0048] Comparative Example 1

[0049] A method for separating and detecting moxifloxacin and its impurities, compared with Example 1, differs in that the detection wavelength is 293 nm, the gradient elution program is as shown in Table 5; and the remaining steps are the same as those in Example 1.

[0050] Table 5 Gradient elution program

[0051]

[0052] In this comparative example, the peak results of moxifloxacin and its impurities are as follows Figure 2 As shown. Figure 2 It can be seen that the impurity K in the separation and detection method of moxifloxacin and its impurities in this comparative example cannot achieve good separation.

[0053] The present invention has been described according to the above embodiments. It should be understood that the above embodiments do not limit the present invention in any form. Any technical solutions obtained by equivalent replacement or equivalent transformation shall fall within the scope of the present invention.

Claims

1. A method for separating and detecting moxifloxacin and its impurities, characterized in that: The following steps are involved: Passing the sample to be tested into high performance liquid chromatography to separate and detect moxifloxacin and its impurities; The high performance liquid chromatography detection conditions include: a chromatographic column with phenylsilane bonded silica gel as filler; the mobile phase includes mobile phase A and mobile phase B, mobile phase A is a buffer solution, and mobile phase B is methanol; the pH of the buffer solution is 2-3; the column temperature is 45°C; the mobile phase flow rate is 1.0-1.8 mL / min; the injection volume is 5-15 μL; the elution method is gradient elution; the detection wavelength is 277 nm; The buffer solution contains tetrabutylammonium hydrogen sulfate, potassium dihydrogen phosphate, phosphoric acid and water; the preparation method of the buffer solution comprises the following steps: taking tetrabutylammonium hydrogen sulfate and potassium dihydrogen phosphate and dissolving them in water, and adjusting the pH with phosphoric acid; the material-liquid ratio of the tetrabutylammonium hydrogen sulfate, potassium dihydrogen phosphate and water is 0.5g:1g:1L; The impurities are impurity A, impurity B, impurity C, impurity D, impurity E, impurity F, impurity ACB-8, impurity ACB-9, impurity ACB-10, impurity I, impurity J, impurity K and impurity L, and the specific structural formula is as follows: ； The gradient elution procedure is: 。 2. The method for separating and detecting moxifloxacin and its impurities according to claim 1, characterized in that: The pH of the buffer solution is 2.45-2.55; the flow rate of the mobile phase is 1.3 mL / min; and the injection volume is 10 μL.

3. The method for separating and detecting moxifloxacin and its impurities according to claim 1, characterized in that: The chromatographic column model is GL Inertsil Phenyl, with dimensions of 250 mm×4.6 mm and 5 μm.

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