Method for detecting related substances in isosorbide mononitrate sodium chloride injection and application thereof
By using octadecyl-bonded silica gel chromatography column and mobile phase gradient elution method in the detection of isosorbide mononitrate sodium chloride injection, the problem of insufficient detection sensitivity and accuracy in the prior art was solved, and more efficient impurity detection and drug quality control were achieved.
Patent Information
- Application Number
- CN202411792743.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-12-07
- Publication Date
- 2025-03-28
AI Technical Summary
In the prior art, when detecting related impurities in isosorbate mononitrate sodium chloride injection, the sensitivity and accuracy cannot meet the requirements of high-quality control, especially in complex substrates, it is difficult to achieve effective separation and quantification.
The separation effect and detection sensitivity of the target compound are significantly improved by optimizing the ratio and elution time of the mobile phase.
The detection sensitivity and accuracy of isosorbate mononitrate, 2-monomerbate and isosorbate nitrate in sodium isosorbate mononitrate injection was improved, and effective control of drug quality was achieved.
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Abstract
Description
Technical Field
[0001] The present application relates to the technical field of drug analysis and detection, and in particular to a method for detecting related substances in isosorbide mononitrate sodium chloride injection and an application thereof. Background Art
[0002] Isosorbide mononitrate sodium chloride injection is a commonly used anti-angina drug and is increasingly used in clinical practice. 2-Isosorbide mononitrate and isosorbide dinitrate are the main process impurities in the production process of isosorbide mononitrate API. The presence of these impurities may affect the stability, safety and efficacy of the drug. Therefore, the content of related impurities in isosorbide mononitrate sodium chloride injection must be strictly controlled.
[0003] At present, the detection methods of related impurities in isosorbide mononitrate sodium chloride injection mainly include: high performance liquid chromatography (HPLC), thin layer chromatography (TLC) and gas chromatography (GC). Among them, HPLC is widely used due to its high sensitivity and good separation ability. However, in actual operation, due to the unreasonable selection of mobile phase and design of elution program, the sensitivity and accuracy of the detection results cannot meet the requirements of high quality control; especially in complex matrices, these methods are difficult to achieve effective separation and accurate quantification of target substances, thus affecting the quality evaluation and clinical application of drugs.
[0004] Therefore, it is urgent to develop a new detection method to improve the detection sensitivity and accuracy of related substances in isosorbide mononitrate sodium chloride injection. Summary of the invention
[0005] In order to improve the detection sensitivity and accuracy of related impurities in isosorbide mononitrate sodium chloride injection, the present application provides a detection method for related substances in isosorbide mononitrate sodium chloride injection and its application.
[0006] In the first aspect, the present application provides a method for detecting related substances in isosorbide mononitrate sodium chloride injection, using the following technical solution: A method for detecting related substances in isosorbide mononitrate sodium chloride injection comprises the following steps: Prepare a mixed reference solution: weigh 2-isosorbide mononitrate reference, isosorbide nitrate reference and isosorbide mononitrate reference respectively, and then use methanol-water as a diluent to prepare a mixed reference solution with a concentration of 0.8-1.2 μg / mL for 2-isosorbide mononitrate reference, isosorbide nitrate reference and isosorbide mononitrate reference; Prepare the test solution: Use isosorbide mononitrate sodium chloride injection with a concentration of 0.18-0.22 mg / mL of isosorbide mononitrate as the test solution; Liquid chromatography detection: the mixed reference solution and the test solution are respectively injected into the liquid chromatograph, and the chromatographic column with octadecyl bonded silica gel as the filler and the mobile phase gradient elution method are used for separation and detection; the mobile phase gradient elution program is: at 0-6 minutes, the mobile phase A is 100%, and the mobile phase B is 0%; at 6-7.5 minutes, the mobile phase A is linearly reduced to 85%, and the mobile phase B is linearly increased to 15%; at 7.5-25 minutes, the mobile phase A is 85%, and the mobile phase B is 15%; at 25-30 minutes, the mobile phase A is linearly increased to 100%, and the mobile phase B is linearly reduced to 0%; at 30-35 minutes, the mobile phase A is 100%, and the mobile phase B is 0%; the mobile phase A and the mobile phase B are both measured in volume percentage; the mobile phase A is methanol-water, and the mobile phase B is ethanol-trimethylpentane.
[0007] The present application provides a method for detecting related substances in isosorbide mononitrate sodium chloride injection. The method can effectively improve the detection sensitivity and accuracy of isosorbide mononitrate, 2-isosorbide mononitrate and isosorbide dinitrate in isosorbide mononitrate sodium chloride injection, and realize the quality control of isosorbide mononitrate sodium chloride injection. Specifically: First, the present application prepares 2-isosorbide mononitrate, isosorbide nitrate and isosorbide mononitrate reference substances into a mixed reference substance solution with consistent concentration using methanol-water as diluent, ensuring the consistency and reliability of the standard sample during the detection process; secondly, an isosorbide mononitrate sodium chloride injection with a concentration of 0.18-0.22 mg / mL of isosorbide mononitrate is used as the test solution, and its concentration is higher than the concentration range specified in the conventional pharmacopoeia, which can make the impurities in the test sample easier to detect, thereby greatly improving the sensitivity and accuracy of the detection; in addition, the present application uses a chromatographic column with octadecyl bonded silica as a filler and a specific mobile phase gradient elution method for separation and detection, and by optimizing the ratio and elution time of the mobile phase, the separation effect and detection sensitivity of each target compound are significantly improved. In particular, the gradient changes of mobile phases A and B make the retention time and peak shape of different compounds on the chromatographic column more ideal, further improving the reliability and accuracy of the detection. In summary, the present application can accurately determine the content of related substances in isosorbide mononitrate sodium chloride injection by precisely controlling the type and concentration of the mixed reference solution and the test solution and adopting the mobile phase gradient elution method for separation and detection, and the detection sensitivity is high and the reliability of the detection results is good.
[0008] Optionally, the volume ratio of methanol to water in the mobile phase A is (20-35): (65-80), and the volume ratio of ethanol to trimethylpentane in the mobile phase B is (10-25): (75-90).
[0009] In some embodiments, the volume ratio of methanol to water in the mobile phase A can be (20-25):(75-80), (20-30):(70-80), (20-35):(65-80), (25-30):(70-75), (25-35):(65-75) or (30-35):(65-70).
[0010] In a specific embodiment, the volume ratio of methanol to water in the mobile phase A can be 20:80, 25:75, 30:70 or 35:65.
[0011] In some embodiments, the volume ratio of ethanol to trimethylpentane in the mobile phase B can be (10-15):(85-90), (10-20):(80-90), (10-25):(75-90), (15-20):(80-85), (15-25):(75-85) or (20-25):(75-80).
[0012] In a specific embodiment, the volume ratio of ethanol to trimethylpentane in the mobile phase B can be 10:90, 15:85, 20:80 or 25:75.
[0013] Optionally, the volume ratio of methanol to water in the mobile phase A is (25-30): (70-75), and the volume ratio of ethanol to trimethylpentane in the mobile phase B is (15-20): (80-85).
[0014] Optionally, the related substances are isosorbide mononitrate, isosorbide dinitrate and isosorbide 2-mononitrate.
[0015] Optionally, in the liquid chromatography detection, the flow rate of the mobile phase is 0.8-1.2 mL / min.
[0016] Optionally, in the liquid chromatography detection, the injection volume is 20-25 μL.
[0017] Optionally, in the liquid chromatography detection, the detection wavelength is 210 nm.
[0018] Optionally, in the liquid chromatography detection, the column temperature is 25-30°C.
[0019] Optionally, the chromatographic column is an Agilent C18 chromatographic column with specifications of 4.6 mm×250 mm and 5 μm.
[0020] Optionally, the limit of quantification of isosorbide mononitrate is 0.0251 μg / mL, and the limit of detection is 0.0075 μg / mL; the limit of quantification of isosorbide dinitrate is 0.0502 μg / mL, and the limit of detection is 0.0151 μg / mL; the limit of quantification of 2-isosorbide mononitrate is 0.0255 μg / mL, and the limit of detection is 0.0077 μg / mL.
[0021] In the present application, the quantitative limit and detection limit of each component in isosorbide mononitrate sodium chloride injection can be as low as the above values, indicating that the detection method of related substances in isosorbide mononitrate sodium chloride injection provided in the present application has good sensitivity and accuracy, and can better achieve the quality control of isosorbide mononitrate sodium chloride injection.
[0022] In a second aspect, the present application provides an application of a method for detecting related substances in isosorbide mononitrate sodium chloride injection in the quality control of isosorbide mononitrate sodium chloride injection.
[0023] In summary, this application has the following beneficial effects: 1. The present application provides a method for detecting related substances in isosorbide mononitrate sodium chloride injection. The method adjusts the concentrations of a mixed reference solution and a test solution, and uses a chromatographic column filled with octadecyl bonded silica gel and a specific mobile phase and a gradient elution program, so that the contents of isosorbide mononitrate, isosorbide nitrate and 2-isosorbide mononitrate in isosorbide mononitrate sodium chloride injection can be quickly and accurately detected, and the detection has good accuracy, high sensitivity and good reliability, and can achieve quality control of isosorbide mononitrate sodium chloride injection.
[0024] 2. The detection method provided in this application can detect the contents of isosorbide mononitrate, isosorbide nitrate and 2-isosorbide mononitrate in isosorbide mononitrate sodium chloride injection. The quantification limit of isosorbide mononitrate is 0.0251 μg / mL, and the detection limit is 0.0075 μg / mL; the quantification limit of isosorbide nitrate is 0.0502 μg / mL, and the detection limit is 0.0151 μg / mL; the quantification limit of 2-isosorbide mononitrate is 0.0255 μg / mL, and the detection limit is 0.0077 μg / mL.
[0025] 3. The present application further controls the volume ratio of the mobile phase solution within the following range: the volume ratio of methanol-water in mobile phase A is (25-30): (70-75), and the volume ratio of ethanol-trimethylpentane in mobile phase B is (15-20): (80-85). The obtained detection method has better repeatability, and can make the RSD value of the isosorbide dinitrate content detection result ≤0.20%, and the RSD value of the 2-isosorbide mononitrate content detection result ≤0.30%. DETAILED DESCRIPTION
[0026] The present application provides a method for detecting related substances in isosorbide mononitrate sodium chloride injection, comprising the following steps: (1) Preparation of mixed reference solution: (1-1) Diluent: Measure 250 mL of methanol and 750 mL of water, mix well, and obtain a diluent.
[0027] (1-2) Isosorbide 2-mononitrate stock solution: Take about 5 mg of 2-isosorbide mononitrate reference substance, accurately weigh it, put it into a 25 mL volumetric flask, add diluent to dissolve and dilute to the scale, shake well, and obtain 0.2 mg / mL 2-isosorbide mononitrate stock solution.
[0028] (1-3) Isosorbide mononitrate stock solution: Take about 5 mg of isosorbide mononitrate reference substance, accurately weigh it, place it in a 25 mL volumetric flask, add diluent to dissolve and dilute to the scale, shake well, and obtain 0.2 mg / mL isosorbide mononitrate stock solution.
[0029] (1-4) Isosorbide dinitrate stock solution: Take about 5 mg of isosorbide dinitrate reference substance, accurately weigh it, place it in a 25 mL volumetric flask, add diluent to dissolve and dilute to the scale, shake well, and obtain 0.2 mg / mL isosorbide dinitrate stock solution.
[0030] (1-5) Accurately measure 1 mL each of 2-isosorbide mononitrate stock solution, isosorbide mononitrate stock solution, and isosorbide dinitrate stock solution, place them in a 200 mL volumetric flask, dilute to the mark with diluent, and shake well to obtain a mixed reference solution with a concentration of 1.0 μg / mL for 2-isosorbide mononitrate reference substance, isosorbide dinitrate reference substance, and isosorbide mononitrate reference substance.
[0031] (2) Preparation of test solution: Isosorbide mononitrate sodium chloride injection with a concentration of 0.12 mg / mL is used as the test solution; (3) Liquid chromatography detection: The mixed reference solution and the test solution were respectively injected into a liquid chromatograph, and the mobile phase gradient elution method was used for separation and detection. The chromatogram was recorded, and the contents of isosorbide mononitrate, isosorbide dinitrate and 2-isosorbide mononitrate were calculated by peak area according to the external standard method; wherein, the liquid chromatography detection conditions were as follows: the injection volume was 20-25 μL, the detection wavelength was 210 nm, and the flow rate of the mobile phase was 0.8-1.2 mL / min; the chromatographic column was an Agilent C18 chromatographic column with a specification of 4.6 mm×250 mm, 5 μm; The mobile phase gradient elution program is: at 0-6 minutes, the mobile phase A is 100%, and the mobile phase B is 0%; at 6-7.5 minutes, the mobile phase A is linearly reduced to 85%, and the mobile phase B is linearly increased to 15%; at 7.5-25 minutes, the mobile phase A is 85%, and the mobile phase B is 15%; at 25-30 minutes, the mobile phase A is linearly increased to 100%, and the mobile phase B is linearly reduced to 0%; at 30-35 minutes, the mobile phase A is 100%, and the mobile phase B is 0%; the mobile phase A and the mobile phase B are both measured in volume percentage; the mobile phase A is methanol-water, and the mobile phase B is ethanol-trimethylpentane; further, the volume ratio of methanol-water in the mobile phase A is (25-30): (70-75), and the volume ratio of ethanol-trimethylpentane in the mobile phase B is (15-20): (80-85).
[0032] In this application, the CAS number of isosorbide mononitrate is 16051-77-7, the batch number is 100694-201703, 99.9%, and it is derived from the China Food and Drug Inspection Institute; the CAS number of 2-isosorbide mononitrate is 16106-20-0, the batch number is 100695-201703, 100.0%, and it is derived from the China Food and Drug Inspection Institute; the CAS number of isosorbide dinitrate is 87-33-2, the batch number is 100250-201004, 99.9%, and it is derived from the China Food and Drug Inspection Institute; the specification of isosorbide mononitrate sodium chloride injection is 100mL, containing 20mg of isosorbide mononitrate and 0.9g of sodium chloride; the raw materials, reagents, solvents, etc. used in this application can all be obtained commercially.
[0033] The present application is further described in detail below in conjunction with embodiments and performance testing experiments.
[0034] Example 1 Example 1 provides a method for detecting related substances in isosorbide mononitrate sodium chloride injection, comprising the following steps: (1) Preparation of mixed reference solution: (1-1) Diluent: Measure 250 mL of methanol and 750 mL of water, mix well, and obtain a diluent.
[0035] (1-2) Isosorbide 2-mononitrate stock solution: Take about 5 mg of 2-isosorbide mononitrate reference substance, accurately weigh it, put it into a 25 mL volumetric flask, add diluent to dissolve and dilute to the scale, shake well, and obtain 0.2 mg / mL 2-isosorbide mononitrate stock solution.
[0036] (1-3) Isosorbide mononitrate stock solution: Take about 5 mg of isosorbide mononitrate reference substance, accurately weigh it, place it in a 25 mL volumetric flask, add diluent to dissolve and dilute to the scale, shake well, and obtain 0.2 mg / mL isosorbide mononitrate stock solution.
[0037] (1-4) Isosorbide dinitrate stock solution: Take about 5 mg of isosorbide dinitrate reference substance, accurately weigh it, place it in a 25 mL volumetric flask, add diluent to dissolve and dilute to the scale, shake well, and obtain 0.2 mg / mL isosorbide dinitrate stock solution.
[0038] (1-5) Accurately measure 1 mL each of 2-isosorbide mononitrate stock solution, isosorbide mononitrate stock solution, and isosorbide dinitrate stock solution, place them in a 200 mL volumetric flask, dilute to the mark with diluent, and shake well to obtain a mixed reference solution with a concentration of 1.0 μg / mL for 2-isosorbide mononitrate reference substance, isosorbide dinitrate reference substance, and isosorbide mononitrate reference substance.
[0039] (2) Preparation of test solution: Isosorbide mononitrate sodium chloride injection with a concentration of 0.2 mg / mL is used as the test solution; (3) Liquid chromatography detection: The mixed reference solution and the test solution were respectively injected into a liquid chromatograph, and the mobile phase gradient elution method was used for separation and detection. The chromatogram was recorded, and the content of isosorbide dinitrate and / or 2-isosorbide mononitrate was calculated by peak area according to the external standard method; wherein, the liquid chromatography detection conditions were as follows: the injection volume was 20 μL, the detection wavelength was 210 nm, and the flow rate of the mobile phase was 1.0 mL / min; the chromatographic column was Agilent C18 chromatographic column, specification is 4.6mm×250mm, 5μm; mobile phase gradient elution program is: 0-6min, mobile phase A is 100%, mobile phase B is 0%; 6-7.5min, mobile phase A linearly decreases to 85%, mobile phase B linearly increases to 15%; 7.5-25min, mobile phase A is 85%, mobile phase B is 15%; 25-30min, mobile phase A linearly increases to 100%, mobile phase B linearly decreases to 0%; 30-35min, mobile phase A is 100%, mobile phase B is 0%; mobile phase A and mobile phase B are both measured in volume percentage; mobile phase A is methanol-water with a volume ratio of 25:75, and mobile phase B is ethanol-trimethylpentane with a volume ratio of 20:80.
[0040] Limit of quantitation, limit of detection and repeatability of limit of quantitation (1) Limit of quantification and limit of detection: Take 1mL of 2-isosorbide mononitrate stock solution, place it in a 20mL volumetric flask, dilute it to the mark with diluent, shake well, and obtain 0.01mg / mL of 2-isosorbide mononitrate quantitative limit stock solution. Take 1mL of isosorbide mononitrate stock solution, place it in a 20mL volumetric flask, dilute it to the mark with diluent, shake well, and obtain 0.01mg / mL of isosorbide mononitrate quantitative limit stock solution. Take 1mL of isosorbide dinitrate stock solution, place it in a 20mL volumetric flask, dilute it to the mark with diluent, shake well, and obtain 0.01mg / mL of isosorbide dinitrate quantitative limit stock solution.
[0041] Accurately measure 1 mL of 2-isosorbide mononitrate quantification limit stock solution, 1 mL of isosorbide mononitrate quantification limit stock solution, and 2 mL of isosorbide dinitrate quantification limit stock solution, place them in the same 20 mL volumetric flask, dilute to the scale with diluent, shake well, and obtain a quantification limit reserve mixed solution; The quantitative limit reserve mixed solution was injected into the liquid chromatograph and tested according to the method provided in Example 1. The quantitative limit reserve mixed solution was diluted step by step according to the signal-to-noise ratio of the quantitative limit reserve mixed solution. When the peak height signal-to-noise ratio S / N ≥ 10:1, the concentration was used as the quantitative limit; when the peak height signal-to-noise ratio S / N ≥ 3:1, the concentration was used as the detection limit. The results are shown in Table 1 below; (1-2) Repeatability of Quantitation Limit Six quantitative limit reserve mixed solutions (1-6) were prepared in parallel, and the detection was carried out according to the liquid chromatography conditions of Example 1, and the RSD value of the peak area was calculated. The results are shown in Table 2 below Table 1 Test results of limit of quantitation and limit of detection Table 2 Test results of repeatability of quantitation limit The test results in Table 1 and Table 2 show that the quantitative limit concentration of 2-isosorbide mononitrate is 0.0255 μg / mL, equivalent to a limit concentration of 2.55%, and a signal-to-noise ratio of 12.6; the detection limit concentration is 0.0077 μg / mL, equivalent to a limit concentration of 0.77%, and a signal-to-noise ratio of 3.9; the quantitative limit repeatability peak area RSD (n=6) is 4.84%; the quantitative limit concentration of isosorbide mononitrate is 0.0251 μg / mL, equivalent to a limit concentration of 2.51%, and a signal-to-noise ratio of 12.2; The detection limit concentration is 0.0075 μg / mL, equivalent to a limit concentration of 0.75%, and a signal-to-noise ratio of 5.6; the quantitative limit repeatability peak area RSD (n=6) is 7.36%; the quantitative limit concentration of isosorbide dinitrate is 0.0502 μg / mL, equivalent to a limit concentration of 5.02%, and a signal-to-noise ratio of 11.3; the detection limit concentration is 0.0151 μg / mL, equivalent to a limit concentration of 1.51%, and a signal-to-noise ratio of 3.7; the quantitative limit repeatability peak area RSD (n=6) is 4.37%. Therefore, it is shown that the detection method provided by the present application can quantitatively detect 2-mononitrate, isosorbide dinitrate and isosorbide mononitrate at a limit concentration of more than 10%, and more than 5% can be effectively detected. The detection method of related substances in isosorbide mononitrate sodium chloride injection provided by the present application has high sensitivity and good repeatability.
[0042] Embodiment 2-7 Examples 2-7 respectively provide a method for detecting related substances in isosorbide mononitrate sodium chloride injection.
[0043] The difference between the above embodiment and embodiment 1 is that mobile phase A and mobile phase B are specifically shown in Table 3 below.
[0044] Table 3 Types and ratios of mobile phases in Examples 2-7 Comparative Example 1 Comparative Example 1 provides a method for detecting related substances in isosorbide mononitrate sodium chloride injection.
[0045] The difference between the comparative example and Example 1 is that the liquid chromatography adopts a mobile phase elution method for separation and detection, and the mobile phase is methanol-water with a volume ratio of 25:75.
[0046] Comparative Example 2 Comparative Example 2 provides a method for detecting related substances in isosorbide mononitrate sodium chloride injection.
[0047] The difference between the comparative example and Example 1 is that the mobile phase gradient elution program is: at 0 min, mobile phase A is 85%, mobile phase B is 15%; 0-15 min, mobile phase A is linearly reduced to 70%, mobile phase B is linearly increased to 30%; 15-20 min, mobile phase A is 70%, mobile phase B is 30%; 20-21 min, mobile phase A is linearly increased to 85%, mobile phase B is linearly reduced to 15%; 21-35 min, mobile phase A is 85%, mobile phase B is 15%; mobile phase A and mobile phase B are both measured in volume percentage; Mobile phase A was 0.1% phosphoric acid aqueous solution; mobile phase B was methanol.
[0048] Comparative Example 3 Comparative Example 3 provides a method for detecting related substances in isosorbide mononitrate sodium chloride injection.
[0049] The difference between the comparative example and Example 1 is that the mobile phase gradient elution program is: at 0-3min, mobile phase A is 100%, and mobile phase B is 0%; at 3-4min, mobile phase A is linearly reduced to 60%, and mobile phase B is linearly increased to 40%; at 4-25min, mobile phase A is 60%, and mobile phase B is 40%; at 25-26min, mobile phase A is linearly increased to 100%, and mobile phase B is linearly reduced to 0%; at 26-35min, mobile phase A is 100%, and mobile phase B is 0%; mobile phase A and mobile phase B are both measured in volume percentage; Mobile phase A was water; mobile phase B was methanol.
[0050] Performance testing A mixed reference solution and a 100% test sample spiked solution (6 portions prepared in parallel) were prepared respectively, and then the mixed reference solution and the test sample solution were injected into a liquid chromatograph, and the detection was performed according to the methods provided in Examples 1-7 and Comparative Examples 1-3, respectively. The contents of the two impurities, isosorbide dinitrate and 2-isosorbide mononitrate, in the 100% test sample spiked solution were obtained according to the chromatogram, and the average value and RSD value of the test results were calculated. The results are shown in Table 4 below.
[0051] Prepare 100% test sample spiked solution: Accurately measure 0.5 mL of 2-isosorbide mononitrate stock solution and isosorbide dinitrate stock solution, place in a 100 mL volumetric flask, dilute to the mark with the test sample solution, shake well, and obtain 100% test sample spiked solution.
[0052] Table 4 Test results obtained by the test methods provided in Examples 1-7 and Comparative Examples 1-3 According to the test results of Table 4, the detection method provided in Examples 1-7 was used to detect 100% of the test sample spiked solution, and the average content of isosorbide nitrate in the obtained test sample spiked solution was 0.520%, and the RSD value was 0.147-0.302%, and the average content of 2-isosorbide mononitrate was 0.502-0.503%, and the RSD value was 0.137-0.359%; while the detection method provided in Comparative Examples 1-3 was used to detect the test sample spiked solution, and the average content of isosorbide nitrate in the obtained test sample spiked solution was 0.482-0.503%, and the RSD value was 1.212-1.517%, and the average content of 2-isosorbide mononitrate was 0.483-0.491%, and the RSD value was 1.121-1.408%. Therefore, it is shown that the detection method for related substances in isosorbide mononitrate sodium chloride injection provided in the present application has high detection accuracy and good repeatability, and can be used to detect the content of 2-isosorbide mononitrate and isosorbide nitrate impurities in isosorbide mononitrate sodium chloride injection, thereby achieving effective quality control.
[0053] From the test results of Examples 1-7, it can be seen that the detection methods provided in Examples 1, 3 and 6 detect the sample spiked solution, and the average content of isosorbide nitrate in the obtained sample spiked solution is 0.520%, and the RSD value is 0.143-0.173% (≤0.20%), and the average content of 2-isosorbide mononitrate is 0.502-0.503%, and the RSD value is 0.137-0.213% (≤0.30%); while the detection methods provided in Examples 2, 4-5 and 7 detect the sample spiked solution, and the average content of isosorbide nitrate in the obtained sample spiked solution is 0.520%, and the RSD value is 0.258-0.302%, and the average content of 2-isosorbide mononitrate is 0.502-0.503%, and the RSD value is 0.318-0.359%. Therefore, it is explained that the present application further controls the volume ratio of methanol-water in mobile phase A within the range of (25-30):(70-75), and the volume ratio of ethanol-trimethylpentane in mobile phase B within the range of (15-20):(80-85), and the obtained detection method can achieve better separation effect between the compounds, better repeatability of the detection method, and better reliability of the detection results.
[0054] Although the present invention has been described in detail above with general descriptions and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements may be made thereto based on the present invention. Therefore, these modifications or improvements made without departing from the spirit of the present invention all fall within the scope of protection claimed by the present invention.
Claims
1. A method for detecting related substances in isosorbide mononitrate sodium chloride injection, characterized in that: The following steps are involved: Prepare a mixed reference solution: weigh 2-isosorbide mononitrate reference, isosorbide nitrate reference and isosorbide mononitrate reference respectively, and then use methanol-water as a diluent to prepare a mixed reference solution with a concentration of 0.8-1.2 μg / mL for 2-isosorbide mononitrate reference, isosorbide nitrate reference and isosorbide mononitrate reference; Prepare the test solution: Use isosorbide mononitrate sodium chloride injection with a concentration of 0.18-0.22 mg / mL of isosorbide mononitrate as the test solution; Liquid chromatography detection: The mixed reference solution and the test solution are respectively injected into a liquid chromatograph, and separation and detection are performed using a chromatographic column filled with octadecyl bonded silica gel and a mobile phase gradient elution method; the mobile phase gradient elution program is: 0-6min, mobile phase A is 100%, and mobile phase B is 0%; 6-7.5min, mobile phase A linearly decreases to 85%, and mobile phase B linearly increases to 15%; 7.5-25min, mobile phase A is 85%, and mobile phase B is 15%; 25-30min, mobile phase A linearly increases to 100%, and mobile phase B linearly decreases to 0%; 30-35min, mobile phase A is 100%, and mobile phase B is 0%; mobile phase A and mobile phase B are both measured in volume percentage; the mobile phase A is methanol-water, and the mobile phase B is ethanol-trimethylpentane.
2. The detection method according to claim 1, characterized in that: The volume ratio of methanol to water in the mobile phase A is (25-30): (70-75), and the volume ratio of ethanol to trimethylpentane in the mobile phase B is (15-20): (80-85).
3. The detection method according to claim 1, characterized in that: The related substances are isosorbide mononitrate, isosorbide dinitrate and 2-isosorbide mononitrate.
4. The detection method according to claim 1, characterized in that: In the liquid chromatography detection, the flow rate of the mobile phase is 0.8-1.2 mL / min.
5. The detection method according to claim 1, characterized in that: In the liquid chromatography detection, the injection volume is 20-25 μL.
6. The detection method according to claim 1, characterized in that: In the liquid chromatography detection, the detection wavelength is 210 nm.
7. The detection method according to claim 1, characterized in that: In the liquid chromatography detection, the column temperature is 25-30°C.
8. The detection method according to claim 1, characterized in that: The chromatographic column is an Agilent C18 chromatographic column with specifications of 4.6 mm×250 mm and 5 μm.
9. The detection method according to claim 1, characterized in that: The limit of quantification of the isosorbide mononitrate is 0.0251 μg / mL, and the limit of detection is 0.0075 μg / mL; the limit of quantification of isosorbide dinitrate is 0.0502 μg / mL, and the limit of detection is 0.0151 μg / mL; the limit of quantification of 2-isosorbide mononitrate is 0.0255 μg / mL, and the limit of detection is 0.0077 μg / mL.
10. Application of the method for detecting related substances in the isosorbide mononitrate sodium chloride injection according to any one of claims 1 to 9 in the quality control of the isosorbide mononitrate sodium chloride injection.