Method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry

Abstract

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Description

Technical Field

[0001] The invention relates to the technical field of biomedical detection, and in particular to a method for detecting related substances in famotidine injection by using liquid chromatography-mass spectrometry. Background Art

[0002] Ranitidine may contain the mutagenic impurity N-nitrosodimethylamine (hereinafter referred to as NDMA). On April 1, 2020, the FDA issued an announcement requiring pharmaceutical companies to immediately withdraw all ranitidine drugs. Famotidine has similar genotoxic and carcinogenic warning groups in its chemical structure as ranitidine. Therefore, it is necessary to study famotidine injection for its potential nitrosamine impurities. Summary of the invention

[0003] In order to solve the above technical problems, the present invention provides a method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry, which has low detection limit and high detection sensitivity.

[0004] According to a method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to a specific embodiment of the present invention, the famotidine injection sample to be tested is detected by liquid chromatography-mass spectrometry; the parameters of the liquid chromatography are: the chromatographic column is C18; the mobile phase A is 0.05%-0.15% formic acid-water solution, and the mobile phase B is methanol; the column temperature is 35-45°C; gradient elution; mass spectrometry ion source parameters:

[0005] First mass spectrometry condition

[0006] Scan Type MRM Ionized form APCI polarity Positive NC 4.0μA Collision gas pressure 8psi Atomizing gas pressure 35psi Auxiliary heating gas pressure 50psi Air curtain pressure 30psi Drying gas temperature 300℃ .

[0007] The present invention screens and optimizes relevant test conditions of liquid chromatography and mass spectrometry in gas chromatography-mass spectrometry, and provides a detection method with the advantages of simple operation, low detection limit, high sensitivity, high repeatability, high accuracy, etc.

[0008] According to some embodiments of the present invention, the liquid chromatography conditions are that mobile phase A is 0.1% formic acid-water solution, mobile phase B is methanol, column temperature is 40° C., and / or the chromatographic column of the liquid chromatography conditions is Poroshell 120EC-C18.

[0009] According to some embodiments of the present invention, the concentration of the sample to be tested of the famotidine injection is 8-12 mg / ml, preferably 10 mg / ml.

[0010] According to some embodiments of the present invention, the related substances and their structures are as follows:

[0011] The structures of the first group of related substances

[0012]

[0013]

[0014] According to some embodiments of the present invention, the detection limit of the related substances is 4.5 ng / ml;

[0015] And / or, the limit of quantification of the related substances is 0.90 ng / ml.

[0016] According to some embodiments of the present invention, the injection volume of the liquid chromatography is 18-22μ; the gradient elution conditions include the following:

[0017] First gradient elution table

[0018] T(min) Flow rate (ml / min) A(％) B(％) 0 0.35-0.45 90 10 1.0 0.35-0.45 90 10 6.5 0.35-0.45 40 60 9.0 0.35-0.45 40 60 10.0 0.35-0.45 5 95 14.0 0.35-0.45 5 95 14.1 0.35-0.45 90 10 17.0 0.35-0.45 90 10 ;

[0019] The mass spectrometer ion source parameters also include:

[0020] First valve switching procedure

[0021]

[0022] The scanning parameters are as follows:

[0023] First MRM scan parameters

[0024]

[0025]

[0026] Note: * indicates quantitative ion.

[0027] According to some embodiments of the present invention, the flow rate of the sample to be tested of the famotidine injection is 0.4 ml / min.

[0028] According to some embodiments of the present invention, the related substances and their structures are as follows:

[0029] The second group of related material structures

[0030]

[0031]

[0032] According to some embodiments of the present invention, the detection limits of the related substances are as follows: the detection limit of NDIPA is 0.66 ng / ml, the detection limit of NIPEA is 0.66 ng / ml, the detection limit of NMBA is 2.4 ng / ml, and the detection limit of NMPA is 0.66 ng / ml; and / or, the quantification limits of the related substances are as follows: the quantification limit of NDIPA is 1.32 ng / ml, the quantification limit of NIPEA is 1.33 ng / ml, the quantification limit of NMBA is 4.80 ng / ml, and the quantification limit of NMPA is 1.33 ng / ml.

[0033] According to some embodiments of the present invention, the injection volume of the liquid chromatography is 48-52 μ; and the gradient elution conditions include the following:

[0034] Second gradient elution table

[0035] T(min) Flow rate (ml / min) A(％) B(％) 0 0.30 90 10 1.0 0.30 90 10 6.5 0.30 40 60 9.0 0.30 40 60 10.0 0.30 5 95 11.5 0.30 5 95 11.6 0.30 90 10 14.0 0.30 90 10 ;

[0036] The mass spectrometer ion source parameters also include:

[0037] Second valve switching procedure

[0038]

[0039] The scanning parameters are as follows:

[0040] Second MRM scan parameters

[0041]

[0042]

[0043] Note: * indicates quantitative ion. DETAILED DESCRIPTION

[0044] The present invention is further described in detail below in conjunction with examples, comparative examples and performance testing experiments. These examples should not be construed as limiting the scope of protection claimed by the present invention.

[0045] According to the method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to a specific embodiment of the present invention, liquid chromatography-mass spectrometry is used to detect the sample to be tested of famotidine injection; so set, liquid chromatography is used as a separation system, and mass spectrometry is used as a detection system. The sample is separated from the mobile phase in the mass spectrum part, and after being ionized, the mass analyzer of the mass spectrometer separates the ion fragments by mass number, and a mass spectrum is obtained through a detector. Liquid chromatography-mass spectrometry embodies the complementarity of the advantages of chromatography and mass spectrometry, combines the high separation ability of chromatography to complex samples with the advantages of MS having high selectivity, high sensitivity and being able to provide relative molecular mass and structural information, and can detect a very small amount of nitrosamine impurities in famotidine injection, thereby improving the detection sensitivity and accuracy of the analytical method. The test result is reliable, and then the quality and safety of famotidine injection are guaranteed.

[0046] The parameters of the liquid chromatography are: the chromatographic column is C18; the mobile phase A is 0.05%-0.15% formic acid-water solution, and the mobile phase B is methanol; the column temperature is 35-45°C; and the gradient elution is performed. In this way, a C18 chromatographic column is used, 0.05%-0.15% formic acid-water solution is used as the mobile phase A, and methanol is used as the mobile phase B. The column temperature is set to 35-45°C, and the separation of related substances in famotidine injection is achieved through gradient elution.

[0047] Mass spectrometer ion source parameters:

[0048] First mass spectrometry condition

[0049] Scan Type MRM Ionized form APCI polarity Positive NC 4.0μA Collision gas pressure 8psi Atomizing gas pressure 35psi Auxiliary heating gas pressure 50psi Air curtain pressure 30psi Drying gas temperature 300℃

[0050] By setting the conditions for mass spectrometry, suitable fragment peaks are obtained.

[0051] According to the detection method of a specific embodiment of the present invention, by adopting the method of liquid chromatography-mass spectrometry, liquid chromatography is used as a separation system, and mass spectrometry is used as a detection system. The sample is separated from the mobile phase in the mass spectrum part, and after being ionized, the mass analyzer of the mass spectrum separates the ion fragments by mass number, and a mass spectrum is obtained through a detector. Liquid chromatography-mass spectrometry embodies the complementarity of the advantages of chromatography and mass spectrometry, and the high separation ability of chromatography to complex samples is combined with the advantages of MS having high selectivity, high sensitivity and being able to provide relative molecular mass and structural information, and a small amount of nitrosamine impurities in famotidine injection can be detected, thereby the detection sensitivity and accuracy of the analytical method can be improved. The test result is reliable, and then the quality and safety of famotidine injection are guaranteed.

[0052] The present invention screens and optimizes relevant test conditions of liquid chromatography and mass spectrometry in gas chromatography-mass spectrometry, and provides a detection method with the advantages of low detection limit, high precision, high repeatability and high accuracy.

[0053] According to some embodiments of the present invention, the liquid chromatography conditions are as follows: mobile phase A is 0.1% formic acid-water solution, mobile phase B is methanol, column temperature is 40° C., and the chromatographic column of the liquid chromatography conditions is Poroshell 120EC-C18. With such settings, the parameters of the chromatographic column are better and a better separation effect can be obtained.

[0054] According to some embodiments of the present invention, the concentration of the sample to be tested of famotidine injection is 8-12 mg / ml, preferably 10 mg / ml. In this way, the concentration of the sample to be tested of famotidine injection in the method of the present invention is appropriate, which is conducive to obtaining better detection results.

[0055] According to some embodiments of the present invention, the relevant substances and their structures are as follows:

[0056] The structures of the first group of related substances

[0057]

[0058]

[0059] With such arrangement, the method of the present invention can detect the above nine impurities in famotidine injection, and the separation effect is good.

[0060] According to some embodiments of the present invention, the detection limit of the related substances is 4.5 ng / ml. With such settings, the detection limit of the detection method of the present invention is low and the detection sensitivity is high.

[0061] According to other embodiments of the present invention, the quantitative limit of the related substances is 0.90 ng / ml. This setting further limits the quantitative limit of the related substances and ensures the accuracy of the detection method of the present invention.

[0062] According to some embodiments of the present invention, the injection volume of the liquid chromatography is 18-22 μ. With such a configuration, the injection volume of the liquid chromatography is appropriate.

[0063] Gradient elution conditions included the following:

[0064] First gradient elution table

[0065]

[0066]

[0067] The mass spectrometer ion source parameters also include:

[0068] First valve switching procedure

[0069]

[0070] The scanning parameters are as follows:

[0071] First MRM scan parameters

[0072]

[0073] Note: * indicates quantitative ion.

[0074] With such configuration, the detection method of the present invention provides clear gradient elution conditions, spectral ion source parameters and scanning parameters, so the detection result is more accurate.

[0075] According to some embodiments of the present invention, the flow rate of the sample to be tested of famotidine injection is 0.4 ml / min. With such a setting, the flow rate of the sample to be tested is more suitable and the test result is more accurate.

[0076] According to some embodiments of the present invention, the relevant substances and their structures are as follows:

[0077] The second group of related material structures

[0078]

[0079] With such a configuration, the method of the present invention can be used to detect the above nine impurities in famotidine injection, and the separation effect is good.

[0080] According to some embodiments of the present invention, the detection limits of the related substances are as follows: the detection limit of NDIPA is 0.66 ng / ml, the detection limit of NIPEA is 0.66 ng / ml, the detection limit of NMBA is 2.4 ng / ml, and the detection limit of NMPA is 0.66 ng / ml. In this way, the detection limit of the detection method of the present invention is low and the detection sensitivity is high.

[0081] The quantitative limits of the related substances are as follows: the quantitative limit of NDIPA is 1.32 ng / ml, the quantitative limit of NIPEA is 1.33 ng / ml, the quantitative limit of NMBA is 4.80 ng / ml, and the quantitative limit of NMPA is 1.33 ng / ml. In this way, the quantitative limits of the related substances are further limited to ensure the accuracy of the detection method of the present invention.

[0082] According to some embodiments of the present invention, the injection volume of the liquid chromatography is 48-52 μ. With such a configuration, the injection volume of the liquid chromatography is appropriate.

[0083] Gradient elution conditions included the following:

[0084] Second gradient elution table

[0085]

[0086]

[0087] The mass spectrometer ion source parameters also include:

[0088] Second valve switching procedure

[0089]

[0090] The scanning parameters are as follows:

[0091] Second MRM scan parameters

[0092]

[0093] Note: * indicates quantitative ion.

[0094] With such configuration, the detection method of the present invention provides clear gradient elution conditions, spectral ion source parameters and scanning parameters, so the detection result is more accurate.

[0095] Refer to FDA literature (Control of Nitrosamine Impurities in Human Drugs Guidance for Industry) [1] With EMA literature (Assessment report Procedure underArticle 5(3)ofRegulation EC(No)7262004Nitrosamine impurities in human medicinal products) [2] It can be seen from references such as , that there is no clear acceptable intake for the nine nitrosamine impurities, including NMOR, NPYR, NDEA, NPIP, NMPHA, NDPA, NEPHA, NDBA, and NDBZA. The European Medicines Agency (EMA) set a temporary daily allowable intake (ADI) value of 18 ng / day for NDSRI, which was later adjusted to 178 ng / day, valid for 12 months until a more accurate ADI value is obtained. The U.S. Food and Drug Administration (FDA) also set a temporary ADI value of 26.5 ng / day for NDSRI. These temporary ADI values ​​are not used for more than 12 months to provide regulators with sufficient time to evaluate the long-term effects of these impurities. The acceptable intakes for NMPA, NDIPA, and NIPEA are all 26.5 ng / day; the acceptable intake for NMBA is 96 ng / day.

[0096] In order to ensure the quality and safety of famotidine injection, combined with the maximum daily dose of famotidine injection of 40 mg / day, the acceptable intake of 9 nitrosamine impurities, including NMOR, NPYR, NDEA, NPIP, NMPHA, NDPA, NEPHA, NDBA, and NDBZA, was set at 18 ng / day, and the limit standard was confirmed to be 0.45 ppm. Based on this limit, the LC-MS method was used to establish a detection method for 9 nitrosamine impurities, including NMOR, NPYR, NDEA, NPIP, NMPHA, NDPA, NEPHA, NDBA, and NDBZA, in famotidine injection, and the system applicability, specificity, linearity and range, detection limit, quantification limit, accuracy, precision, stability, and durability of the method were verified, confirming that the method is suitable for determining the content of 9 nitrosamine impurities, including NMOR, NPYR, NDEA, NPIP, NMPHA, NDPA, NEPHA, NDBA, and NDBZA, in famotidine injection. Under the final selected conditions, the system suitability, specificity, linearity and range, detection limit, quantification limit, accuracy, stability and durability of the method were verified, confirming that the method is suitable for the determination of N-nitrosodimethylamine content in famotidine raw materials and injection. Referring to the maximum daily dose of famotidine injection of 40mg / day, the limit standards of NMPA, NDIPA and NIPEA were confirmed to be 0.6625ppm; the limit standard of NDMA was 2.4ppm. Based on this limit, the LC-MS method was used to establish the detection method of NMPA, NMBA, NDIPA and NIPEA in famotidine injection, and the system suitability, specificity, linearity and range, detection limit, quantification limit, accuracy, precision, stability and durability of the method were verified, confirming that the method is suitable for the determination of NMPA, NMBA, NDIPA and NIPEA content in famotidine injection.

[0097] Specific Exemplary Embodiments The detection method of the present invention is explained below by means of specific exemplary embodiments.

[0098] Example 1

[0099] A method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry, wherein the famotidine injection sample to be tested is detected by liquid chromatography-mass spectrometry;

[0100] 2.1 Liquid chromatography conditions:

[0101] Chromatographic column: Agilent Poroshell 120EC-C18 (2.1×150mm, 2.7μm);

[0102] Mobile phase A: 0.1% formic acid-water solution; Mobile phase B: methanol;

[0103] Column temperature: 40°C; Injection volume: 50 μl.

[0104] The gradient elution conditions were as follows:

[0105] Table 1. Gradient elution table

[0106] T(min) Flow rate (ml / min) A(％) B(％) 0 0.30 90 10 1.0 0.30 90 10 6.5 0.30 40 60 9.0 0.30 40 60 10.0 0.30 5 95 11.5 0.30 5 95 11.6 0.30 90 10 14.0 0.30 90 10

[0107] 2.2 Mass spectrometry ion source parameters:

[0108] Table 2. Mass spectrometry conditions

[0109]

[0110] 2.3 Scan parameters:

[0111] Table 3. MRM scan parameters

[0112]

[0113] Note: * indicates quantitative ion.

[0114] 3. Solution preparation, calculation formula and acceptance criteria:

[0115] 3.1 Solution preparation

[0116] 3.1.1. Blank solution: 10% methanol.

[0117] 3.1.2 Test solution: Take 1 ml of famotidine injection and put it into a liquid phase vial to obtain a concentration of 10 mg / ml.

[0118] 3.1.3. Reference substance stock solution

[0119] 3.1.3.1. Reference stock solution I: Accurately weigh appropriate amounts of the four nitrosamine compounds and place them in 10 ml volumetric flasks. Dilute to the mark with 10% methanol and shake well to obtain the reference stock solutions I of the four nitrosamine compounds.

[0120] Table 4. Reference substance stock solution I

[0121]

[0122] 3.1.3.2. Reference stock solution II: Accurately measure appropriate amount of each reference stock solution I into different 20 ml volumetric bottles, dilute to the scale with 10% methanol, shake well, and obtain the four nitrosamine compound reference stock solutions II.

[0123] Table 5. Reference substance stock solution II

[0124] Source solution name Measured volume (μl) Dilution volume (ml) Concentration (ng / ml) Preparation solution name NMPA stock solution Ⅰ 20 20 1088.304 NMPA stock solution Ⅱ NMBA stock solution Ⅰ 20 20 1089.000 NMBA stock solution Ⅱ NDIPA stock solution Ⅰ 20 20 1141.712 NDIPA stock solution Ⅱ NIPEA stock solution Ⅰ 20 20 1065.312 NIPEA stock solution Ⅱ

[0125] 3.1.3.3. Intermediate concentration stock solution: Accurately measure an appropriate amount of each reference substance stock solution II into the same 20 ml volumetric flask, dilute to the mark with 10% methanol, and shake well to obtain the 400% limit concentration stock solution.

[0126] Table 6. Intermediate concentration stock solutions

[0127]

[0128] 3.1.4 Detection limit solution: Accurately measure 0.25 ml of 400% limit concentration solution into the same 10 ml volumetric flask, dilute to the mark with 10% methanol, and shake well to obtain the detection limit solution. See Table 7 for details.

[0129] Table 7. Detection limit solutions

[0130]

[0131] 3.1.5 Standard curve solution: Accurately measure appropriate amounts of 400% limit concentration solution into different volumetric bottles, dilute to the scale with 10% methanol, and shake well to obtain a series of standard curve solutions of different concentrations. See Table 8 to Table 9 for details.

[0132] Table 8. Standard curve solutions

[0133] Solution name Transfer 400% limit concentration volume (ml) Fixed volume (ml) STD1(LOQ) 0.50 10 STD2 1.25 10 STD3 2.50 10 STD4 3.75 10 STD5 5.00 10

[0134] Table 9. Standard curve solution concentrations

[0135]

[0136] 3.1.6 System suitability solution: Take the "100% limit concentration reference solution (STD3 solution)" under "3.1.5" and perform analysis 6 times in succession.

[0137] 3.1.7 Detection limit and quantification limit solutions

[0138] Detection limit solution: Take the "LOD solution" under "3.1.4" and inject it three times continuously for analysis;

[0139] Limit of quantitation solution: Take the “LOQ solution” under “3.1.5” and inject it 6 times continuously for analysis.

[0140] 3.1.8. Specific solution

[0141] Blank solution: Take the “blank solution” under “3.1.1”;

[0142] Test solution: Take the “test solution” under “3.1.2”;

[0143] Reference substance positioning solution: Accurately measure appropriate amount of reference substance stock solution II into different 10ml volumetric bottles, make up to the blank solution volume to the mark, and shake well.

[0144] Table 10. Reference solution

[0145]

[0146] 100% limit concentration mixed control solution: Take the "100% limit concentration reference solution (STD3 solution)" under "3.1.5".

[0147] The above solutions were injected once for analysis.

[0148] 3.1.9 Stable solution

[0149] Test solution: Take the “test solution” under “3.1.2”;

[0150] 100% limit concentration reference solution: Take the "100% limit concentration reference solution (STD3 solution)" under "3.1.5";

[0151] The above solutions were placed at room temperature for different time periods and injected once for analysis.

[0152] 3.2 Calculation formula

[0153] 3.2.1 Concentration calculation

[0154] C 供试品 =(yb) / k

[0155] C 供试品 : Concentration of target compound in solution, ng / ml (the result can be directly calculated by the instrument software workstation);

[0156] y: response value of target compound in solution;

[0157] b: intercept of standard curve;

[0158] k: slope of the standard curve.

[0159] 3.2.2 Calculation of content

[0160]

[0161] X 含量 : The content of target compound in the test sample, ppm;

[0162] Cx: detection concentration of target compound in the test sample, ng / ml;

[0163] Specification: 20mg / 2ml.

[0164] 3.3 Acceptance criteria

[0165] According to the standard curve method, NDIPA shall not exceed 0.6625ppm, NIPEA shall not exceed 0.6625ppm, NMBA shall not exceed 2.4ppm, and NMPA shall not exceed 0.6625ppm.

[0166] 4. Verification results:

[0167]

[0168]

[0169]

[0170] Example 2

[0171] Example 2 provides a method for detecting related substances in famotidine injection using liquid chromatography-mass spectrometry.

[0172] The difference between Example 2 and Example 1 is that:

[0173] The parameters of the liquid chromatography were as follows: mobile phase A was 0.05% formic acid-water solution, and mobile phase B was methanol;

[0174] The column temperature is 35° C.; the concentration of the sample to be tested of famotidine injection is 8 mg / ml; and the injection volume of the liquid chromatography is 18 μ.

[0175] Specificity test results: same as in Example 1.

[0176] Example 3

[0177] Example 3 provides a method for detecting related substances in famotidine injection using liquid chromatography-mass spectrometry.

[0178] The difference between Example 3 and Example 1 is that:

[0179] The parameters of the liquid chromatography were as follows: mobile phase A was 0.15% formic acid-water solution, and mobile phase B was methanol;

[0180] The column temperature is 45°C; the concentration of the famotidine injection sample to be tested is 12 mg / ml; and the injection volume of the liquid chromatography is 22 μ.

[0181] Specificity test results:

[0182] The results showed that it was the same as in Example 1.

[0183] Example 4

[0184] A method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry, wherein the famotidine injection sample to be tested is detected by liquid chromatography-mass spectrometry;

[0185] Chromatographic column: Agilent Poroshell 120EC-C18 (2.1×150mm, 2.7μm);

[0186] Mobile phase A: 0.1% formic acid-water solution; Mobile phase B: methanol;

[0187] Column temperature: 40°C; Injection volume: 20 μl.

[0188] The gradient elution conditions were as follows:

[0189] Table 1. Gradient elution table

[0190] T(min) Flow rate (ml / min) A(％) B(％) 0 0.40 90 10 1.0 0.40 90 10 6.5 0.40 40 60 9.0 0.40 40 60 10.0 0.40 5 95 14.0 0.40 5 95 14.1 0.40 90 10 17.0 0.40 90 10

[0191] 2.2 Mass spectrometry ion source parameters:

[0192] Table 2. Mass spectrometry conditions

[0193]

[0194] 2.3 Scan parameters:

[0195] Table 3. MRM scan parameters

[0196]

[0197]

[0198] Note: * indicates quantitative ion.

[0199] 3. Solution preparation, calculation formula and acceptance criteria:

[0200] 3.1 Solution preparation

[0201] 3.1.1. Blank solution: 10% methanol.

[0202] 3.1.2 Test solution: Take 1 ml of famotidine injection and put it into a liquid phase vial to obtain a concentration of 10 mg / ml.

[0203] 3.1.3. Reference substance stock solution

[0204] 3.1.3.1. Reference substance stock solution I: Accurately measure appropriate amounts of the 12 nitrosamine mixed standards and place them in a 10 ml volumetric flask. Dilute to the mark with 10% methanol and shake well to obtain reference substance mixed stock solution I.

[0205] Table 4. Reference substance stock solution I

[0206]

[0207] 3.1.3.2. Reference substance stock solution II: Accurately measure an appropriate amount of reference substance mixed stock solution I into a 10 ml volumetric flask, dilute to the mark with 10% methanol, and shake well to obtain reference substance mixed stock solution II.

[0208] Table 5. Reference substance stock solution II

[0209]

[0210]

[0211] 3.1.4. Detection limit solution: Accurately measure 0.045 ml of reference substance mixed stock solution II into a 10 ml volumetric flask, dilute to the mark with 10% methanol, and shake well to obtain the detection limit solution.

[0212] Table 6. Detection limit solutions

[0213]

[0214] 3.1.5 Standard curve solution: Accurately measure appropriate amounts of reference substance mixed stock solution II into different volumetric bottles, dilute to the mark with 10% methanol, and shake well to obtain a series of standard curve solutions of different concentrations.

[0215] Table 7. Standard curve solutions

[0216] Solution name Volume of reference substance mixed stock solution II (ml) Fixed volume (ml) STD1(LOQ) 0.090 10.0 STD2 0.225 10.0 STD3 0.450 10.0 STD4 0.675 10.0 STD5 0.900 10.0

[0217] Table 8. Standard curve solution concentration-1

[0218]

[0219] Table 9. Standard Curve Solution Concentration-2

[0220]

[0221]

[0222] 3.1.6 System suitability solution: Take the "100% limit concentration reference solution (STD3 solution)" under "3.1.5" and perform analysis 6 times in succession.

[0223] 3.1.7 Detection limit and quantification limit solutions

[0224] Detection limit solution: Take the "LOD solution" under "3.1.4" and inject it three times continuously for analysis;

[0225] Limit of quantitation solution: Take the “LOQ solution” under “3.1.5” and inject it 6 times continuously for analysis.

[0226] 3.1.8. Specific solution

[0227] Blank solution: Take the “blank solution” under “3.1.1”;

[0228] Test solution: Take the “test solution” under “3.1.2”;

[0229] 100% limit concentration mixed control solution: Take the "100% limit concentration reference solution (STD3 solution)" under "3.1.5".

[0230] The above solutions were injected once for analysis.

[0231] 3.1.9 Stable solution

[0232] Test solution: Take the “test solution” under “3.1.2”;

[0233] 100% limit concentration reference solution: Take the "100% limit concentration reference solution (STD3 solution)" under "3.1.5";

[0234] The above solutions were placed at room temperature for different time periods and injected once for analysis.

[0235] 3.2 Calculation formula

[0236] The concentration and content calculations are as in Example 1.

[0237] 3.3 Acceptance criteria

[0238] According to the standard curve method, NMOR shall not exceed 0.45ppm, NPYR shall not exceed 0.45ppm, NDEA shall not exceed 0.45ppm, NPIP shall not exceed 0.45ppm, NMPHA shall not exceed 0.45ppm, NDPA shall not exceed 0.45ppm, NEPHA shall not exceed 0.45ppm, NDBA shall not exceed 0.45ppm, and NDBZA shall not exceed 0.45ppm.

[0239] 4. Verification results:

[0240]

[0241]

[0242]

[0243]

[0244] Example 5

[0245] Example 5 provides a method for detecting related substances in famotidine injection using liquid chromatography-mass spectrometry.

[0246] The difference between Example 5 and Example 4 is that:

[0247] The parameters of the liquid chromatography were as follows: mobile phase A was 0.05% formic acid-water solution, and mobile phase B was methanol;

[0248] The column temperature is 35° C.; the concentration of the sample to be tested of famotidine injection is 8 mg / ml; and the injection volume of the liquid chromatography is 48 μ.

[0249] Specificity test results: same as Example 4.

[0250] Example 6

[0251] Example 6 provides a method for detecting related substances in famotidine injection using liquid chromatography-mass spectrometry.

[0252] The difference between Example 6 and Example 4 is that:

[0253] The parameters of the liquid chromatography were as follows: mobile phase A was 0.15% formic acid-water solution, and mobile phase B was methanol;

[0254] The column temperature is 45°C; the concentration of the sample to be tested of famotidine injection is 12 mg / ml; and the injection volume of the liquid chromatography is 52 μ.

[0255] Specificity test results:

[0256] The results showed that it was the same as in Example 4.

[0257] 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 famotidine injection by liquid chromatography-mass spectrometry, characterized in that: Using liquid chromatography-mass spectrometry to detect the sample to be tested of the famotidine injection; The parameters of the liquid chromatography are as follows: the chromatographic column is C18; Mobile phase A was 0.05%-0.15% formic acid-water solution, and mobile phase B was methanol; Column temperature is 35-45°C; Gradient elution; The mass spectrometer ion source parameters are shown in the following table: First mass spectrometry condition 。 2. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 1, characterized in that: The conditions of the liquid chromatography are as follows: mobile phase A is 0.1% formic acid-water solution, mobile phase B is methanol, column temperature is 40° C., and / or, The chromatographic column of the liquid chromatography condition is Poroshell 120EC-C18.

3. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 1, characterized in that: The concentration of the sample to be tested of the famotidine injection is 8-12 mg / ml, preferably 10 mg / ml.

4. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 1, characterized in that: The related substances and their structures are as follows: The structures of the first group of related substances 5. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 4, characterized in that: The detection limits of the related substances were all 4.5 ng / ml; And / or, the limit of quantification of the related substances is 0.90 ng / ml.

6. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 4, characterized in that: The injection volume of the liquid chromatography is 18-22μ; Gradient elution conditions included the following: First gradient elution table ； The mass spectrometer ion source parameters also include: First valve switching procedure The scanning parameters are as follows: First MRM scan parameters Note: * indicates quantitative ion.

7. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 4, characterized in that: The flow rate of the sample to be tested of the famotidine injection is 0.4 ml / min.

8. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 1, characterized in that: The related substances and their structures are as follows: Second related substance structure 9. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 8, characterized in that: The detection limits of the related substances are as follows: the detection limit of NDIPA is 0.66 ng / ml, the detection limit of NIPEA is 0.66 ng / ml, the detection limit of NMBA is 2.4 ng / ml, and the detection limit of NMPA is 0.66 ng / ml; And / or, the quantification limits of the related substances are as follows: the quantification limit of NDIPA is 1.32 ng / ml, the quantification limit of NIPEA is 1.33 ng / ml, the quantification limit of NMBA is 4.80 ng / ml, and the quantification limit of NMPA is 1.33 ng / ml.

10. The method for detecting related substances in famotidine injection by liquid chromatography-mass spectrometry according to claim 1, characterized in that: The injection volume of the liquid chromatography is 48-52μ; Gradient elution conditions included the following: Second gradient elution table ； The mass spectrometer ion source parameters also include: Second valve switching procedure The scanning parameters are as follows: Second MRM scan parameters Note: * indicates quantitative ion.

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