Method for detecting impurities in nitrosamine of hydrobromic acid potioxetine by liquid chromatography-mass spectrometry

Abstract

The invention discloses a method for detecting impurities in nitrosamine of vortioxetine hydrobromide by liquid chromatography-mass spectrometry. According to the method, an ultra-high performance liquid chromatograph-mass spectrometer is adopted, and octadecylsilane chemically bonded silica is used as a filler; the chromatographic conditions are as follows: the column temperature is 35-45 DEG C, the temperature of a sample introduction disc is 6-8 DEG C, the flow velocity is 0.3-0.5 ml / min, and the sample introduction amount is 2-20 microliters; and eluting with a gradient mobile phase. Mass spectrum conditions are as follows: a triple quadrupole tandem mass spectrometer is used for detection, an ion source is an electrospray ion source, and a positive ion scanning mode is adopted. Calculating the content of the impurity G by adopting an external standard method. The method for analyzing the impurity G in the vortioxetine hydrobromide, established by the invention, is strong in specificity, high in stability, high in accuracy, high in precision and strong in durability, and can be used as a method for detecting the content of the impurity G in the vortioxetine hydrobromide.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical analysis technology, specifically relating to a method for detecting nitrosamine impurities in vortioxetine hydrobromide using liquid chromatography-mass spectrometry. Background Technology

[0002] Vortioxetine hydrobromide is used to treat depression in adults.

[0003] Impurity G (1-(2-((2,4-dimethylphenyl)thio)phenyl)-4-nitrosopiperazine) is a nitrosamine genotoxic impurity. Summary of the Invention

[0004] The purpose of this invention is to provide an analytical method for detecting impurity G using ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS / MS), accurately detecting the content of impurity G in vortioxetine hydrobromide products, and achieving quality control.

[0005] The method for analyzing impurity G is as follows: using octadecylsilane-bonded silica gel as the packing material; column temperature of 35-45℃, injection plate temperature of 6-8℃, flow rate of 0.3-0.5 ml / min, and injection volume of 2-20 μl; Chromatographic conditions: Mobile phase A: 0.1% formic acid solution; Mobile phase B: 0.1% formic acid acetonitrile solution; Elution with gradient mobile phases, with a mobile phase volume ratio of 100%, the gradient of the mobile phases is set as follows:

[0006] Mass spectrometry conditions: Mass spectrometry detector used, electrospray ionization positive ion mode (ESI) + Multiple reaction monitoring (MRM) was performed, with mass-to-charge ratios (m / z) of 328.1→298.1 selected as quantitative ion pairs and 328.1→256.1 as qualitative ion pairs for detection. Specific mass spectrometry conditions were set as follows:

[0007] The specific steps of the method are as follows: (1) The solvent is water and acetonitrile in a 1:1 ratio; (2) Test solution: Take an appropriate amount of this product, weigh it accurately, dissolve it in solvent and dilute it quantitatively to prepare a solution containing 0.8 mg of vortioxetine per ml; (3) Reference stock solution: Take an appropriate amount of impurity G reference standard, accurately weigh it, dissolve it in acetonitrile and quantitatively dilute it to prepare a solution containing 0.8 μg of impurity G per 1 ml; (4) Reference solution: Accurately measure an appropriate amount of the reference stock solution and dilute it quantitatively with a solvent to prepare a solution containing 16 ng of impurity G per 1 ml; (5) Determination: Accurately measure the reference solution and the test solution, inject them into the ultra-high performance liquid chromatography-mass spectrometry instrument, record the chromatograms, and calculate the content of impurity G by peak area according to the external standard method.

[0008] The analytical method for impurity G in vortioxetine hydrobromide established in this invention has high specificity, high stability, high accuracy, high precision, and high robustness, and can be used as a method for detecting the content of impurity G in vortioxetine hydrobromide. Attached Figure Description

[0009] Figure 1 This is a linear relationship graph for impurity G. Detailed Implementation

[0010] The following will provide a detailed description of the quality control method for impurity G in vortioxetine hydrobromide according to the present invention through examples. However, this should not be construed as limiting the scope of the claims of the present invention to the following examples. All technologies implemented based on the above content of the present invention fall within the scope of the present invention. Example 1

[0011] Column: ACQUITY UPLC HSS T3 (2.1×100mm, 1.8μm) The chromatographic conditions were as follows: column temperature 35℃; injection plate temperature 6℃; flow rate 0.4 ml / min; injection volume 2 μl; mobile phase A: 0.1% formic acid solution; mobile phase B: 0.1% formic acid acetonitrile solution; elution was performed using a gradient mobile phase, with the gradient set as follows (based on a 100% volume ratio):

[0012] Mass spectrometry conditions: Mass spectrometry detector used, electrospray ionization positive ion mode (ESI) + Multiple reaction monitoring (MRM) was performed, with mass-to-charge ratios (m / z) of 328.1→298.1 selected as quantitative ion pairs and 328.1→256.1 as qualitative ion pairs for detection. Specific mass spectrometry conditions were set as follows:

[0013] Specificity and System Applicability Solution preparation: Solvent: Water-acetonitrile (1:1).

[0014] Reference solution: Accurately weigh approximately 4 mg of impurity G reference standard and place it in a 50 ml volumetric flask. Add acetonitrile and shake to dissolve impurity G. Dilute to the mark with acetonitrile and mix well. Accurately measure 0.5 ml and place it in a 50 ml volumetric flask. Dilute to volume with acetonitrile and mix well. Accurately measure 1 ml and place it in a 50 ml volumetric flask. Dilute to volume with solvent and mix well.

[0015] Test solution: Accurately weigh about 50.8 mg of this product, place it in a 50 ml volumetric flask, add solvent to dissolve and dilute to the mark, shake well, and the solution is ready.

[0016] result Specific results

[0017] System applicability results

[0018] The specificity and system applicability results meet the requirements.

[0019] Limit of detection and limit of quantitation Solution preparation: Solvent: Water-acetonitrile (1:1).

[0020] Reference solution: Accurately weigh approximately 4 mg of impurity G reference standard and place it in a 50 ml volumetric flask. Add acetonitrile and shake to dissolve impurity G. Dilute to the mark with acetonitrile and mix well. Accurately measure 0.5 ml and place it in a 50 ml volumetric flask. Dilute to volume with acetonitrile and mix well. Accurately measure 1 ml and place it in a 50 ml volumetric flask. Dilute to volume with solvent and mix well.

[0021] Limit of Quantitation Solution: Accurately measure 1 ml of the reference solution into a 50 ml volumetric flask, dilute to volume with solvent, and shake well.

[0022] Detection limit solution: Accurately measure 5 ml of the quantitation limit solution and place it in a 10 ml volumetric flask. Dilute to volume with solvent and shake well.

[0023] result Detection limit results

[0024] Quantitative limit results

[0025] The results of the detection limit and quantitation limit experiments meet the detection requirements.

[0026] Linearity and Range Solution preparation Solvent: Water-acetonitrile (1:1).

[0027] Linear stock solution: Accurately weigh approximately 4 mg of impurity G reference standard and place it in a 50 ml volumetric flask. Add acetonitrile and shake to dissolve impurity G. Dilute to the mark with acetonitrile and shake well. Accurately measure 0.5 ml of the solution and place it in a 50 ml volumetric flask. Dilute to volume with acetonitrile and shake well.

[0028] Linear solution 1: Accurately measure 0.1 ml of the linear stock solution, place it in a 50 ml volumetric flask, dilute to the mark with solvent, and shake well to obtain the solution.

[0029] Linear solution 2: Accurately measure 0.2 ml of the linear stock solution, place it in a 50 ml volumetric flask, dilute to the mark with solvent, and shake well to obtain the solution.

[0030] Linear solution 3: Accurately measure 0.5 ml of the linear stock solution, place it in a 20 ml volumetric flask, dilute to the mark with solvent, and shake well to obtain the solution.

[0031] Linear solution 4: Accurately measure 1 ml of the linear stock solution, place it in a 10 ml volumetric flask, dilute to the mark with solvent, and shake well to obtain the solution.

[0032] Linear solution 5: Accurately measure 1.5 ml of the linear stock solution, place it in a 20 ml volumetric flask, dilute to the mark with solvent, and shake well to obtain the solution.

[0033] Linear solution 6: Accurately measure 2 ml of the linear stock solution, place it in a 10 ml volumetric flask, dilute to the mark with solvent, and shake well to obtain the solution.

[0034] result linear relationship of impurity G

[0035] The linear relationship of impurity G is good.

[0036] Solution stability Solution preparation: Solvent: Water-acetonitrile (1:1).

[0037] Reference solution: Accurately weigh approximately 4 mg of impurity G reference standard and place it in a 50 ml volumetric flask. Add acetonitrile and shake to dissolve impurity G. Dilute to the mark with acetonitrile and mix well. Accurately measure 0.5 ml and place it in a 50 ml volumetric flask. Dilute to volume with acetonitrile and mix well. Accurately measure 1 ml and place it in a 50 ml volumetric flask. Dilute to volume with solvent and mix well.

[0038] 100% spiked test solution: Accurately weigh about 50.8 mg of this product, place it in a 50 ml volumetric flask, add 1 ml of reference stock solution, dissolve and dilute to the mark with solvent, and shake well to obtain the solution.

[0039] result Stability results of reference solution

[0040] Stability results of 100% spiked test solution

[0041] Conclusion: At 6℃, the reference solution was stable for 15.8 hours; the 100% spiked test solution was stable for 16 hours.

[0042] Accuracy and repeatability Solution preparation Solvent: Water-acetonitrile (1:1).

[0043] Reference solution: Accurately weigh approximately 4 mg of impurity G reference standard and place it in a 50 ml volumetric flask. Add acetonitrile and shake to dissolve impurity G. Dilute to the mark with acetonitrile and mix well. Accurately measure 0.5 ml and place it in a 50 ml volumetric flask. Dilute to volume with acetonitrile and mix well. Accurately measure 1 ml and place it in a 50 ml volumetric flask. Dilute to volume with solvent and mix well.

[0044] 50% spiked test solution: Accurately weigh approximately 50.8 mg of this product and place it in a 50 ml volumetric flask. Add 0.5 ml of the reference stock solution, dissolve and dilute to the mark with solvent, and shake well. Prepare three parallel solutions.

[0045] 100% Spiked Test Solution: Accurately weigh approximately 50.8 mg of this product and place it in a 50 ml volumetric flask. Add 1 ml of the reference stock solution, dissolve and dilute to the mark with solvent, and shake well. Prepare 6 parallel solutions. (The first 3 solutions are used as 100% spiked test solution accuracy solutions, and all 6 solutions are used as repeatability solutions).

[0046] 150% spiked test solution: Accurately weigh approximately 50.8 mg of this product and place it in a 50 ml volumetric flask. Add 1.5 ml of the reference stock solution, dissolve and dilute to the mark with solvent, and shake well. Prepare three parallel solutions.

[0047] Accuracy results

[0048] Repeatability results

[0049] The accuracy and repeatability results meet the testing requirements.

[0050] intermediate precision Solution preparation Solvent: Water-acetonitrile (1:1).

[0051] Reference solution: Accurately weigh approximately 4 mg of impurity G reference standard and place it in a 50 ml volumetric flask. Add acetonitrile and shake to dissolve impurity G. Dilute to the mark with acetonitrile and mix well. Accurately measure 0.5 ml and place it in a 50 ml volumetric flask. Dilute to volume with acetonitrile and mix well. Accurately measure 1 ml and place it in a 50 ml volumetric flask. Dilute to volume with solvent and mix well.

[0052] Intermediate precision solution: Accurately weigh approximately 50.8 mg of this product and place it in a 50 ml volumetric flask. Add 1 ml of the reference stock solution, dissolve and dilute to the mark with solvent, and shake well. Prepare 6 parallel solutions.

[0053] result Intermediate precision results

[0054] The intermediate precision results meet the testing requirements and the precision is good.

[0055] Durability Solution preparation Solvent: Water-acetonitrile (1:1).

[0056] Reference solution: Accurately weigh approximately 4 mg of impurity G reference standard and place it in a 50 ml volumetric flask. Add acetonitrile and shake to dissolve impurity G. Dilute to the mark with acetonitrile and mix well. Accurately measure 0.5 ml and place it in a 50 ml volumetric flask. Dilute to volume with acetonitrile and mix well. Accurately measure 1 ml and place it in a 50 ml volumetric flask. Dilute to volume with solvent and mix well.

[0057] 100% spiked test solution: Accurately weigh about 50.8 mg of this product, place it in a 50 ml volumetric flask, add 1 ml of reference stock solution, dissolve and dilute to the mark with solvent, and shake well to obtain the solution.

[0058] result Solvent durability results

[0059] Durability results of reference solution

[0060] Robustness results of 100% spiked test solution

[0061] The durability results meet the testing requirements.

[0062] The above are merely preferred embodiments of the present invention. It should be noted that researchers in the art can make various modifications and alterations without departing from the inventive concept of the present invention, and these modifications and alterations all fall within the protection scope of the present invention.

Claims

1. A method for detecting nitrosamine impurities in vortioxetine hydrobromide using liquid chromatography-mass spectrometry, characterized in that, Chromatographic conditions: Octadecylsilane-bonded silica gel was used as the stationary phase; 0.1% formic acid aqueous solution was used as mobile phase A; and 0.1% formic acid acetonitrile solution was used as mobile phase B, with gradient elution as shown in the table below; column temperature was 35-45℃; injection plate temperature was 6-8℃; flow rate was 0.3-0.5 ml / min; and injection volume was 2-20 μl. ， Mass spectrometry conditions: Detection was performed using a triple quadrupole tandem mass spectrometer with an electrospray ionization (ESI) source in positive ion scanning mode. Selected reaction monitoring (MRM) was used, with mass-to-charge ratios (m / z) of 328.1–298.1 selected as quantitative ion pairs and 328.1–256.1 as qualitative ion pairs. Specific mass spectrometry conditions are shown in the table below. 。 2. The method as described in claim 1, characterized in that, The specific steps of the method are as follows: (1) The solvent is water and acetonitrile in a 1:1 ratio; (2) Test solution: Take an appropriate amount of this product, weigh it accurately, dissolve it in solvent and dilute it quantitatively to prepare a solution containing 0.8 mg of vortioxetine per ml; (3) Reference stock solution: Take an appropriate amount of impurity G reference standard, accurately weigh it, dissolve it in acetonitrile and quantitatively dilute it to prepare a solution containing 0.8 μg of impurity G per 1 ml; (4) Reference solution: Accurately measure an appropriate amount of the reference stock solution and dilute it quantitatively with a solvent to prepare a solution containing 16 ng of impurity G per 1 ml; (5) Determination: Accurately measure the reference solution and the test solution, inject them into the ultra-high performance liquid chromatography-mass spectrometry instrument, record the chromatograms, and calculate the content of impurity G by peak area according to the external standard method.

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