Analysis method for determining halogenated alkane impurities in tigliptin hydrobromide

Abstract

The invention discloses an analysis method for determining halogenated alkane impurities in tigliptin hydrobromide, and relates to the technical field of quality analysis. In order to solve the problem of detection of halogenated alkane impurities in tigliptin hydrobromide, the invention provides a convenient, efficient and accurate detection method. The method comprises the following steps: preparing a test solution and an impurity reference solution, and detecting by adopting a gas chromatographic method; the method can be used for accurately determining the content of halogenated alkane in tigliptin hydrobromide, plays a guiding role in development of a synthesis process, and contributes to establishment of quality standards of tigliptin hydrobromide bulk drugs.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of pharmaceutical analysis, in particular to an analysis method for determining halogenated alkane impurities in teneligliptin hydrobromide hydrate. BACKGROUND

[0002] Teneligliptin hydrobromide hydrate, the chemical name of which is {(2S, 4S)-4-[4-(3-methyl-1-phenyl-1H-pyrazol-5-yl)piperazin-1-yl]pyrrolidin-2-yl}(1, 3-thiazolidin-3-yl)methanone bis-hemihydrobromide hydrate, has the following chemical structural formula: DPP-4 inhibitors are one of the main drug types for treating diabetes. Such drugs can not only inhibit the inactivation of glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), increase the levels of endogenous GLP-1 and GIP, and promote the release of insulin from islet beta cells, but also inhibit the secretion of glucagon from islet alpha cells, increase the level of insulin, reduce blood glucose, and are less likely to induce hypoglycemia and increase body weight.

[0003] According to the production process of teneligliptin hydrobromide hydrate, hydrogen bromide is used in the synthesis process. Hydrogen bromide reacts with alcohol reagents to generate bromomethane, 2-bromopropane, chloromethane, chloroethane, bromoethane, and 2-chloropropane impurities. These impurities contain a mutagenic warning structure and belong to mutagenic impurities. Therefore, it is necessary to control bromomethane, 2-bromopropane, chloromethane, chloroethane, bromoethane, and 2-chloropropane in teneligliptin hydrobromide hydrate. Therefore, it is very important to develop an analysis method for detecting halogenated alkane impurities in teneligliptin hydrobromide hydrate. At present, the quality standard of teneligliptin hydrobromide hydrate is only collected in IP, and is not collected in USP, BP, EP, JP, and Chinese Pharmacopoeia. However, mutagenic impurities are not controlled in IP. In order to ensure the quality of drug research and production, it is of great significance to develop an accurate and effective analysis method for determining halogenated alkane impurities in teneligliptin hydrobromide hydrate for the quality control of drug products. SUMMARY

[0004] The purpose of the present application is to provide an analysis method for determining halogenated alkane impurities in teneligliptin hydrobromide hydrate. The method uses a gas chromatography system for analysis. The method can detect the content of bromomethane, 2-bromopropane, chloromethane, chloroethane, bromoethane, and 2-chloropropane in teneligliptin hydrobromide hydrate, and is convenient for the quality control of teneligliptin hydrobromide hydrate.

[0005] To achieve the purpose of the above-mentioned application, the present application adopts the following technical solutions: A method for determining halogenated alkane impurities in tigilazone hydrobromide, comprising the following steps: (1) Preparation of solution A, diluent: N, N-dimethylformamide-water (50:50) B, control solution: take bromomethane, 2-bromopropane, chloromethane, chloroethane, 2-chloropropane respectively, accurately weigh, dissolve with diluent to obtain the control solution; C, test solution: take tigilazone hydrobromide sample, dissolve with diluent.

[0006] (2) Determination method Take the blank solvent, control solution and test solution into the gas chromatograph respectively, and the chromatographic conditions are as follows: Chromatographic column: 6% cyanopropyl phenyl-94% dimethyl polysiloxane as stationary liquid; initial temperature is 40-60℃, maintained for 5-15 minutes, increased to 210-250℃ at the rate of 20-30℃ per minute, maintained for 5-10 minutes; injection port temperature is 160-200℃; detector temperature is 220-280℃; headspace bottle equilibrium temperature is 80-90℃, equilibrium time is 20-40 minutes; carrier gas flow rate is 3-8ml per minute; split ratio is 3:1-7:1.

[0007] Compared with the prior art, the beneficial effects of the present application are: The detection method of halogenated alkane impurities in tigilazone hydrobromide is not collected in the pharmacopoeia of each country. The present application provides a detection method of halogenated alkane impurities in tigilazone hydrobromide, which has strong specificity, high sensitivity, good accuracy, high precision and good repeatability, can effectively determine the halogenated alkane impurities in tigilazone hydrobromide, and is used for detection and monitoring in the production process of tigilazone hydrobromide, which is helpful for the establishment of the quality standard of the raw material drug. BRIEF DESCRIPTION OF DRAWINGS

[0008] Fig. 1 It is the blank solution chromatogram of the method for determining halogenated alkane impurities in tigilazone hydrobromide of the present application; Fig. 2 It is the control solution chromatogram of the method for determining halogenated alkane impurities in tigilazone hydrobromide of the present application; Fig. 3 It is the test solution detection chromatogram of the method for determining halogenated alkane impurities in tigilazone hydrobromide of the present application. DETAILED DESCRIPTION

[0009] With reference to the drawings of the embodiments of the present application, the technical solutions in the embodiments of the present application will be described clearly and completely. Obviously, the described embodiments are only a part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments of the present application, all other embodiments obtained by those of ordinary skill in the art without creative work fall within the scope of the present application.

[0010] With reference to the drawings of the embodiments of the present application, the technical solutions in the embodiments of the present application will be described clearly and completely. Obviously, the described embodiments are only a part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments of the present application, all other embodiments obtained by those of ordinary skill in the art without creative work fall within the scope of the present application. Figs. 1-3 The present application provides a technical solution: With reference to the drawings of the embodiments of the present application, the technical solutions in the embodiments of the present application will be described clearly and completely. Obviously, the described embodiments are only a part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments of the present application, all other embodiments obtained by those of ordinary skill in the art without creative work fall within the scope of the present application. Figs. 1-3 The present application provides a technical solution: Embodiment 1: Specificity: 1. Instruments and reagents: Shimadzu GC-2030 gas chromatograph, and Waters Empower3 (network version) chromatographic workstation configured with the analysis instrument.

[0011] N,N-dimethylformamide (chromatographic grade): reagent specification: N,N-dimethylformamide (chromatographic grade, purity ≥ 99.9%); experimental water is ultrapure water (resistivity ≥ 18.2 MΩ·cm, 25℃); chloromethane, bromomethane, chloroethane, 2-chloropropane, 2-bromopropane reference substance; 2. Chromatographic conditions: Chromatographic column: 6% cyanopropyl phenyl-94% dimethyl polysiloxane (or similar polarity) as stationary liquid (Agilent DB-624, 60m x 0.53mm, 3.0μm); Detector: FID; Detector temperature: 250℃; Injection port temperature: 180℃; Carrier gas flow rate: 5ml / min; Split ratio: 5:1; Headspace bottle equilibrium temperature: 85℃; Headspace bottle equilibrium time: 25 minutes; Temperature program table: In this embodiment, by selecting a chromatographic column with 6% cyanopropyl phenyl-94% dimethyl polysiloxane (or similar polarity) as the stationary liquid, and cooperating with the above temperature program (initial 50℃ for 10 minutes, increased to 230℃ at 25℃ / min for 5 minutes), headspace equilibrium conditions (85℃ for 25 minutes) and carrier gas parameters (flow rate 5ml / min, split ratio 5:1), the target halogenated alkane impurities (chloromethane, bromomethane, chloroethane, 2-chloropropane, 2-bromopropane) and other components in the blank solvent and test sample matrix are effectively separated, thereby achieving the purpose of specific detection.

[0012] 3. Preparation of related solutions: (1) Diluent: N,N-dimethylformamide-water (50:50); (2) Each impurity stock solution: Take chloromethane, chloroethane, bromomethane, 2-chloropropane, 2-bromopropane, accurately weigh, respectively placed in different volumetric flasks, dissolved and diluted to the mark with diluent, shake well. (Each concentration is about 0.5 mg / ml); (3) Impurity mixed stock solution: accurately take the appropriate amount of each impurity stock solution, place it in the same volumetric flask, dilute to the mark with diluent, shake well. (Each concentration is about 50 μg / ml); (4) Reference solution: accurately take the appropriate amount of impurity mixed stock solution, place it in the volumetric flask, dilute to the mark with diluent, shake well. (Each concentration is about 5 μg / ml); (5) Test solution: take a suitable amount of tigergliptin hydrobromide, accurately weigh, place it in a volumetric flask, dissolve and dilute to the mark with diluent, shake well. (The concentration is about 10 mg / ml).

[0013] 4. Determination method: Accurately take (1) diluent, (4) reference solution and (5) test solution under the preparation of the above related solutions, respectively headspace injection, record the chromatogram, the test results are shown in Table 1, the chromatogram is shown in Fig. 1 , Fig. 2 and Fig. 3 .

[0014] Table 1 Specificity test results: The above test results show that the separation degree between each chromatographic peak is greater than 1.5, indicating that the method has good specificity.

[0015] Example 2: Quantitative limit and detection limit: Quantitative limit solution: accurately take the appropriate amount of each impurity stock solution under item (2) of Example 1, place it in a volumetric flask, dilute to the mark with diluent, shake well. (Chloromethane 0.025 μg / ml, bromomethane 0.075 μg / ml, chloroethane 0.025 μg / ml, 2-chloropropane 0.025 μg / ml, 2-bromopropane 0.077 μg / ml) Detection limit solution: accurately take 10 ml of the quantitative limit solution, place it in a 20 ml volumetric flask, dilute to the mark with diluent, shake well. (Chloromethane 0.0125 μg / ml, bromomethane 0.0375 μg / ml, chloroethane 0.0125 μg / ml, 2-chloropropane 0.0125 μg / ml, 2-bromopropane 0.0385 μg / ml) This embodiment determines the limit of quantitation (signal-to-noise ratio ≥10) and limit of detection (signal-to-noise ratio ≥3) for each target impurity by gradually diluting the target haloalkane impurity stock solution and combining the chromatographic conditions and headspace injection method set in the invention, thus verifying the sensitivity of the method to meet the requirements of the invention for detecting low-concentration mutagenic impurities.

[0016] The headspace vials are 20 ml in size. After each solution is transferred to the headspace vial, it is immediately sealed and detected according to the chromatographic conditions set in Example 1 (flame ionization detector, injection port temperature 180°C, detector temperature 250°C, etc.). Accurately measure the above-mentioned limit of quantitation solution and limit of detection solution and inject them into the headspace, record the chromatograms, and the results are shown in Table 2.

[0017] Table 2 Results of Limit of Quantitation and Limit of Detection: The results show that the quantitation limits of each halogenated hydrocarbon impurity are all less than 50% of the reporting limit (2.5 μg / ml), with a signal-to-noise ratio greater than 10; the signal-to-noise ratio of the detection limit is greater than 3, and the sensitivity meets the requirements.

[0018] Example 3: Linearity and Range: Linear solution: Accurately measure the impurity mixed stock solution under Example 1 (3) according to Table 3, place it in the corresponding volumetric flask, dilute to the mark with diluent, and shake well.

[0019] The linear solutions corresponding to the limit of quantitation are prepared according to the method for preparing the limit of quantitation solutions in Example 2. The specific dilution logic for solutions L1-L5 is as follows: accurately measure the corresponding volume of the mixed stock solution of target haloalkane impurities (each with a concentration of approximately 50 μg / ml) under Example 1, dilute to the mark with diluent, and shake well to obtain the linear solutions of the corresponding concentrations.

[0020] Table 3. Preparation of linear solutions: Determination method: The linear solutions L1 to L5 were accurately measured and injected into the headspace, and the chromatograms were recorded. Linear regression analysis was performed with concentration as the abscissa (x) and peak area as the ordinate (y) to obtain the linear equation and correlation coefficient. The determination results are shown in Table 4.

[0021] This embodiment involves preparing linear solutions of target haloalkane impurities with different concentration gradients and performing headspace sampling detection under the chromatographic conditions set by the invention. The linear relationship between peak area and concentration is examined, providing a mathematical basis for the accurate quantification of impurity content and achieving the core objective of quantitative detection of haloalkane impurities in the invention.

[0022] Table 4. Results of linearity and range tests: The above results show that the peak area and concentration of each impurity exhibit a good linear relationship within the range from the limit of quantitation to 0.10% of the concentration of the test sample solution, with correlation coefficients (r) greater than 0.990.

[0023] Example 4: Accuracy: Reference solution: Take the reference solution (4) under Example 1 above as the reference solution.

[0024] Test solution: Take the test solution (5) under Example 1 above as the test solution.

[0025] Accuracy Sample Solution: Weigh an appropriate amount of ticagliptin hydrobromide according to Table 5 below, place it in the corresponding volumetric flask, add an appropriate amount of diluent to dissolve it, and then accurately measure an appropriate amount of the impurity mixed stock solution under (3) of Example 1 above, place it in the corresponding volumetric flask, dilute it to the mark with diluent, shake well, and prepare 3 sample solutions in parallel for each level.

[0026] Table 5. Preparation methods of accuracy sample solutions: Assay: Accurately measure the reference solution, test solution, and accuracy sample solution under the accuracy section and inject them into the headspace separately. Record the chromatograms and calculate the recovery rate according to the external standard method for impurities: Recovery rate (%) = (measured amount - background amount) / added amount × 100%, where 'measured amount' is the impurity concentration × solution volume in the accuracy sample solution, 'added amount' is the concentration × added volume of the added impurity stock solution, and 'background amount' is the measured amount of impurities in the test solution (in this example, no impurities were detected in the test solution, so the background amount is 0). The recovery rate results of each haloalkane impurity are shown in Tables 6 to 10.

[0027] This embodiment performs headspace sampling detection under the chromatographic conditions determined in the invention, calculates the recovery rate using the external standard method for impurities, verifies the accuracy of the method, ensures the reliability of the determination results of the content of the target mutagenic impurities in the test sample, and meets the requirements for accurate detection of impurities in drug quality control.

[0028] Table 6. Results of Chloromethane Recovery Test: Table 7 Results of the brominated methane recovery test: Table 8 Results of chloroethane recovery test: Table 9. Results of 2-chloropropane recovery test: Table 10 Results of 2-Bromopropane Recovery Test: Results and Analysis: At three levels (50% limit concentration, 100% limit concentration, and 150% limit concentration), the recoveries calculated by peak area using the external standard method for impurities were 97.18%, 101.02%, 95.33%, 98.28%, and 98.07%, respectively. The RSDs were 9.86%, 4.51%, 6.98%, 6.55%, and 5.23%, respectively, which met the requirements. The accuracy test results of this method also met the requirements.

Claims

1. An analytical method for determining halogenated alkane impurities in ticagliptin hydrobromide, wherein the halogenated alkane impurities include chloromethane, bromomethane, chloroethane, bromoethane, 2-chloropropane, and 2-bromopropane introduced during production and storage, characterized in that: It is detected using gas chromatography, which includes the following steps: (1) Gas chromatography conditions: Chromatographic column: 6% cyanopropylphenyl-94% dimethylpolysiloxane as stationary phase; initial temperature 40–60℃, maintained for 5–15 minutes, then increased to 210–250℃ at a rate of 20–30℃ per minute, maintained for 5–10 minutes; injection port temperature 160–200℃; detector temperature 220–280℃; headspace vial equilibration temperature 80–90℃, equilibration time 20–40 minutes; carrier gas flow rate 3–8 ml per minute; split ratio 3:1–7:

1. (2) Preparation of reference solution; (3) Prepare the test solution; (4) Determination method: Accurately measure the headspace sample of the reference solution and the test solution, record the chromatogram, and calculate the content of haloalkanes in the test solution by peak area according to the external standard method for impurities.

2. The analytical method for determining haloalkanes in ticagliptin hydrobromide as described in claim 1, characterized in that: The haloalkane impurities include at least one of the following: chloromethane, bromomethane, chloroethane, bromoethane, 2-chloropropane, and 2-bromopropane.

3. The analytical method for determining haloalkanes in ticagliptin hydrobromide as described in claim 1, characterized in that: The initial temperature in step (1) is 40-60°C; maintain for 5-15 minutes.

4. The analytical method for determining haloalkanes in ticagliptin hydrobromide as described in claim 1, characterized in that: Step (1) involves raising the temperature to 210-250°C at a rate of 20-30°C per minute and maintaining it for 5-10 minutes.

5. The analytical method for determining haloalkanes in ticagliptin hydrobromide as described in claim 1, characterized in that: In step (1), the injection port temperature is 160-200℃ and the detector temperature is 220-280℃.

6. The analytical method for determining haloalkanes in ticagliptin hydrobromide as described in claim 1, characterized in that: The headspace flask equilibration temperature in step (1) is 80-90°C; the equilibration time is 20-40 minutes.

7. The analytical method for determining haloalkanes in ticagliptin hydrobromide as described in claim 1, characterized in that: The carrier gas flow rate in step (1) is 3-8 ml per minute; the split ratio is 3:1-7:

1.

8. The analytical method for determining haloalkanes in ticagliptin hydrobromide as described in claim 1, characterized in that: The concentration of the reference solution prepared in step (2) is 0.15–50 μg / ml.

9. The analytical method for determining haloalkanes in ticagliptin hydrobromide as described in claim 1, characterized in that: The concentration of the test solution prepared in step (3) is 0.3 to 100 mg / ml.

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