Method for detecting p-chlorobenzhydryl piperazine in levocetirizine hydrochloride particles
The separation and determination of impurity A in levocetirizine hydrochloride particles by high performance liquid chromatography solved the problem of difficulty in detection, and achieved improvement in quality control and guarantee of pharmaceutical safety.
Patent Information
- Application Number
- CN202510235397.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-02-28
- Publication Date
- 2025-05-30
AI Technical Summary
The detection of impurity A (p-chlorodiphenylpiperazine) in levocetirizine hydrochloride particles is difficult to detect, affecting the quality control of the preparation.
High performance liquid chromatography was used to separate and measure the separation and determination of levocetirizine hydrochloride and impurity A from the octadecylsilane-bonded silica gel chromatography column using a mixed solution of phosphoric acid and acetonitrile-methanol solution as the mobile phase.
The accurate and reliable detection of impurity A in levocetirizine hydrochloride particles is achieved, the quality control level of the preparation is improved, and the safety of pharmaceuticals is ensured.
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Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of pharmaceutical analysis, and particularly relates to a method for detecting p-chlorodiphenylmethylpiperazine in levocetirizine hydrochloride granules. Background Art
[0002] Levocetirizine hydrochloride is a highly effective selective histamine H1 receptor antagonist, which is widely used in the treatment of various allergic reactions. This drug is mainly used clinically to improve skin and mucosal allergic symptoms such as allergic rhinitis, urticaria, and angioneurotic edema. In addition, it can effectively relieve allergic reactions during colds, providing significant symptom relief for patients.
[0003] Levocetirizine hydrochloride granules usually take effect 1 hour after administration, and the effect can last for 24 hours. It only needs to be taken once a day, which is convenient to use. And as a granule drug, it is easy to dissolve and is very suitable for children or adults with difficulty swallowing. The study of drug-related substances is one of the key issues in the quality study of preparations. Due to factors such as the synthesis process of levocetirizine hydrochloride raw materials, the production process of levocetirizine hydrochloride granule preparations, and the storage process, impurity A will be produced in levocetirizine hydrochloride granules. Therefore, the analysis of impurity A in levocetirizine hydrochloride granules is of great significance for the production and quality control of levocetirizine hydrochloride granules. Summary of the Invention
[0004] In view of this, the present invention provides a method for detecting p-chlorodiphenylmethylpiperazine in levocetirizine hydrochloride granules. Through the method of the present invention, the separation and determination of levocetirizine hydrochloride and related substances can be achieved, thereby effectively realizing the quality control of levocetirizine hydrochloride granules.
[0005] The technical solution of the present invention is realized as follows:
[0006] A method for detecting p-chlorodiphenylmethylpiperazine in levocetirizine hydrochloride granules, comprising the following steps: using high performance liquid chromatography to determine the related substances in the levocetirizine hydrochloride granules; the chromatographic column in the method is a chromatographic column filled with octadecylsilane-bonded silica gel; the mobile phase is a mixed solution of mobile phase A and mobile phase B, using an aqueous phosphoric acid solution as mobile phase A and an acetonitrile-methanol solution as mobile phase B, and linear gradient elution is adopted. The concentration of mobile phase A is adjusted within the range of 75% v / v - 20% v / v, the concentration of mobile phase B is adjusted within the range of 80% v / v - 25% v / v, and the elution time is 60 - 70 min.
[0007] Further, the aqueous phosphoric acid solution is obtained by adjusting the pH of the aqueous solution to 2.8 - 3.0 with phosphoric acid; the volume ratio of acetonitrile to methanol in the acetonitrile-methanol solution is 70 - 80:20 - 30.
[0008] Further, the linear gradient elution program is as follows:
[0009]
[0010] Further, the specifications of the chromatographic column are Inertsil ODS-3, 4.6 mm × 250 mm, 3 μm or a chromatographic column with equivalent efficiency; the column temperature of the chromatographic column is 30-40 °C; the flow rate of the mobile phase is 0.8-1.0 mL / min.
[0011] Further, the detection wavelength of the ultraviolet detector in the high performance liquid chromatography is 220-240 nm; the injection volume in the high performance liquid chromatography is 10-20 μL.
[0012] Further, the preparation method of the test solution is as follows: Take the test sample, weigh accurately, place it in a volumetric flask, dissolve it ultrasonically with the solvent, allow it to stand to room temperature, dilute it to the scale with the solvent, shake well, filter, and take the subsequent filtrate to obtain the test solution; the solvent is composed of acetonitrile and water mixed in a volume ratio of 45-50:50-55.
[0013] Further, the preparation method of the levocetirizine hydrochloride reference solution is as follows: Take the levocetirizine hydrochloride reference substance, weigh accurately, dissolve it with the solvent and quantitatively dilute it to prepare a solution with a levocetirizine hydrochloride concentration of 2-4 μg / mL to obtain the levocetirizine hydrochloride reference solution; the solvent is composed of acetonitrile and water mixed in a volume ratio of 45-50:50-55.
[0014] Further, the preparation of the impurity A reference solution is as follows: Take the impurity A reference substance, weigh accurately, dissolve it with the solvent and quantitatively dilute it to prepare a solution with an impurity A concentration of 10-12 μg / mL to obtain the impurity A reference solution; the solvent is composed of acetonitrile and water mixed in a volume ratio of 45-50:50-55.
[0015] Further, the blank solution is composed of acetonitrile and water mixed in a volume ratio of 45-50:50-55; the impurity A is p-chlorodiphenylmethylpiperazine.
[0016] Further, the preparation method of the system suitability solution is as follows: Weigh accurately the levocetirizine hydrochloride reference substance and lactose, place them in a volumetric flask, dissolve them ultrasonically after adding the solvent, allow it to stand to room temperature, accurately add the impurity A reference stock solution, dilute it to the scale with the solvent, shake well, filter, and take the subsequent filtrate to obtain the system suitability solution.
[0017] Furthermore, the system suitability solution was prepared as follows: Weigh accurately 10 mg of levocetirizine hydrochloride reference substance and about 4 g of lactose, place them in the same 50-ml volumetric flask, add an appropriate amount of solvent, sonicate to dissolve levocetirizine hydrochloride, allow to stand at room temperature, accurately add 3 ml of impurity A reference substance solution, dilute to the mark with the solvent, shake well, filter, and take the subsequent filtrate; the solvent is a mixture of acetonitrile and water in a volume ratio of 50:50.
[0018] Compared with the prior art, the beneficial effects of the present invention are as follows:
[0019] The present invention provides a method for detecting p-chlorodiphenylmethylpiperazine in highly efficient levocetirizine hydrochloride granules, which can accurately and reliably separate and determine levocetirizine hydrochloride and related substances (impurity A: p-chlorodiphenylmethylpiperazine), thereby effectively realizing the quality control of levocetirizine hydrochloride granules and improving pharmaceutical safety. Specific embodiments
[0020] In order to better understand the technical content of the present invention, specific examples are provided below to further illustrate the present invention.
[0021] Unless otherwise specified, the experimental methods used in the embodiments of the present invention are all conventional methods.
[0022] Unless otherwise specified, the materials, reagents, etc. used in the embodiments of the present invention can all be obtained from commercial channels.
[0023] The manufacturer of the levocetirizine hydrochloride granules of the present invention is: Kangzhi Pharmaceutical Co., Ltd., the product name is: levocetirizine hydrochloride granules, and the brand is: Kangzhitai.
[0024] The related substance in the present invention is impurity A: p-chlorodiphenylmethylpiperazine, and the chemical structural formula is:
[0025]
[0026] Example 1 - Method for detecting p-chlorodiphenylmethylpiperazine in levocetirizine hydrochloride granules
[0027] The related substances in the levocetirizine hydrochloride granules were determined by high performance liquid chromatography; the related substance is impurity A: p-chlorodiphenylmethylpiperazine;
[0028] The preparation method of the test solution is as follows: Take the test sample, weigh accurately, place it in a volumetric flask, add the solvent and sonicate to dissolve, allow to stand at room temperature, dilute to the mark with the solvent, shake well, filter, and take the subsequent filtrate to obtain the test solution;
[0029] The preparation method of levocetirizine hydrochloride reference solution is as follows: Take levocetirizine hydrochloride reference substance, weigh it accurately, dissolve it with a solvent and quantitatively dilute it to prepare a solution with a levocetirizine hydrochloride concentration of 2 μg / mL, that is, the levocetirizine hydrochloride reference solution is obtained;
[0030] The preparation of impurity A reference solution is as follows: Take impurity A reference substance, weigh it accurately, dissolve it with a solvent and quantitatively dilute it to prepare a solution with an impurity A concentration of 10 μg / mL, that is, the impurity A reference solution is obtained;
[0031] The solvents in the test solution, levocetirizine hydrochloride reference solution and impurity A reference solution are all composed of acetonitrile and water mixed in a volume ratio of 50:50;
[0032] The blank solution is composed of acetonitrile and water mixed in a volume ratio of 50:50.
[0033] The chromatographic conditions in the high performance liquid chromatography analysis are as follows: The chromatographic column is a chromatographic column filled with octadecylsilane bonded silica gel, and the specifications of the chromatographic column are Inertsil ODS-3, 4.6 mm × 250 mm, 3 μm; The mobile phase is a mixed solution of mobile phase A and mobile phase B. The phosphoric acid aqueous solution obtained by adjusting the pH of the aqueous solution to 2.9 with phosphoric acid is mobile phase A, and the acetonitrile-methanol solution obtained by mixing acetonitrile and methanol in a volume ratio of 80:20 is mobile phase B; The flow rate of the mobile phase is 0.8 ml / min; The column temperature is 40 °C; The detection wavelength of the ultraviolet detector is 230 nm; The injection volume is 20 μL;
[0034] Perform linear gradient elution according to the following table:
[0035]
[0036] Analyze according to the above method, record the chromatogram, and verify the above test method.
[0037] 1.1 Specificity
[0038] Table 1 Results of specificity experiment determination
[0039] Retention time (min) Theoretical plate number USP Resolution Impurity A 16.069 40995 - Levocetirizine Hydrochloride 25.289 216545 64.951
[0040] As can be seen from Table 1, the elution peak order is impurity A and levocetirizine hydrochloride in turn, and the resolution between the two is greater than 1.5. The peak resolution of levocetirizine hydrochloride and impurity A in the chromatogram is good and the specificity is strong.
[0041] 1.2 Quantitation limit and detection limit
[0042] Dilute the levocetirizine hydrochloride reference solution and the impurity A reference solution step by step, perform the detection according to the method of Example 1, record the chromatogram, and determine the detection limit of each related substance according to the signal-to-noise ratio S / N = 3. Finally, determine the quantitation limit according to the detection result according to the signal-to-noise ratio S / N = 10. The results are shown in Table 2 below.
[0043] Table 2 Detection Limits and Quantitation Limits of Each Related Substance
[0044] Related substances Detection limit ug / mL Quantitation limit ug / mL Levocetirizine Hydrochloride 0.016 0.047 Impurity A 0.012 0.020
[0045] The detection limit and quantitation limit results of each related substance of the method of the present invention all meet the verification requirements.
[0046] 1.3 Accuracy
[0047] Take the levocetirizine hydrochloride reference substance, weigh it accurately, dissolve it with a solvent and quantitatively dilute it to prepare sample solutions with impurity A concentrations of 8 μg / mL, 10 μg / mL, and 12 μg / mL. There are 3 portions for each concentration. The solvent is a mixture of acetonitrile and water in a volume ratio of 50:50. Take the impurity A reference substance, weigh it accurately, dissolve it with a solvent and quantitatively dilute it to prepare sample solutions with impurity A concentrations of 8 μg / mL, 10 μg / mL, and 12 μg / mL. There are 3 portions for each concentration. The solvent is a mixture of acetonitrile and water in a volume ratio of 50:50. Perform high-performance liquid chromatography analysis according to the method of Example 1 above. At the same time, use the blank solution as a control, record the chromatogram respectively, calculate the recovery rate by the external standard method and calculate the RSD value. The results are shown in the table below.
[0048] Table 3 Results of the Accuracy Test for Related Substances
[0049]
[0050]
[0051] The above results show that the recovery rates of the samples at three concentrations of levocetirizine hydrochloride are 99.69 - 104.50%, and the RSD is 1.05%. The recovery rates of the samples at three concentrations of impurity A are 99.72 - 104.97%, meeting the requirements, and the RSD is 2.66% which is less than 5%. It shows that the recovery rate of the method of the present invention is good and the accuracy of this method is good.
[0052] 1.4 Precision
[0053] 1.4.1 Repeatability
[0054] According to the method of Example 1 above, inject the levocetirizine hydrochloride reference solution, the impurity A reference solution, and the test solution for detection respectively. Each sample is tested 6 times, and the results are averaged. The results are shown in the table below.
[0055] Table 4 Results of Repeatability Experiment
[0056]
[0057] As can be seen from the above results, for 6 parallel determinations, the RSDs of levocetirizine hydrochloride and impurity A are both less than 5%. The results of the repeatability test meet the verification requirements, indicating that this method has good repeatability.
[0058] 1.4.2 Intermediate Precision
[0059] The contents of levocetirizine hydrochloride and impurity A were detected by experiment operator A and experiment operator B respectively at different times using different liquid chromatographs according to the method of Example 1, and the results of the intermediate precision test were given.
[0060] Table 5 Results of Intermediate Precision Test
[0061]
[0062]
[0063] As can be seen from the above table, when detected by different experiment operators at different times and using different instruments, the RSD values of levocetirizine hydrochloride and impurity A are both less than 5%. The detection precision of the method of Example 1 of the present invention is good.
[0064] 1.5 Robustness
[0065] 1.5.1 Solution Stability
[0066] The test solution, the reference solution of levocetirizine hydrochloride, and the reference solution of impurity A were respectively placed in a refrigerator for refrigeration for 24 h, 48 h, 7 d, 10 d, 20 d, and placed at room temperature for 24 h, and then chromatographic injection analysis was carried out according to the above method of the present invention. The results show that when the test solution, the reference solution of levocetirizine hydrochloride, and the reference solution of impurity A are stored in the above environment, the RSDs of the peak areas of levocetirizine hydrochloride and impurity A are both less than 1%, and the solution stability is good.
[0067] 1.5.2 Degradation Test
[0068] Take the test solution, the reference solution of levocetirizine hydrochloride, and the reference solution of impurity A as sample solutions for oxidation (take 1 ml of each sample solution into a beaker, add hydrogen peroxide with a mass concentration of 30% to make the concentration of the sample solution 3%, and measure after standing for 1 h), acid degradation (take 1 ml of each sample solution into a beaker and add 100 μL of 1 mol / L -1 hydrochloric acid solution, and after standing for 1 h, 3 h, and 6 h, add 1 mol / L -1100 μL of sodium hydroxide solution was taken, mixed well, and then 20 μL was precisely measured and injected into the solution for analysis. At the same time, a blank comparison was made), alkali degradation (1 mL of the sample solution was taken into a beaker and 1 mol.L -1 100 μL of hydrochloric acid sodium hydroxide solution was added, and after standing for 1 h, 3 h, and 6 h, 1 mol.L -1 100 μL of hydrochloric acid solution was added, mixed well, and then 20 μL was precisely measured and injected into the solution for analysis. At the same time, a blank comparison was made), light exposure (the sample solution was irradiated under ultraviolet light at 365 nm for 30 min, 1 h, and 4 h), high temperature (the sample solution was placed at 60 °C for 3 h). After being destroyed under different conditions, the method of Example 1 of the present invention was used for detection. The results showed that after being destroyed under the above conditions, levocetirizine hydrochloride and impurity A could be well separated, and the purity of the main peak met the requirements. Under the above conditions, the recovery rate of levocetirizine hydrochloride was between 98.46% and 100.5%, and the recovery rate of impurity A was between 95.73% and 100.4%, indicating that the method of the present invention had good stability.
[0069] Comparative Example 1
[0070] The difference between this comparative example and the example was that the composition and flow rate of the mobile phase were changed, and the rest was the same as in Example 1. Mobile phase A of this comparative example was a 0.05 mol / L potassium dihydrogen phosphate buffer solution, and mobile phase B was methanol; the flow rate of the mobile phase was 1.5 mL / min.
[0071] The results showed that impurity A was affected by adjacent impurity peaks and could not be effectively separated.
[0072] Comparative Example 2
[0073] The difference between this comparative example and Example 1 was that the gradient elution process was changed, and the rest was the same as in Example 1.
[0074] The linear gradient elution program of this comparative example was:
[0075] .
[0077] The results showed that levocetirizine hydrochloride and impurity A could not be well separated and were also easily interfered by the solvent peak. The above results indicated that the method of the present invention could effectively detect the related substance p-chlorodiphenylmethylpiperazine in levocetirizine hydrochloride granules.
[0078] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included in the protection scope of the present invention.
Claims
1. A method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules, characterized in that: The following steps are involved: The method adopts high performance liquid chromatography to determine the related substances in the levocetirizine hydrochloride particles; the chromatographic column in the method is a chromatographic column with octadecylsilane bonded silica gel as a filler; the mobile phase is a mixed solution of mobile phase A and mobile phase B, phosphoric acid aqueous solution is used as mobile phase A, acetonitrile-methanol solution is used as mobile phase B, linear gradient elution is adopted, the concentration of mobile phase A is adjusted within the range of 75% v / v-20% v / v, the concentration of mobile phase B is adjusted within the range of 80% v / v-25% v / v, and the elution time is 60-70 minutes.
2. The method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules according to claim 1, characterized in that: The phosphoric acid aqueous solution is obtained by adjusting the pH value of the aqueous solution to 2.8-3.0 with phosphoric acid; and the volume ratio of acetonitrile to methanol in the acetonitrile-methanol solution is 70-80:20-30.
3. The method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules according to claim 1, characterized in that: The linear gradient elution program is:
4. The method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules according to claim 1, characterized in that: The specification of the chromatographic column is Inertsil ODS-3, 4.6mm×250mm, 3μm or a chromatographic column with equivalent performance; the column temperature of the chromatographic column is 30-40°C; and the flow rate of the mobile phase is 0.8-1.0mL / min.
5. The method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules according to claim 1, characterized in that: The detection wavelength of the ultraviolet detector in the high performance liquid chromatography is 220-240 nm; the injection volume in the high performance liquid chromatography is 10-20 μL.
6. The method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules according to claim 1, characterized in that: The preparation method of the test solution is as follows: take the test sample, accurately weigh it, place it in a volumetric bottle, add a solvent to dissolve it ultrasonically, place it to room temperature, dilute it to the scale with the solvent, shake it well, filter it, and take the filtrate to obtain the test solution; the solvent is a mixture of acetonitrile and water in a volume ratio of 45-50:50-55.
7. The method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules according to claim 1, wherein the preparation method of the levocetirizine hydrochloride reference solution is as follows: taking a levocetirizine hydrochloride reference substance, accurately weighing it, adding a solvent to dissolve it and quantitatively diluting it to prepare a solution with a levocetirizine hydrochloride concentration of 2-4 μg / mL, thereby obtaining the levocetirizine hydrochloride reference substance solution; the solvent is prepared by mixing acetonitrile and water in a volume ratio of 45-50:50-55.
8. The method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules according to claim 1, characterized in that: The preparation of the impurity A reference solution is as follows: take the impurity A reference, accurately weigh it, add a solvent to dissolve it and quantitatively dilute it to make a solution with an impurity A concentration of 10-12 μg / mL, thereby obtaining the impurity A reference solution; the solvent is a mixture of acetonitrile and water in a volume ratio of 45-50:50-55.
9. The method for detecting p-chlorobenzhydrylpiperazine in levocetirizine hydrochloride granules according to claim 1, characterized in that: The blank solution is prepared by mixing acetonitrile and water in a volume ratio of 45-50:50-55; the impurity A is p-chlorobenzhydrylpiperazine.