CN103412064A - Method for detecting impurities of DL-2-Chlorophenylglycine through high performance liquid chromatograph - Google Patents
Method for detecting impurities of DL-2-Chlorophenylglycine through high performance liquid chromatograph Download PDFInfo
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- CN103412064A CN103412064A CN2013103164314A CN201310316431A CN103412064A CN 103412064 A CN103412064 A CN 103412064A CN 2013103164314 A CN2013103164314 A CN 2013103164314A CN 201310316431 A CN201310316431 A CN 201310316431A CN 103412064 A CN103412064 A CN 103412064A
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Abstract
The invention discloses a method for detecting impurities of DL-2-Chlorophenylglycine through high performance liquid chromatograph. The method specifically comprises the following steps: (1), determining that the chromatographic column is c18/250*4.6mm/5 mmu m, the column temperature is 30 DEG C, the wavelength is 254 nm, the flow rate is 0.8 mL/min, and the sample size is 10 mmu L; (2) mobile phase A detection: mixing 20 mmol/L of potassium dihydrogen phosphate solution with the volume of 95% and methanol with the volume of 5%, adjusting the pH value to be 2.5 with phosphate; and mobile phase B detection: mixing acetonitrile with the volume of 50% and methanol with the volume of 50%; (3) measuring and recording chromatogram. Through the method provided by the invention, all impurities of DL-2-Chlorophenylglycine, in particular to chlorobenzene glycine (impurity B) and 0-chlorphenyl glycinamide (impurity C), can be better separated, and the repeatability is good; the successful development of the method has important realistic significance in raw material control and quality assurance in production of bulk drug of clopidogrel.
Description
Technical field
The invention belongs to the organic analysis technical field, particularly a kind of method that detects the o-chlorobenzene glycine impurities of one of clopidogrel raw materials by high performance liquid chromatography.
Background technology
Bisulfate clopidogrel (Clopidogrel Hydrogensulfate) is the RA233 of new generation of being succeeded in developing in 1986 by French Sai Nuofei (Sanofi) company, can be used for prevention of arterial atherosis and the treatment acute myocardial infarction AMI, commodity are called " Plavix ".This product took the lead in, in U.S. listing, entering subsequently the multinational markets such as Europe, North America, Australia, Singapore in March, 1998, and August calendar year 2001 at Discussion on Chinese Listed.It belongs to the Thienopyridines medicine, is a kind of novel adenosine diphosphate (ADP) receptor antagonist, because have good security and suppress fast hematoblastic advantage, becomes one of antiplatelet drug of widespread use thereby replaced ticlopidine.
The preparation method of clopidogrel has a variety of, and the o-chlorobenzene glycine (S) of wherein take is the reactions steps such as esterification, fractionation, condensation, cyclization and salify as the synthetic route of initiation material.
In order to improve the quality of pharmaceutical production, according to pharmaceutical production standard and product standard, as the regulation of American Pharmacopeia (USP) and European Pharmacopoeia (EP), need to carry out effective analysis and control to the quality of primary raw material o-chlorobenzene glycine (S).Analyze preparation and the degradation process of o-chlorobenzene glycine, several impurity compositions that may exist are:
Current, analyze and detect o-chlorobenzene glycine (S) and mostly by efficient liquid-phase chromatography method, realize, its chromatographic process is generally: chromatographic column (C18/250 * 4.6mm/5 μ m), column temperature (25 ℃), wavelength (254nm), flow velocity (1.0mL/min) and sample size (20 μ L); Selected mobile phase is: water/acetonitrile/tetrahydrochysene furan (85/9/6), and wherein water is the aqueous dibasic potassium phosphate solution 800mL of 20mmol/L, hydrochloric acid is adjusted to pH=2.5, and to be diluted to volume be 1000mL.
Detect according to the method described above substrate o-chlorobenzene glycine (S) sample that contains p-chlorophenylglycine (impurity B) and adjacent chlorobenzene glycine amide (impurity C), discovery can only show an impurity peaks, and namely impurity B can not be separated (seeing Fig. 1) well with C under this testing conditions.
In order to verify the above results, reference substance mixing sample introduction by p-chlorophenylglycine (impurity B) and adjacent chlorobenzene glycine amide (impurity C), the result obtained still is unimodal (seeing Fig. 2), further proved thus the high-efficiency liquid chromatography method for detecting of current general o-chlorobenzene glycine (S), for impurity, separate, especially for p-chlorophenylglycine (impurity B) and separating of adjacent chlorobenzene glycine amide (impurity C) having major defect, even under this system, can't effectively isolate these two impurity.
Summary of the invention
In order to overcome the defect of prior art, the object of the present invention is to provide a kind of improved high performance liquid chromatography that utilizes to detect the method for preparing clopidogrel raw material o-chlorobenzene glycine (S) impurities.This detection method can be separated the plurality of impurities of o-chlorobenzene glycine (S) effectively, particularly realizes efficient separation of p-chlorophenylglycine (impurity B) and adjacent chlorobenzene glycine amide (impurity C).
To achieve these goals, the present invention mainly provides following technical scheme: a kind of method that detects o-chlorobenzene glycine impurity by high performance liquid chromatography is characterized in that described detection method comprises the steps: (1) selection definite chromatographic column, column temperature, wavelength, flow velocity and sample size; (2) preparation mobile phase definite gradient program; And chromatogram is measured and recorded in (3).
In addition, the present invention also comprises following attached technical scheme:
Described chromatographic column is C18/250 * 4.6mm/5 μ m.
Described column temperature 30-50 ℃, preferably 30 ℃.
Described mensuration wavelength is 245-260nm, preferably 254nm.
Described flow velocity is 0.5-1.5mL/min, preferably 0.8mL/min.
Described mobile phase A is after the potassium dihydrogen phosphate of the 20mmol/L of 90-99% volume mixes with the methyl alcohol of 10-1% volume, to use phosphoric acid adjust pH to 2.5, and preferably the potassium dihydrogen phosphate of the 20mmol/L of 95% volume is used phosphoric acid adjust pH to 2.5 after with the methyl alcohol of 5% volume, mixing.
Described Mobile phase B is 25-75% volumes of acetonitrile and 75-25% volumes methanol, preferably 50% volumes of acetonitrile and 50% volumes methanol.
The gradient time-program(me) of described mobile phase is 100%-0%/0-40min.
Than prior art, the involved in the present invention high performance liquid chromatography that utilizes detects the method for o-chlorobenzene glycine (S) impurities of one of clopidogrel raw materials.This detection method can be separated the impurity of o-chlorobenzene glycine effectively, particularly realize efficient separation of p-chlorophenylglycine (impurity B) and adjacent chlorobenzene glycine amide (impurity C), the method degree of separation is high, favorable reproducibility, has important practical significance for the control of the raw material in the clopidogrel production of raw medicine and quality assurance.
The accompanying drawing explanation
Fig. 1 measures the high-efficient liquid phase chromatogram of the o-chlorobenzene glycine (S) that contains impurity B and C according to a kind of traditional detection method
Fig. 2 measures the high-efficient liquid phase chromatogram of impurity B and impurity C reference substance potpourri according to another kind of traditional detection method.
Fig. 3 is the gradient time-program(me) corresponding to the mobile phase of detection method of the present invention.
Fig. 4 measures the high-efficient liquid phase chromatogram of the o-chlorobenzene glycine (S) that contains impurity B and C according to detection method of the present invention.
Fig. 5 measures the high-efficient liquid phase chromatogram of impurity B and impurity C reference substance potpourri according to detection method of the present invention.
Fig. 6 measures the high-efficient liquid phase chromatogram of the o-chlorobenzene glycine (S) that contains impurity A, B, C, D, E and F according to the inventive method.
Embodiment
Below will set forth the impurity that how detects o-chlorobenzene glycine by high performance liquid chromatography by several concrete testing processes and spectrogram.
(1) chromatographic condition
Chromatographic column: C18,250 * 4.6mm, 5 μ m
Column temperature: 30 ℃
Wavelength: 254nm
Flow velocity: 0.8mL/min
Sample size: 10 μ L
(2) mobile phase
Mobile phase A: the potassium dihydrogen phosphate of the 20mmol/L of 95% volume, the methyl alcohol of 5% volume, use phosphoric acid adjust pH to 2.5 after mixing.Mobile phase B: the methyl alcohol of the acetonitrile of 50% volume and 50% volume.
The gradient time-program(me) can be referring to Fig. 3.
(3) record collection of illustrative plates
Embodiment mono-:
With reference to Fig. 4, in o-chlorobenzene glycine (S), contain p-chlorophenylglycine (impurity B) and adjacent chlorobenzene glycine amide (impurity C).
Embodiment bis-:
With reference to Fig. 5, impurity reference substance checking: p-chlorophenylglycine (impurity B) and adjacent chlorobenzene glycine amide (impurity C).
Embodiment tri-:
With reference to Fig. 6, contain impurity A, B, C, D, E and F in o-chlorobenzene glycine (S).
It is pointed out that above-mentioned preferred embodiment only is explanation technical conceive of the present invention and characteristics, its purpose is to allow the person skilled in the art can understand content of the present invention and implement according to this, can not limit the scope of the invention with this.All equivalences that Spirit Essence is done according to the present invention change or modify, within all should being encompassed in protection scope of the present invention.
Claims (8)
1. a method that detects o-chlorobenzene glycine impurity by high performance liquid chromatography, is characterized in that described detection method comprises the steps:
(1) selection definite chromatographic column, column temperature, wavelength, flow velocity and sample size;
(2) preparation mobile phase definite gradient program; And
(3) measure and record chromatogram.
2. by high performance liquid chromatography, detect according to claim 1 the method for o-chlorobenzene glycine impurity, it is characterized in that: described chromatographic column is C18/250 * 4.6mm/5 μ m.
3. by high performance liquid chromatography, detect according to claim 1 the method for o-chlorobenzene glycine impurity, it is characterized in that: described column temperature 30-50 ℃.
4. by high performance liquid chromatography, detect according to claim 1 the method for o-chlorobenzene glycine impurity, it is characterized in that: described mensuration wavelength is 245-260nm.
5. by high performance liquid chromatography, detect according to claim 1 the method for o-chlorobenzene glycine impurity, it is characterized in that: described flow velocity is 0.5-1.5mL/min.
6. by high performance liquid chromatography, detect according to claim 1 the method for o-chlorobenzene glycine impurity, it is characterized in that: described mobile phase A is with phosphoric acid adjust pH to 2.5 after the potassium dihydrogen phosphate of the 20mmol/L of 90-99% volume mixes with the methyl alcohol of 10-1% volume.
7. by high performance liquid chromatography, detect according to claim 1 the method for o-chlorobenzene glycine impurity, it is characterized in that: described Mobile phase B is 25-75% volumes of acetonitrile and 75-25% volumes methanol.
8. by high performance liquid chromatography, detect according to claim 1 the method for o-chlorobenzene glycine impurity, it is characterized in that: the gradient time-program(me) of described mobile phase is 100%-0%/0-40min.
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Cited By (4)
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| CN106220634A (en) * | 2016-08-03 | 2016-12-14 | 苏州立新制药有限公司 | Related substances F and G of pemetrexed disodium and preparation thereof and detection method |
| CN106596799A (en) * | 2017-01-20 | 2017-04-26 | 阳泉煤业（集团）有限责任公司 | Method for detecting glycine and impurities thereof by high performance liquid chromatography |
| CN108760940A (en) * | 2018-08-03 | 2018-11-06 | 安徽省金楠医疗科技有限公司 | A kind of bisulfate clopidogrel detection method |
| CN114689737A (en) * | 2021-12-31 | 2022-07-01 | 浙江车头制药股份有限公司 | Analysis method of S-o-chlorophenyl glycine methyl ester tartrate related substances |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106220634A (en) * | 2016-08-03 | 2016-12-14 | 苏州立新制药有限公司 | Related substances F and G of pemetrexed disodium and preparation thereof and detection method |
| CN106596799A (en) * | 2017-01-20 | 2017-04-26 | 阳泉煤业（集团）有限责任公司 | Method for detecting glycine and impurities thereof by high performance liquid chromatography |
| CN108760940A (en) * | 2018-08-03 | 2018-11-06 | 安徽省金楠医疗科技有限公司 | A kind of bisulfate clopidogrel detection method |
| CN114689737A (en) * | 2021-12-31 | 2022-07-01 | 浙江车头制药股份有限公司 | Analysis method of S-o-chlorophenyl glycine methyl ester tartrate related substances |
| CN114689737B (en) * | 2021-12-31 | 2023-12-08 | 浙江车头制药股份有限公司 | Analysis method of S-o-chlorophenylglycine methyl tartrate related substances |
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Effective date of registration: 20190308 Address after: 255000 Jinyang Road, Gaoqing Economic Development Zone, Zibo City, Shandong Province Patentee after: Shandong Lixin Pharmaceutical Co., Ltd. Address before: 215151 No. 21 Tangxi Road, Suzhou High-tech Zone, Jiangsu Province Patentee before: Suzhou Lixin Pharmacy Co., Ltd. |
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Denomination of invention: Method for detecting impurities of DL-2-Chlorophenylglycine through high performance liquid chromatograph Effective date of registration: 20190422 Granted publication date: 20150520 Pledgee: Hengfeng Bank Co., Ltd. Zibo Branch Pledgor: Shandong Lixin Pharmaceutical Co., Ltd. Registration number: 2019370000082 |
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Date of cancellation: 20201111 Granted publication date: 20150520 Pledgee: Hengfeng Bank Co.,Ltd. Zibo Branch Pledgor: Shandong Lixin Pharmaceutical Co.,Ltd. Registration number: 2019370000082 |