CN101393185B - Method for analytically separating clopidogrel and enantiomer thereof by HPLC method - Google Patents
Method for analytically separating clopidogrel and enantiomer thereof by HPLC method Download PDFInfo
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- CN101393185B CN101393185B CN 200710122128 CN200710122128A CN101393185B CN 101393185 B CN101393185 B CN 101393185B CN 200710122128 CN200710122128 CN 200710122128 CN 200710122128 A CN200710122128 A CN 200710122128A CN 101393185 B CN101393185 B CN 101393185B
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Abstract
The invention discloses a mensuration method for analyzing and separating a clopidogrel midbody and an enantiomer of the clopidogrel midbody. In the method, a cellulose-type chiral column is adopted; and a mixed solution of acetonitrile and a phosphoric acid/phosphate buffer (the buffer is regulated to a certain PH value) is used as a mobile phase; and the clopidogrel or enantiomeric impurity containing clopidogrel preparation can be rapidly separated and analyzed.
Description
Technical field
The invention belongs to and relate to a kind of high performance liquid chromatography, especially a kind of high performance liquid chromatography of separating clopidogrel intermediate and enantiomter thereof of analyzing.
Background technology
Clopidogrel is a kind of anticoagulant, and the structural formula of a kind of intermediate in its preparation process is
Molecular formula is C
16H
16CINO
2S, chemistry (S)-α by name-(2-chlorphenyl)-6, the 7-dihydro-thiophene is [3,2-c]-pyridine-5 (4H)-methyl acetate also, contain 1 asymmetric carbon atom in the molecule of this intermediate, in the process by directed synthesising target compound clopidogrel, need to control the content of its enantiomter.
For the enantiomter impurity of clopidogrel intermediate, in the middle of the medicine building-up process, need carry out quality control.The separation that contains the enantiomter of asymmetric carbon atom is the difficult point of quality control in the synthetic and preparation process of chiral drug always, realizes that the quality control aspect synthetic and the preparation process that is separated in the clopidogrel medicine of clopidogrel intermediate and corresponding isomeride thereof has realistic meaning.
Summary of the invention
The object of the present invention is to provide a kind of efficient liquid-phase chromatography method that separates clopidogrel intermediate and enantiomter impurity thereof of analyzing, thereby realize clopidogrel intermediate and its enantiomter separate impurities mensuration.
The applicant finds, with the cellulose family chiral column, be that the mixed solution of phosphoric acid/phosphate buffer of 2-4 is the phase that flows with acetonitrile and pH value, clopidogrel intermediate and enantiomter thereof effectively can be separated, thereby can accurately control the quality of clopidogrel.Method of the present invention can be analyzed simply, quickly and accurately and separate clopidogrel intermediate and enantiomter impurity thereof.
Further, (the cellulose family chiral column of DACEL150mm * 4.6mm) can obtain better to analyze separating effect to select OD-RH for use.OD-RH cellulose family chiral column is to be filler with (cellulose iii (3,5-xylyl carbamate)).
Further, the phosphate in phosphoric acid/phosphate buffer of the present invention is selected from ammonium dihydrogen phosphate (ADP) or sodium dihydrogen phosphate, most preferably ammonium dihydrogen phosphate (ADP).
It is the mixed solution of 25: 75 acetonitriles and phosphoric acid/phosphate buffer that flowing in the method for the present invention selected volume ratio mutually for use.
The mobile pH value of middle phosphoric acid/phosphate buffer mutually in the method for the present invention is 2.5.
Analysis separation method of the present invention, can realize in accordance with the following methods:
(1) it is an amount of to get the clopidogrel intermediate sample, with methyl alcohol or isopropyl alcohol sample dissolution, is mixed with the sample solution of the chloride pyrrole Gray of every 1mL 0.1mg～0.5mg.
(2) flow rate of mobile phase being set is 0.4～1.2mL/min, and the detection wavelength is 205～235nm, and the optimum detection wavelength is 225nm, and the chromatographic column column oven is 20～40 ℃, and the optimum temperature of chromatographic column column temperature is room temperature.
(3) the sample solution 2-50 μ L that gets (1) injects liquid chromatograph, the analysis of finishing clopidogrel intermediate and enantiomter thereof with separate.
Wherein:
High performance liquid chromatograph: Tianjin, island: LC-10ATvp, SPD-M10Avp, SCL-10Avp, DGU-12A
Chromatographic column: the OD-RH (chiral chromatographic column of DACEL150mm * 4.6mm)
Phase flows: acetonitrile-(0.01mol/L sodium dihydrogen phosphate, phosphoric acid,diluted is transferred pH to 2.5)=25: 75
Flow velocity: 1.0mL/min
Detect wavelength: 225nm
Column temperature: room temperature
Sampling volume: 10 μ L
The present invention adopts cellulose family OD-RH, and (chiral chromatographic column of DACEL150mm * 4.6mm) can effectively be analyzed and separate clopidogrel intermediate and enantiomter (impurity) thereof; Select the phased soln sample that flows, guaranteed the stability of solution; Select sampling volume 10 μ L, column temperature is room temperature, has improved the symmetry of chromatographic peak.The invention solves the raw material that contains clopidogrel intermediate and enantiomter thereof and analysis and the separation problem of preparation, thereby guaranteed the quality controllable of clopidogrel and preparation thereof.
Description of drawings
Fig. 1 embodiment 1, the HPLC figure of acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid transfer pH2.5)=25: 75;
Fig. 2 embodiment 2, the HPLC figure of acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid transfer pH3.0)=25: 75
Fig. 3 embodiment 3, the HPLC figure of acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid transfer pH4.0)=25: 75
Fig. 4 embodiment 4, the HPLC figure of acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid transfer pH3.5)=30: 70
Embodiment:
Instrument and condition
High performance liquid chromatograph: Tianjin, island: LC-10ATvp, SPD-M10Avp, SCL-10Avp, DGU-12A;
Chromatographic column: OD-RH (DACEL150mm * 4.6mm);
Phase flows: acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid is transferred pH2.5)=25: 75
Flow velocity: 1.0mL/min;
Column temperature: room temperature;
Sampling volume: 10 μ L.
Detect wavelength: 225nm;
Experimental procedure
Take by weighing each 5mg of clopidogrel intermediate and enantiomter thereof respectively, place the 25mL volumetric flask, add mobile phased soln and dilution and put scale, shake up, as biased sample solution.And take by weighing each 5mg of its enantiomter of clopidogrel intermediate respectively, and place 2 25mL volumetric flasks respectively, add mobile phased soln and dilution and put scale, shake up, as qualitative contrast solution.
Get each contrast solution and biased sample solution respectively, carry out efficient liquid phase chromatographic analysis by above-mentioned condition, the record chromatogram.The results are shown in accompanying drawing 1, No. 1 peak is clopidogrel among the figure, and No. 2 peaks are isomeride, and the clopidogrel main peak can be separated fully with isomeride under this condition as can be seen, and the clopidogrel main peak is about 33.9min.
Instrument and condition
High performance liquid chromatograph: Tianjin, island: LC-10ATvp, SPD-M10Avp, SCL-20Avp, DGU-12A;
Chromatographic column: OD-RH (DACEL150mm * 4.6mm);
Phase flows: acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid is transferred pH3.0)=25: 75;
Flow velocity: 1.0mL/min;
Detect wavelength: 225nm;
Column temperature: room temperature;
Sampling volume: 10 μ L.
Experimental procedure
Take by weighing each 5mg of clopidogrel intermediate and enantiomter thereof respectively, place the 25mL volumetric flask, add mobile phased soln and dilution and put scale, shake up, as biased sample solution.And take by weighing each 5mg of its enantiomter of clopidogrel intermediate respectively, and place 2 25mL volumetric flasks respectively, add mobile phased soln and dilution and put scale, shake up, as qualitative contrast solution.
Get each contrast solution and biased sample solution respectively, carry out efficient liquid phase chromatographic analysis by above-mentioned condition, the record chromatogram.The results are shown in accompanying drawing 2, No. 1 peak is clopidogrel among the figure, and No. 2 peaks are isomeride, and the clopidogrel main peak can be separated fully with isomeride under this condition as can be seen, and the clopidogrel main peak is about 33.9min.
Embodiment 3
Instrument and condition
High performance liquid chromatograph: Tianjin, island: LC-10ATvp, SPD-M10Avp, SCL-10Avp, DGU-12A;
Chromatographic column: OD-RH (DACEL150mm * 4.6mm);
Phase flows: acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid is transferred pH4.0)=25: 75;
Flow velocity: 1.0mL/min;
Detect wavelength: 225nm;
Column temperature: room temperature;
Sampling volume: 10 μ L.
Experimental procedure
Take by weighing each 5mg of clopidogrel intermediate and enantiomter thereof respectively, place the 25mL volumetric flask, add mobile phased soln and dilution and put scale, shake up, as biased sample solution.And take by weighing each 5mg of its enantiomter of clopidogrel intermediate respectively, and place 2 25mL volumetric flasks respectively, add mobile phased soln and dilution and put scale, shake up, as qualitative contrast solution.
Get each contrast solution and biased sample solution respectively, carry out efficient liquid phase chromatographic analysis by above-mentioned condition, the record chromatogram.The results are shown in accompanying drawing 3, No. 1 peak is clopidogrel among the figure, and No. 2 peaks are isomeride, and the clopidogrel main peak can be separated fully with isomeride under this condition as can be seen, and the clopidogrel main peak is about 33.9min.
Embodiment 4
Instrument and condition
High performance liquid chromatograph: Tianjin, island: LC-10ATvp, SPD-M10Avp, SCL-10Avp, DGU-12A;
Chromatographic column: OD-RH (DACEL150mm * 4.6mm);
Phase flows: acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid is transferred pH3.5)=30: 70;
Flow velocity: 1.0mL/min;
Detect wavelength: 225nm;
Column temperature: room temperature;
Sampling volume: 10 μ L.
Experimental procedure
Take by weighing each 5mg of clopidogrel intermediate and enantiomter thereof respectively, place the 25mL volumetric flask, add mobile phased soln and dilution and put scale, shake up, as biased sample solution.And take by weighing each 5mg of its enantiomter of clopidogrel intermediate respectively, and place 2 25mL volumetric flasks respectively, add mobile phased soln and dilution and put scale, shake up, as qualitative contrast solution.
Get each contrast solution and biased sample solution respectively, carry out efficient liquid phase chromatographic analysis by above-mentioned condition, the record chromatogram.The results are shown in accompanying drawing 4, No. 1 peak is clopidogrel among the figure, and No. 2 peaks are isomeride, as can be seen under this condition the clopidogrel main peak can with the isomeride baseline separation, and the clopidogrel main peak is about 29.2min.
Claims (4)
1. a high-efficient liquid phase chromatogram technique analysis separates the method for clopidogrel intermediate and enantiomter thereof, it is characterized in that:
High performance liquid chromatograph: Tianjin, island: LC-10ATvp, SPD-M10Avp, SCL-10Avp, DGU-12A;
Chromatographic column: OD-RH (DACEL150mm * 4.6mm);
Phase flows: acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid is transferred pH2.5)=25: 75;
Flow velocity: 1.0mL/min;
Column temperature: room temperature;
Sampling volume: 10 μ L;
Detect wavelength: 225nm;
Experimental procedure:
Take by weighing each 5mg of clopidogrel intermediate and enantiomter thereof respectively, place the 25mL volumetric flask, add mobile phased soln and dilution and put scale, shake up, as biased sample solution; And take by weighing each 5mg of its enantiomter of clopidogrel intermediate respectively, and place 2 25mL volumetric flasks respectively, add mobile phased soln and dilution and put scale, shake up, as qualitative contrast solution;
Get each contrast solution and biased sample solution respectively, carry out efficient liquid phase chromatographic analysis by above-mentioned condition, the record chromatogram.
2. a high-efficient liquid phase chromatogram technique analysis separates the method for clopidogrel intermediate and enantiomter thereof, it is characterized in that:
Tianjin, high performance liquid chromatograph island: LC-10ATvp, SPD-M10Avp, SCL-10Avp, DGU-12A;
Chromatographic column: OD-RH (DACEL150mm * 4.6mm);
Phase flows: acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid is transferred pH3.0)=25: 75;
Flow velocity: 1.0mL/min;
Detect wavelength: 225nm;
Column temperature: room temperature:
Sampling volume: 10 μ L;
Experimental procedure:
Take by weighing each 5mg of clopidogrel intermediate and enantiomter thereof respectively, place the 25mL volumetric flask to add to flow phased soln and dilution to put scale, shake up, as biased sample solution; And take by weighing each 5mg of its enantiomter of clopidogrel intermediate respectively, and place 2 25mL volumetric flasks respectively, add mobile phased soln and dilution and put scale, shake up, as qualitative contrast solution;
Get each contrast solution and biased sample solution respectively, carry out efficient liquid phase chromatographic analysis by above-mentioned condition, the record chromatogram.
3. a high-efficient liquid phase chromatogram technique analysis separates the method for clopidogrel intermediate and enantiomter thereof, it is characterized in that:
High performance liquid chromatograph: island rule: LC-10ATvp, SPD-M10Avp, SCL-10Avp, DGU-12A;
Chromatographic column: OD-RH (DACEL150mm * 4.6mm);
Phase flows: acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid is transferred pH4.0)=25: 75;
Flow velocity 1.0mL/min;
Detect wavelength: 225nm;
Column temperature: room temperature;
Sampling volume: 10 μ L;
Experimental procedure:
Take by weighing each 5mg of clopidogrel intermediate and enantiomter thereof respectively, place the 25mL volumetric flask, add mobile phased soln and dilution and put scale, shake up, as biased sample solution; And take by weighing each 5mg of its enantiomter of clopidogrel intermediate respectively, and place 2 25mL volumetric flasks respectively, add mobile phased soln and dilution and put scale, shake up, as qualitative contrast solution;
Get each contrast solution and biased sample solution respectively, carry out efficient liquid phase chromatographic analysis by above-mentioned condition, the record chromatogram.
4. a high-efficient liquid phase chromatogram technique analysis separates the method for clopidogrel intermediate and enantiomter thereof, it is characterized in that:
High performance liquid chromatograph: Tianjin, island: LC-10ATvp, SPD-M10Avp, SCL-10Avp, DGU-12A;
Chromatographic column: OD-RH (DACEL150mm * 4.6mm);
Phase flows; Acetonitrile-(0.01mol/L ammonium dihydrogen phosphate (ADP), phosphoric acid is transferred pH3.5)=30: 70;
Flow velocity: 1.0mL/min;
Detect wavelength: 225nm;
Column temperature: room temperature;
Sampling volume: 10 μ L;
Experimental procedure:
Take by weighing each 5mg of clopidogrel intermediate and enantiomter thereof respectively, place the 25mL volumetric flask, add mobile phased soln and rare sample and put scale, shake up, as biased sample solution; And take by weighing each 5mg of its enantiomter of clopidogrel intermediate respectively, and place 2 25mL volumetric flasks respectively, add mobile phased soln and dilution and put scale, shake up, as qualitative contrast solution;
Get each contrast solution and biased sample solution respectively, carry out efficient liquid phase chromatographic analysis by above-mentioned condition, the record chromatogram.
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| CN102043023B (en) * | 2009-10-22 | 2014-05-14 | 北京万全阳光医学技术有限公司 | Method for measuring materials associated with Ropinirole hydrochloride by high performance liquid chromatography |
| CN103308636B (en) * | 2013-04-28 | 2014-04-09 | 山东信立泰药业有限公司 | Quality control method of D-(+)-alpha-(2-thiofuran ethylamino)-alpha-(2-chlorphenyl) methyl acetate or salt thereof and application of D-(+)-alpha-(2-thiofuran ethylamino)-alpha-(2-chlorphenyl) methyl acetate or salt thereof in clopidogrel production |
| CN103412064B (en) * | 2013-07-25 | 2015-05-20 | 苏州立新制药有限公司 | Method for detecting impurities of DL-2-Chlorophenylglycine through high performance liquid chromatograph |
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| Title |
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| 张青山等.氯吡格雷中间体的合成.《精细化工》.2004,第21卷(第3期),195-196. * |
| 艾又生等.HPLC法测定硫酸氯毗格雷片的含量.《中国药师》.2006,第9卷(第12期),1100-1101. * |
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