CN101221147A - Method for detecting Raltitrexed enantiomer by capillary tube electrophoresis - Google Patents
Method for detecting Raltitrexed enantiomer by capillary tube electrophoresis Download PDFInfo
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- CN101221147A CN101221147A CNA2007101319762A CN200710131976A CN101221147A CN 101221147 A CN101221147 A CN 101221147A CN A2007101319762 A CNA2007101319762 A CN A2007101319762A CN 200710131976 A CN200710131976 A CN 200710131976A CN 101221147 A CN101221147 A CN 101221147A
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Abstract
The invention relates to a method for detecting a raltitrexed enantiomer by adopting a capillary electrophoresis method which calculates the limitation of the raltitrexed to the enantiomer in the raltitrexed according to a self-contrast method of main components by separating the raltitrexed and the enantiomer through the capillary electrophoresis; uses a solution with the volume ratio of 1 to 1 between methanol and water for preparing a sample solution; an electrophoresis buffer liquid of the capillary electrophoresis is Tris-H3PO4 with a pH value between 6.0 to 10; adds a chiral selection agent and sodium dodecyl sulfate into the buffer liquid. The method overcomes the defects that the use of a chiral flowing phase and a chiral column for separating the enantiomer is difficult and has higher cost because the molecule structures of the raltitrexed not only comprises carboxyl, but also comprises amidogen. The method has the advantages of good separating effect, no pollution, low cost and being suitable for checking the raltitrexed enantiomer.
Description
Technical field
The present invention relates to the method that a kind of capillary electrophoresis detects the Raltitrexed enantiomter.
Background technology
The inspection of chiral drug enantiomter method commonly used has chiral hplc, by in moving phase, adding chiral selector or adopting chiral chromatographic column to separate inspection, often separating effect is bad for the former, specificity is not strong, latter's chromatographic column is more expensive, cost is higher, and chromatographic column in use post is imitated very easily to descend and is caused separating effect bad, not only contain carboxyl but also contain amino in Raltitrexed and this body structure of isomeride thereof in addition, stranded easily in the molecule in the liquid-phase chromatographic analysis process, increased difficulty to separation; The inspection requirements sample that vapor-phase chromatography is used for the chiral drug enantiomter has certain volatility and thermal stability, and use has limitation very much.
Summary of the invention
For for addressing the above problem, the invention provides the method that a kind of capillary electrophoresis detects the Raltitrexed enantiomter, mainly be the single enantiomter of separating and detect Raltitrexed.
The present invention is achieved in that it mainly comprises the preparation of sample, Capillary Electrophoresis condition and analytical approach; It with Raltitrexed and stage enantiomer separation thereof, calculates the limit of its enantiomter in the Raltitrexed by Capillary Electrophoresis by main composition Self-control method; Use methyl alcohol: the preparation of water=1: 1 (volume ratio) solution is equipped with need testing solution, and the electrophoretic buffer of Capillary Electrophoresis is Tris-H
3PO
4, this pH of buffer value is 6.0～10, adds chiral selector and lauryl sodium sulfate in this damping fluid.
Damping fluid Tris-H
3PO
4Preparation be trishydroxymethylaminomethane (Tris) solution with 0.05～0.1M, be the phosphorus acid for adjusting pH value of 0.1M with concentration, the pH value of damping fluid is preferably 8.Chiral selector is alpha-cyclodextrin, beta-schardinger dextrin-, sulfonation beta-schardinger dextrin-, HP-, gamma-cyclodextrin or carboxymethyl-beta-cyclodextrin.The concentration range of chiral selector in damping fluid is 2～20mM, and chiral selector is preferably carboxymethyl-beta-cyclodextrin, and its concentration is 8mM.The deposition condition parameter is: capillary pipe length length overall 48.5cm, effective length 40cm, capillary inner diameter are 50 μ m, and working voltage is 15～30kV, and capillary temperature is 20～30 ℃, fluid mechanics sample introduction 50mbar 5 seconds.Preferred working voltage 20kV, 25 ℃ of temperature capillaceous.Electrode: by '+' to ', the detection wavelength is 228nm; The concentration that adds dodecyl base sodium sulphate in buffer solution is 0.01～0.1M, and preferred concentration is 30mM.Use methyl alcohol: water=1: 1 (volume ratio) solution preparation Raltitrexed chirality thing analytic sample, test sample concentration between 50～200 μ g/ml, the concentration 1 μ g/ml～100 μ g/ml of enantiomter.The computing method of Raltitrexed mapping allosome are preparation Raltitrexed need testing solution and the reference substance solution that contains the Raltitrexed of 1% Raltitrexed enantiomter reference substance, the difference sample introduction, the corresponding peak area of mapping structure body must not be greater than the peak area of enantiomter in the contrast solution in the Raltitrexed need testing solution.
In order to guarantee the reappearance of testing result, when kapillary uses for the first time, wash activation to it, the program of flushing is to use the NaOH of 0.1mol/ to wash earlier 15 minutes, then with high purity water flushing 10 minutes, damping fluid flushing 5 minutes.After this washed 3 minutes with running buffer before the each run.
That the inspection that capillary electrophoresis is used for the chiral drug isomeride has is efficient, quick, easy and simple to handle, environmental friendliness, amount of samples are few, cost is low, Raltitrexed and enantiomter thereof can baseline separation etc. advantage, its higher separation efficiency makes to separate selects the less enantiomter of coefficient can obtain satisfied separating effect.The kind of selective agent that is used for the Capillary Electrophoresis chiral resolution is more, wherein uses the most extensively with cyclodextrin, and obtains easily, has broad application prospects in chiral resolution.
Embodiment
Embodiment 1
Experiment is carried out on the Agilent 3D CE of U.S. Agilent Technologies company type capillary electrophoresis apparatus, fluid mechanics sample introduction, kapillary effective length 40cm, the fused quartz capillary column of internal diameter 50 μ m.The process for preparation of sample is as follows:
Trishydroxymethylaminomethane (Tris) solution of preparation 75mmol/L is that the phosphoric acid of 0.1M is regulated pH to 8.0 as damping fluid with concentration, adds the 8mmol/L carboxymethyl-beta-cyclodextrin (lauryl sodium sulfate of CM-β-CD) and 30mmol/L.Use methyl alcohol: water=1: 1 (volume ratio) solution is prepared Raltitrexed (s-raltitrexed) solution and Raltitrexed and its enantiomter (r-and s-raltitrexed) mixed solution respectively, and Raltitrexed concentration is 50 μ g/ml.Wash 3min with buffer solution successively before the sample introduction.Experimental temperature: 25 ℃, detect wavelength: 228nm, fluid mechanics sample introduction 50mbar5 second; The results are shown in Figure 1.
Deposition condition is with embodiment 1.Use methyl alcohol: the preparation of water=1: 1 (volume ratio) solution, prepare serial Raltitrexed (s-raltitrexed) and Raltitrexed enantiomter (r-raltitrexed) solution respectively, sample introduction, the recording responses value is the electrophoretogram of 3 times of baseline noises, be the detectability of Raltitrexed and enantiomter thereof, its detection is limited to 1 μ m/ml.The results are shown in Figure 2
Tris-phosphoric acid solution with 50mmol/L～200mol/L is a damping fluid, is that the phosphoric acid of 0.1M is regulated pH to 8.0 with concentration, adds 8mmol/L ethyloic beta-schardinger dextrin-(CM-β-CD) and the lauryl sodium sulfate of 30mmol/L; Other the results are shown in Figure 3 with embodiment 1.
1, a kind of inspection method that is used for chiral drug Raltitrexed enantiomter.
2, its method is a high performance capillary electrophoresis
The present invention is used to check the single enantiomter of Raltitrexed, and method is simple, the separation efficiency height, and velocity of separation is fast, and sample size is few, and analysis cost is low.General some reagent commonly used that only need, all reagent dosages are all less, and are more much lower than analyzing cost with the high performance liquid chromatogram method.Its useful effect mainly shows:
(1) amount of samples is few: each required sample of sample introduction has only several nL;
(2) environmental friendliness: used damping fluid is aqueous solution, need not with an organic solvent, than the more environmental protection of HPLC method;
(3) analysis cost is low: required capillary column and reagent are much lower with respect to HPLC chiral column cost.
Fig. 1 is the testing result figure of embodiment one
Fig. 2 is the testing result figure of embodiment two
Fig. 3 is the testing result figure of embodiment three
1, a kind of inspection method that is used for chiral drug Raltitrexed enantiomter.
2, its method is a high performance capillary electrophoresis
The present invention is used to check the single enantiomter of Raltitrexed, and method is simple, the separation efficiency height, and velocity of separation is fast, and sample size is few, and analysis cost is low.General some reagent commonly used that only need, all reagent dosages are all less, and are more much lower than analyzing cost with the high performance liquid chromatogram method.Its useful effect mainly shows:
(1) amount of samples is few: each required sample of sample introduction has only several nL;
(2) environmental friendliness: used damping fluid is aqueous solution, need not with an organic solvent, than the more environmental protection of HPLC method;
(3) analysis cost is low: required capillary column and reagent are much lower with respect to HPLC chiral column cost.
Claims (7)
1. the method for a capillary electrophoresis detection Raltitrexed enantiomter mainly comprises the preparation of sample, Capillary Electrophoresis condition and analytical approach; It with Raltitrexed and stage enantiomer separation thereof, calculates the limit of its enantiomter in the Raltitrexed by Capillary Electrophoresis by main composition Self-control method; Use methyl alcohol: the preparation of water=1: 1 (volume ratio) solution is equipped with need testing solution, and the electrophoretic buffer of Capillary Electrophoresis is Tris-H
3PO
4, this pH of buffer value is 6.0～10, adds chiral selector and lauryl sodium sulfate in this damping fluid.
2. a kind of capillary electrophoresis according to claim 1 detects the method for Raltitrexed enantiomter, it is characterized in that: damping fluid Tris-H
3PO
4Preparation be trishydroxymethylaminomethane (Tris) solution with 0.05～0.1M, be the phosphorus acid for adjusting pH value of 0.1M with concentration.
3. a kind of capillary electrophoresis according to claim 1 detects the method for Raltitrexed enantiomter, it is characterized in that: the concentration range of chiral selector in damping fluid is 2～20mM, and wherein chiral selector is alpha-cyclodextrin, beta-schardinger dextrin-, sulfonation beta-schardinger dextrin-, HP-, gamma-cyclodextrin or carboxymethyl-beta-cyclodextrin.
4. a kind of capillary electrophoresis according to claim 1 detects the method for Raltitrexed enantiomter, it is characterized in that: the deposition condition parameter is: capillary pipe length length overall 48.5cm, effective length 40cm, capillary inner diameter is 50 μ m, working voltage is 15～30kV, and capillary temperature is 20～30 ℃, fluid mechanics sample introduction 50mbar 5 seconds, electrode: to '-', the detection wavelength is 228nm by '+'; The concentration that adds dodecyl base sodium sulphate in buffer solution is 0.01～0.1M.
5. a kind of capillary electrophoresis according to claim 1 detects the method for Raltitrexed enantiomter, it is characterized in that: use methyl alcohol: water=1: 1 (volume ratio) solution preparation Raltitrexed chirality thing analytic sample, test sample concentration between 50～200 μ g/ml, the concentration 1 μ g/ml～100 μ g/ml of enantiomter.
6. a kind of capillary electrophoresis according to claim 1 detects the method for Raltitrexed enantiomter, it is characterized in that: described working voltage 20kv; 25 ℃ of temperature capillaceous, the concentration of lauryl sodium sulfate is 30mM., and the pH value of damping fluid is 8, and chiral selector is a carboxymethyl-beta-cyclodextrin, and its concentration is 8mM.
7. detect the method for Raltitrexed enantiomter according to the described a kind of capillary electrophoresis of claim, it is characterized in that: the computing method of Raltitrexed mapping allosome are preparation Raltitrexed need testing solution and the reference substance solution that contains the Raltitrexed of 1% Raltitrexed enantiomter reference substance, the difference sample introduction, the corresponding peak area of mapping structure body must not be greater than the peak area of enantiomter in the contrast solution in the Raltitrexed need testing solution.
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| CN102095775A (en) * | 2010-12-01 | 2011-06-15 | 江南大学 | Method for efficiently separating and detecting theanine enantiomer by capillary electrophoresis |
| CN102661990A (en) * | 2012-05-03 | 2012-09-12 | 中国科学院化学研究所 | Application of cationic amino acid ion liquid to chiral ligand exchange capillary electrophoresis |
| CN102809613A (en) * | 2011-06-03 | 2012-12-05 | 河北以岭医药研究院有限公司 | Determination method of fingerprint of Poecilobdella manillensis |
| CN101726529B (en) * | 2009-12-07 | 2013-04-17 | 浙江工业大学 | Novel method for splitting 2,4,5-dichlorprop enantiomers by capillary electrophoresis apparatus |
| CN104297316A (en) * | 2014-10-24 | 2015-01-21 | 常州大学 | Chiral sensor based on beta-cyclodextrin and preparation method thereof |
| CN104965018A (en) * | 2015-07-03 | 2015-10-07 | 湖北博凯医药科技有限公司 | Method for resolution of racemization 2-chloropropionic acid by adopting capillary electrophoresis separation-diode array detection technique |
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| CN107167535A (en) * | 2017-07-20 | 2017-09-15 | 南京普氟生物检测技术有限公司 | A kind of method that reversed phase liquid chromatography detects Raltitrexed enantiomter |
| CN109283262A (en) * | 2017-07-21 | 2019-01-29 | 南京正大天晴制药有限公司 | Pass through the method for high efficiency liquid chromatography for separating and determining Raltitrexed and its impurity |
| CN111678998A (en) * | 2020-04-30 | 2020-09-18 | 扬子江药业集团有限公司 | Separation and detection method for enantiomers in raltitrexed |
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| US6316613B1 (en) * | 1997-07-25 | 2001-11-13 | Beckman Coulter, Inc. | Chiral separation of pharmaceutical compounds with charged cyclodextrins using capillary electrophoresis |
| CN1100261C (en) * | 1997-11-29 | 2003-01-29 | 中国科学院大连化学物理研究所 | Capillary electrophoretic dismantling analysis method of chiral compound and its special reagent kit |
| JP3285832B2 (en) * | 1998-11-25 | 2002-05-27 | 三菱重工業株式会社 | Highly sensitive analysis method for dioxins |
| DE10042451A1 (en) * | 2000-08-29 | 2002-03-14 | Studiengesellschaft Kohle Mbh | High-throughput method for determining the enantiomeric unit of chiral compounds |
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| CN101726529B (en) * | 2009-12-07 | 2013-04-17 | 浙江工业大学 | Novel method for splitting 2,4,5-dichlorprop enantiomers by capillary electrophoresis apparatus |
| CN102095775A (en) * | 2010-12-01 | 2011-06-15 | 江南大学 | Method for efficiently separating and detecting theanine enantiomer by capillary electrophoresis |
| CN102809613A (en) * | 2011-06-03 | 2012-12-05 | 河北以岭医药研究院有限公司 | Determination method of fingerprint of Poecilobdella manillensis |
| CN102809613B (en) * | 2011-06-03 | 2015-04-15 | 河北以岭医药研究院有限公司 | Determination method of fingerprint of Poecilobdella manillensis |
| CN102661990A (en) * | 2012-05-03 | 2012-09-12 | 中国科学院化学研究所 | Application of cationic amino acid ion liquid to chiral ligand exchange capillary electrophoresis |
| CN104297316A (en) * | 2014-10-24 | 2015-01-21 | 常州大学 | Chiral sensor based on beta-cyclodextrin and preparation method thereof |
| CN104297316B (en) * | 2014-10-24 | 2016-11-02 | 常州大学 | A kind of chiral sensor based on beta-schardinger dextrin-and preparation method thereof |
| CN104965018A (en) * | 2015-07-03 | 2015-10-07 | 湖北博凯医药科技有限公司 | Method for resolution of racemization 2-chloropropionic acid by adopting capillary electrophoresis separation-diode array detection technique |
| CN106124600A (en) * | 2016-07-06 | 2016-11-16 | 黑龙江中医药大学 | Utilize capillary electrophoresis technique in Fructus Forsythiae 8 kinds of methods that lignan component quantitative determines |
| CN106124600B (en) * | 2016-07-06 | 2018-08-07 | 黑龙江中医药大学 | Using capillary electrophoresis technique to 8 kinds of lignan components are quantitative determined in Fructus Forsythiae method |
| CN107167535A (en) * | 2017-07-20 | 2017-09-15 | 南京普氟生物检测技术有限公司 | A kind of method that reversed phase liquid chromatography detects Raltitrexed enantiomter |
| CN107167535B (en) * | 2017-07-20 | 2020-02-04 | 南京和曦生物科技有限公司 | Method for detecting raltitrexed enantiomer by reversed phase liquid chromatography |
| CN109283262A (en) * | 2017-07-21 | 2019-01-29 | 南京正大天晴制药有限公司 | Pass through the method for high efficiency liquid chromatography for separating and determining Raltitrexed and its impurity |
| CN109283262B (en) * | 2017-07-21 | 2019-08-06 | 南京正大天晴制药有限公司 | Pass through the method for high efficiency liquid chromatography for separating and determining Raltitrexed and its impurity |
| CN111678998A (en) * | 2020-04-30 | 2020-09-18 | 扬子江药业集团有限公司 | Separation and detection method for enantiomers in raltitrexed |
| CN111678998B (en) * | 2020-04-30 | 2021-11-09 | 扬子江药业集团有限公司 | Separation and detection method for enantiomers in raltitrexed |
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