CN107764890A - A kind of differentiation detection method of Ezetimibe enantiomter - Google Patents
A kind of differentiation detection method of Ezetimibe enantiomter Download PDFInfo
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Abstract
The invention discloses a kind of differentiation detection method of Ezetimibe enantiomter, using beta cyclodextrin and Cu ions as part, for with Ezetimibe (S, R, S), (R, S, R) complex compound of different stable forms is formed, and it is identified differentiation using mass spectrography.The present invention highly shortened Ezetimibe enantiomter (S, R, S), (R, S, R) detection time, by enantiomter (S, R, S), (R, S, R analysis time) foreshortens to 2 3min, easy operating process, and it is very good to distinguish repeatability；Sample size is few needed for differentiation, only needs 200 μ l or so, cost is very low.The present invention distinguishes for the medicine enantiomter and provides better method, while provides new thinking to distinguish the analysis of the larger multichiral center enantiomter of difficulty.
Description
Technical field
The present invention relates to Pharmaceutical Analysis technical field, and in particular to a kind of Ezetimibe enantiomter (S, R, S), (R,
S, R) differentiation detection method.
Background technology
Ezetimibe is that cholesterol suppresses absorbent, suitable for treating primary hypercholesterolemia, homozygote familial
Hypercholesterolemia (HoFH) and homozygote Sitosterolemia (or phytosterol mass formed by blood stasis), its molecular formula are C24H21F2NO3, chemistry
Entitled 1- (4- fluorophenyls) -3 (R)-[(the S)-hydroxypropyls of 3- (4- fluorophenyls) -3
Base] -4 (S)-(4- hydroxyphenyls) -2- azetidines (azetidine) ketone, structural formula is：
Contain 3 chiral centres in the molecule, multiple optical isomers be present, it is miscellaneous for the optical isomer of Ezetimibe
Matter, quality control must be carried out among pharmaceutical synthesis process.The differentiation of optical isomer containing asymmetric carbon atom is always chirality
The difficult point of quality control in pharmaceutical synthesis and production process, realize the differentiation of Ezetimibe and its optical isomer in Ezetimibe
There is realistic meaning in terms of the synthesis of medicine and the quality control of production process.It is different for the molecule of multichiral center, diastereomeric
Structure body differentiation difficulty is low, and enantiomter differentiation difficulty is larger, therefore to the research of such medicine enantiomter differentiating method
It is significant.
At present, the differentiating method of conventional Ezetimibe enantiomter is traditional chromatography, as LC, HPLC, CE,
CEC, SFC etc., but these methods are required for longer differentiation time and larger sample size, and practical operation is brought not
Just.Mass spectrum has the characteristics of quick, accurate, sensitive, from after the differentiation for chiral isomer in 1977 first, now
Great development is arrived, the method for main differentiation chiral isomer includes mass spectrography and tandem mass spectrometry.The former include again it is main-
Object method, ion-molecule reaction method and diastereoisomer dissociation method.At present Ezetimibe enantiomerism is distinguished with mass spectrography
Body yet there are no document report.
The content of the invention
It is an object of the present invention to provide a kind of Ezetimibe enantiomter (S, R, S), the differentiation detection method of (R, S, R),
To solve the deficiencies in the prior art.
The present invention uses following technical scheme：
A kind of differentiation detection method of Ezetimibe enantiomter, using beta-schardinger dextrin and Cu ions as part, use
In the complex compound with Ezetimibe (S, R, S), the different stable forms of (R, S, R) formation, and it is identified using mass spectrography
Distinguish.
Further, the final concentration of 20-25 μ g/ml of beta-schardinger dextrin；First add a small amount of DMSO whole to beta-schardinger dextrin during configuration
Dissolving, again with methanol dilution are stand-by.
Further, Cu ions are divalence, final concentration of 12-12.5 μ g/ml；During configuration solvent is used as by the use of distilled water.
Further, mass spectrographic sample injection method is pin pump direct injected, prevents complex compound before being collisionally dissociated just
Influenceed by external environment.
Further, the flow velocity of pin pump direct injected is 20 μ l/min, acquisition time 1min, and mass spectrum true temp is
250 DEG C, charge-mass ratio scope sets 300-2000.
Further, when second order mses experiment is carried out in mass spectrum, target parent ion [Cu (Ezetimibe) is directly inputted2
(β-CD)]2+Charge-mass ratio (m/z=1007.8) enter second order mses, collision energy is adjusted to 0 to find parent ion.
Further, it is 14-16, acquisition time 40ms to carry out collision energy during second order mses experiment.
Beneficial effects of the present invention：
The invention provides a kind of new differentiation for Ezetimibe enantiomter (S, R, S), (R, S, R) to detect
Method, using beta-schardinger dextrin and divalence Cu ions as part, according to diastereoisomer liberation characteristic difference, with mass spectrography come
Ezetimibe and its enantiomter are distinguished, and optimal structure existing for the complex system is built by the way of computer simulation
Type, deeply understand that it distinguishes mechanism.
The present invention highly shortened Ezetimibe enantiomter (S, R, S), (R, S, R) detection time, and mapping is different
Structure body (S, R, S), the analysis time of (R, S, R) foreshorten to 2-3min, easy operating process, and distinguish repeatability very
It is good；Sample size is few needed for differentiation, only needs 200 μ l or so, cost is very low.The present invention is distinguished for the medicine enantiomter and provided
Better method, while provide new thinking to distinguish the analysis of the larger multichiral center enantiomter of difficulty.
Brief description of the drawings
Fig. 1 is that (A is side view, B to bow for the dimensional structure diagrams of SRS- Ezetimibes+beta-schardinger dextrin+Cu ionic systems
View)；
Fig. 2 is that (A is side view, B to bow for the dimensional structure diagrams of RSR- Ezetimibes+beta-schardinger dextrin+Cu ionic systems
View)；
Fig. 3 is the second order mses figure of SRS- Ezetimibes+beta-schardinger dextrin+Cu ionic systems；
Fig. 4 is the second order mses figure of RSR- Ezetimibes+beta-schardinger dextrin+Cu ionic systems.
Embodiment
The present invention is done with reference to embodiment and accompanying drawing and further explained.The following example is merely to illustrate this hair
It is bright, but it is not used to limit the practical range of the present invention.
Embodiment 1
A. sample configures：Accurately weigh 1mg RSR- Ezetimibes, 1mg SRS- Ezetimibes, 1mg CuSO4Solid with
And 10mg beta-schardinger dextrins.RSR- Ezetimibes and SRS- Ezetimibes are respectively with methanol dilution to 50 μ g/ml；CuSO4Solid
1mg/ml is diluted to pure water；Beta-schardinger dextrin is first dissolved to invisible solid (DMSO is no more than 200 μ l) with a small amount of DMSO, then
With methanol dilution to 1mg/ml, again with methanol is diluted to 50 μ g/ml.Take the μ g/ml RSR- Ezetimibes of 200 μ l 50 and 200 μ
The μ g/ml beta-schardinger dextrins of l 50 mix, and add 5 μ l 1mg/ml CuSO4The aqueous solution obtains sample 1；SRS- Ezetimibes are matched somebody with somebody with above-mentioned
The method of putting obtains sample 2.
B. mass spectrum is distinguished：By above-mentioned 2 samples respectively by way of pin pump direct injected, in Agilient Q-ToF
6550 high-resolution mass spectrometers persistently inject mass spectrum, acquisition time 1min with 20 μ l/min speed, and mass spectrum true temp is 250
DEG C, charge-mass ratio scope sets 300-2000；Collision energy is set to search out parent ion [Cu for 0 on second order mses figure
(Ezetimibe)2(β-CD)]2+(m/z=1007.8) collision energy, is then gradually increased again, until drawing with notable difference
Fragment ion mass spectrogram, collision energy 14-16, acquisition time 40ms, as shown in Figure 3, Figure 4.Fig. 3 is SRS- according to folding wheat
The second order mses figure of cloth+beta-schardinger dextrin+Cu ionic systems, parent ion are [Cu (Ezetimibe)2(β-CD)]2+(m/z=
1007.8) fragment ion caused by is respectively [Ezetimibe-H2O+H]+(m/z=392.15), [Ezetimibe+H]+(m/z
=410.15), [Cu (β-CD)-H]+And [Cu (Ezetimibe) (β-CD)-H] (m/z=1196.3)+(m/z=1605.5).Figure
4 be the second order mses figure of RSR- Ezetimibes+beta-schardinger dextrin+Cu ionic systems, caused fragment ion and SRS- Ezetimibes+
Beta-schardinger dextrin+Cu ionic systems are identical, still [Ezetimibe-H2O+H]+
(m/z=392.15) abundance of this fragment ion has difference therewith.Fig. 3, Fig. 4 Mass spectrometry experiments result, can
To find out under identical collision energy, [Ezetimibe-H2O+H]+(m/z=392.15) abundance of this fragment ion is according to folding
Wheat cloth (S, R, S) is lower than Ezetimibe (R, S, R), shows that the former combines more stable, it was confirmed that following theoretical calculations
As a result.
C. computer simulation method structure complex compound preferred configuration：With the DFT (Density functionals in computer simulation method
Method) calculated, wherein free multiplicity is 2, system charge number is that 2, Cu ions use Lanl2DZ Pseudopotential basis sets, and other are former
Son uses 6-31G* base groups.The preferred configuration that 2 kinds of complex compounds may possess in mass spectrographic vacuum environment is calculated, intuitively
Ground shows the differentiation mechanism of Ezetimibe enantiomter, as shown in Figure 1, 2.Fig. 1 is calculated using computer simulation experiment
The dimensional structure diagram of the SRS- Ezetimibes gone out+beta-schardinger dextrin+Cu ionic systems, 2 molecule SRS- Ezetimibes one pass through
Beta-schardinger dextrin cavity is worn, forming Cu-O keys with Cu ions is complexed；One is located at beta-schardinger dextrin periphery, and hydrogen bond knot is formed with its hydroxyl
Close.Fig. 2 is the stereochemical structure of the RSR- Ezetimibes+beta-schardinger dextrin+Cu ionic systems calculated using computer simulation experiment
Schematic diagram, 2 molecule RSR- Ezetimibes are respectively positioned at beta-schardinger dextrin hole, and Hydrogenbond is formed with its hydroxyl.Comparison diagram 1, Fig. 2
Understand, the mode of Ezetimibe (S, R, S), (R, S, R) and ligand binding is different, Ezetimibe (S, R, S) meeting and Cu
Ion forms coordinate bond, merely will more firmly and stably by Hydrogenbond than Ezetimibe (R, S, R).
The present embodiment distinguishes Ezetimibe (S, R, S), (R, S, R) enantiomter using mass spectrography, and the used time only needs 2-
3min, very quickly, also it is aided with the Research on experimental methods of computer simulation its mechanism in addition, is provided for the differentiation of chiral isomer
New thinking.
Claims (7)
1. a kind of differentiation detection method of Ezetimibe enantiomter, it is characterised in that made using beta-schardinger dextrin and Cu ions
For part, for forming the complex compound of different stable forms with Ezetimibe (S, R, S), (R, S, R), and mass spectrography is utilized to it
Differentiation is identified.
2. the differentiation detection method of Ezetimibe enantiomter according to claim 1, it is characterised in that beta-schardinger dextrin
Final concentration of 20-25 μ g/ml；A small amount of DMSO is first added all to be dissolved to beta-schardinger dextrin during configuration, again with methanol dilution is stand-by.
3. the differentiation detection method of Ezetimibe enantiomter according to claim 1, it is characterised in that Cu ions are
Divalence, final concentration of 12-12.5 μ g/ml；During configuration solvent is used as by the use of distilled water.
4. the differentiation detection method of Ezetimibe enantiomter according to claim 1, it is characterised in that it is mass spectrographic enter
Quadrat method is pin pump direct injected, prevents complex compound from just being influenceed before being collisionally dissociated by external environment.
5. the differentiation detection method of Ezetimibe enantiomter according to claim 1, it is characterised in that pin pump is direct
The flow velocity of sample introduction is 20 μ l/min, and acquisition time 1min, mass spectrum true temp is 250 DEG C, and charge-mass ratio scope sets 300-
2000。
6. the differentiation detection method of Ezetimibe enantiomter according to claim 1, it is characterised in that in mass spectrum
When carrying out second order mses experiment, target parent ion [Cu (Ezetimibe) is directly inputted2(β-CD)]2+Charge-mass ratio (m/z=
1007.8) enter second order mses, collision energy is adjusted to 0 to find parent ion.
7. the differentiation detection method of Ezetimibe enantiomter according to claim 1, it is characterised in that carry out two level
Collision energy is 14-16, acquisition time 40ms during Mass spectrometry experiments.
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2022017547A1 (en) * | 2020-07-21 | 2022-01-27 | 宁波大学 | Beta cyclodextrin-based, aminobenzene sulfonic acid positional isomer analysis reagent and method |
| CN114034759A (en) * | 2020-07-21 | 2022-02-11 | 中国计量科学研究院 | Reagent and method for analyzing chiral structure of ibuprofen drug molecule |
| WO2022017548A3 (en) * | 2020-07-21 | 2022-03-03 | 宁波大学 | Reagent and method for chiral structure analysis of ibuprofen molecules |
| CN115453004A (en) * | 2022-10-08 | 2022-12-09 | 南京科默生物医药有限公司 | Method for detecting related substances in ezetimibe atorvastatin calcium tablets |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2022017547A1 (en) * | 2020-07-21 | 2022-01-27 | 宁波大学 | Beta cyclodextrin-based, aminobenzene sulfonic acid positional isomer analysis reagent and method |
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| WO2022017549A3 (en) * | 2020-07-21 | 2022-03-03 | 中国计量科学研究院 | Reagent and method used for molecular chiral structural analysis of drug ibuprofen |
| WO2022017548A3 (en) * | 2020-07-21 | 2022-03-03 | 宁波大学 | Reagent and method for chiral structure analysis of ibuprofen molecules |
| CN115453004A (en) * | 2022-10-08 | 2022-12-09 | 南京科默生物医药有限公司 | Method for detecting related substances in ezetimibe atorvastatin calcium tablets |
| CN115453004B (en) * | 2022-10-08 | 2023-10-13 | 南京科默生物医药有限公司 | Detection method of related substances in ezetimibe atorvastatin calcium tablet |
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