CN105954447B - A method of lactone enantiomter is stood by HPLC Analyze & separate benzoyl section - Google Patents
A method of lactone enantiomter is stood by HPLC Analyze & separate benzoyl section Download PDFInfo
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- CN105954447B CN105954447B CN201610574784.8A CN201610574784A CN105954447B CN 105954447 B CN105954447 B CN 105954447B CN 201610574784 A CN201610574784 A CN 201610574784A CN 105954447 B CN105954447 B CN 105954447B
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- Prior art keywords
- lactone
- benzoyl section
- enantiomter
- benzoyl
- hexane
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- 150000002596 lactones Chemical class 0.000 title claims abstract description 107
- 125000003236 benzoyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C(*)=O 0.000 title claims abstract description 43
- 238000000034 method Methods 0.000 title claims abstract description 24
- 238000004128 high performance liquid chromatography Methods 0.000 title claims abstract description 11
- -1 (±)-benzoyl Chemical group 0.000 claims description 66
- 239000012452 mother liquor Substances 0.000 claims description 30
- 238000012360 testing method Methods 0.000 claims description 28
- 239000007788 liquid Substances 0.000 claims description 18
- 239000012085 test solution Substances 0.000 claims description 18
- 238000001514 detection method Methods 0.000 claims description 16
- 238000002474 experimental method Methods 0.000 claims description 15
- 238000005070 sampling Methods 0.000 claims description 15
- 239000000203 mixture Substances 0.000 claims 3
- 238000000926 separation method Methods 0.000 abstract description 7
- 230000003287 optical effect Effects 0.000 abstract description 6
- 238000004458 analytical method Methods 0.000 abstract description 4
- 239000012046 mixed solvent Substances 0.000 abstract description 4
- 238000002360 preparation method Methods 0.000 abstract description 4
- 229920000856 Amylose Polymers 0.000 abstract description 2
- 239000000523 sample Substances 0.000 description 40
- 229960000935 dehydrated alcohol Drugs 0.000 description 29
- 239000012071 phase Substances 0.000 description 23
- 239000000243 solution Substances 0.000 description 20
- QKGYJVXSKCDGOK-UHFFFAOYSA-N hexane;propan-2-ol Chemical compound CC(C)O.CCCCCC QKGYJVXSKCDGOK-UHFFFAOYSA-N 0.000 description 13
- KFZMGEQAYNKOFK-UHFFFAOYSA-N isopropyl alcohol Natural products CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 description 12
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 11
- 229940079593 drug Drugs 0.000 description 9
- 239000003814 drug Substances 0.000 description 9
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 8
- 229940127293 prostanoid Drugs 0.000 description 6
- 150000003814 prostanoids Chemical class 0.000 description 6
- 239000012535 impurity Substances 0.000 description 5
- 239000012488 sample solution Substances 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 4
- 239000000377 silicon dioxide Substances 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 238000004587 chromatography analysis Methods 0.000 description 3
- 230000007812 deficiency Effects 0.000 description 3
- 150000002148 esters Chemical group 0.000 description 3
- 150000002240 furans Chemical class 0.000 description 3
- 150000003180 prostaglandins Chemical class 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 125000001231 benzoyloxy group Chemical group C(C1=CC=CC=C1)(=O)O* 0.000 description 2
- 239000000945 filler Substances 0.000 description 2
- 125000004029 hydroxymethyl group Chemical group [H]OC([H])([H])* 0.000 description 2
- 238000013507 mapping Methods 0.000 description 2
- 229940094443 oxytocics prostaglandins Drugs 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- KXDHJXZQYSOELW-UHFFFAOYSA-M Carbamate Chemical compound NC([O-])=O KXDHJXZQYSOELW-UHFFFAOYSA-M 0.000 description 1
- 230000005526 G1 to G0 transition Effects 0.000 description 1
- OBRRYUZUDKVCOO-FVCCEPFGSA-N [(3ar,4s,5r,6as)-4-(hydroxymethyl)-2-oxo-3,3a,4,5,6,6a-hexahydrocyclopenta[b]furan-5-yl] benzoate Chemical compound O([C@H]1[C@@H]([C@H]2CC(=O)O[C@H]2C1)CO)C(=O)C1=CC=CC=C1 OBRRYUZUDKVCOO-FVCCEPFGSA-N 0.000 description 1
- 239000013543 active substance Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 125000000051 benzyloxy group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])O* 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005194 fractionation Methods 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 125000001449 isopropyl group Chemical group [H]C([H])([H])C([H])(*)C([H])([H])[H] 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- SYSQUGFVNFXIIT-UHFFFAOYSA-N n-[4-(1,3-benzoxazol-2-yl)phenyl]-4-nitrobenzenesulfonamide Chemical class C1=CC([N+](=O)[O-])=CC=C1S(=O)(=O)NC1=CC=C(C=2OC3=CC=CC=C3N=2)C=C1 SYSQUGFVNFXIIT-UHFFFAOYSA-N 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/60—Construction of the column
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N2030/022—Column chromatography characterised by the kind of separation mechanism
- G01N2030/027—Liquid chromatography
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- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
Abstract
The invention discloses a kind of methods for using amylose class chiral chromatographic column to found lactone enantiomter by HPLC Analyze & separate benzoyl section as mobile phase using positive mixed solvent.This method can simple, accurate, efficient analysis separation benzoyl section found lactone enantiomter, to realize that the quality for founding lactone enantiomter to benzoyl section controls.Lactone founds in the benzoyl section that the present invention can also be used to the preparation single optical activity of high-purity.
Description
Technical field
The present invention relates to a kind of chromatography separation methods, separate benzoyl especially by high-efficient liquid phase chromatogram technique analysis
The method that lactone enantiomter is found by section.
Technical background
Prostaglandin (Prostaglandins, abridge PGs) be it is a kind of it is important there is the active substance of endogenous physiological,
It is widely present in each tissue in vivo, is the important medium of various physiological processes.Since content is extremely low in human body, clinically
Prostanoid drug mainly by it is artificial synthesized come.Lactone (Corey lactone benzoate) founds in benzoyl section
The common starting material of synthesis of prostaglandins drug, molecular formula C15H16O5, chemical name are as follows: 5- (benzoyloxy) hexahydro -4-
(hydroxymethyl) -2H- cyclopentano [b] furans -2- ketone, common name are as follows: lactone, structural formula found in benzoyl section are as follows:
, theoretically there are multiple optical isomers, but according to its reaction mechanism and conjunction in molecule chiral centre containing there are four
Infer at method, only exist a pair of of enantiomter, is i.e. (3aS, 4R, 5S, 6aR) -5- (benzoyloxy) hexahydro -4- (hydroxyl
Methyl) -2H- cyclopentano [b] furans -2- ketone (lactone founds in (+)-benzoyl section) and (3aR, 4S, 5R, 6aS) -5- (benzoxy
Base) hexahydro -4- (hydroxymethyl) -2H- cyclopentano [b] furans -2- ketone (lactone founds in (-)-benzoyl section), mapping is different each other for the two
Structure body impurity continues to participate in reaction in the next steps, forms a variety of corresponding impurity, influences the quality of prostanoid drug.Cause
This, how Analyze & separate this to enantiomter, improve prostanoid drug quality, it is ensured that the purity of prostanoid drug
And safety, it is the current technical issues that need to address.
102532079 B of Chinese invention patent CN discloses a kind of preparation method of vertical lactone of benzoyl section.This method from
Ke Linei esterdiol sets out, and is reflected by acylated, ester exchange reaction obtains product.
The fractionation of chiral drug enantiomter and the determination of absolute configuration are one of crucial importance and long-standing ask
Topic, in numerous methods for obtaining optical isomer, the high performance liquid chromatography separation method comprising chiral stationary phase is most commonly seen.
Chinese invention patent ZL201310652110.1 discloses one kind (-)-benzoyl section and founds lactone and its optical siomerism
Body Analyze & separate method, this method select Daicel AD-H chiral column, found lactone enantiomter to benzoyl section and analyze
And separation.
The Analyze & separate technique study of lactone enantiomter, only spy are stood by rare benzoyl section in currently available technology
There is also Silica Surfaces in some problems and deficiency, such as chromatographic column to coat amylose-three-(3,5- dimethyl in rope research
Carbanilate) filler and benzoyl section stand lactone enantiomter interaction difference it is smaller, stereoselectivity compared with
It is low, so that the vertical lactone enantiomter of benzoyl section is unable to get and efficiently separates.
Therefore, it is necessary to a kind of new method for capableing of effective vertical lactone enantiomter of Analyze & separate benzoyl section is established,
Impurity content is controlled, and then improves the prostanoid drug quality of synthesis.
Summary of the invention
The purpose of the invention is to overcome the deficiencies of the prior art and provide a kind of to use new chiral chromatographic column with positive
Mixed solvent is the HPLC method of mobile phase, and this method being capable of accurate, the vertical lactone enantiomerism of efficient analysis separation benzoyl section
Body, to realize that the quality for founding lactone enantiomter to benzoyl section controls, while it is single to can also be used to preparation high-purity
Lactone founds in the benzoyl section of optical activity.
The present invention provides a kind of methods for founding lactone enantiomter by HPLC Analyze & separate benzoyl section, using color
Composing filler is the chiral chromatographic column that Silica Surface is coated with amylose-three (s)-Alpha-Methyl carbanilate, and is used
Normal phase chromatography separation determination.
Mobile phase used in normal phase chromatography of the present invention can be n-hexane/isopropyl alcohol mixed solvent, n-hexane/anhydrous
One of alcohol mixed solvent；
Further, the volume ratio of n-hexane/isopropanol used in the present invention is 90:10~80:20, preferably
90:10~85:15；
Further, the volume ratio of n-hexane/dehydrated alcohol used in the present invention is 95:5~84:16, preferably
92:8~85:15.
Analyze & separate method of the present invention, can realize in accordance with the following methods:
(1) lactone flowing phased soln founds in (±)-benzoyl section, be configured to 1mL containing (±)-benzoyl section stand lactone 1~
The test solution of 10 μ g；Sample volume is 1~20 μ L, even more preferably 5~10 μ L；
(2) setting n-hexane/isopropanol flow rate of mobile phase be 0.8~1.5mL/min, preferably 0.9~1.5mL/min,
N-hexane/dehydrated alcohol flow rate of mobile phase is 0.3~1.5mL/min, preferably 0.4~1.0mL/min；Detection wavelength is 210
~235nm is more preferably 230nm；Column temperature is room temperature；
(3) made 5~10 μ L of test solution injection chiral chromatographic column (250 × 4.6mm, 5.0 μm) of step (1) is taken, it is complete
At benzoyl section stand lactone enantiomter analysis with separate.
It innovative point of the invention and has the beneficial effect that
1. using amylose-three (s)-Alpha-Methyl carbanilate chiral chromatographic column；Using more preferably flowing
Phase；Selective flow phased soln sample selects preferably sample volume；Select preferably wavelength；Ensure that the stability of sample,
The symmetry of chromatographic peak is improved, and has preferable separating effect.Implement according to the method for the present invention, (+)-benzoyl Ke Linei
Two successive appearances of enantiomter of lactone, separating degree >=1.5, effective Analyze & separate benzoyl found in ester and (-)-benzoyl section
Lactone enantiomter is found by section.
2. the Silica Surface that the present invention uses is coated with amylose-three (s)-Alpha-Methyl phenyl compared with previous research
The AS-H chiral chromatographic column of carbamate is coated with (3, the 5- dimethylphenylamino formic acid of amylose-three with Silica Surface
Ester) AD-H amylose class chiral column compare, to benzoyl section stand lactone enantiomter stereoselectivity it is higher, separation
Degree is higher, effectively can found lactone enantiomter to benzoyl section by Analyze & separate.
3. the Analyze & separate method is quick, accurate and efficient, it can be ensured that lactone enantiomter is found by benzoyl section
Quality controllability；Analyze & separate method of the present invention, the benzoyl section that can also be used to the preparation single optical activity of high-purity are vertical
Lactone.
4. the technology overcomes the deficiencies in the prior art, solves the Analyze & separate that lactone enantiomter is found by benzoyl section
Problem can effectively control target product and its enantiomter impurity content, exempt enantiomter impurity to subsequent synthesis
The interference of reaction improves prostanoid drug quality, it is ensured that drug safety.
It can be seen that the present invention develop it is a kind of using new chiral chromatographic column Analyze & separate benzoyl section stand lactone mapping
The HPLC method of isomers is very necessary.
Detailed description of the invention
Fig. 1: HPLC figure, chromatographic condition: chromatographic column: Daicel AD-H (250 × 4.6mm, 5.0 μm)；Mobile phase: just oneself
Alkane-dehydrated alcohol (90:10)；Column temperature: 30 DEG C.
Fig. 2: HPLC figure, chromatographic condition: chromatographic column: BC chiral AS-H (, 250mm × 4.6mm, 5 μm)；Mobile phase:
N-hexane-dehydrated alcohol (92:8)；Column temperature: room temperature.
Specific embodiment
It is a kind of to the present invention that lactone enantiomter is stood by HPLC Analyze & separate benzoyl section below with reference to embodiment
Method is described further, but protection scope of the present invention is not limited to embodiment.
Embodiment 1
Instrument and condition
Chromatographic column: Daicel AD-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (90:10)
Column temperature: 30 DEG C
Flow velocity: 1.0mL/min
Sample concentration: 10 μ g/mL
Sampling volume: 2 μ L
Detection wavelength: 220nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-isopropanol (90:10)
Scale is dissolved and be diluted to, is shaken up, as test sample mother liquor.Precision is drawn 100 μ L of test sample mother liquor and is placed in 10mL volumetric flask,
N-hexane-isopropanol (90:10) solution constant volume is configured to (±)-benzoyl section that concentration is 10 μ g/mL and stands lactone confession to scale
Test sample solution draws 2 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: see attached drawing 1, lactone founds in (+)-benzoyl section and two enantiomters of lactone are found by (-)-benzoyl section
Separating degree be 0.9, benzoyl section stand lactone enantiomter cannot efficiently separate.
Embodiment 2
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-isopropanol (90:10)
Column temperature: room temperature
Flow velocity: 1.0mL/min
Sample concentration: 10 μ g/mL
Sampling volume: 1 μ L
Detection wavelength: 210nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-isopropanol (90:10)
Scale is dissolved and be diluted to, is shaken up, as test sample mother liquor.Precision is drawn 100 μ L of test sample mother liquor and is placed in 10mL volumetric flask,
N-hexane-isopropanol (90:10) solution constant volume is configured to (±)-benzoyl section that concentration is 10 μ g/mL and stands lactone confession to scale
Test sample solution draws 1 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.7, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 3
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-isopropanol (80:20)
Column temperature: room temperature
Flow velocity: 0.8mL/min
Sample concentration: 5 μ g/mL
Sampling volume: 5 μ L
Detection wavelength: 235nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-isopropanol (80:20)
Scale is dissolved and be diluted to, is shaken up, as test sample mother liquor.Precision is drawn 50 μ L of mother liquor and is placed in 10mL volumetric flask, n-hexane-
It is molten to be configured to the vertical lactone test sample of (±)-benzoyl section that concentration is 5 μ g/mL to scale for isopropanol (80:20) solution constant volume
Liquid draws 5 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 4
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-isopropanol (90:10)
Column temperature: room temperature
Flow velocity: 1.5mL/min
Sample concentration: 1 μ g/mL
Sampling volume: 10 μ L
Detection wavelength: 235nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-isopropanol (90:10)
Scale is dissolved and be diluted to, is shaken up, as test sample mother liquor.Precision is drawn 10 μ L of mother liquor and is placed in 10mL volumetric flask, n-hexane-
It is molten to be configured to the vertical lactone test sample of (±)-benzoyl section that concentration is 1 μ g/mL to scale for isopropanol (90:10) solution constant volume
Liquid draws 10 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 5
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-isopropanol (85:15)
Column temperature: room temperature
Flow velocity: 0.9mL/min
Sample concentration: 1 μ g/mL
Sampling volume: 20 μ L
Detection wavelength: 230nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-isopropanol (85:15)
Scale is dissolved and be diluted to, is shaken up, as test sample mother liquor.Precision is drawn 10 μ L of mother liquor and is placed in 10mL volumetric flask, n-hexane-
It is molten to be configured to the vertical lactone test sample of (±)-benzoyl section that concentration is 1 μ g/mL to scale for isopropanol (85:15) solution constant volume
Liquid draws 20 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 2.25, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 6
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-isopropanol (88:12)
Column temperature: room temperature
Flow velocity: 1.0mL/min
Sample concentration: 10 μ g/mL
Sampling volume: 1 μ L
Detection wavelength: 230nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-isopropanol (88:12)
Scale is dissolved and be diluted to, is shaken up, as test sample mother liquor.Precision draw 100 μ L of mother liquor be placed in 10mL volumetric flask, just oneself
Alkane-isopropanol (88:12) solution constant volume is configured to (±)-benzoyl section that concentration is 10 μ g/mL and founds lactone test sample to scale
Solution draws 1 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 7
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (95:5)
Column temperature: room temperature
Flow velocity: 1.5mL/min
Sample concentration: 5 μ g/mL
Sampling volume: 10 μ L
Detection wavelength: 210nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-dehydrated alcohol (95:
5) it dissolves and is diluted to scale, shake up, as test sample mother liquor.Precision draw 50 μ L of mother liquor be placed in 10mL volumetric flask, just oneself
Alkane-dehydrated alcohol (95:5) solution constant volume is configured to (±)-benzoyl section that concentration is 5 μ g/mL and founds lactone test sample to scale
Solution draws 10 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 8
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (84:16)
Column temperature: room temperature
Flow velocity: 0.4mL/min
Sample concentration: 5 μ g/mL
Sampling volume: 5 μ L
Detection wavelength: 220nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-dehydrated alcohol (84:
16) it dissolves and is diluted to scale, shake up, as test sample mother liquor.Precision draw 50 μ L of mother liquor be placed in 10mL volumetric flask, just oneself
Alkane-dehydrated alcohol (84:16) solution constant volume is configured to (±)-benzoyl section that concentration is 5 μ g/mL and founds lactone for examination to scale
Product solution draws 5 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 9
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (92:8)
Column temperature: room temperature
Flow velocity: 0.8mL/min
Sample concentration: 1 μ g/mL
Sampling volume: 20 μ L
Detection wavelength: 230nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-dehydrated alcohol (92:
8) it dissolves and is diluted to scale, shake up, as test sample mother liquor.Precision draw 10 μ L of mother liquor be placed in 10mL volumetric flask, just oneself
Alkane-dehydrated alcohol (92:8) solution constant volume is configured to (±)-benzoyl section that concentration is 1 μ g/mL and founds lactone test sample to scale
Solution draws 20 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: see attached drawing 2, lactone founds in (+)-benzoyl section and two enantiomters of lactone are found by (-)-benzoyl section
Separating degree be 1.6, benzoyl section stand lactone enantiomter can be kept completely separate.
Embodiment 10
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (85:15)
Column temperature: room temperature
Flow velocity: 0.3mL/min
Sample concentration: 10 μ g/mL
Sampling volume: 2 μ L
Detection wavelength: 230nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-dehydrated alcohol (85:
15) it dissolves and is diluted to scale, shake up, as test sample mother liquor.Precision is drawn 100 μ L of mother liquor and is placed in 10mL volumetric flask, just
Hexane-dehydrated alcohol (85:15) solution constant volume is configured to (±)-benzoyl section that concentration is 10 μ g/mL and stands lactone confession to scale
Test sample solution draws 2 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.7, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 11
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (85:15)
Column temperature: room temperature
Flow velocity: 0.4mL/min
Sample concentration: 10 μ g/mL
Sampling volume: 5 μ L
Detection wavelength: 235nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-dehydrated alcohol (85:
15) it dissolves and is diluted to scale, shake up, as test sample mother liquor.Precision is drawn 100 μ L of mother liquor and is placed in 10mL volumetric flask, just
Hexane-dehydrated alcohol (85:15) solution constant volume is configured to (±)-benzoyl section that concentration is 10 μ g/mL and stands lactone confession to scale
Test sample solution draws 5 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 12
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (87:13)
Column temperature: room temperature
Flow velocity: 0.8mL/min
Sample concentration: 1 μ g/mL
Sampling volume: 10 μ L
Detection wavelength: 230nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-dehydrated alcohol (87:
13) it dissolves and is diluted to scale, shake up, as test sample mother liquor.Precision draw 10 μ L of mother liquor be placed in 10mL volumetric flask, just oneself
Alkane-dehydrated alcohol (87:13) solution constant volume is configured to (±)-benzoyl section that concentration is 1 μ g/mL and founds lactone for examination to scale
Product solution draws 10 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 13
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (85:15)
Column temperature: room temperature
Flow velocity: 0.6mL/min
Sample concentration: 10 μ g/mL
Sampling volume: 5 μ L
Detection wavelength: 230nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-dehydrated alcohol (85:
15) it dissolves and is diluted to scale, shake up, as test sample mother liquor.Precision is drawn 100 μ L of mother liquor and is placed in 10mL volumetric flask, just
Hexane-dehydrated alcohol (85:15) solution constant volume is configured to (±)-benzoyl section that concentration is 10 μ g/mL and stands lactone confession to scale
Test sample solution draws 5 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Embodiment 14
Instrument and condition
Chromatographic column: BC chiral AS-H (250 × 4.6mm, 5.0 μm)
Mobile phase: n-hexane-dehydrated alcohol (88:12)
Column temperature: room temperature
Flow velocity: 0.8mL/min
Sample concentration: 5 μ g/mL
Sampling volume: 10 μ L
Detection wavelength: 230nm
Experimental procedure
Precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, n-hexane-dehydrated alcohol (88:
12) it dissolves and is diluted to scale, shake up, as test sample mother liquor.Precision draw 50 μ L of mother liquor be placed in 10mL volumetric flask, just oneself
Alkane-dehydrated alcohol (88:12) solution constant volume is configured to (±)-benzoyl section that concentration is 5 μ g/mL and founds lactone for examination to scale
Product solution draws 10 μ L of test solution and enters liquid chromatogram, records chromatogram.
Experimental result: lactone founds in (+)-benzoyl section and the separating degree of two enantiomters of lactone is found by (-)-benzoyl section
It is 1.6, benzoyl section founds lactone enantiomter and can be kept completely separate.
Claims (1)
1. a kind of method for founding lactone enantiomter by HPLC Analyze & separate benzoyl section, comprising the following steps:
Instrument and condition:
Chromatographic column: BC chiral AS-H, 250 × 4.6mm, 5.0 μm；
Mobile phase: volume ratio is n-hexane-isopropyl alcohol mixture of 85:15；
Column temperature: room temperature；
Flow velocity: 0.9mL/min；
Sample concentration: 1 μ g/mL；
Sampling volume: 20 μ L；
Detection wavelength: 230nm；
Experimental procedure: precision weighs (±)-benzoyl section and founds lactone 10mg, is placed in 10mL volumetric flask, is 85:15 with volume ratio
N-hexane-isopropyl alcohol mixture dissolve and be diluted to scale, shake up, as test sample mother liquor；Precision draws 10 μ L of mother liquor
It is placed in 10mL volumetric flask, the n-hexane-isopropyl alcohol mixture constant volume to scale for being 85:15 with volume ratio is configured to concentration
Lactone test solution is found for (±)-benzoyl section of 1 μ g/mL, 20 μ L of test solution is drawn and enters liquid chromatogram, record color
Spectrogram.
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| CN107064336B (en) * | 2017-02-28 | 2023-06-06 | 长春百纯和成医药科技有限公司 | Method for separating enantiomer of Colalactone diol by pre-column derivatization analysis |
| CN107085058A (en) * | 2017-05-12 | 2017-08-22 | 成都丽凯手性技术有限公司 | A kind of PHBA enantiomters method for detecting impurities |
| CN108956808B (en) * | 2018-06-04 | 2021-07-20 | 吉林医药学院 | A kind of HPLC method analysis and preparation method of diacetylcorilactone enantiomer |
| CN108693272B (en) * | 2018-06-04 | 2021-03-12 | 吉林百纯化学科技有限公司 | Method for analyzing and preparing N- (p-toluenesulfonyl) -L-alanine and enantiomer thereof by HPLC method |
| CN112229923B (en) * | 2020-09-30 | 2022-11-11 | 东北制药集团股份有限公司 | Method for detecting 15-ketone and related substances thereof by adopting high performance liquid chromatography |
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