CN107321329A - A kind of preparation method of Cichoric acid trace integral post - Google Patents

A kind of preparation method of Cichoric acid trace integral post Download PDF

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CN107321329A
CN107321329A CN201710622279.0A CN201710622279A CN107321329A CN 107321329 A CN107321329 A CN 107321329A CN 201710622279 A CN201710622279 A CN 201710622279A CN 107321329 A CN107321329 A CN 107321329A
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cichoric acid
cichoric
integral post
trace
acid
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CN107321329B (en
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刘照胜
孙亚坤
阿吉艾克拜尔·艾萨
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Xinjiang Technical Institute of Physics and Chemistry of CAS
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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
    • B01J20/26—Synthetic macromolecular compounds
    • B01J20/268—Polymers created by use of a template, e.g. molecularly imprinted polymers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01D—SEPARATION
    • B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
    • B01D15/08—Selective adsorption, e.g. chromatography
    • B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
    • B01D15/22—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to the construction of the column
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/28—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties
    • B01J20/28014—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof characterised by their form or physical properties characterised by their form
    • B01J20/2805—Sorbents inside a permeable or porous casing, e.g. inside a container, bag or membrane
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
    • B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
    • B01J20/30—Processes for preparing, regenerating, or reactivating
    • B01J20/3092—Packing of a container, e.g. packing a cartridge or column
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C67/00—Preparation of carboxylic acid esters
    • C07C67/48—Separation; Purification; Stabilisation; Use of additives

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  • Organic Chemistry (AREA)
  • Analytical Chemistry (AREA)
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  • Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)

Abstract

The present invention relates to a kind of preparation method of Cichoric acid trace integral post, this method is using Cichoric acid as template molecule, while zinc acetate or cobalt acetate are added, withN,NDimethylformamide, dimethyl sulfoxide (DMSO) and the methyl imidazolium tetrafluoroborate of 1 butyl 3 do pore-foaming agent, function monomer, crosslinking agent or initiator are added, then ultrasonic dissolution removes dissolved oxygen and initiation reaction, finally obtains selective Cichoric acid molecular engram integral column.The method of the invention template consumption is few, greatly reduces the trace cost of Cichoric acid.Successfully synthesize Cichoric acid trace integral post in stainless steel tube, and blank control post and carry out chromatographic performance optimization.Test result indicates that, imprinting factor reaches as high as 24.81.This method is prepared simply, and high molecular polymer durability is good, is isolating and purifying for Cichoric acid（Purity is higher than 98.5%）There is provided a kind of cost-effective method.

Description

A kind of preparation method of Cichoric acid trace integral post
Technical field
The present invention relates to a kind of preparation method of Cichoric acid trace integral post, the imprinted polymer shows good to Cichoric acid Good selectivity, available for isolating and purifying for Cichoric acid.
Background technology
Cichoric acid is one of a kind of important immune active ingredient in witloof cauline leaf, and pharmacological research shows in recent years, witloof Acid has enhancing immunologic function and antiinflammatory action, and can suppress hyaluronidase, protect collagen from can cause degraded from By the influence of base, also with HIV1 and HIV2 integrases are suppressed, promote the effect of insulin releasing, therefore, Cichoric acid has very Big medical value, in the market is big to Cichoric acid demand.
At present, the method that traditional extraction separated and prepared purifying cichoric acid mainly has：Supercritical CO2Extraction, macropore Resin method, counter current chromatography etc..CN102060706A is reported by supercritical CO2Extraction carries Echinacea to ethanol and extracted Take, add salt to disperse afterwards, alkalization is eluted, eluent is concentrated, be dried to obtain chrysanthemum with anion exchange resin and cationic resin column Lettuce acid, it is cumbersome although this method can obtain the larger Cichoric acid of purity.CN103641716A reports Echinacea After the processes such as ultrasonic wave extraction, vacuum-concentrcted, macroreticular resin loading, deionized water rinsing, elution, vacuum-concentrcted The Cichoric acid that purity is 22% is obtained, what is obtained is only Cichoric acid runic thing, it is impossible to reach medicinal requirements.CN101148410A is reported Road is extracted by ethanol, macroporous resin adsorption elution, high speed adverse current chromatogram purifying obtain purity larger Cichoric acid, but work Skill is complicated.In summary, the shortcomings of traditional technique has condition harshness, cost height, cycle length, small selectivity.Therefore, one Plant simple, cheap, quick purification process particularly important.
Molecular engram be it is a kind of be based on " key-lock-principle ", by mimic biology cell between enzyme-substrate or antibody-antigene Interaction, the artificial synthesized emerging technology material with specific selectivity.The method very simple of molecular engram, and Easily operated, the raw material needed in real work has：Function monomer, template, pore-foaming agent and crosslinking agent etc..In certain bar (such as low temperature and irradiance or heating) initiated polymerization under part, finally again removes molecular template with such as extraction or through acid-hydrolyzed method Remove.So just obtain matching and had to it hole of fine selectivity completely with template molecule on three dimensions, so as to So that template molecule to be enriched with certain matrix.SPE (SPE) is to utilize solid absorbent by the mesh in fluid sample Compound absorption is marked, is separated with the matrix and interfering compound of sample, elution is then used again, separation and enrichment mesh is reached Mark the separating and purifying technology of compound.In addition, molecular engram material has good compatibility and single-minded selectivity, tolerance is high Temperature, high pressure, soda acid and organic solvent, prepare simplicity and are easy to preserve, therefore in SPE, chromatographic isolation analysis, film point From, be widely used in terms of analogue enztme, biomimetic sensor.
But the special construction of witloof acid molecule causes the molecule to form intramolecular hydrogen bond, this effect and the template molecule Interaction between function monomer forms competition, often slackens the recognition performance of the molecularly imprinted polymer of preparation, reduces it The effect of practical application.For example, Saad et al. exists " Preparation and application of molecularly imprinted polymer for isolation of chicoric acid from Chicorium intybus L..medicinal plant”(Analytica Chimica Acta.2015,877:80-89.) with Cichoric acid (CA) for template Molecule, 4-vinylpridine (4-VP) is function monomer, and ethylene glycol dimethacrylate (EDMA) is crosslinking agent, and template point The mol ratio of son, function monomer and crosslinking agent is 1:4:20, it is polymerize using body and precipitation polymerization, has synthesized CA molecular engrams and gathered Compound, Selective Separation goes out Cichoric acid from the analog of Cichoric acid, but sterling Cichoric acid is not obtained from runic thing. Therefore the high Cichoric acid imprinted polymer of synthesis of selective is the significant challenge in a separation actual sample.
The content of the invention
The mesh of the present invention is that this method is using Cichoric acid as mould there is provided a kind of preparation method of Cichoric acid trace integral post Plate molecule, while it is zinc acetate or cobalt acetate to add metal ion, with DMF, dimethyl sulfoxide (DMSO) and 1- fourths Base -3- methyl imidazolium tetrafluoroborates do pore-foaming agent, add function monomer, crosslinking agent and initiator, and then ultrasonic dissolution is removed Dissolved oxygen and initiation reaction are gone, the Cichoric acid molecular engram integral column with high selectivity is finally obtained.The method of the invention Template consumption is few, greatly reduces the trace cost of Cichoric acid.Cichoric acid trace is successfully synthesized in stainless steel tube Integral post, and blank control post and carry out chromatographic performance optimization.Test result indicates that, imprinting factor reaches as high as 24.81.Should Method is prepared simply, and high molecular polymer durability is good, is that isolate and purify (purity is higher than 98.5%) of Cichoric acid provides A kind of cost-effective method.
A kind of preparation method of Cichoric acid trace integral post of the present invention, follows these steps to carry out：
A, by 1.19-23.72mg Cichoric acids and 2.30-18.35mg metal ion it is that zinc acetate or cobalt acetate are dissolved in 240 μ L DMF, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methylimidazole tetrafluoro boric acids In salt, 452-1118 μ L GDMA and 64 μ L 4-vinylpridine are then added, 20mg is added Radical initiator azodiisobutyronitrile, it is ultrasonic 10-20 minutes, remove dissolving in oxygen, after solution completely mix after, will Solution is transferred in long 100mm, diameter 4.6mm stainless steel column, is quickly sealed the two ends of stainless steel column, permanent in temperature 60 C Reacted 18 hours in tepidarium；
B, stainless steel column taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 Methanol-acetic acid 150-200mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and flushing obtains Cichoric acid print after finishing Mark integral post.
The metal ion added in step a is zinc acetate.
A kind of preparation method of Cichoric acid trace integral post of the present invention, the chromatographic performance that this method passes through integral post Evaluate, measure compared to blank control post, Cichoric acid trace integral post is stronger to the retention property of Cichoric acid；Experimental result table Bright, imprinting factor reaches as high as 24.81, and the present invention is isolated and purified (purity is higher than 98.5%) available for Cichoric acid.
Brief description of the drawings
Fig. 1 is Cichoric acid of the present invention in the Cichoric acid trace integral post and its blank control post using zinc acetate as metal ion On different chromatographic behaviors, wherein 1 be blank control pillar, 2 be Cichoric acid trace integral post；
Fig. 2 is Cichoric acid of the present invention in the Cichoric acid trace integral post and its blank control post using cobalt acetate as metal ion On different chromatographic behaviors, wherein 1 be blank control pillar, 2 be Cichoric acid trace integral post；
Fig. 3 is present invention Cichoric acid and its analog in the Cichoric acid trace integral post using zinc acetate as metal ion Chromatogram retains figure, wherein 1 is caffeic acid, 2 be chlorogenic acid, and 3 be Cryptochlorogenic acid, and 4 be corilagin, and 5 be the caffeoyl Kuis of 4,5- bis- Peaceful acid, 6 be Cichoric acid；
Fig. 4 is that present invention chromatogram of Cichoric acid and its analog on the blank control post using zinc acetate as metal ion is protected Figure is stayed, wherein 1 is caffeic acid, 2 be chlorogenic acid, and 3 be Cryptochlorogenic acid, and 4 be Isochlorogenic acid C, and 5 be corilagin, 6 For Cichoric acid.
Embodiment
With reference to specific embodiment, the present invention is further elaborated on.
Embodiment 1
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 11.86mg Cichoric acids and 9.18mg metal ion it is that zinc acetate is dissolved in 240 μ L N, N- dimethyl methyls In acid amides, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, 745 μ L are then added GDMA and 64 μ L 4-vinylpridine, add the 20mg isobutyl of radical initiator azo two Nitrile, ultrasound 10 minutes removes the oxygen in dissolving, and after solution is mixed completely, solution is transferred into long 100mm, diameter 4.6mm Stainless steel column in, quickly the two ends of stainless steel column are sealed, in temperature 60 C water bath with thermostatic control react 18 hours；
B, stainless steel column taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 Methanol-acetic acid 150mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and it is whole that flushing obtains Cichoric acid trace after finishing Scapus.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined, with thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 27.30.
Embodiment 2
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 11.86mg Cichoric acids and 9.18mg metal ion it is that zinc acetate is dissolved in 240 μ L N, N- dimethyl methyls In acid amides, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, 560 μ L are then added GDMA and 64 μ L 4-vinylpridine, add the 20mg isobutyl of radical initiator azo two Nitrile, ultrasound 15 minutes removes the oxygen in dissolving, and after solution is mixed completely, solution is transferred into long 100mm, diameter 4.6mm Stainless steel column in, quickly the two ends of stainless steel column are sealed, in temperature 60 C water bath with thermostatic control react 18 hours；
B, stainless steel column taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 Methanol-acetic acid 200mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and it is whole that flushing obtains Cichoric acid trace after finishing Scapus.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time t of the Cichoric acid on trace post is determinedR, with thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 24.46.
Embodiment 3
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 11.86mg Cichoric acids and 9.18mg metal ion it is that zinc acetate is dissolved in 240 μ L N, N- dimethyl methyls In acid amides, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, 452 μ L are then added GDMA and 64 μ L 4-vinylpridine, finally add 20mg radical initiator azo two Isobutyronitrile, ultrasound 20 minutes, removing is dissolved in oxygen therein, and after solution is mixed completely, solution is transferred into stainless steel column In (long 100mm, diameter 4.6mm), the two ends of stainless steel column are quickly sealed, are reacted 18 hours in temperature 60 C water bath with thermostatic control；
B, the stainless steel column of synthesis taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 methanol-acetic acid 160mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and flushing can obtain witloof after finishing Sour trace integral post.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined, with thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 29.83.
Embodiment 4
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 11.86mg Cichoric acids and 9.18mg metal ion it is that zinc acetate is dissolved in 240 μ L N, N- dimethyl methyls In acid amides, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, 1118 μ L are then added GDMA and 64 μ L 4-vinylpridine, finally add 20mg radical initiator azo two Isobutyronitrile, ultrasound 20 minutes, removing is dissolved in oxygen therein, and after solution is mixed completely, solution is transferred into stainless steel column In (long 100mm, diameter 4.6mm), the two ends of stainless steel column are quickly sealed, are reacted 18 hours in temperature 60 C water bath with thermostatic control；
B, the stainless steel column of synthesis taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 methanol-acetic acid 160mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and flushing can obtain witloof after finishing Sour trace integral post.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined, with thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 13.43.
Embodiment 5
Situ aggregation method prepares Cichoric acid trace integral post：
A, the N that the metal ion zinc acetate of 11.86mg Cichoric acids and 4.59mg is dissolved in 240 μ L, N- dimethyl formyls In amine, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, then add 745 μ L's The 4-vinylpridine of GDMA and 64 μ L, the radical initiator azo two for finally adding 20mg is different Butyronitrile, ultrasound 15 minutes, removing is dissolved in oxygen therein, after solution is mixed completely, and it is (long that solution is transferred into stainless steel column 100mm, diameter 4.6mm) in, the two ends of stainless steel column are quickly sealed, are reacted 18 hours in temperature 60 C water bath with thermostatic control；
B, the stainless steel column of synthesis taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 methanol-acetic acid 180mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and flushing can obtain witloof after finishing Sour trace integral post.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined.With thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 18.61.
Embodiment 6
Situ aggregation method prepares Cichoric acid trace integral post：
A, the N that the metal ion cobalt acetate of 11.86mg Cichoric acids and 2.30mg is dissolved in 240 μ L, N- dimethyl formyls In amine, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, then add 745 μ L's The 4-vinylpridine of GDMA and 64 μ L, the radical initiator azo two for finally adding 20mg is different Butyronitrile, ultrasonic 10-20 minutes, removing is dissolved in oxygen therein, and after solution is mixed completely, solution is transferred into stainless steel column In (long 100mm, diameter 4.6mm), the two ends of stainless steel column are quickly sealed, are reacted 18 hours in temperature 60 C water bath with thermostatic control；
B, the stainless steel column of synthesis taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 methanol-acetic acid 200mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and flushing can obtain witloof after finishing Sour trace integral post.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined.With thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 22.88.
Embodiment 7
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 11.86mg Cichoric acids and 18.35mg metal ion it is that zinc acetate is dissolved in 240 μ L N, N- dimethyl methyls In acid amides, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, 745 μ L are then added GDMA and 64 μ L 4-vinylpridine, finally add 20mg radical initiator azo two Isobutyronitrile, ultrasonic 10-20 minutes, removing is dissolved in oxygen therein, and after solution is mixed completely, solution is transferred into stainless steel In post (long 100mm, diameter 4.6mm), the two ends of stainless steel column are quickly sealed, 18 are reacted in temperature 60 C water bath with thermostatic control small When；
B, the stainless steel column of synthesis taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 methanol-acetic acid 190mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and flushing can obtain witloof after finishing Sour trace integral post.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined.With thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 16.34.
Embodiment 8
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 11.86mg Cichoric acids and 5.90mg metal ion it is that cobalt acetate is dissolved in 240 μ L N, N- dimethyl methyls In acid amides, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, 745 μ L are then added GDMA and 64 μ L 4-vinylpridine, add the 20mg isobutyl of radical initiator azo two Nitrile, ultrasound 10 minutes removes the oxygen in dissolving, and after solution is mixed completely, solution is transferred into long 100mm, diameter 4.6mm Stainless steel column in, quickly the two ends of stainless steel column are sealed, in temperature 60 C water bath with thermostatic control react 18 hours；
B, stainless steel column taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 Methanol-acetic acid 150mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and it is whole that flushing obtains Cichoric acid trace after finishing Scapus.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined, with thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 3.41.
Embodiment 9
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 1.19mg Cichoric acids and 9.18mg metal ion it is that zinc acetate is dissolved in 240 μ L N, N- dimethyl formyls In amine, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, then add 745 μ L's The 4-vinylpridine of GDMA and 64 μ L, adds the 20mg isobutyl of radical initiator azo two Nitrile, ultrasound 10 minutes removes the oxygen in dissolving, and after solution is mixed completely, solution is transferred into long 100mm, diameter 4.6mm Stainless steel column in, quickly the two ends of stainless steel column are sealed, in temperature 60 C water bath with thermostatic control react 18 hours；
B, stainless steel column taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 Methanol-acetic acid 150mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and it is whole that flushing obtains Cichoric acid trace after finishing Scapus.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined, with thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 1.20.
Embodiment 10
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 23.72mg Cichoric acids and 9.18mg metal ion it is that zinc acetate is dissolved in 240 μ L N, N- dimethyl methyls In acid amides, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, 745 μ L are then added GDMA and 64 μ L 4-vinylpridine, add the 20mg isobutyl of radical initiator azo two Nitrile, ultrasound 10 minutes removes the oxygen in dissolving, and after solution is mixed completely, solution is transferred into long 100mm, diameter 4.6mm Stainless steel column in, quickly the two ends of stainless steel column are sealed, in temperature 60 C water bath with thermostatic control react 18 hours；
B, stainless steel column taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 Methanol-acetic acid 150mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and it is whole that flushing obtains Cichoric acid trace after finishing Scapus.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined, with thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 9.77.
Embodiment 11
Situ aggregation method prepares Cichoric acid trace integral post：
A, by 5.93mg Cichoric acids and 9.18mg metal ion it is that zinc acetate is dissolved in 240 μ L N, N- dimethyl formyls In amine, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, then add 745 μ L's The 4-vinylpridine of GDMA and 64 μ L, adds the 20mg isobutyl of radical initiator azo two Nitrile, ultrasound 10 minutes removes the oxygen in dissolving, and after solution is mixed completely, solution is transferred into long 100mm, diameter 4.6mm Stainless steel column in, quickly the two ends of stainless steel column are sealed, in temperature 60 C water bath with thermostatic control react 18 hours；
B, stainless steel column taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 Methanol-acetic acid 150mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and it is whole that flushing obtains Cichoric acid trace after finishing Scapus.
Chromatographic performance evaluation is carried out to Cichoric acid trace integral post with high performance liquid chromatography：UV absorption wavelength is set 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with volume ratio 9:1 acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time tR of the Cichoric acid on trace post is determined, with thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in trace post Retention factors reach 7.19.
Embodiment 12
Situ aggregation method prepares Cichoric acid blank control integral post：
A, by 9.18mg metal ion it is that zinc acetate is dissolved in 240 μ L DMF, the two of 1200 μ L In methyl sulfoxide and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates, 745 μ L dimethacrylate second is then added The 4-vinylpridine of diol ester and 64 μ L, finally adds 20mg radical initiator azodiisobutyronitrile, ultrasonic 10-20 Minute, removing is dissolved in oxygen therein, and after solution is mixed completely, solution is transferred into stainless steel column (long 100mm, diameter In 4.6mm), the two ends of stainless steel column are quickly sealed, are reacted 18 hours in temperature 60 C water bath with thermostatic control；
B, the stainless steel column of synthesis taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 methanol-acetic acid 150-200mL mixed liquor rinses integral post, and flow velocity is 1.0mL/min, and flushing can obtain after finishing Cichoric acid trace integral post.
Chromatographic performance evaluation is carried out to blank control integral post integral post with high performance liquid chromatography：UV absorption ripple is set Long 326nm, flow velocity is 1.0mL/min, 25 DEG C of column temperature, with acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) (9:1,v/v) Cichoric acid trace integral post is rinsed to baseline values, sample introduction, retention time t of the Cichoric acid on trace post is determinedR.With thousand points One of acetone demarcate the dead time t of pillar0, k=(t are distinguished according to formulaR-t0)/t0, Cichoric acid is calculated in blank pair 1.10 are reached according to integral post retention factors, less than the retention on Cichoric acid trace post, is shown in the presence of a small amount of template, chrysanthemum Lettuce acid is by success blot, and imprinting factor passes through formula IF=kMIP/kNIPCalculate, reached 24.81, imprinting effect shows in Fig. 1.
Embodiment 13
The selection performance evaluation of Cichoric acid trace integral post and its blank control post：
Cichoric acid trace integral post selects performance evaluation：UV absorption wavelength 326nm is set, and flow velocity is 1.0mL/min, post 25 DEG C of temperature, with acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) (9:1, v/v) Cichoric acid trace integral post is rinsed to base Line level, difference sample introduction chlorogenic acid, Cryptochlorogenic acid, caffeic acid, Isochlorogenic acid C, corilagin obtains Cichoric acid Chromatogram of the analog in trace integral post, as shown in Fig. 2 reservation of the Cichoric acid trace integral post to Cichoric acid is stronger, Efficiently separating for Cichoric acid and its analogue can be realized；
Blank control post selects performance evaluation：UV absorption wavelength 326nm is set, and flow velocity is 1.0mL/min, column temperature 25 DEG C, with acetonitrile-acetate buffer salt (pH 3.6,200mmol/L) (9:1, v/v) Cichoric acid trace integral post is rinsed to baseline water Flat, difference sample introduction chlorogenic acid, Cryptochlorogenic acid, caffeic acid, Isochlorogenic acid C, corilagin obtains the class of Cichoric acid Like chromatogram of the thing in trace integral post, as shown in figure 3, blank control integral post post can not realize Cichoric acid and its structure class Efficiently separated like thing.

Claims (2)

1. a kind of preparation method of Cichoric acid trace integral post, it is characterised in that follow these steps to carry out：
A, by 1.19-23.72mg Cichoric acids and 2.30-18.35mg metal ion it is that zinc acetate or cobalt acetate are dissolved in 240 μ L DMF, 1200 μ L dimethyl sulfoxide (DMSO) and 2468 μ L 1- butyl -3- methyl imidazolium tetrafluoroborates In, 452-1118 μ L GDMA and 64 μ L 4-vinylpridine are then added, adds 20mg's Radical initiator azodiisobutyronitrile, it is ultrasonic 10-20 minutes, the oxygen in dissolving is removed, will be molten after solution is mixed completely Liquid is transferred in long 100 mm, the mm of diameter 4.6 stainless steel column, is quickly sealed the two ends of stainless steel column, in temperature 60 C Reacted 18 hours in water bath with thermostatic control；
B, stainless steel column taken out from water-bath, load onto column cap, be connected on high pressure pump, with volume ratio 9:1 methanol- Acetic acid 150-200 mL mixed liquor rinses integral post, and flow velocity is 1.0 mL/min, and flushing obtains Cichoric acid trace after finishing Integral post.
2. a kind of preparation method of Cichoric acid trace integral post as described in right 1, it is characterised in that the metal added in step a Ion is zinc acetate.
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CN112275266A (en) * 2020-10-21 2021-01-29 中国科学院新疆理化技术研究所 Preparation method of scopoletin imprinted monolithic column
CN112275265A (en) * 2020-10-21 2021-01-29 中国科学院新疆理化技术研究所 Preparation method of syringin imprinted monolithic column

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CN112275266A (en) * 2020-10-21 2021-01-29 中国科学院新疆理化技术研究所 Preparation method of scopoletin imprinted monolithic column
CN112275265A (en) * 2020-10-21 2021-01-29 中国科学院新疆理化技术研究所 Preparation method of syringin imprinted monolithic column

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