CN113274993A - Preparation method of silica gel matrix chromatographic packing for separating strong-polarity drugs - Google Patents
Preparation method of silica gel matrix chromatographic packing for separating strong-polarity drugs Download PDFInfo
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- CN113274993A CN113274993A CN202110570901.4A CN202110570901A CN113274993A CN 113274993 A CN113274993 A CN 113274993A CN 202110570901 A CN202110570901 A CN 202110570901A CN 113274993 A CN113274993 A CN 113274993A
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- silica gel
- chromatographic packing
- porous silica
- gel matrix
- drying
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- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 title claims abstract description 90
- 239000000741 silica gel Substances 0.000 title claims abstract description 89
- 229910002027 silica gel Inorganic materials 0.000 title claims abstract description 89
- 239000003814 drug Substances 0.000 title claims abstract description 44
- 229940079593 drug Drugs 0.000 title claims abstract description 41
- 238000012856 packing Methods 0.000 title claims abstract description 32
- 239000011159 matrix material Substances 0.000 title claims abstract description 22
- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 239000004005 microsphere Substances 0.000 claims abstract description 37
- 239000000463 material Substances 0.000 claims abstract description 22
- 239000002777 nucleoside Substances 0.000 claims abstract description 17
- 150000003833 nucleoside derivatives Chemical class 0.000 claims abstract description 16
- 238000000926 separation method Methods 0.000 claims abstract description 14
- 238000013375 chromatographic separation Methods 0.000 claims abstract description 11
- 238000006116 polymerization reaction Methods 0.000 claims abstract description 11
- GRYLNZFGIOXLOG-UHFFFAOYSA-N Nitric acid Chemical compound O[N+]([O-])=O GRYLNZFGIOXLOG-UHFFFAOYSA-N 0.000 claims abstract description 10
- 229910017604 nitric acid Inorganic materials 0.000 claims abstract description 10
- 239000003431 cross linking reagent Substances 0.000 claims abstract description 9
- 239000003999 initiator Substances 0.000 claims abstract description 9
- 239000011148 porous material Substances 0.000 claims abstract description 7
- 238000002791 soaking Methods 0.000 claims abstract description 3
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 claims description 42
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 32
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 30
- 238000005406 washing Methods 0.000 claims description 26
- OZAIFHULBGXAKX-UHFFFAOYSA-N 2-(2-cyanopropan-2-yldiazenyl)-2-methylpropanenitrile Chemical group N#CC(C)(C)N=NC(C)(C)C#N OZAIFHULBGXAKX-UHFFFAOYSA-N 0.000 claims description 22
- 238000001035 drying Methods 0.000 claims description 22
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 21
- 238000010438 heat treatment Methods 0.000 claims description 17
- 238000000034 method Methods 0.000 claims description 16
- 229960002555 zidovudine Drugs 0.000 claims description 12
- HBOMLICNUCNMMY-XLPZGREQSA-N zidovudine Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](N=[N+]=[N-])C1 HBOMLICNUCNMMY-XLPZGREQSA-N 0.000 claims description 11
- 230000004048 modification Effects 0.000 claims description 10
- 238000012986 modification Methods 0.000 claims description 10
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- HRPVXLWXLXDGHG-UHFFFAOYSA-N Acrylamide Chemical compound NC(=O)C=C HRPVXLWXLXDGHG-UHFFFAOYSA-N 0.000 claims description 8
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 claims description 8
- 239000000178 monomer Substances 0.000 claims description 8
- KVNYFPKFSJIPBJ-UHFFFAOYSA-N 1,2-diethylbenzene Chemical compound CCC1=CC=CC=C1CC KVNYFPKFSJIPBJ-UHFFFAOYSA-N 0.000 claims description 7
- 230000004913 activation Effects 0.000 claims description 7
- 239000002245 particle Substances 0.000 claims description 7
- 238000002444 silanisation Methods 0.000 claims description 7
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 6
- CERQOIWHTDAKMF-UHFFFAOYSA-N Methacrylic acid Chemical compound CC(=C)C(O)=O CERQOIWHTDAKMF-UHFFFAOYSA-N 0.000 claims description 6
- 239000003153 chemical reaction reagent Substances 0.000 claims description 6
- 239000008367 deionised water Substances 0.000 claims description 6
- 229910021641 deionized water Inorganic materials 0.000 claims description 6
- 238000010828 elution Methods 0.000 claims description 6
- 235000019441 ethanol Nutrition 0.000 claims description 6
- 230000007935 neutral effect Effects 0.000 claims description 6
- 239000012429 reaction media Substances 0.000 claims description 6
- 238000010992 reflux Methods 0.000 claims description 6
- 239000006228 supernatant Substances 0.000 claims description 6
- DBCAQXHNJOFNGC-UHFFFAOYSA-N 4-bromo-1,1,1-trifluorobutane Chemical compound FC(F)(F)CCCBr DBCAQXHNJOFNGC-UHFFFAOYSA-N 0.000 claims description 5
- STVZJERGLQHEKB-UHFFFAOYSA-N ethylene glycol dimethacrylate Substances CC(=C)C(=O)OCCOC(=O)C(C)=C STVZJERGLQHEKB-UHFFFAOYSA-N 0.000 claims description 5
- 239000002126 C01EB10 - Adenosine Substances 0.000 claims description 4
- OKKRPWIIYQTPQF-UHFFFAOYSA-N Trimethylolpropane trimethacrylate Chemical compound CC(=C)C(=O)OCC(CC)(COC(=O)C(C)=C)COC(=O)C(C)=C OKKRPWIIYQTPQF-UHFFFAOYSA-N 0.000 claims description 4
- 229960005305 adenosine Drugs 0.000 claims description 4
- 239000012156 elution solvent Substances 0.000 claims description 4
- BDAGIHXWWSANSR-UHFFFAOYSA-N methanoic acid Natural products OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 claims description 4
- 125000004368 propenyl group Chemical group C(=CC)* 0.000 claims description 3
- OSWFIVFLDKOXQC-UHFFFAOYSA-N 4-(3-methoxyphenyl)aniline Chemical compound COC1=CC=CC(C=2C=CC(N)=CC=2)=C1 OSWFIVFLDKOXQC-UHFFFAOYSA-N 0.000 claims description 2
- VGGSQFUCUMXWEO-UHFFFAOYSA-N Ethene Chemical compound C=C VGGSQFUCUMXWEO-UHFFFAOYSA-N 0.000 claims description 2
- 239000005977 Ethylene Substances 0.000 claims description 2
- 239000003795 chemical substances by application Substances 0.000 claims description 2
- 235000019253 formic acid Nutrition 0.000 claims description 2
- 229960001627 lamivudine Drugs 0.000 claims description 2
- JTEGQNOMFQHVDC-NKWVEPMBSA-N lamivudine Chemical compound O=C1N=C(N)C=CN1[C@H]1O[C@@H](CO)SC1 JTEGQNOMFQHVDC-NKWVEPMBSA-N 0.000 claims description 2
- 229920000642 polymer Polymers 0.000 abstract description 4
- 238000006243 chemical reaction Methods 0.000 abstract description 3
- 229910052710 silicon Inorganic materials 0.000 abstract description 3
- 239000010703 silicon Substances 0.000 abstract description 3
- 230000003213 activating effect Effects 0.000 abstract 1
- 230000000877 morphologic effect Effects 0.000 abstract 1
- 239000003960 organic solvent Substances 0.000 abstract 1
- XDLMVUHYZWKMMD-UHFFFAOYSA-N 3-trimethoxysilylpropyl 2-methylprop-2-enoate Chemical compound CO[Si](OC)(OC)CCCOC(=O)C(C)=C XDLMVUHYZWKMMD-UHFFFAOYSA-N 0.000 description 8
- 239000002904 solvent Substances 0.000 description 6
- 230000005526 G1 to G0 transition Effects 0.000 description 4
- 230000009286 beneficial effect Effects 0.000 description 4
- 239000000969 carrier Substances 0.000 description 4
- RQFQJYYMBWVMQG-IXDPLRRUSA-N chitotriose Chemical compound O[C@@H]1[C@@H](N)[C@H](O)O[C@H](CO)[C@H]1O[C@H]1[C@H](N)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)N)[C@@H](CO)O1 RQFQJYYMBWVMQG-IXDPLRRUSA-N 0.000 description 4
- 239000004088 foaming agent Substances 0.000 description 4
- 239000011259 mixed solution Substances 0.000 description 4
- 235000010724 Wisteria floribunda Nutrition 0.000 description 3
- 229930182494 ginsenoside Natural products 0.000 description 3
- RQBBFKINEJYDOB-UHFFFAOYSA-N acetic acid;acetonitrile Chemical compound CC#N.CC(O)=O RQBBFKINEJYDOB-UHFFFAOYSA-N 0.000 description 2
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 2
- 238000012512 characterization method Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 229920000344 molecularly imprinted polymer Polymers 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 1
- UFNDONGOJKNAES-UHFFFAOYSA-N Ginsenoside Rb1 Natural products CC(=CCCC(C)(OC1OC(COC2OC(CO)C(O)C(O)C2O)C(O)C(O)C1O)C3CCC4(C)C3C(O)CC5C6(C)CCC(OC7OC(CO)C(O)C(O)C7OC8OC(CO)C(O)C(O)C8O)C(C)(C)C6CC(O)C45C)C UFNDONGOJKNAES-UHFFFAOYSA-N 0.000 description 1
- CDXSJGDDABYYJV-UHFFFAOYSA-N acetic acid;ethanol Chemical compound CCO.CC(O)=O CDXSJGDDABYYJV-UHFFFAOYSA-N 0.000 description 1
- 229960004150 aciclovir Drugs 0.000 description 1
- 229960000643 adenine Drugs 0.000 description 1
- 229930013930 alkaloid Natural products 0.000 description 1
- 125000000304 alkynyl group Chemical group 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- -1 and then Substances 0.000 description 1
- IVRMZWNICZWHMI-UHFFFAOYSA-N azide group Chemical group [N-]=[N+]=[N-] IVRMZWNICZWHMI-UHFFFAOYSA-N 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 125000003636 chemical group Chemical group 0.000 description 1
- 239000007810 chemical reaction solvent Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- SVWLIIFHXFGESG-UHFFFAOYSA-N formic acid;methanol Chemical compound OC.OC=O SVWLIIFHXFGESG-UHFFFAOYSA-N 0.000 description 1
- 229940089161 ginsenoside Drugs 0.000 description 1
- GZYPWOGIYAIIPV-JBDTYSNRSA-N ginsenoside Rb1 Chemical compound C([C@H]1O[C@H]([C@@H]([C@@H](O)[C@@H]1O)O)O[C@@](C)(CCC=C(C)C)[C@@H]1[C@@H]2[C@@]([C@@]3(CC[C@H]4C(C)(C)[C@@H](O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O5)O[C@H]5[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O5)O)CC[C@]4(C)[C@H]3C[C@H]2O)C)(C)CC1)O[C@@H]1O[C@H](CO)[C@@H](O)[C@H](O)[C@H]1O GZYPWOGIYAIIPV-JBDTYSNRSA-N 0.000 description 1
- TXEWRVNOAJOINC-UHFFFAOYSA-N ginsenoside Rb2 Natural products CC(=CCCC(OC1OC(COC2OCC(O)C(O)C2O)C(O)C(O)C1O)C3CCC4(C)C3C(O)CC5C6(C)CCC(OC7OC(CO)C(O)C(O)C7OC8OC(CO)C(O)C(O)C8O)C(C)(C)C6CCC45C)C TXEWRVNOAJOINC-UHFFFAOYSA-N 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 239000012535 impurity Substances 0.000 description 1
- 238000002329 infrared spectrum Methods 0.000 description 1
- 239000002608 ionic liquid Substances 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 238000004811 liquid chromatography Methods 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 238000001000 micrograph Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 125000003835 nucleoside group Chemical group 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 238000004366 reverse phase liquid chromatography Methods 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 229960001203 stavudine Drugs 0.000 description 1
- 125000000391 vinyl group Chemical group [H]C([*])=C([H])[H] 0.000 description 1
- 229920002554 vinyl polymer Polymers 0.000 description 1
- 229940120938 zidovudine and lamivudine Drugs 0.000 description 1
Images
Classifications
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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
- B01J20/281—Sorbents specially adapted for preparative, analytical or investigative chromatography
-
- 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
- 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
- B01J20/28016—Particle form
- B01J20/28021—Hollow particles, e.g. hollow spheres, microspheres or cenospheres
-
- 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
- B01J20/281—Sorbents specially adapted for preparative, analytical or investigative chromatography
- B01J20/282—Porous sorbents
-
- 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
- B01J20/281—Sorbents specially adapted for preparative, analytical or investigative chromatography
- B01J20/282—Porous sorbents
- B01J20/283—Porous sorbents based on silica
-
- 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
- B01J20/281—Sorbents specially adapted for preparative, analytical or investigative chromatography
- B01J20/286—Phases chemically bonded to a substrate, e.g. to silica or to polymers
-
- 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
- G01N30/60—Construction of the column
- G01N30/6095—Micromachined or nanomachined, e.g. micro- or nanosize
Abstract
The invention belongs to the field of preparation of novel chromatographic packing, and discloses a preparation method of silica gel matrix chromatographic packing for chromatographic separation of strong-polarity drugs, which comprises the following steps: soaking silica gel in dilute nitric acid, and activating silicon hydroxyl on the surface of the silica gel; modifying double bonds on the surface of the silica gel; taking nucleoside drugs as template molecules to prepare a prepolymerization system; adding an initiator, a cross-linking agent and double-bond modified silica gel into the prepolymer system to perform imprinting polymerization reaction on the surface of the silica gel; and (3) eluting the silica gel obtained by the polymerization reaction by using an organic solvent, and removing template molecules to obtain the porous silica gel microspheres coated by the imprinted polymer. The invention combines the excellent selectivity of the molecular imprinting polymer with the properties of the silica gel material such as good pore structure, morphological characteristics, mechanical strength and the like, and controls the reaction conditions, so that the obtained chromatographic packing has high chromatographic separation column efficiency, high separation selectivity, good chromatographic peak symmetry and good separation reproducibility.
Description
Technical Field
The invention belongs to the field of preparation of novel chromatographic packing, and particularly relates to a preparation method of silica gel matrix chromatographic packing for chromatographic separation of strong-polarity drugs.
Background
Recently, with the continuous development of the biomedical industry, strong polar drugs (ginsenosides, nucleosides, sugar amino acids, alkaloids, and the like) attract extensive attention of researchers. The method has important significance for separating and analyzing strong-polarity drugs, improving the drug safety, analyzing the drug metabolic process and the like.
High performance liquid chromatography is most commonly used among many analytical detection methods for highly polar drugs today, however, it is difficult to efficiently separate them by conventional reverse phase chromatography columns. Therefore, the development of novel chromatographic separation materials is of great significance. At present, in order to solve the difficult problem of chromatographic separation of strong polar drugs, researchers have developed a series of novel chromatographic stationary phases, including novel chromatographic separation materials such as ionic liquid modified porous silica gel, carbon point modified porous silica gel, and polymer modified porous silica gel, for example, CN101757897A discloses a preparation method of chitosan oligosaccharide hydrophilic chromatographic stationary phases, which adopts "link chemistry" as a bonding reaction method to bond chitosan oligosaccharide, firstly, a terminal alkynyl is introduced on the surface of silica gel, and then, water, methanol or a mixture of these solvents is used as a reaction solvent to bond chitosan oligosaccharide modified with an azide group to the surface of silica gel, so as to obtain the chitosan oligosaccharide hydrophilic chromatographic stationary phase.
However, strongly polar drugs are still difficult to isolate efficiently. Therefore, the development of a novel liquid chromatography stationary phase with unique selectivity is of great significance for solving the problems.
Disclosure of Invention
In view of the above-mentioned disadvantages of the prior art, the present invention aims to provide a method for preparing a silica gel matrix chromatographic packing for chromatographic separation of strongly polar drugs.
In order to achieve the above purpose, the invention provides a preparation method of silica gel matrix chromatographic packing for chromatographic separation of strong polar drugs, which adopts the following technical scheme:
a method for preparing silica gel matrix chromatographic packing for separating strong polar drugs comprises the following steps:
1) silica gel selection and activation treatment: selecting porous silica gel microspheres with the particle size of 2-5 microns and the pore diameter of 15-30 nm as a carrier, soaking the porous silica gel microspheres by using dilute nitric acid with the mass fraction of 8-20%, washing the porous silica gel microspheres by using deionized water until the supernatant is neutral, washing the porous silica gel microspheres for 1-3 times by using absolute ethyl alcohol, and drying the porous silica gel microspheres;
2) silica gel surface modification double bond: adding an ethylene-based or propenyl silanization reagent into anhydrous toluene serving as a reaction medium, heating and refluxing for 8-24 h, washing with the anhydrous toluene and ethanol or methanol in sequence to obtain double-bond modified porous silica gel microspheres, and drying;
3) constructing a prepolymerization system: selecting one or more than two nucleoside drugs as template molecules, methacrylic acid or acrylamide as a functional monomer, and methanol, ethanol or acetonitrile as a pore-forming agent, and carrying out prepolymerization for 5-20 h at-5-10 ℃ to obtain a prepolymerization system;
4) polymerization reaction: slowly adding (one drop in liquid for about 1-5 seconds and 10 mg in solid for about 1-5 seconds) an initiator, a cross-linking agent and double-bond modified porous silica gel microspheres into a prepolymerization system, and reacting for 12-48 h at 30-70 ℃;
5) elution of template molecules: eluting the chromatographic separation material for 6-18 times by using an elution solvent, and drying for later use.
Preferably, in the step 1), a vacuum oven is adopted for drying, the temperature of the vacuum oven is set to be 50-100 ℃, and the time is 5-16 h.
Preferably, in the step 2), the toluene is adopted for washing for 2-5 times, and the methanol or ethanol is adopted for washing for 2-5 times.
Preferably, in the step 2), heating and drying are adopted, wherein the heating and drying temperature is 50-80 ℃, and the time is 6-16 h.
Preferably, in the step 4), the initiator is Azobisisobutyronitrile (AIBN); the crosslinking agent is one or more of trimethylolpropane Trimethacrylate (TRIM), Ethylene Glycol Dimethacrylate (EGDMA) or Diethylbenzene (DVB).
Preferably, the drying temperature in the step (5) is 50-70 ℃, and the time is 8-24 h.
The activation of the dilute nitric acid in the step 1) can effectively reduce metal impurities on the surface of the silica gel material, increase the content of silicon hydroxyl on the surface of the silica gel, and is beneficial to subsequent silanization modification and imprinting polymerization reaction on the surface of the porous silica gel.
In the step 2), modification of vinyl or propenyl is beneficial to guiding the imprinting polymerization reaction to be carried out on the surface of the porous silica gel orderly and controllably, and is beneficial to carrying out end capping treatment on silicon hydroxyl on the surface of the silica gel. Is beneficial to effectively improving the chromatographic separation performance of the material.
In the step 3), the nucleoside medicament can be obtained by market purchase, and can be made in China or imported, and three conditions are required to be met: 1) the stability of the molecule is good, and the molecule does not contain chemical groups which can prevent or influence polymerization reaction; 2) the molecule has carboxyl, hydroxyl and other groups capable of interacting with the functional monomer; 3) the template molecules have stronger rigidity in the imprinting process and are convenient to elute.
In some preferred embodiments of the present invention, the nucleoside drug is any one or a combination of zidovudine, lamivudine or adenosine.
In the step 3), methacrylic acid or acrylamide is used as a functional monomer, the methacrylic acid has carboxyl, the acrylamide has amino, and the methacrylic acid and the acrylamide can respectively act with template molecules to form a molecularly imprinted polymer cavity, and the formed polymers respectively have different charge characteristics, so that different separation effects can be realized.
The elution solvent in the step 5) is one or more than two of methanol, acetonitrile, acetic acid, formic acid, water and other solvents.
In the step 1), dilute nitric acid with the mass fraction of 8-20% is adopted to soak the porous silica gel microspheres for 1-5 hours.
The porous silica gel microspheres can be obtained commercially, and can be imported or made in China.
Compared with the prior art, the invention takes porous silica gel microspheres as a carrier, selects nucleoside drugs as template molecules, and provides a preparation method of a novel silica gel matrix chromatographic packing by means of a surface imprinting technology.
Drawings
Other features, objects and advantages of the invention will become more apparent upon reading of the detailed description of non-limiting embodiments with reference to the following drawings:
FIG. 1 is a schematic diagram of the preparation of silica gel matrix chromatographic packing for strong polar drug separation of example 1;
FIG. 2 is a scanning electron microscope image of the porous silica gel microspheres (bare silica gel carrier) (a) used in example 1 and the surface imprinted chromatographic packing (b) of the nucleoside drug zidovudine;
FIG. 3 is an infrared spectrum of the porous silica gel microspheres (a), the surface imprinted chromatographic packing (b) of the nucleoside drug zidovudine and the non-imprinted material (c) used in example 1;
FIG. 4 is a chromatogram separation chart of C18 packing (a), nucleoside drug zidovudine surface imprinting chromatogram packing (b) and non-imprinting material for four nucleoside drugs (C);
FIG. 5 is the chromatogram separation chart of the chromatographic packing of surface imprinting of nucleoside drug zidovudine against ginsenosides.
FIG. 6 is the reproducibility of the continuous separation result of nucleoside drugs by the surface imprinted chromatographic packing of the nucleoside drugs zidovudine.
Detailed Description
The present invention will be described in detail with reference to specific examples. The following examples will assist those skilled in the art in further understanding the invention, but are not intended to limit the invention in any way. It should be noted that variations and modifications can be made by persons skilled in the art without departing from the spirit of the invention. All falling within the scope of the present invention.
Example 1
As shown in fig. 1, a method for preparing a silica gel matrix chromatographic packing for separating a strong polar drug comprises the following steps:
(1) silica gel selection and activation treatment: porous silica gel microspheres (purchased from FuJi, Japan) with the particle size of 5 mu m and the pore diameter of 20nm are selected as carriers, are soaked in dilute nitric acid with the mass fraction of 15% for 2 hours, are washed by deionized water until the supernatant is neutral, are washed by absolute ethyl alcohol for 3 times, and are dried in vacuum for 8 hours at the temperature of 55 ℃.
(2) Silica gel surface modification double bond: adding a silanization reagent gamma-methacryloxypropyltrimethoxysilane (gamma-MPS, KH570) into anhydrous toluene serving as a reaction medium, heating and refluxing for 24h, washing for 1 time by using the anhydrous toluene and washing for 3 times by using anhydrous ethanol in sequence to obtain double-bond modified porous silica gel microspheres, and heating for 8h and drying at the temperature of 55 ℃.
(3) Constructing a prepolymerization system: zidovudine is selected as a template molecule, methacrylic acid is taken as a functional monomer, methanol is taken as a pore-foaming agent, and prepolymerization is carried out for 12h at 4 ℃.
(4) Polymerization reaction: slowly adding an initiator Azobisisobutyronitrile (AIBN), a crosslinking agent Ethylene Glycol Dimethacrylate (EGDMA) and double-bond modified (double-bond functionalized) porous silica gel microspheres into a prepolymerization system under the reaction condition of 50 ℃ for 12 hours, then continuously heating to 60 ℃ and continuously reacting for 24 hours;
(5) elution of template molecules: eluting the solvent with a methanol-formic acid mixed solution with a ratio of 9:1, washing the material for 6 times, washing the material with absolute ethanol for 2 times, and drying at 55 ℃ for 8h for later use.
The characterization result shows that the particle size of the obtained material is uniformly distributed at 5 μm, and the material has a better spherical shape (see figure 2); the infrared characterization result proves that the molecularly imprinted polymer is successfully modified on the surface of the porous silica gel (see figure 3). The obtained nucleoside medicament zidovudine surface imprinted chromatographic packing can realize baseline separation on four nucleoside medicaments (1-adenine, 2-acyclovir, 3-stavudine and 4-zidovudine), the separation column efficiency on four targets is respectively 15,300, 19,300, 17,100 and 12,600 theoretical plate number per meter, and the separation result is superior to that of C18 and non-imprinted materials (see figure 4). In addition, the obtained imprinted material can realize baseline separation on ginsenoside compounds, see fig. 5, wherein the separation column efficiency on ginsenoside Rb1 can reach 8,600 theoretical plate number per meter; after the obtained imprinted material is continuously used for 17 days, the column effect, the retention factor, the peak area, the chromatographic peak shape symmetry and the like are good, and the good stability is shown, which is shown in figure 6.
Example 2
A method for preparing silica gel matrix chromatographic packing for separating strong polar drugs comprises the following steps:
(1) silica gel selection and activation treatment: porous silica gel microspheres (purchased from FuJi, Japan) with the particle size of 5 mu m and the pore diameter of 15nm are selected as carriers, the porous silica gel microspheres are soaked by dilute nitric acid with the mass fraction of 10 percent for 1.5h, then the porous silica gel microspheres are washed by deionized water until the supernatant is neutral, and then the porous silica gel microspheres are washed by absolute ethyl alcohol for 2 times and are dried in vacuum for 8h at the temperature of 60 ℃.
(2) Silica gel surface modification double bond: anhydrous toluene is taken as a reaction medium, a silanization reagent gamma-methacryloxypropyltrimethoxysilane (gamma-MPS) is added, heating and refluxing are carried out for 12 hours, the anhydrous toluene is adopted for washing for 2 times, and the anhydrous ethanol is adopted for washing for 2 times in sequence, so that double-bond modified porous silica gel microspheres are obtained, and the double-bond modified porous silica gel microspheres are heated for 8 hours and dried at the temperature of 60 ℃.
(3) Constructing a prepolymerization system: zidovudine and lamivudine are selected as double-template molecules, acrylamide is used as a functional monomer, acetonitrile is used as a pore-foaming agent, and prepolymerization is carried out for 16h at the temperature of 8 ℃.
(4) Polymerization reaction: slowly adding an initiator Azobisisobutyronitrile (AIBN), a cross-linking agent Diethylbenzene (DVB) and double-bond functionalized porous silica gel microspheres into a prepolymerization system, reacting for 15 hours under the condition of 58 ℃, then continuously heating to 60 ℃, and continuously reacting for 18 hours; (5) elution of template molecules: eluting the solvent with ethanol-acetic acid mixed solution at a ratio of 8:3, washing the material for 15 times, washing the material with anhydrous ethanol for 1 time, and drying at 65 ℃ for 18h for later use.
Example 3
A method for preparing silica gel matrix chromatographic packing for separating strong polar drugs comprises the following steps:
(1) silica gel selection and activation treatment: porous silica gel microspheres (purchased from Tianjin Borui) with the particle size of 3 mu m and the pore diameter of 16nm are selected as carriers, are soaked for 1h by using dilute nitric acid with the mass fraction of 20 percent, are washed by deionized water until the supernatant is neutral, are washed by absolute ethyl alcohol for 1 time, and are dried in vacuum for 5h at the temperature of 100 ℃.
(2) Silica gel surface modification double bond: adding a silanization reagent gamma-methacryloxypropyltrimethoxysilane (gamma-MPS) into anhydrous toluene serving as a reaction medium, heating and refluxing for 8 hours, washing for 5 times by using the anhydrous toluene and washing for 5 times by using anhydrous ethanol to obtain double-bond modified porous silica gel microspheres, and heating for 7 hours at 70 ℃ for drying.
(3) Constructing a prepolymerization system: zidovudine and adenosine are selected as double-template molecules, acrylamide is used as a functional monomer, acetonitrile is used as a pore-foaming agent, and prepolymerization is carried out for 16h at the temperature of 8 ℃.
(4) Polymerization reaction: slowly adding an initiator Azobisisobutyronitrile (AIBN), a crosslinking agent trimethylolpropane Trimethacrylate (TRIM) and double-bond functionalized porous silica gel microspheres into a prepolymerization system, reacting for 12h under the condition of 60 ℃, and then continuously heating to 70 ℃ and continuously reacting for 12 h;
(5) elution of template molecules: eluting the solvent with acetonitrile-acetic acid mixed solution at a ratio of 6:1, washing the material for 10 times, washing the material with absolute ethanol for 2 times, and drying at 70 ℃ for 15h for later use.
Example 4
A method for preparing silica gel matrix chromatographic packing for separating strong polar drugs comprises the following steps:
(1) silica gel selection and activation treatment: porous silica gel microspheres (purchased from Fuji of Japan) with the particle size of 5 microns and the pore diameter of 30nm are selected as carriers, are soaked for 3 hours by using dilute nitric acid with the mass fraction of 15 percent, are washed by deionized water until the supernatant is neutral, are washed by absolute ethyl alcohol for 1 time, and are dried in vacuum for 5 hours at the temperature of 100 ℃.
(2) Silica gel surface modification double bond: adding a silanization reagent gamma-methacryloxypropyltrimethoxysilane (gamma-MPS) into anhydrous toluene serving as a reaction medium, heating and refluxing for 8 hours, washing for 5 times by using the anhydrous toluene and washing for 5 times by using anhydrous ethanol to obtain double-bond modified porous silica gel microspheres, and heating for 7 hours at 70 ℃ for drying.
(3) Constructing a prepolymerization system: zidovudine and adenosine are selected as double-template molecules, acrylamide is used as a functional monomer, acetonitrile is used as a pore-foaming agent, and prepolymerization is carried out for 16h at the temperature of-5 ℃.
(4) Polymerization reaction: slowly adding an initiator Azobisisobutyronitrile (AIBN), a crosslinking agent trimethylolpropane Trimethacrylate (TRIM) and double-bond functionalized porous silica gel microspheres into a prepolymerization system, reacting for 12h under the condition of 30 ℃, and then continuously heating to 65 ℃ and continuously reacting for 12 h;
(5) elution of template molecules: eluting the solvent with acetonitrile-acetic acid mixed solution at a ratio of 6:1, washing the material for 10 times, washing the material with absolute ethanol for 2 times, and drying at 70 ℃ for 15h for later use.
The foregoing description of specific embodiments of the present invention has been presented. It is to be understood that the present invention is not limited to the specific embodiments described above, and that various changes and modifications may be made by one skilled in the art within the scope of the appended claims without departing from the spirit of the invention.
Claims (9)
1. A preparation method of silica gel matrix chromatographic packing for separating strong polar drugs is characterized by comprising the following steps:
1) silica gel selection and activation treatment: selecting porous silica gel microspheres with the particle size of 2-5 microns and the pore diameter of 15-30 nm as a carrier, soaking the porous silica gel microspheres by using dilute nitric acid with the mass fraction of 8-20%, washing the porous silica gel microspheres by using deionized water until the supernatant is neutral, washing the porous silica gel microspheres for 1-3 times by using absolute ethyl alcohol, and drying the porous silica gel microspheres;
2) silica gel surface modification double bond: adding an ethylene-based or propenyl silanization reagent into anhydrous toluene serving as a reaction medium, heating and refluxing for 8-24 h, washing with the anhydrous toluene and ethanol or methanol in sequence to obtain double-bond modified porous silica gel microspheres, and drying;
3) constructing a prepolymerization system: selecting one or more than two nucleoside drugs as template molecules, methacrylic acid or acrylamide as a functional monomer, and methanol, ethanol or acetonitrile as a pore-forming agent, and carrying out prepolymerization for 5-20 h at-5-10 ℃ to obtain a prepolymerization system;
4) polymerization reaction: slowly adding an initiator, a cross-linking agent and double-bond modified porous silica gel microspheres into a prepolymerization system, and reacting for 12-48 h at 30-70 ℃;
5) elution of template molecules: eluting the chromatographic separation material for 6-18 times by using an elution solvent, and drying for later use.
2. The preparation method of the silica gel matrix chromatographic packing for separating the strong polar drugs according to claim 1, wherein in the step 1), the drying is performed by using a vacuum oven, and the temperature of the vacuum oven is set to be 50-100 ℃ for 5-16 h.
3. The method for preparing silica gel matrix chromatographic packing for separating strong polar drugs in claim 1, wherein in the step 2), the toluene is used for washing 2-5 times, and the methanol or ethanol is used for washing 2-5 times.
4. The method for preparing the silica gel matrix chromatographic packing for separating the strong polar drugs according to claim 1, wherein in the step 2), heating and drying are adopted, wherein the heating and drying temperature is 50-80 ℃, and the time is 6-16 h.
5. The method for preparing silica gel matrix chromatographic packing for strong polar drug separation according to claim 1, wherein in the step 4), the initiator is azobisisobutyronitrile; the cross-linking agent is one or more of trimethylolpropane trimethacrylate, ethylene glycol dimethacrylate or diethylbenzene.
6. The method for preparing the silica gel matrix chromatographic packing for separating the strong polar drugs according to claim 1, wherein the drying temperature in the step (5) is 50-70 ℃ and the drying time is 8-24 h.
7. The method for preparing the silica gel matrix chromatographic packing for the separation of the strong polar drugs according to any one of claims 1 to 6, wherein the nucleoside drugs are any one or a combination of zidovudine, lamivudine or adenosine.
8. The method for preparing silica gel matrix chromatographic packing for separating strong polar drugs according to claim 1, wherein the elution solvent in the step 5) is any one or two or more of methanol, acetonitrile, acetic acid, formic acid or water.
9. The method for preparing the silica gel matrix chromatographic packing for separating the strong polar drugs according to claim 1, wherein in the step 1), the porous silica gel microspheres are soaked in dilute nitric acid with a mass fraction of 8-20% for 1-5 h.
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| CN114797808A (en) * | 2022-05-11 | 2022-07-29 | 烟台大学 | Fluorine-containing liquid chromatography packing for polar drug separation and preparation method thereof |
| CN114832799A (en) * | 2022-06-01 | 2022-08-02 | 中国科学院兰州化学物理研究所 | Preparation and application of carbon-point bonded silica gel chromatographic stationary phase based on eutectic solvent |
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