capillary columns
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Preparation of organic polymer monolithic columns in fused silica capillaries was aimed at fast gradient separation of proteins. For this purpose, polymerization in situ procedure was optimized, using ethylene dimetacrylate and butyl metacrylate monomers with azobisisobutyronitrile as initiator of the polymerization reaction in presence of non-aqueous porogen solvent mixtures composed of 1-propanol and 1,4-butanediol. The separation of proteins in totally monolithic capillary columns was compared with the chromatography on a new type of "hybrid interparticle monolithic" capillary columns, prepared by in situ polymerization in capillary packed with superficially porous spherical beds, 37-50 microm. The "hybrid" columns showed excellent stability and improved hydrodynamic flow properties with respect to the "totally" monolithic capillary columns. The separation selectivity is similar in the two types of columns. The nature of the superficially porous layer (bare silica or bonded C18 ligands) affects the separation selectivity less significantly than the porosity (density) of the monolithic moiety in the interparticle space, controlled by the composition of the polymerization mixture. The retention behaviour of proteins on all prepared columns is consistent with the reversed-phase gradient elution theory.
In this paper, the concentration of N-isopropylacrylamide in the polymerization mixture has been varied to prepare several polymethacrylate monolithic capillary columns. Polymer monoliths combining N-isopropylacrylamide with zwitterion monomer, as well as various dimethacrylate crosslinking monomers have been prepared and characterized. Uracil, thiourea, phenol, toluene, ethylbenzene, propylbenzene, and butylbenzene have been used to characterize retention of prepared capillary columns in the mobile phases with 40-95% of acetonitrile and at working temperatures ranging from 25 to 60°C. By an optimization of six-parameter polynomial models we have found that the retention of small molecules is affected mainly by the concentration of the acetonitrile in the mobile phase with very low contribution of working temperature and combined effect of acetonitrile concentration and temperature. Concentration of the mobile phase controlled also enthalpy of the retention. On the other hand, entropic contribution was almost insensitive to the change of the mobile phase composition, especially for mobile phases containing more than 60% of acetonitrile.
... Capillary Electrophoresis-Mass Spectrometry 16 -- 1.4. ... ... The Status of Capillary Surfaces in Capillary Electrophoresis: Influence on Electroosmotic -- Flow 483 ... ... Capillary Isoelectric Focusing 712 -- 20.6. Capillary Gel Electrophoresis 713 -- 20.7. ... ... On-Capillary Assays 904 -- 27.4. ... ... On-Line Capillary Microreactors-Capillary -- Electrophoresis Systems 909 -- 27.5. ...
Chemical analysis ; 146
1047 s.
- Klíčová slova
- elektroforéza kapilární,
- Konspekt
- Biochemie. Molekulární biologie. Biofyzika
- NLK Obory
- fyzika, biofyzika
- biomedicínské inženýrství
This study introduces a silica-based monolith in a capillary format (0.1 mm × 100 mm) as a support for immobilization of liposomes and its characterization in immobilized liposome chromatography. Silica-based monolithic capillary columns prepared by acidic hydrolysis of tetramethoxysilane in the presence of polyethylene glycol and urea were modified by (3-aminopropyl)trimethoxysilane, whereby amino groups were introduced to the monolithic surface. These groups undergo reaction with glutaraldehyde to form an iminoaldehyde, allowing covalent binding of pre-formed liposomes containing primary amino groups. Two types of phospholipid vesicles were used for column modification; these were 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphatidyl choline with and without 1,2-diacyl-sn-glycero-3-phospho-L-serine. The prepared columns were evaluated under isocratic separation conditions employing 20mM phosphate buffer at pH 7.4 as a mobile phase and a set of unrelated drugs as model analytes. The liposome layer on the synthesized columns significantly changed the column selectivity compared to the aminopropylsilylated monolithic stationary phase. Monolithic columns modified by liposomes were stable under the separation conditions, which proved the applicability of the suggested preparation procedure for the synthesis of capillary columns dedicated to study analyte-liposome interactions. The column efficiency originating from the silica monolith was preserved and reached, e.g., more than 120,000 theoretical plates/m for caffeine as a solute.
The temperature effects during the sol-gel process and ageing of the silica-based monolith on the structure and separation efficiency of the capillary columns (100microm i.d., 150mm) for HPLC separations were studied. The tested columns were synthesized from a mixture of tetramethoxysilane, polyethylene glycol and urea under the acidic conditions. The temperature was varied from 40 degrees C to 44 degrees C and formation of bypass channels between the silica mold and the capillary wall was examined. The temperature of 43 degrees C was estimated as optimal for preparation of efficient silica capillary columns which were subsequently modified by octadecyldimethyl-N,N-diethylaminosilane or covered by poly(octadecyl methacrylate) and tested using standard mixture of alkylbenzenes under the isocratic conditions.
- MeSH
- benzenové deriváty chemie MeSH
- koncentrace vodíkových iontů MeSH
- mikroskopie elektronová rastrovací MeSH
- močovina chemie MeSH
- oxid křemičitý chemie MeSH
- polyethylenglykoly chemie MeSH
- silany chemie MeSH
- teplota MeSH
- vysokoúčinná kapalinová chromatografie metody MeSH
- změna skupenství MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The synthesis and characterization of large-bore silica-based monolithic capillary columns (0.32mm×150mm) are presented. Columns were prepared by acidic hydrolysis of a mixture containing tetramethoxysilane (TMOS) and 1,2-bis(trimethoxysilyl)ethane (BTME) in different molar ratios in the presence of polyethylene glycol and urea. The monoliths were modified by zwitterionic monomer [2-(methacryloyloxy)ethyl]-dimethyl-(3-sulfopropyl)-ammonium hydroxide via 3-(trimethoxysilyl)propyl methacrylate. Prepared stationary phases were evaluated by scanning electron microscopy and chromatographic separation of nucleobases and their derivatives in the HILIC mode. The best chromatographic results were obtained with the column prepared from the reaction mixture containing BTME and TMOS in a 1:4 molar ratio. The permeability of such column reached 1.68×10(-14)m(2) and the efficiency, expressed as a height equivalent of the theoretical plate, did not exceed 10.5μm for the tested compounds. The columns were successfully applied to HILIC separation of native and labeled oligosaccharides and glycans released from bovine ribonuclease B and human immunoglobulin G.
- MeSH
- chromatografie kapalinová přístrojové vybavení metody MeSH
- ethan analogy a deriváty chemie MeSH
- hmotnostní spektrometrie s elektrosprejovou ionizací MeSH
- hydrofobní a hydrofilní interakce MeSH
- imunoglobulin G metabolismus MeSH
- lidé MeSH
- methakryláty chemie MeSH
- mikroskopie elektronová rastrovací MeSH
- oligosacharidy analýza izolace a purifikace MeSH
- organické sloučeniny křemíku chemie MeSH
- oxid křemičitý chemie MeSH
- ribonukleasy metabolismus MeSH
- skot MeSH
- trimethylsilylové sloučeniny chemie MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- skot MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
This study introduces zwitterionic monolithic capillary columns intended for isocratic and gradient HILIC separations. Silica-based monolithic capillary columns (100 μm × 150 mm) prepared by acidic hydrolysis of tetramethoxysilane in the presence of polyethylene glycol and urea were modified by a sulfoalkylbetaine zwitterion ([2-(methacryloyloxy)ethyl]-dimethyl-(3-sulfopropyl)-ammonium hydroxide) to HILIC stationary phase. The prepared columns were evaluated under the isocratic and gradient separation conditions using a standard mixture containing nucleic acid bases, nucleosides, and 2-deoxynucleosides. Mobile phases contained high concentration of acetonitrile (95-85%, v/v) and 5-50mM ammonium acetate or ammonium formate in the pH range of 3-6. The synthesized columns showed a long-term stability under the separation conditions while the high permeability and efficiency originating from dual structure of the silica monolith were preserved. The relative standard deviations (RSDs) for the retention times of tested solutes were lower than 2% under the isocratic conditions and lower than 3.5% under the gradient conditions.
- MeSH
- acetonitrily chemie MeSH
- betain analogy a deriváty chemie MeSH
- chromatografie kapalinová přístrojové vybavení metody MeSH
- hydrofobní a hydrofilní interakce MeSH
- nukleosidy chemie izolace a purifikace MeSH
- nukleové kyseliny chemie izolace a purifikace MeSH
- oxid křemičitý chemie MeSH
- reprodukovatelnost výsledků MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
