Yufeng Shen

Poly(ethylene-propylene glycol)-modified fused-silica columns for capillary electrophoresis using epoxy resin as intermediate coating

Abstract An epoxy resin was used for fused-silica capillary column pretreatment before coating with a hydrophilic polymer for capillary electrophoresis. The unique properties of the epoxy resins allow the deposition of a dense polymer layer on the column surface using the dynamic coating method. By using a free-radical cross-linking initiator, a hydrophilic polymer coating can be subsequently immobilized over the resin covered surface. The coating is of sufficient thickness and hydrophilicity to reduce protein adsorption and minimize electro-osmotic flow. Basic protein separations were achieved with high efficiency and reproducibility; one thousand consecutive runs on a single column gave an R.S.D. value of less than 2% in migration time. The coating was stable throughout the pH range 4–11. The coating provides a simple, stable, and reproducible means for deactivation of fused-silica columns for capillary elettrophoresis.

High speed solvating gas chromatography using packed capillaries containing sub-5 μm particles

Abstract In this study, fused-silica capillaries of 250 μm I.D. were packed with spherical porous and nonporous octadecyl bonded silica particles having diameters of 1.5 and 3 μm. These capillaries were used with CO 2 as mobile phase at elevated temperatures. At the column inlet, the mobile phase was a supercritical fluid, while at the column outlet, it was a gas. The mobile phase gradually changed from a supercritical fluid to a gas within the column. Because the mobile phase exhibited solvating ability for the analytes being separated, in contrast to conventional gas chromatography (GC) in which the mobile phase is only a carrier, this variation has been named “solvating gas chromatography” (SGC). Using this technique, capillaries packed with small particles produced column efficiencies as high as 1200 plates s −1 for retained solutes. Therefore, packed capillary SGC is the method of choice to carry out high speed GC separations. The effects of column inlet pressure, column length and particle characteristics on mobile phase linear velocity, retention factor, column efficiency, speed of analysis and resolution were investigated.

Capillary electrophoresis using diol-bonded fused-silica capillaries

In this paper, 3-glycidoxypropyltrimethoxysilane was used to produce diol-bonded capillaries at room temperature for capillary electrophoresis (CE). A variety of standard reference compounds and authentic biological samples including ribonucleotides, peptides and proteins were used to test the columns. It was found that greatly suppressed electroosmotic flow was measured over a pH range of 3-10. Lower than 1.6% relative standard deviation (> 10 runs) in migration time was observed for the analysis of test proteins. For real samples of ribonucleotides in tumor cell extracts, approximately 1 million theoretical plates and excellent peak shapes were obtained. The high column efficiency and symmetrical peaks allowed the separation of samples with only 0.6% maximum difference in migration times. The diol-bonded fused-silica capillary columns were stable when used in a pH range of 2-8 under typical CE conditions. The column preparation method involved a simple dynamic coating procedure at room temperature, greatly simplifying the more typical static coating methods that require vacuum pumps and ovens.

Elevated temperature liquid chromatography using reversed‐phase packed capillary columns

Reversed-phase packed capillary column liquid chromatography (LC) was investigated at column temperatures of 24°C, 40°C, 60°C, and 80°C. For a mixed mobile phase containing acetonitrile and water (72:28, v/v), the viscosity of the mobile phase decreased 2.2 times and the diffusion rate of the solute increased 2.6 times in the range of temperatures investigated. The faster diffusion rate of naphthalene in the mobile phase at 80°C resulted in a 2.6-fold faster optimum linear velocity and a 2.5-fold decrease in the C term of the van Deemter equation compared to values obtained at 24°C. Better efficiencies were obtained at faster than optimum linear velocities at elevated temperatures, while at the same time retention times were much shorter than at room temperature. It was also found that temperature could have a better or worse effect on selectivity, depending on the retention factors, enthalpies, and entropies of the compounds analyzed. The pressure drop decreased by one half at an elevated temperature of 80°C compared with that at room temperature for the mobile phase investigated. It is estimated that smaller particles or longer columns could be used at elevated temperatures in an LC system to obtain even higher total plate numbers.

Packed capillary column supercritical fluid chromatography using SE-54 polymer encapsulated silica

A new method of preparing stationary phases for packed capillary column supercritical fluid chromatography (SFC) is presented. Surface deactivation of silica particles was carried out by dehydrocondensation of the silicon hydride groups in polymethylhydrosiloxane with the silanol groups on the silica surface. The deactivated particles were then coated with a thin film of SE-54 stationary phase. The coated layer was immobilized by a crosslinking reaction between the methyl groups of the surface-bonded polymethylhydrosiloxane and the SE-54 stationary phase using dicumyl peroxide as a free radical initiator. With these two reactions, the polar groups on the silica surface were more completely capped than with bonding only a monomolecular polymeric layer on the silica surface. The SFC performance of the newly developed packing materials was evaluated using a standard polarity mixture, a series of fatty acid methyl esters, a peppermint oil, and several high-molecular-mass and complex polymers.

High-efficiency solvating gas chromatography using packed capillaries.

In this study, column efficiency in packed capillary column solvating gas chromatography (SGC) was investigated. Long (>3 m) fused silica capillaries with an inner diameter of 250 μm were packed with 10 and 15 μm spherical porous (300 Å) octadecyl bonded silica particles using a CO(2) slurry packing method. A 336 cm × 250 μm i.d. fused silica capillary containing 10 μm particles provided a total column efficiency of 264 000 plates (k = 0.41), corresponding to a reduced plate height of 1.27, using CO(2) as the mobile phase at a column inlet pressure of 260 atm. A minimum plate height of 12.7 μm and a maximum plate number per unit time of 813 plates/s were obtained using packed capillary SGC. Retention factors were dependent on the column inlet pressure but independent of the pressure gradient along the column. Gasoline and diesel samples were separated under SGC conditions, and the results were comparable to those obtained using typical open tubular column gas chromatography.

Cyclodextrin polymer encapsulated particles for supercritical fluid chromatography

Cyclodextrin polysiloxane encapsulated particles were prepared for packed capillary column supercritical fluid chromatography. Silica particles were deactivated using polymethylhydrosiloxane through a dehydrocondensation reaction, and then coated and crosslinked with a cyclodextrin-substituted polymethylsiloxane. Using a fluidized bed coating method, nonagglomerating polymer coated particles were obtained even when a high loading of cyclodextrin polymer (25% w/w) was used. The polarities and enantiomeric selectivites of the packing materials were investigated for a wide variety of chiral test solutes, including hydrocarbon, alcohol, ketone, ester, lactone, diol, acid and amine, using neat supercritical CO2 as mobile phase. The effects of temperature and pressure on polarity and enantiomeric selectivity were also investigated.

Reversed-phase liquid chromatography of proteins and peptides using multimodal copolymer-encapsulated silica

Multimodal copolymer-encapsulated particles for liquid chromatography were prepared by bonding 1-octadecene and unsaturated carboxylic acids on silica particles (5 microm diameter, 300 A pores) for liquid chromatography of proteins. These multimodal copolymer-encapsulated particles can provide both hydrophobic and hydrogen bonding interactions with polar compounds. The chromatographic performance of these multimodal copolymer-encapsulated particles for peptide and protein separations was evaluated under reversed-phase conditions. Compared with typical C8-bonded silica, polymer-encapsulated particles were more stable in acidic mobile phases and provided better recoveries, especially for large proteins (Mr>0.5 x 10(6)). Totally hydrophobic polymer-encapsulated particles were found to produce broad peaks for proteins, and significant improvements were observed by introducing hydrophilic groups (-COOH) onto the polymer-encapsulated surface to form a multimodal phase. For the reversed-phase liquid chromatography of peptides and proteins, improved selectivity and increased solute retention were found using the multimodal polymer-encapsulated particles. More peaks were resolved for the separation of complex peptide mixtures such as protein digests using the multimodal polymer-encapsulated particles as compared to totally hydrophobic polymer-encapsulated particles.

Electrochromatographic contributions in capillary electrophoresis of biomolecules

Abstract In this study, electrically driven separations of charged biomolecules including ribonucleotides and peptides were investigated using various surface modified columns. These columns included cationic, neutral and anionic polymeric-coated columns. Experiments demonstrated that for neutral polymer-coated columns, solutes eluted according to their electrophoretic mobilities. Using surface charged columns, the elution of charged solutes was determined by their electrophoretic mobilities, electroosmotic flow rates and interactions between charged solutes and the charged column inner wall. Retention factors were used to investigate the interactions between solutes and the column inner wall. Column efficiency measurements for charged solutes on charged surface columns were also examined, and the results were compared with those obtained using neutral polymer-coated columns. It was found that separations of multivalent biomolecules were difficult using charged surface columns because of either strong adsorption or high electrophoretic mobilities.

Performance of polyethylenimine‐coated particles in packed capillary column supercritical fluid chromatography

Strongly basic polyethylenimines (PEI) were coated on both untreated and diol-bonded silica particles. The resultant basic surface was found suitable for the separation of strongly basic alkylamines such as α-phenylethylamines with a pKb of ca. 4, under supercritical fluid chromatography (SFC) conditions with neat carbon dioxide. Interactions between the coated PEI and the silanol groups on the silica surface provided a stable coating. The acidic silica surface was neutralized by the basic amine functionalities in the PEI. Preliminary surface deactivations before coating with the basic PEI and subsequent end capping with hexamethyldisilazane (HMDS) were found necessary to obtain inert packing materials. However, it was found that the basic particle surface was not suitable for the separation of proton-donor compounds, including some weakly acidic phenols (pKa ca. 10). The effects of temperature on retention factors and peak shapes were studied. The stability of the PEI-coated particles was investigated by measuring the loss of organic carbon content of the particles after washing with supercritical CO2. It was found that the strong basicity and the extremely high molecular weight of the coated PEI produced stable coatings under SFC conditions.