Effects of the column pressure drop in packed-column supercritical-fluid chromatography

Summary Columns packed with small particles (i.e. 3 to 10μm) may be used for fast separations in supercritical-fluid chromatography (SFC). The analysis times required for such packed columns are typically an order of magnitude smaller than what may be achieved with contemporary open-tubular (capillary) columns (25 to 100μm i.d.). However, packed columns give rise to much higher pressure drops over the column. The maximum allowable pressure drop will determine how many plates can be achieved with packed columns. In this paper the effects are described of the pressure drop on retention (capacity factor) and efficiency (number of plates) for packed columns of various lengths and internal diameters, packed with three different octadecylsilica materials. Carbon dioxide has been used as the mobile phase under several different sets of pressure and temperature conditions.     

An investigation of 3μm reverse-phase octadecylsilane packing materials for high-performance liquid chromatography

Summary The efficiency of high-performance liquid chromatography columns packed with 5 m and 3 m reverse-phase octadecylsilane packing materials has been evaluated using four test mixtures. Both stopped flow and valve injection have been used and typical efficiencies are given.     

Hypercrosslinked polystyrene as a novel type of high-performance liquid chromatography column packing material. Mechanisms of retention

An experimental material, Chromalite 5HGN (Purolite, UK), that represents hypercrosslinked polystyrene as a new type of neutral stationary phase for HPLC was examined. The material contains no functional groups, but is compatible with any kind of nonpolar and highly polar mobile phase, and even with water. It is chemically resistant and thermally stable. When using aqueous organic mobile phases, Chromalite 5HGN works similar to standard C18 reversed-phase packings, but is characterized by much greater hydrophobicity and, sometimes, unusual selectivity. When using nonpolar mobile phases, i.e. under "quasi normal-phase" conditions, the retention is mostly governed by the interactions between pi-electronic systems of the adsorbent and adsorbate. Adding highly polar, even hydrophilic solvents into the mobile phase, leads to a shift of retention times toward the "reversed-phase" kind of chromatography, which gives an additional possibility in fine tuning the column selectivity.     

Preparation and evaluation of carbon coated alumina as a high surface area packing material for high performance liquid chromatography

The retention of polar compounds, the separation of structural isomers and thermal stability make carbonaceous materials very attractive stationary phases for liquid chromatography (LC). Carbon clad zirconia (C/ZrO₂), one of the most interesting, exhibits unparalleled chemical and thermal stability, but its characteristically low surface area (20–30 m²/g) limits broader application as a second dimension separation in two-dimensional liquid chromatography (2DLC) where high retentivity and therefore high stationary phase surface area are required. In this work, we used a high surface area commercial HPLC alumina (153 m²/g) as a support material to develop a carbon phase by chemical vapor deposition (CVD) at elevated temperature using hexane vapor as the carbon source. The loading of carbon was varied by changing the CVD time and temperature, and the carbon coated alumina (C/Al₂O₃) was characterized both physically and chromatographically. The resulting carbon phases behaved as a reversed phase similar to C/ZrO₂. At all carbon loadings, C/Al₂O₃ closely matched the unique chromatographic selectivity of carbon phases, and as expected the retentivity was increased over C/ZrO₂. Excess carbon – the amount equivalent to 5 monolayers – was required to fully cover the oxide support in C/Al₂O₃, but this was less excess than needed with C/ZrO₂. Plate counts were 60,000–76,000/m for 5 μm particles. Spectroscopic studies (XPS and FT-IR) were also conducted; they showed that the two materials were chemically very similar.     

The contribution of the pressure drop to analyte retention in coupled column supercritical fluid chromatography of lipids

Summary This paper reports the qualitative and quantitative effects of the column pressure drop on the retention of lipid components in a serially coupled capillary column SFC system. The contribution of the pressure drop consists of two components, the density effect and the flow effect. The magnitude of the flow effect,i. e. the change in retention which results from changes in the flow-rate when column pressures are changed, is determined by the difference in single column analyte k values. The effect will be positive compared with the uncorrected retention values when the column with largest k value is closest to the injector. With the columns in reversed order, the effect will be negative. The contribution from the density effect always resulted in larger coupled column k values and was in most instances of more significance than the flow effect component. Values calculated with and without pressure drop correction have been compared and it has been shown that for most of the eighteen model lipid compounds investigated, the deviations from the experimental retention factors were smaller when pressure drop corrections were made.     

The effects of the column pressure drop on retention and efficiency in packed and open tubular supercritical fluid chromatography

The effects of the pressure drop across the column on retention and efficiency in SFC have been studied. Numerical methods are described which enable the prediction of hold-up time and pressure drop in both packed and open tubular columns. Predictions of both hold-up time and pressure drop are in good agreement with experimental data. The density gradient along the column can be calculated using the numerical methods and a procedure is described which enables the calculation of the overall capacity factors of the solutes from the density profile in the column. Significant variations of the capacity factor are observed along the column. The effect of the density gradient along the column on local diffusivity and dispersion is studied. The column efficiency in systems with significant pressure drops is affected by changes in: the linear velocity of the mobile phase; the diffusion coefficients; and the capacity factors of the solutes along the column. The overall efficiency of the chromatographic system can be calculated if, as is the case for open tubular columns, adequate plate height equations are available.     

The use of microparticulate guard columns in reverse-phase high-performance liquid chromatography

Summary The use of mciroparticulate guard columns in conjunction with high efficiency reverse-phase columns has been evaluated in terms of system efficiency and peak symmetry using three standard test mixtures. The effects of linking tube dimensions, particle size of guard column packing and the use of low or zero dead volume couplings have been investigated. Both valve and stopped-flow injection techniques have been used. Recommendations are given for the most efficient use of guard columns.     

Influence of temperature in reverse-phase high-performance liquid chromatography with gradient elution

The influence of temperature on the retention of several species separated by reverse-phase liquid chromatography by gradient elution is shown to be of enough importance to warrant careful control of temperature if reproducible results are to be obtained. The smaller the particle size in the column, the greater the effect of temperature, and therefore the control should be greater. Likewise, it has been verified that for a given solvent gradient, independent of its complexity, there is a linear relation between ln k′ and 1/T, which also occurs in separations by isocratic elution. Dufek's equation can be adjusted perfectly to the experimental data obtained from gradient elutions, and may be used in the simulation and optimization of gradient chromatographic processes.     

Preparative high-performance liquid chromatography of peptides on a new reversed-phase packing material, Kromasil C18

Preparative reversed-phase high-performance liquid chromatography has found wide use in the production of peptides for pharmaceutical formulations. Purity of the substance and overall economy of the chromatographic system are the most important criterias. In this sense optimized, silica particles and production process with capability to separately control parameters important to chromatography, are essential to high-performance chromatography. Kromasil C18 packing material was tested and evaluated in respect of its selectivity, flow and pressure properties, resolution, load capacity, recovery, adsorption effects, mechanical strength and chemical degradation.     

Magnesium oxide microspheres as a packing material for the separation of basic compounds in normal-phase liquid chromatography

Uniform monodisperse magnesium oxide microspheres with a high surface area have been prepared by a facile seed-induced precipitation. By characterizing these particles with scanning electron microscopy and N(2) physisorption techniques, the results demonstrate that these magnesium oxide microspheres have an average particle diameter of 9.5 microm, a specific surface area of 211.7 m(2)g(-1), a total pore volume of 0.76 mL g(-1), and an average pore diameter of 143 A. The chromatographic properties of these microspheres have been investigated in normal-phase mode for the separation of various basic compounds including aniline, quinoline, and pyridine derivatives. In contrast to conventional silica, the magnesium oxide particles exhibit unique selectivity and retention property for the separation of the tested basic compounds, and these microspheres are promising as an alternative new packing material for high-performance liquid chromatography.     

Thermodynamics of the sorption of water-soluble vitamins in reverse-phase high performance liquid chromatography

The thermodynamics of the sorption of certain water-soluble vitamins on a C18 reverse phase from water-acetonitrile solutions of different compositions is studied. The thermodynamic characteristics of the investigated chromatographic systems are calculated. The dependences of standard molar enthalpy and changes in entropy when the sorbate transfers from the bulk solution to the surface layer on the concentration of the organic component in the mobile phase are analyzed. The boundaries for applying the main retention models describing the sorption of the investigated compounds are discussed.     

Use of alumina with anchored polymer coating as packing material for reversed-phase high-performance liquid chromatography

A new material with anchored polymer coating on alumina can be used as a packing material for the reversed-stationary phase of HPLC columns. The coatings are formed by in situ polymerization of maleic acid adsorbed on alumina with 1-octadecene, with cross-linking by 1,4-divinylbenzene. The chromatographic performance of this material was studied and compared with that of 20–30 μm octadecyl-bonded silica particles and a commercial column packed with 5 μm octadecyl-bonded silica. The results indicate that this new material is a promising alternative to silica-based reversed-phase packings due to its competitive performance and stability in acidic and basic media.     

Retention in gas chromatography when there is a large pressure drop in different sections of a coupled column

Methods providing control of the selectivity of a coupled column were compared. The method of vacuum barochromatography has been developed. This method allows the selectivity of a coupled column to be controlled over a broad range of the Rohrschneider scale using medium carrier-gas pressures. The possibility of identifying the components of a complex mixture based on chromatographic spectra was considered.Translated fromIzvestiya Akademii Nauk. Seriya Khimicheskaya, No. 12, pp. 2941–2947, December, 1996.     

Influence of the packing heterogeneity on the performance of liquid chromatography supports

We report on a series of plate height and flow resistance data obtained via computational fluid dynamics simulations in a simplified two-dimensional (2D) mimic of real packed bed and monolithic columns. By varying the external porosity (0.4 < epsilon < 0.8) and the degree of packing randomness, a good qualitative insight in the relationship between the packing porosity and heterogeneity and the general chromatographic performance parameters is obtained, unbiased by any differences in phase retention factor k', mobile phase diffusivity or viscosity or intra-skeleton porosity. The results provide a quantitative support for the use of domain size reduced plate heights as a means to compare the performance of chromatographic beds with a different porosity, as it was found that packings with a similar degree of packing heterogeneity yield very similar domain size reduced h(min)-values, nearly completely independent of the porosity. The study also clearly shows that the presence of preferential flow paths (inevitably accompanied by the presence of more clustered regions) leads to a decrease of the flow resistance, but also leads to a strong increase of the band broadening if supports with the same porosity epsilon and the same radial width are compared. For the presently considered 2D system, the flow resistance reduction is too small to overcome the corresponding strong increase in band broadening, such that the presence of preferential flow paths always leads to an overall increase of the separation impedance.     

Copolymer of Di (methacryloyloxymethyl) naphthalene and divinylbenzene as a column packing for high-performance liquid chromatography

Summary A new porous polyaromatic ester packing was synthetized for high performance liquid chromatography. The relationship between retention and chain length of the members of homologous series of alkylbenzenes, N-alkylanilines, alkylarylethers, alkylbenzoates and alkylarylketones on this new stationary phase using different eluents was investigated. Using the alkylarylketone scale the retention indices of the homologues and test compounds were calculated. The results were compared with those obtained for poly (styrene-divinylbenzene) polymers. For both types of packing the first members of each homologous series gave non-linear behaviour. The methylene group index increments are different for the studied homologous series; thus there is no simple additivity of the retention indices. The efficiency of the porous polymeric columns is a function of the capacity factor of the solute and the organic component of the eluent.     

Development of dual gradient column in liquid chromatography

The dual gradient column, in which both the chemical property of the stationary phase and the flow velocity in the mobile phase are heterogeneous longitudinally along the column, is developed to obtain the mobile phase gradient-like elution in an isocratic condition. Here, the step-wise dual gradient columns were prepared by connecting an inlet column (I.D. 50 microm, packed with ODS) serially to an outlet column (I.D. 100-200 microm, packed with the mixture of ODS and C1 [9:1]). The retention behavior of alkylbenzenes was able to be controlled in the dual gradient column depending on the variation in the flow velocity. Moreover, the change in retention behavior induced by the flow velocity variation for the dual gradient columns was quite different from that by the variation in organic modifier content of the mobile phase in isocratic elution for a single gradient column and can induce the similar effect with an ordinary gradient elution in a mobile phase composition.     

Control of column temperature in reversed-phase liquid chromatography

When separations by reversed-phase liquid chromatography (RP-LC) are carried out at temperatures other than ambient, resulting retention times and bandwidths can depend on the equipment used. As a result, an RP-LC separation that is adequate when carried out on one LC system may prove inadequate when the separation is repeated on a second system. In the present study, various temperature-related problems which can result in a failure of method transfer for non-ambient RP-LC methods were examined. Means for correcting for such effects and thereby ensuring method transferability are described.     

Application of column liquid chromatography (HPLC) to special problems in food chemistry a laboratory note

Summary Liquid chromatography has been used for the determination of amino acids, sugars and biogenic amines in food. Some special problems, for example, determination of patuline in apple juice, hyoscyamine and scopolamine in french beans preserves, were also solved by HPLC.