Stationary phases commonly used

There are two types of stationary phases commonly used in exclusion chromatography silica gel and micro-reticulated cross-linked polystyrene gels. A third type of exclusion media is comprised of the Dextran gels. Dextran gels are produced by the action of certain bacteria on a sucrose substrate. They consist of framework of glucose units that can form a gel in aqueous solvents that have size exclusion properties. Unfortunately the gels are mechanically weak and thus, cannot tolerate the high pressures necessary for HPLC and, as a consequence, are of very limited use to the analyst. 

In stationary phases commonly used in RPC the picket fence configuration is not likely because of the relatively low surface concentration of th ligates, oi. < S p.m6l/m. This value is much smaller tl n the surface concenlrutbn in compressed monolayers of compounds ccjfntaining alkyl chains with small polar terminal groups-(hydroxyl, carboxyl, amino groups) that was found to be 8 )ittiol/m at the Water sUrftice (///). 

Recently, reversed-phase partition chromatography has become the method of choice for both qualitative and quantitative analysis of carotenoids. The stationary phases commonly used are those with C,8-bonded chains (ODS) their performances are influenced by the extent of endcap-... 

A system for assessing the effectiveness of chromatographic systems by calculating the discriminating power has been described (A. C. Moffat et al.y J. Chromat., 1974, 90, 1-7). Using this method, a number of stationary phases commonly used in toxicology have been examined (A. C. Moffat etal.y... 

Another approach to improving resolution is to use thin films of stationary phase. Capillary columns used in gas chromatography and the bonded phases commonly used in HPLC provide a significant decrease in plate height due to the reduction of the Hs term in equation 12.27. 

Virtually all current research in SFC utilizes either small bore packed columns with particles of 5-10 micrometers in diameter optimized for use in liquid chromatography or narrow bore, fused silica open tubular columns with Immobilized phases similar to those used in gas chromatography. In the latter case columns of saaller internal diameter, 10-100 micrometers, shorter lengths (generally less than 20 m with 1-10 m being the most common length), and more firmly crosslinked stationary phases are used by coaparison with standard columns for gas chromatography. In all... 

There have been many reported chiral stationary phases for use in both packed and capillary gas chromatography. Most of these phases are of the carbonyl-bis-L-valine isopropyl ester, diamide, and peptide phase types. The most common phase is Chirasil-Val from Alltech Applied Science Laboratories (State College, PA). This phase is ideal for the separation of a variety of enantiomers including amino acids, sugars, amines, and peptides. The phase is composed of L-valine-tert-butylamide linked through a car-oxamide group to a polysiloxane backbone every seven dimethylsiloxane units apart. 

Porous microparticles are the most common stationary phase particles used in modern HPLC. The role of pore size is a critical one, as the pores provide the surface with which the sample interacts. Particles with small pores exhibit a high surface area and therefore have greater retention. Large molecules like proteins, however, may be excluded from the small pores, and for those molecules a packing with a larger pore size is preferable. The difference between porous particles, pellicular particles, and porous microparticles is illustrated in Figure 3.19. Porous particles are seldom used owing to low efficiencies and are not discussed further. 

Intermolecular forces were discussed in Chapter 3 and extended to GC stationary phases in Chapter 8. Rohrschneider, followed by McReynolds, investigated the nature of GC stationary phases by using a few common chemicals as probes. Their retention on a given liquid reflected the extent of their interaction with the stationary phase. By choosing probes with selective interactions, they could determine a set of numbers that characterize the liquids under study. 

A column consisting of a deactivated silica-based stationary phase is used for the packed-column mode. A packed column allows larger volumes of sample solvent to be injected, thus improving sensitivity. Generally, the column dimensions are 1 x 100-250 mm and the particle size is 5 / m. Commercial SFC instruments are also available that will handle the classical 4.6 x 150-mm or 250-mm columns. With the introduction of electronically controlled variable restrictors to control the back pressure, the packed columns are becoming increasingly more popular. This feature allows the independent flow and pressure control of mobile phases, thus helping in rapid optimization of selectivities. Some of the commonly used packed columns are as follows ... 

Conventional thin-layer chromatography (TLC) in our experience, known under the name planar chromatography, uses horizontal or vertical glass or Teflon chambers for the development of chromatograms. As stationary phases, commonly known adsorbents or supports based on silica gel, aluminium oxide, magnesium silica, cellulose, and so forth are used particle sizes are about 20 jitm. The migration of the mobile phase is based on the phenomenon of capillary forces. This chromatographic method is described, in detail, in other sections of this volume. 

The most common mobile phases used for reversed-phase chromatography involve combinations of either methanol or acetonitrile and buffered water (like phosphate buffered, pH 7). There are other organic solvents that are used in conjunction with buffered water in reversed-phase chromatography, e.g., other alcohols or THF, but these are less commonly used. Similarly to normal phase, pH can be used to increase/decrease the interactions between the solutes and the stationary phase. Common additives are trifluoroacetic acid and diethylamine. As with normal-phase chromatography, the mobile phase must be chosen carefully. Generally, it is understood that the mobile phase should have a low viscosity, be LTV transparent, and be completely miscible. 

Gas chromatography is often divided into categories based on the type of stationary phase used. Gas-liquid chromatography (GLC) implements a porous, inert solid support that is coated with a viscous, nonvolatile liquid phase. On the other hand, gas-solid chromatography (GSC) uses a solid adsorbent as the stationary phase. Klee offers these general rules-of-thumb for selection of stationary phase materials use solid adsorbents to separate room-temperature gases, liquid stationary phases to separate room-temperature liquid and solid mixtures, polar phases for polar solutes, and nonpolar phases for nonpolar solutes. Table 1 lists common liquid- and solid-stationary phase materials available for use in capillary columns. Barry cross-refers numerous column materials from nine different manufacturers. ... 

Adsorption chromatography exploits differences in the relative affinity of solutes for a solid adsorbent used as the stationary phase. Common stationary phase materials for adsorption chromatography are porous silica gel, activated alumina, activated carbon, magnesium oxide, carbonates, and highly cross-linked polymers such as styrene-divinylbenzene and methac-rylates. The chemical natures of these adsorbent stationary phase materials make them well suited for separations of solute mixtures that differ in polarity and chemical functionality. For example, silica is an acidic adsorbent that retains basic compounds to a greater extent than nonbasic ones. In contrast, alumina... 

In reversed-phase chromatography (RPC), a relatively nonpolar stationary phase is used, with a polar mobile phase such as methanol, acetonitrile, tetrahydro-furan, water, or usually a mixture of water with one of the organic solvents. The organic solvent is called the modifier, and acetonitrile is the most common one. The water content is varied for adjusting the polarity. Methanol is used for acidic compounds and acetonitrile for basic compounds. Tetrahydrofuran is used for those with large dipoles. These solvents are UV transparent and have low viscosity. The most common bonded phases are n-octyldecyl (Cig) or n-decyl (Cg) chains, or phenyl groups. Polar reversed-phase columns such as polyethylene glycol (PEG)... 

Series coupling of columns containing the same stationary phase is used to enhance efficiency and with different stationary phases to fine tune selectivity [8]. Series coupling of packed columns became popular after it was demonstrated that the column pressure drop did not limit the total column efficiency to the extent that had been predicted (section 7.4.2). Serial coupling of 2 or 3 standard columns is practical for routine applications and provides a total plate count in excess of 50,000. There is no theory for selectivity optimization for coupled packed columns but suitable conditions often can be estimated from separations on the individual columns. Effective selectivity changes require that the coupled columns have different retention properties. A number of practical examples for the separation of polymer additives, polycyclic aromatic hydrocarbons, phytic acid impurities and enatiomers have been described [137,195,196]. Series coupling of open tubular columns with different stationary phases is less common, but changes in selectivity are predictable, at least when the pressure drop is low [197]. 

In general, one can divide size exclusion chromatography into two categories SEC in organic solvents and SEC in aqueous solvents. For polymers that are soluble in organic solvents, styrene-divinylbenzene stationary phases are used for the most part. For packings that are soluble in aqueous eluents, -hydrophilic methacrylates are most commonly used. 

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