Partition chromatography

Separation is based on solute partitioning between two liquid phases. 

Ion-exchange chromatography has many clinical applications, including the separation of amino acids, peptides, proteins, nucleotides, oligonucleotides, and nucleic acids. Another important application of ion-exchange chromatography is the separation and removal of inorganic ions from aqueous mixtures. Thus most water purification units used to prepare deionized water for the laboratory contain mixed-bed columns of cation and anion resins (see Chapter 1). 

Partition chromatography is categorized as either GLC or liquid-liquid chromatography (LLC). LEG is further categorized as either normal phase or reversed phase. For normal-phase LLC a polar liquid is used as the stationary phase, and a relatively nonpolar solvent or solvent mixture is used as the mobile phase. In reversed-phase partition chromatography, the stationary phase is nonpolar, and the mobile phase is relatively polar.  

Ion-suppression and ion-pair chromatography are two forms of reversed-phase chromatography used to separate ionic solutes. 

With ion-suppression chromatography, the ionic character of a weakly acidic or basic analyte is neutralized or suppressed through modification of the mobile phase pH. By neutralizing its ionic group, the solute is less polar and 

The extracting solvent in this scenario is the chromatographic mobile phase, while the sample solvent is the stationary phase. Liquid-liquid partition chromatography is based on this idea. The mobile phase is a liquid that moves through a liquid stationary phase as the mixture components partition or distribute themselves between the two phases and become separated. The separation mechanism is thus one of the dissolving of the mixture components to different degrees in the two phases according to their individual solubilities in each. 

Since the separation depends on the relative solubilities of the components in the two phases, the polarities of the components and of the stationary and mobile phases are important to consider. If the stationary phase is somewhat polar, it will retain polar components more than it will nonpolar components, and thus 

FIGURE 11.7 An illustration of how two mixture components separate, by the movement of an extracting solvent across a stationary sample solvent, due to the differing solubilities of the components in the two phases. 

FIGURE 11.8 An illustration of partition chromatography. A thin liquid film chemically bonded to the surface of finely divided solid particles is the stationary phase. 

The mobile phase for partition chromatography can also be a gas (GLC). In this case, however, the mixture components solubility in the mobile phase is not an issue—rather, their relative vapor pressures are important. This idea will be expanded in Chapter 12. 

For more polar components, the bonded normal phase supports are particularly useful and have benefits over adsorption chromatography on silica by allowing a rapid response to solvent composition changes. Other modes of partition chromatography, such as Ugand-exchange and affinity chromatography, have found particular applications for the resolution of water-soluble biomolecules. 

When cellulose is used as a stationary phase, with water or aqueous organic solvents as eluents, the separation of substances is by partition between the eluting mixture and the water adsorbed on the column. This is similar to the cellulose in paper chromatography. 

The most widely used type of HPl.C is panitiim chromatography, in which the stationary phase is a second liquid that is immiscible with the liquid mobile phase. In the past, most of the applications have been to nonionic, polar compounds of low to moderate molecular mass (usually --. TOIX)). Recently, however, methods have been developed (deriv atizatipn and ion pairing) that have e.xtended partition separations to ionic cumpound.s. 

The supports for the majority of bonded-phase packings for partition chromatography are prepared from rigid silica, or silica-based, compositions. These solids are formed as uniform, porous, mechanically sturdy particles commonly having diameters of 1.5-10 pm. with 3- and 5-pm particles being most common. The surface of fully hydrolyzed silica (hydrolyzed by heating with 0,1 M HCI for a day or two) is made up of chemically reactive silanol group.s, Thai is. 

Normal phase chromaiography l.ou. poiant mobile phase 

Reversed pltase chromatography High-polariiy niobilc phase 

FIGURE 28-14 Relationship between polarity and elution limes lor normal-phase and reversed-phase chromatography, 

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Van Den Dool, H. and P.D. Kratz (1963), Generalization of the retention index system including linear temperature programmed gas-liquid partition chromatography . J. Chromatogr, Vol. 11, p. 463. 

Schematics showing the basis of separation in (a) adsorption chromatography, (b) partition chromatography, (c) ion-exchange chromatography, (d) size-exciusion chromatography, and (e) eiectrophoresis. For the separations in (a), (b), and (d) the soiute represented by the soiid circie ( ) is the more strongiy retained.

Most immunoassay kits and many commercial immunoassay analyzers are based on heterogenous EIA or FIA. These include an immunoassay system that uses FIA linked to radial partition chromatography of the antibody—antigen complex (39) a system that uses antibody-coated tubes for enzyme immunoassay of a variety of hormones and dmgs (40) and a system that uses either a sandwich or competitive FIA format to measure a variety of analytes (41). 

Halomycins. The halomycins are a group of four antibiotics produced by M.icromonospora halophjtica and separated by partition chromatography on Chromosorb W coated with formamide (19). Further purification was accompHshed using preparative dc (212). 

Somatostatin [38916-34-6] M 1637.9, [a]p -36 (c 0.57, 1% AcOH). A tetradecapeptide which is purified by gel filtration on Sephadex G-25, eluting with 2N AcOH, and then by liquid partition chromatography on Sepahdex G-25 using n-BuOH-AcOH-H20 (4 1 5) and has Rp= 0.4. It is a brain growth hormone releasing-inhibiting factor which has also been synthesised. [Burgus et al. Proc Natl Acad Sci USA 70 684 1973, Sorantakis and McKinley Biochem Biophys Res Commun 54 234 1973 Hartridt et al. Pharmazie 37 403 1982.]... 

The great leap forward for chromatography was the seminal work of Martin and Synge (7) who in 1941 replaced countercurrent liquid-liquid extraction by partition chromatography for the analysis of amino acids from wool. Martin also realized that the mobile phase could be a gas rather than a liquid, and with James first developed (8) gas chromatography (GC) in 1951, following the gas-phase adsorption-chromatographic separations of Phillips (9). 

G. Haugaard and T. D. Kroner, Partition chromatography of amino acids with applied voltage , 7. Am. Chem. Soc. 70 2135-2137 (1948). 

Paper strip chromatography showed approximately equal amounts of substrate and a more polar product (A -pregnadiene-9a-fluoro-11/3,l6a,17a,21-tetrol-3,20-dione 16,21-diacetate) together with very small amounts of two less polar products. Partition chromatography of 0.25 gram of the residue (diatomaceous earth column system 2 parts ethyl acetate,... 

A. P. Eoueault (Ed.), Centrifugal Partition Chromatography, Chromatographie Seienee Series, Vol. 68, Mareel Dekker, New York (1995)... 

An interesting consequence of selective sorption is that conditions for partition chromatography arise which may enhance the normal ion exchange separation factors. This aspect has been utilised by Korkisch34 for separation of inorganic ions by the so-called combined ion exchange-solvent extraction method (CISE). 

Palladium, D. of as dimethylglyoximate. (g) 463 as nioximate, (g) 474 by EDTA, (ti) 329 Paper chromatography 229 see Thin layer chromatography Parallax errors due to, 85 Parallel determinations 132 Partial ionic equations 850, (T) 851 Partition chromatography 13. 217 Partition coefficient 162 Patton and Reeder s indicator 317, 328 Peptisation 419. 421... 

PCS. Partition chromatography on starch column 196, 261). MBA. Microbiological assay. 

The extracted crude luciferin was purified by Anderson s benzoy-lation method mentioned above (Anderson, 1935), followed by partition chromatography on a cellulose powder column using a mixture of ethyl acetate-ethanol-water (5 2 3 Tsuji, 1955) as the solvent, again in a hydrogen atmosphere. The luciferin obtained (3 mg) was highly pure by paper chromatography, and appeared suitable for crystallization. I... 

Figure 7. Infrared spectrum of pyrethrin I Isolated after dual partition chromatography First partition column. Celite-acetonitrile-hexane Second partition column. Silicic acid-nitromethane-hexane (with 5% acetone). Corresponds to peak 3 of gas chromatographic separation of pyrethrum mixture...

Paper Chromatography, Paper Partition Chromatography. See under Chromatography In Vol 3, C289-L to C298-L... 

In gas-liquid partition chromatography (GLPC), the stationary phase is a liquid that coats the particles in the tube or the walls of the tube. Often the tube itself is very narrow and long, perhaps 100 m, and has to be coiled (Fig. 4). Solutes are separated, as in liquid chromatography, by their relative solubility in the gas and liquid phases. In... 

Separation by adsorption chromatography takes place preferentially as a result of hydrogen bonding or dipole-dipole interactions. Hence, separation of mixtures of substances on silica gel layers by lipophilic solvents primarily takes place according to polarity differences. Further separation within a polarity group can then be achieved either two-dimensionally or off-line by partition chromatography on anotho TLC plate (Fig. 4). 

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