Preparative-scale chromatography layers

Dry-column chromatography is a variant of preparative-scale thin-layer chromatography with similar resolution but a higher sample capacity. A glass column or Nylon tube... 

Dry-column chromatography is not a widely used today. Preparative-scale thin-layer chromatography or flash chromatography is generally preferred. Although separations are fast, the recovery of separated zones is slow and labor intensive compared with elution methods. 

Nonionic surfactants, including EO-PO block copolymers, may be readily separated from anionic surfactants by a simple batch ion exchange method [21] analytical separation of EO-PO copolymers from other nonionic surfactants is possible by thin-layer chromatography (TLC) [22,23] and paper chromatography [24], and EO-PO copolymers may themselves be separated into narrow molecular weight fractions on a preparative scale by gel permeation chromatography (GPC) [25]. 

Chromatographic development chambers for analytical pirrposes are commercially available in several different sizes. The most commonly used ones are rectangiflar glass tanks with inner dimensions of 21 X21 X9 cm, and they can be used to develop two plates simultaneously in the preparative scale. Even bigger tanks are available for much larger plates, for preparative layer chromatography. The width of the chamber should be varied depending on the size and the number of plates to be developed. 

Clark et al. [53] subjected primaquine to metabolic studies using microorganisms. A total of 77 microorganisms were evaluated for their ability to metabolize primaquine, of these, 23 were found to convert primaquine to one or more metabolites (thin-layer chromatography analysis). Preparative scale fermentation of primaquine with four different microorganisms resulted in the isolation of two metabolites, identified as 8-(3-carboxy-l-methylpropylamino)-6-methoxyquinoline and 8-(4-acetamido-l-methylbutylamino)-6-methoxyquinoline. The structures of the metabolites were proposed, based primarily on a comparison of the 13C NMR spectra of the acetamido metabolite and the methyl ester of the carboxy metabolite with that of primaquine. The structures of both metabolites were confirmed by direct comparison with authentic samples. 

Thin-layer chromatography (TLC) is one of the most popular and widely used separation techniques because of its ease of use, cost-effectiveness, high sensitivity, speed of separation, as well as its capacity to analyze multiple samples simultaneously. It has been applied to many disciplines including biochemistry [1,2], toxicology [3,4], pharmacology [5,6], environmental science [7], food science [8,9], and chemistry [10,11]. TLC can be used for separation, isolation, identification, and quantification of components in a mixture. It can also be utilized on the preparative scale to isolate an individual component. A large variety of TLC equipment is available and discussed later in this chapter. 

The more recent applications of open-column chromatography in fat-soluble vitamin assays utilize liquid-solid (adsorption) chromatography using gravity-flow glass columns dry-packed with magnesia, alumina, or silica gel. Such columns enable separations directly comparable with those obtained by thin-layer chromatography to be carried out rapidly on a preparative scale. 

The traditional procedures are similiar to analytical methods with the major differences being the use of thicker sorbent layers. These procedures are generally faster and more convenient than classical column chromatography [6,49]. Preparative scale TLC usually has decreased analyte resolution compared to analytical TLC. 

Exploratory thin-layer separations are commonly used as guides in selecting the best separation conditions for corresponding preparative scale separations on columns. In this connection it should be possible to establish optimum sample size as well, using thin-layer chromatography. 

Although many of the products of the reaction were very polar, as shown by HPLC and thin-layer chromatography, we were able to determine some of the products by extracting the aqueous solution of a preparative-scale reaction done at pH 8 with diethyl ether and analyzing the extract by gas chromatography-mass spectroscopy (GC-MS). The principal ether-soluble product was 2-hydroxy-l,4-naphthoquinone (lawsone, 4). The mass spectrum of lawsone was sufficiently characteristic to distinguish it clearly from other hydroxynaphthoquinone isomers such as Juglone (11). Lawsone has also been reported previously as a product of photolysis of 1-naphthol in alkaline solution (12). 

Novel analytical techniques such as forced-flow planar chromatography (FFPC) and optimum pressure laminar chromatography (OPLC) are other additions to ever-refined tools for separation on a preparative scale, wherein small amounts of complex mixtures may be separated more efficiently on thin-layer chromatography plates operating at fast medium-pressure development with continuous collection of mobile phase at the end of chromatographic plates (Nyredy, 20(X), 2003). 

Table 5.1 illustrates the differences in scale of operations between normal analytical TLC, preparative layer chromatography and preparative column chromatography. Separations which have been successfully achieved by analytical TLC may be transferred directly to the preparative layer, because the same sorption media with the same grain size are used in each technique. However, it is not always possible to transfer directly, separations which have been successfully achieved by analytical TLC to preparative column chromatography with equal success because appreciably different adsorbent grain sizes are used in the two techniques. 

Analytical TLC Preparative layer chromatography (laboratory scale) Preparative column chromatography (laboratory scale)... 

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