Paper chromatography Chambers

There is no other facet where thin-layer chromatography reveals its paper-chromatographic ancestry more clearly than in the question of development chambers (Fig. 56). Scaled-down paper-chromatographic chambers are still used for development to this day. From the beginning these possessed a vapor space, to allow an equilibration of the whole system for partition-chromatographic separations. The organic mobile phase was placed in the upper trough after the internal space of the chamber and, hence, the paper had been saturated, via the vapor phase, with the hydrophilic lower phase on the base of the chamber. 

Obtain a paper chromatography developing chamber (similar to that shown in Figure 11.12). Pour 70% isopropyl alcohol (the mobile phase) into the chamber to a depth of 2 cm. Then, place the paper sheet in the chamber as shown in Figure 11.12. Cover the chamber. 

Whatman 3MM paper, 20 X 20 cm Amino acid standard solutions, 1% in water Chromatography chambers... 

Tfmt layer chromatography is similar to paper chromatography except that a coated glass or plastic plate is used instead of paper, and the results are visualized via an iodine vapor chamber. 

The different methods of arranging the filter paper in the chromatographic chamber have been applied to sugar separations descending, ascending,39 40 and circular or horizontal development.41 These methods and other techniques will be discussed below. Detailed descriptions of the equipment needed for paper chromatography are readily available.8... 

There are two popular LC techniques in which the stationary bed is supported on a planar surface rather than in a column paper chromatography (PC) and thin-layer chromatography (TLC). PC preceded TLC by some 10 to 15 years, and a large number of excellent separations were devised for it. But beginning about 1956, it was found that TLC could also be used for most of these separations and that it was faster, more reproducible, more versatile, and more convenient. As a result, most laboratories have abandoned the use of PC with its large cumbersome glass chambers. Those who have not, continue to use PC because they feel that the original PC methods are superior or because of the lower cost of PC. 

Figure 21-7. Equilibrium chambers for paper chromatography. Left, glass jar type. Right, Stainless steel type. (Courtesy - A. H. Thomas Scientific Company, Swedesboro, NJ)...

The filter paper is placed in the chamber and the vessel is closed by placing the lid securely in its top. Ample time for vapour equilibration is allowed before the strip is lowered (by some means, not involving opening the Figure 111 Set-up of ascendlng paper chromatography. 

Figure 21.10. Developing chambers for paper chromatography. A Ascending development B Descending development. Note that similar chambers are used in TLC. From D. Abbott and R. S. Andrews, An Introduction to C3iromatography, Boston Houghton Mifflin, 1965, by permission of the publisher.

Samples are spotted on the plate, which is developed in a solvent chamber as described for paper chromatography. The solvents used for TLC are the same ones used in column chromatography on the same stationary phase. 

The group solution for analysis is best applied in 2 or 3 different concentrations alongside one another at the start points. It is possible in this way to detect traces of an element and also to eliminate occlusions through excessive concentrations of another element. It is also always an advantage to carry out parallel chromatography of the ions expected ia the chromatogram. The chromatography chambers in aU work are lined with filter paper to ensure a constant vapour phase (CS). 

In the case of thin-layer chromatography there is frequently no wait to establish complete equilibrium in the chamber before starting the development. The chamber is usually lined with a U-shaped piece of filter paper and tipped to each side after adding the mobile phase so that the filter paper is soaked with mobile phase and adheres to the wall of the chamber. As time goes on the mobile phase evaporates from the paper and would eventually saturate the inside of the chamber. 

Over 50 different pyridazin-3-ones were evaluated for biological activity in a wheat (Triticum aestivum L.) test system described previously (1). Briefly, seeds were germinated in 9-cm petri dishes on three layers of filter paper. Pyridazinones were dissolved in acetone and the filter papers were impregnated with 1 ml of acetone solution. After the soluent evaporated, 10 ml of distilled water were added to form an inhibitor concentration of 100 yM. Seeds were planted directly on the moist papers and germinated for 4 days in a controlled environment chamber on a 16-hr photoperiod with 27+lC day temperature and 21+lC night temperature. Light intensity from both fluorescent and incandescent bulbs was 28 klux at dish level. Lipids were extracted and recovered from 1 g of lyophilized shoot tissue, separated into membrane and non-membrane lipids, and analyzed by gas chromatography as described (1). 

The procedure of paper and thin-layer chromatography. A Application of the sample. B Setting plate in solvent chamber. C Movement of solvent by capillary action. D Detection of separated components and calculation of Rf. 

A typical chamber for paper and thin-layer chromatography. 

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