Gradient elution in TLC

Gradient development. Gradient developments have been used to achieve separation of complex mixtures such as plant extracts, dyes, etc. However, their application to lipid separation is not very common. Golkiewicz (1996) discusses in detail stationary phase gradients and mobile phase gradients, the theory behind solvent selection, automated techniques and applications of gradient elution in TLC. 

Figure 3 Devices for gradient elution in TLC. (Reprinted from Ref. 7 with permission.)...

Selectivity of the separation in TLC is achieved by various of the aforementioned techniques (e.g. multiple development, gradient elution, sequence TLC, AMD, HPPLC or OPLC). Multidimensional TLC methods are described in Section 7.4.4. 

Burger, K. TLC-PMD, thin-layer chromatography with gradient-elution in comparison to HPLC. Fresenius. Z. Anal. Chem. 1984, 318 (3 ), 228. 

The application of gradient development in TLC greatly improves the separation efficiency, especially in the case of complex samples containing components of wide range of polarity. Gradient elution can separate all components in a single run. 

Successful separations of many complex mixtures by HPLC gradient elution have demonstrated the utility of this technique (1-5). In contrast to HPLC, gradient development in TLC has been applied relatively rarely, owing to the rather complex devices required for the generation of reproducible gradients and lack of a simple theory of gradient development. Niederwieser and Honegger (6,7) have systematized many experimental results and outlined some theoretical problems. 

This equation, although originating from the plate theory, must again be considered as largely empirical when employed for TLC. This is because, in its derivation, the distribution coefficient of the solute between the two phases is considered constant throughout the development process. In practice, due to the nature of the development as already discussed for TLC, the distribution coefficient does not remain constant and, thus, the expression for column efficiency must be considered, at best, only approximate. The same errors would be involved if the equation was used to calculate the efficiency of a GC column when the solute was eluted by temperature programming or in LC where the solute was eluted by gradient elution. If the solute could be eluted by a pure solvent such as n-heptane on a plate that had been presaturated with the solvent vapor, then the distribution coefficient would remain sensibly constant over the development process. Under such circumstances the efficiency value would be more accurate and more likely to represent a true plate efficiency. 

Thin-layer chromatography (TLC) is used both for characterization of alcohol sulfates and alcohol ether sulfates and for their analysis in mixtures. This technique, combined with the use of scanning densitometers, is a quantitative analytical method. TLC is preferred to HPLC in this case as anionic surfactants do not contain strong chromophores and the refractive index detector is of low sensitivity and not suitable for gradient elution. A recent development in HPLC detector technology, the evaporative light-scattering detector, will probably overcome these sensitivity problems. 

The problem of the separation of samples containing components of widely different polarities is difficult because of general elution. This can be solved by use of gradient elution. As has been observed, in TLC separation of plant extracts, gradient elution markedly improves the separation of spots owing to stronger displacement effects... 

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