Chapter 3: Sample Pre-treatment

1 Introduction

Pre-treatment of samples before application to the TLC/HPTLC layer rarely requires more than a few basic steps. The quality of commercially available TLC and HPTLC layers is such that most analytes can be more easily distinguished from impurities than was possible with "home made" plates. Usually the presence of sample contaminants does not cause a problem in TLC. Whereas the repeated injection of contaminants on to an HPLC column can quickly render it useless, TLC or HPTLC plates are normally only used once and are generally less sensitive to contamination. When the impure sample is applied to the sorbent layer as a spot or band, both the components of interest and the interfering impurities are deposited together. Once development begins, the contaminants are often left behind at or near the origin whilst the components of interest migrate in the direction of flow of the solvent front. If the sample solvent is mainly aqueous or viscous, then dilution with an organic polar solvent, like methanol, ethanol or acetonitrile, will aid application to the layer. The sample solution is then able to wet the surface and penetrate into the sorbent effectively. The application will take on a regular spherical shape in the case of spots and a fine, sharp, well-defined line in the case of bands. Filtering of the sample solution is also an important step that can improve the result of the eventual chromatogram. As the sample volumes will be small, simple syringe filters of 0.45 or 0.2 ?