Chemical Reactions on the Layer

Most chemical reactions on the planar chromato-graphic layer are carried out after development of the chromatogram. This is described as post-chromato-graphic visualization and is particularly useful when the presence of 'unknowns' in the sample need to be detected. However, it is also possible, as will be shown later, to carry out a pre-chromatographic de-rivatization before chromatographic development and this can have significant advantages. As post-chromatographic visualization is the major method of detection, it is this technique that will be given consideration first.

Post-chromatographic visualization Post-chromato-graphic identification of chromatographic zones can be achieved by spraying or dipping the TLC/HPTLC layer with a universal or group-specific reagent. Sometimes 'destaining' techniques can improve sensitivity where such reagents are employed. Where complex mixtures of large numbers of analytes of differing functionality are involved, sequencing reactions can be used to locate groups of compounds progressively.

The most used and dependable reagents are discussed in detail below, however, this represents only a small fraction of the detection reagents available, and the literature should be consulted for a more comprehensive review. In choosing the best detection reagent, the following points should be borne in mind:

1. Sensitivity of detection.

2. Selectivity of the reagent for the analytes of interest.

3. Background effects, particularly where plates are to be scanned spectrophotometrically.

4. Stability of detection reagent.

5. Stability of the chromatogram after chemical or thermal treatment.

6. Ease of preparation of the spraying or dipping reagent.

7. Hazards associated with the use of particular detection reagent.

Universal chemical reagents

Iodine vapour/solution Iodine reacts chemically with unsaturated compounds, which is rarely reversible on silica gel layers. Also irreversible oxidations, electrophilic substitutions, addition reactions, and the formation of charged transfer complexes have been observed. The so-called 'iodine reaction' is possibly an oxidation with the initial formation of a radical cation as shown in the following reaction equation:

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