Post Chromatographic Derivatization

It is an inherent advantage of planar chromatography that fractions are stored on the plate and can readily be derivatized after chromatography in order to render them detectable, to improve detection limit, or to change selectively properties of sample components.

Liquid derivatizing agents can be transferred onto the plate by spraying or dipping. Provided the reagent is suitable, dipping is the preferred technique.

The Chromatogram Immersion device by Camag is an example of an instrument that allows proper execution of the dipping technique. The chromato-graphic plate must be immersed and withdrawn at a uniform speed to avoid tide marks, which could interfere with densitometric evaluation. By maintaining a defined immersion time, derivatization conditions can be standardized.

Spraying has to be used when two reagent solutions have to be applied in sequence without intermediate drying. Diazotization followed by coupling is an example. There are several sprayers on the market, from simple laboratory atomizer to electropneumatic TLC sprayers.

Whenever reagents are sprayed onto a plate, an efficient dust and mist-removing device should be used to protect laboratory personnel against poisonous or irritating sprays and solvent vapours. Com-merical equipment ensures the complete removal of excess spray from the atomizer and spray particles rebounding from the TLC plate. There is no deflection of the spray jet before it reaches the chromato-gram, an effect often occurring in a normal laboratory fume hood.

In most cases, the derivatization reaction needs to be completed by heat treatment. Heating the chromatogram plate uniformly and reproducibly at the desired temperature can be accomplished with a heater specifically designed for this purpose.

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