Quantitative Determination of Alkaloids with TLC

The spots are photographed (black and white) in daylight or UV light, either directly or after detection with some reagent. The size and intensity of the photographed spots are then evaluated photometrically. Poethke and Kinze [177] have attempted photometric evaluation directly on the layer but the limits of error are rather broad in this procedure. 

The zone and adsorbent are scraped off the plate and transferred quantitatively to an extractor. Methanol is generally used for the extraction, the volume being then made up to a definite amount. The substance itself is determined polarographically, colorimetricaUy (after conversion into a coloured derivative) or, easily the best, spectrophotometricaUy at the wave length in the UV where the substance has a characteristic adsorption maximum. A blank on the adsorbent is necessary in the spectrophotometric method. The amount of substance can then be calculated frpm the extinction coefficient. This combination of TLC and spectrophotometric determination of the eluted substance, is a method of wide analytical potentiality in biology and chemistry. 

Radioactively labelled substances may be radiometricaUy evaluated directly or also after scraping off the plate (see Chapter I, p. 155). 

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