Thin-Layer Chromatography

• The irradiation of the sample must be diffuse.
• Monochromatic radiation must be employed for the analysis, so that the diffraction
and refraction phenomena in the layer shall be as uniform as possible. This
also means that the radiation reaching the detector in reflectance retains its
"color value" and only changes in its intensity. This would not be the case for
polychromatic light; since a certain proportion of the light is absorbed during
the determination the composition of the light would change and would,
amongst other things, alter the sensitivity of the photomultiplier to the remaining
light.
• Mirror reflection (= regular reflection) must not occur.
• The layer thickness must be large in comparison with the wavelength employed
so that no radiation can penetrate right through the layer and escape measurement.
• The particles must be randomly distributed in the layer to avoid interference
effects.
• The particles making up the adsorbent must be very much smaller in size than
the thickness of the chromatographic layer.
These general requirements also apply to adsorbents laden with substance. All
these requirements are not fulfilled to the same extent in thin-layer chromatography.
So the KUBELKA-MUNK function does not apply without qualification.
For this reason it is understandable that numerous empirical functions have been
proposed as substitutes for use in practical analysis [5, 74].
deactivation);
emitting the absorbed energy instantan
of fluorescence [3, 75, 76].
Emission
Fig. 27: Schematic representation of the relat
emission of the molecules — m and m' are tfc
numbers [4].