When TLC is performed, compounds are generally identified using the retardation factor (Rf),
which is relatively constant for a particular compound in a given solvent. The Rf is calculated using
the equation:
Equation 12.1Because its value is relatively constant, the Rf value can be used to identify unknown compounds.
This technique is most frequently performed on a small scale to identify unknown compounds. It
can also be used on a larger scale as a means of purification, a technique called preparative TLC. As
the large plate develops, the larger spot of sample splits into bands of individual compounds, which
can then be scraped off and washed to yield pure compounds.
COLUMN CHROMATOGRAPHY
The principles behind column chromatography are the same as for thin-layer chromatography,
although there are some differences. First, column chromatography uses an entire column filled
with silica or aluminum beads as an adsorbent, allowing for much greater separation. The setup for
this is shown in Figure 12.6. In addition, thin-layer chromatography uses capillary action to move the
solvent up the plate, whereas column chromatography uses gravity to move the solvent and
compounds down the column. To speed up the process, one can force the solvent through the
column using gas pressure, a technique called flash column chromatography. In column
chromatography, the solvent polarity can also be changed to help elute the desired compound.