Organic Chemistry Laboratory Techniques, 2016a

2.3.A.4 C THE BANDS ARE ELUTING UNEVENLY
If the components of a mixture are colored, it may be obvious
when the bands elute in a crooked manner. This is most likely
due to the column being clamped at a slight diagonal.
If the column is clamped in a slanted manner, components will
travel in a slanted manner (Figure 2.67). This may cause
separation problems if the components have a similar R f .
There is no way to fix this problem midway through a column,
but if the components have very different R f values, the slanted
bands may have no effect on the separation. In the future, be
sure to check that the column is perfectly vertical in both the
side-to-side and front-back directions.
Figure 2.67: Effect of a crooked column on separation.
2.3.B MICROSCALE (PIPETTE) COLUMNS
A macroscale column is much too big for very small quantities of material (< 20 mg). Instead, a column
can be constructed using a disposable Pasteur pipette.
In order to get good separation, it is ideal if the desired component has an R f around 0.35 and is separated
from other components by at least 0.2 R f units. 11 If the spots to be separated are very close (< 0.2 ΔR f ), it’s
best if the middle of the spots has an R f of 0.35. An R f near 0.35 is ideal because it is slow enough that
stationary-mobile phase equilibration can occur, but fast enough to minimize band widening from diffusion.
2.3.B.1 STEP-BY-STEP PROCEDURES
Figure 2.68: a) TLC plate of purple dye, b) Elution with a pipette column.
The column pictured in this section shows purification of a drop of dilute purple food dye (made from 1
drop red dye, 1 drop blue dye and 15 drops water). The dye is separated as best as possible into its three
components: blue, red and pink (as seen in the TLC plate of Figure 2.68a). A 2.5” column of silica gel is
used and eluted with a solution made from a 1:3:1 volume ratio of 6 M NH 4 OH:1-pentanol:ethanol.
11 W.C. Still, M. Kahn, A. Mitra, J. Org. Chem., Vol.43, No.14, 1978.
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