Page 181 - Multidimensional Chromatography

174 Multidimensional Chromatography

if, as a result of the use of mobile phases of different composition, the interactive
forces which bring about retention are different for the two consecutive develop-
ments. A good separation can be obtained when the surface area of the plate over
which the spots are spread is relatively large (Figure 8.3(d)).
Thus, a 2-D separation can be seen as 1-D displacement operating in two dimen-
sions. The 2-D TLC separation is of no interest if selection of the two mobile phases
is not appropriate. With this in mind, displacement in either direction can be either
selective or non-selective. A combination of two selective displacements in 2-D TLC
will lead to the application of different separating mechanisms in each direction. As
an extreme, if the solvent combinations are the same (S T1 S T2 ; S V1 S V2 ) or very
similar (S T1 S T2 ; S V1 S V2 ), the compounds to be separated will be poorly
resolved or even unresolved, and as a result a diagonal pattern will be obtained. In
such circumstances, a slight increase in resolution might occur, because of an
increase by a factor of √2 in the distance of migration of the zone (4).
The point at which the sample is spotted can be regarded as the origin of a coordi-
nate system (9). The process of development is performed in two steps; the first in
the direction of the x-axis to a distance L x . After evaporation of the solvents used, the
second development will be performed in the direction of the y-axis to a distance L y .
The positions of the compounds after development in the x-direction depend on the
S T and S V values of the first mobile phase being applied. Similarly, the migration dis-
tances of the individual compounds also depend on the total solvent strength and
total selectivity of the second mobile phase. After development in the x-direction, the
ordinates of all compounds are zero. After development in the y-direction, their
abscissa values follow from their positions on the x-axis after the first development.
The final positions of the spots are thus determined by the coordinates x(i) and y(i),
which can be expressed as follows:

(8.1)
R fxy(i) R fx(i) , R fy(i)
The principle of 2-D TLC separation is illustrated schematically in Figure 8.4. The
multiplicative law for 2-D peak capacity emphasizes the tremendous increase in
resolving power which can be achieved; in theory, this method has a separating
2
capacity of n , where n is the one-dimensional peak capacity (9). If this peak capac-
ity is to be achieved, the selectivity of the mobile phases used in the two different
directions must be complementary.
For two reasons, the peak capacity in 2-D TLC is less than the product of those of
two one-dimensional developments (10). First, the sizes of the spots of the com-
pounds being separated are always larger in the second development than was the ini-
tial sample spot. Secondly, during the second development the spots spread laterally
and must therefore be separated with a resolution greater than unity at the beginning
of the second development if they are to have a resolution of unity at the end (1). The
separation efficiency can be increased by performing multiple development in one or
n
both directions. Therefore, 2-D TLC combined with D is a promising route to real
improvements in planar chromatography in the future (see Section 8.13 below).