Focus
How to get a headstart on new
(a)
b)
(c)
Solvent
Front
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projects. Figure 1.
Spot reconcentration mechanism
Spot Width
Migration Distance
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Figure 2.
Spot width vs. migration distance in HPTLC
PTH-AAs from a complex mixture demonstrated that HPTLC is capable of handling a fairly thorny analytical problem, since this separation is hardly a trivial one. Poole and his colleagues performed the separation on silica gel plates using five development steps with four changes of mobile phase. The derivatives were identified by scanning densitometry, the total analysis requiring less than one hour. And even the two unseparated derivatives, alanine and tryptophan, could be baseline resolved in an alternative HPTLC solvent system. One might think that a procedure such as this, in which five separate development steps are performed, would lead to excessive spot broadening and a consequent degradation of resolution. In fact, this is not the case, due to a phenomenon known as the spot reconcentration mechanism (see Figure 1). The spot in part (a) of the figure represents a component separated out in a single development step. The horizontal line represents an advancing solvent front in a second development step with a different solvent. In part (b), the solvent front has advanced just past the original spot, and the component involved has been reconcentrated into a spot of smaller area. In part (c) the new spot has migrated up the plate somewhat. In a protracted single-solvent unidimensional development process (Fig-
794 A · ANALYTICAL CHEMISTRY, VOL. 54, NO. 7, JUNE 1982
ure 2), a spot broadens linearly as it moves up the plate. With multiple development, on the other hand, a relatively constant spot width can be maintained due to the spot reconcentration that occurs during each cycle. Prospects. According to Fenimore and Davis (reference above), market demands will likely stimulate the development of HPTLC plates with tighter particle size distributions and even smaller average particle sizes. In addition, improved microcomputercontrolled densitometers with refined optical systems will probably be developed for HPTLC. The latest commercial developments in HPTLC were recently reviewed by Thomas H. Maugh II in Science (1982,226,161-63). An exciting recent development is the growing popularity of reversed phase stationary phases for HPTLC. In normal adsorption HPTLC, the mobile phase is less polar than the stationary phase (usually silica gel). In reversed phase TLC, it is the other way around. The reversed phase adsorbent is produced, as in LC, by reacting silica gel with an organochlorosilane to generate a C-8 or C-18 hydrophobic layer on the bead. The availability of reversed phases should make it possible for HPTLC to make further inroads into a domain that was once the sole province of its more glamorous relative, LC. Stuart A. Borman
