Thin Layer Chromatography - Elution Development in Thin Layer Chromatography 2
Finally, distribution will take place in section (Z) between pure solvent (C) and a surface covered (and consequently deactivated) with solvent (C). Even this is an over-simplification, as the composition of the mobile phase in each section will not be constant but the concentration of adsorbed solvent will decrease along the plate. Furthermore, as the separation progresses, the length of sections (X), (Y) and (Z) will continually increase. Such a system is extremely difficult to treat in a precise manner theoretically, particularly as the boundaries are nowhere near as sharp as those drawn in figure 2. In fact, the overall effect is as though the separation was carried out sequentially on three separate sections of a plate each section having a different stationary phase and mobile phase. In each section, the separation will then be achieved by elution development but the overall effect will be a form of gradient elution. The complexity of the system increases with the number of solvents used and, of course, their relative concentrations. The composition of the eluting solvent is basically the concentration profile that would result from the frontal analysis of the mobile phase solvent mixture. This somewhat complicated elution procedure does not in any way affect the accuracy or precision of any quantitative assays but tends to make the optimum choice of phase system sometimes extremely difficult to predict.
The elution process can be simplified considerably by pre-conditioning the plate with solvent vapor from the mobile phase before the separation is started. Unfortunately, this only partly reduces the adsorption effect, as the equilibrium between the solvent vapor and the adsorbent surface will not necessarily be the same as that between the liquid solvent and the surface. It is clear that by forming a gradient by the frontal analysis of the mobile phase and carefully choosing the solvent mixture, very delicate pseudo-gradients can be formed which, in no small measure, accounts for the versatility, popularity, and success of TLC.