plate rotated 90° and redeveloped in the orthogonal direction. If the same solvent is used for both developments, only a slight increase in resolution can be anticipated (a factor of >/2). This in crease will correspond to the increased migration distance for the sample. More efficient separation requires the resolved sample to be distributed over the entire plate surface, necessitating complementary separation mecha nisms for the orthogonal development that may be challenging to achieve. Difficulties in scanning two-dimen sional chromatograms with slit-scan ning densitometers and in the con struction of a two-dimensional forcedflow instrument have prevented twodimensional TLC from reaching its true potential as a separation tool. The potential of a chromatographic system to provide a certain separation can be estimated from its separation number, referred to as the spot capaci ty in TLC (i.e., the number of spots resolved with a resolution of unity that can be placed between the sample spot at the origin and the spot of an unretained compound) (17, 26). An exact spot capacity value is more difficult to calculate than the equivalent value for a column system because it depends on many experimental variables that are not well understood. For a single, unidimensional devel opment using capillary flow-controlled conditions, it is not difficult to achieve a spot capacity between 10 and 20, but it is very difficult to reach 25 and prac tically impossible to exceed 30. To ob tain separation numbers greater than 25, very long plates and prohibitively long separation times would be needed. (In HPLC, separation numbers around 150 can be achieved; in exceptional cases, a value of 500 can be obtained.) In theory, the spot capacity of forcedflow TLC should be identical to that of column chromatography. In practice, however, shorter bed lengths and limit ed inlet pressures compared with those used in column chromatography set a practical limit of about 80 with com mercially available equipment (with a plate length of 30 cm, a particle size of 5 μτα, and a pressure drop of 36 atm). For two-dimensional TLC under capil lary controlled-flow conditions it should be easy to achieve a spot capaci ty of 100-250, but it will be difficult to reach 400 and nearly impossible to ex ceed 500 except in very favorable cir cumstances. If forced-flow develop ment is used in the first direction and elution in the second, a spot capacity of a few thousand is achievable. This rep resents an order of magnitude increase in separation potential over column systems, but the experimental difficul
ties in implementing such a separation system are formidable (27). The confident analysis of moderateto-complex mixtures requires a large separation capacity that is most readily obtained by using multidimensional and multimodal separation techniques (21,23). In multimodal separations two different separation techniques are combined via an interface that ideally allows independent and optimized use of the two separation techniques. Ex amples of multimodal techniques in clude GC/TLC, HPLC/TLC, and SFC/ TLC. Spectroscopic techniques such as IR, Raman, and mass spectrometry can be used in alternative multimodal ap proaches in which partially separated sample zones are further resolved based on differences in their spectro scopic properties (23, 28). The TLC plate can function as a storage medium, enabling spectroscopic evaluation that is free of time constraints and timeconsuming techniques such as signal averaging to improve sensitivity. Interface requirements for coupling microcolumn HPLC or SFC to TLC are comparatively simple. With microbore HPLC columns that operate at flow rates of 5-100 /iL/min, the complete
column effluent can be deposited on the TLC plate using a modified sprayjet band applicator (29,30). For exam ple, Jaenchen used this approach to separate a mixture of 56 pesticides by a combination of reversed-phase microbore HPLC and normal-phase auto mated multiple development (31). More than a decade ago, Stahl (32) de scribed an apparatus for supercritical fluid extraction (SFE) and deposition onto a TLC plate, which used a very fine heated capillary (25-50 μνα) as the interface. Decompression of the super critical fluid occurs with rapid cooling that favors the deposition process without inhibiting the evaporation of the decompressed fluid. The current strong interest in SFE should encour age réévaluation of StahPs early work, particularly if spectroscopic evaluation for structural identification is required. The TLC plate in this case is a much more convenient fluid elimination interface for spectroscopy than some of the on-line approaches currently under development. Certain spectroscopic techniques such as lowtemperature fluorescence line-narrowing spectroscopy, surface-enhanced Raman spectroscopy, and liquid sur-
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ANALYTICAL CHEMISTRY, VOL. 61, NO. 22, NOVEMBER 15, 1989 · 1261 A
