overpressure layer chromatography (oplc) is further developed for the online preparative isolation of natural compounds. Its use is demonstrated for the rapid isolation of one or two compounds in less than 1 h as well as for the isolation of more substances from plant extracts andor prepurified fractions which require a few hours. Depending on the separation problem, sample sizes between 50 mg and 0.5 g may be applied to 20x20 cm silica plates. Principles of the method, practical considerations, and advantages of this techique over other available methodologies are discussed.For many years, the most commonly used method in preparative planar chromatography has been traditional layer chromatography, requiring removal and subsequent elution of substance zones (1). Sequential thin layer chromatography (stlc) with locally and temporally variable solvent application (2,3) has also been useful for preparative purposes (4), even though it is an off-line method. One major advance in preparative planar chromatography was the development of centrifugally accelerated layer chromatography (clc), allowing on-line separations to be made. It has been shown, however, to be rather inefficient when used to solve complex separation problems ( 5 ) .Overpressure layer chromatography (oplc) is a forced-flow technique developed by Tyihak, et af. (6)(7)(8) in which the vapor phase is eliminated by completely covering the sorbent layer with an elastic membrane under external pressure. The separation is thereby carried out in a closed system with parameters that may be controlled. To date, there is information available on preparative oplc separations only in the off-line mode We now report the use of oplc on a 20-and 40-cm distance for preparative on-line (9-11). separation of naturally occurring compounds.
METHODSThe chromatographic plate becomes a planar column by sealing the plate all around and by draining off the solvent. The eluent passes a detector system and is fractionated by an automatic collector. The flow scheme is depicted on Figure 1. MOBILE PHASE IWLEl I
SummaryThis paper describes the fundamental factors causing the "disturbing zone" in OPLC. This phenomenon of distorted substance zones appears when the OPLC separation is started with a dry layer and is due to interaction between the gas that is physically bound to the surface of the sorbent (ma) and the gas molecules dissolved in the eluent (mis). Possibilities for elimination of the adverse effectsof disturbing zones ultimatelydepend on the establishment of operating conditions that inhibit their initial formation. Since modification of the location of the disturbing zone is only possible within a very narrow range, the sole solution to this problem is to conduct a pre-run, which may be carried out with hexane in the case of separation of apolar compounds. For separation of polar substances, the pre-run may also be performed with hexane or with a solvent of the mobile phase in which the components are unable to migrate. The selection of this solvent may be considered during the optimization of the eluent system.
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