Preparative On-Line Overpressure Layer Chromatography (OPLC): a New Separation Technique for Natural Products

Abstract

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

Separation of Cucurbitacins by Overpressure Layer Chromatography fromHemsleya gigantha1

“…In order to optimize the mobile phase composition for a single development, the PRISMA method was then used (19,20 (7,9,11,(22)(23)(24)(25). OPLC has been shown to be a useful separation technique in natural products research for both analytical and preparative purposes (20,21,26,27). Using this technique, we are now able to separate cucurbitacins having an unsaturated side-chain from their corresponding 23,24-dihydro derivatives.…”

“…sample B could be separated into 3 and 4 in 20 mm. Based on these analytical results, a larger quantity (50mg each) of A and B was finally purified by using the OPLC on-line preparative method(21).Thefour isolates were then identified as 23,24-dihydrocucurbitacin-F-25-O-acetate (1), cucurbitacin-F-25-O-acetate (2), cucurbitacin-F (3), and 23,24-dihydrocucurbitacin-F (4), by comparison with authentic samples and/or by spectroscopic analysis ('H-and '3C-NMR, MS)…”

New planar chromatographic techniques in natural product analysis

“…The use of OPLC as an on-line preparative method is reported for the separation of cannabinoids (Oroszlan et al, 1987), poppy alkaloids, furocoumarins and secoiridoid glycosides (Nyiredy et al, 1986b), and for cytotoxic canthin-6-one alkaloids, epimeric cardiac glycosides (Erdelmeier et al, 1987), quassinoids and Podophyflum lignans (Erdelmeier et a f . , 1986).…”

“…, 1986). This method has also proven useful for the rapid isolation or purification of single, major constituents from natural sources (Nyiredy et al, 1986b;Erdelmeier et al, 1987;Meksuriyen and Cordell, 1988;Fullas et a f . , 1989).…”

Recent Developments in the Isolation and Structure Elucidation of Naturally Occurring Iridoid Compounds

“…However, early systematic studies were reported by Cornelis et al (95) and Bock et al (96). The structure of secogalioside (42), for example, was assigned by chemical correlation with sweroside (17) using 1H-and 13C-NMR data as well as NOB experiments, thus allowing the full structure determination including conformation as well as the absolute configuration (96). Furthermore, it has been shown that compounds like morroniside (43) exist as a mixture of C-7 anomers in solution leading to mutarotational equilibrium.…”