Two-Dimensional Thin-Layer Chromatographic Determination of Phenolic Antioxidants from Eupatorium cannabinum Extracts on Cyano-Bonded Polar Stationary Phases

Two-dimensional thin layer chromatography on cyano-bonded polar stationary phase was performed to optimize the separation of some antioxidant phenolic compounds from Eupatorium cannabinum extracts. Propan-2-ol mixed with n-heptane and ethyl acetate mixed with n-heptane were used as non-aqueous mobile phases in normal phase separations (1st direction of development in 2-dimensional high-performance thin-layer chromatography (2D-HPTLC mode)) and methanol mixed with water was used as a mobile phase in reversed phase (2nd direction of development in 2D-HPTLC mode). The plates were sprayed by use of Merck TLC sprayer using 2-(diphenylboryoxy)-ethylamine and PEG4000 (Merck, Darmstadt, Germany) or DPPH and photographed in CAMAG Cabinet UV lamp at 254 nm and 365 nm by use of Fuji 8 mpx camera. The plots R-F non-aqueous mobile phase vs R-F aqueous mobile phase were prepared to find optimal orthogonal 2D-TLC systems for the separation of investigated test compounds and then extracts from Eupatorium cannabinum. Satisfactory separations of antioxidant phenolic compounds in E. cannabinum extracts were obtained by use of optimized 2D-HPTLC systems.