Kieselgel 60 f254 tlc plates

Thin layer chromatography (TLC) plates (Merck, silica gel 60 F₂₅₄) were used to separate the extracts into chemical constituents. The TLC plates were prepared in duplicate and developed under saturated conditions in different mobile solvent systems according to Kotze and Eloff [36 (link)]: Ethylacetate: methanol: water (EMW) (40:5.4:4) polar/neutral; Chloroform: ethylacetate: formic acid (CEF) (20:16:4) intermediate polarity/acidic and Benzene: ethanol: ammonium hydroxide (BEA) (36:4:0.4) non-polar/basic. An aliquot of 10 μℓ of extract (representing 100 μg of the extract) was placed in a line c, 0.8 cm long and was separated by TLC using Merck, Kieselgel 60 F₂₅₄ TLC plates in a closed, saturated TLC tank. For the qualitative evaluation of a given substance, the R_f value (retention factor) of chromatograms was used as the parameter for comparison. The R_f value of a substance is the ratio of the distance moved by the compound from its origin to the movement of the solvent from the origin. The TLC plates were then sprayed with the Vanillin-sulphuric acid spray reagent (0.1 g vanillin, 28 ml methanol, 1 ml sulphuric acid) for the detection of higher alcohols, phenols, steroids and essential oils [37 ]. The plates were heated at 105 °C until the colours of chromatograms were optimally developed.