A high-performance thin-layer chromatographic method with densitometric detection has been used to determine the convallatoxine content of extracts from the various parts (flowers, leaves, and underground parts such as the rhizomes and buds on the rhizomes) of the plant. Plant extracts were separated on thin layers of silica gel Si 60F254 by multiple gradient development. The convallatoxine content was determined by densitometry and the results were evaluated statistically.
Flavonoid glycosides are much more polar than their aglycones and the two groups of compounds are difficult to separate by planar chromatography owing to the ‘general elution problem’ — glycosides require development with mobile phases of higher eluent strength. The problem of separating both groups can be solved by gradient elution or by incremental gradient multiple development in its simplest form — double development. The latter technique is illustrated for nine glycosides and seven aglycones by development along two-thirds of the plate length with a strong mobile phase and, after evaporation of the solvent, development to the full distance with a weaker mobile phase. The two stages of development are illustrated by densitograms.
In this paper we report the possibilities and advantages of HPTLC for investigation of the acid hydrolysis of six flavonoid glycosides: the 3-arabinoside of quercetin (avicularin), the 3-glucoside of quercetin (isoquercitrin), the 3-rhamnoglucoside of quercetin (rutin), the 7-glucoside of apigenin, the 7-rhamnoglucoside of hesperetin (hesperidin) and the 7-rhamnoglucoside of naringenin (naringin). The results obtained are discussed in the terms of the chemical structures of analyzed compounds. Quantitative determinations of flavonoids were performed by use of densitometry.
Gradient thin-layer chromatography and densitometry have been used for qualitative and quantitative analysis of caffeic acid in some Dipsacaceae family plants. The presence of caffeic acid was determined in complex plant extracts before and after acid hydrolysis.
Densitometric HPTLC and HPLC have been used for quantification of p-coumaric and protocatechuic acids in an ethereal fraction from a methanolic extract of Aquilegia vulgaris L. HPLC analysis was performed on an RP-18 column with methanol-water-formic acid 25:75:0.5 (υ/υ) as mobile phase. Thin layer chromatography was performed on Si60 F254 HPTLC plates with mixtures of heptane, dichloromethane, diisopropyl ether, formic acid, and water as mobile phases. Satisfactory separation of the phenolic acids was achieved by use of the multiple gradient development technique. The quantities of p-coumaric and protocatechuic acids determined by HPLC were 0.374 and 2.283 mg g−1 dry plant material, respectively; HPTLC results were somewhat higher — 0.396 and 2.584 mg g−1, respectively. The precision of both methods, expressed as relative standard deviation, was satisfactory. The methods are useful for quality control of Aquilegia vulgaris extracts.