TLC-image analysis of non-chromophoric tuberostemonine alkaloid derivatives in Stemona species

A simple, selective, precise, and accurate thin-layer chromatographic (TLC) image analytical method was developed and validated for simultaneous quantification of the major components in the root extracts of Stemona tuberosa (tuberostemonine, tuberostemonine N and neotuberostemonine)), and S. phyllantha (tuberostemonine and tuberostemonine A). The analysis was performed by TLC on silica gel 60 F254 aluminum plates using a mixture of dichloromethane: ethyl acetate: methanol: ammonium hydroxide (50:45:4:1) as mobile phase. Post-derivatization was employed by dipping the TLC plate into Dragendorff's reagent to visualize the spots. Image analysis of the scanned TLC plate was performed to detect the contents of tuberostemonine derivatives. The polynomial regression data for the calibration plots showed good linear relationships within the concentration range of 2-7 microg/spot. The method gave satisfactory precision, accuracy, selectivity and could simultaneously quantify tuberostemonine, tuberostemonine A, tuberostemonine N and neotuberostemonine. Dried powdered roots of S. tuberosa grown in Thailand contained 1.31 +/- 0.28, 1.63 +/- 0.18 and 1.24 +/- 0.27% tuberostemonine, tuberostemonine N, and neotuberostemonine (dry weight), respectively, while S. phyllantha roots contained 1.39 +/- 0.14% tuberostemonine and 0.39 +/- 0.08% tuberostemonine A (dry weight). The proposed method was simple, inexpensive, and more accessible to apply for many local authorities and small laboratories.