Quantitation of lupeol from stem bark extract of Betula alnoides Buch.-Ham. ex D.Don by two validated RP-HPLC and TLC-densitometric methods

Abstract

To control the quality of Betula alnoides stem bark (BAS), lupeol, which is a promising compound found in BAS, has been determined by reverse phase high performance liquid chromatography (RP-HPLC) and thin-layer chromatography (TLC)-densitometry. These methods were developed and validated according to the International Council for Harmonization (ICH) and the Association of Official Agricultural Chemists (AOAC) guidelines. RP-HPLC was achieved within 10 min using C8 column under an isocratic elution of acetonitrile at a flow rate of 1.0 mL/min. Lupeol was detected at a UV wavelength of 200 nm. TLC-densitometry performed on TLC silica gel plates was developed by chloroform at 10 cm of distance. Lupeol was detected under visible wavelength at 525 nm after anisaldehyde-sulfuric acid derivatization. Both developed methods exhibited good linearity, specificity, limits of detection and quantitation, accuracy and precision, and robustness. The validated methods were effectively applied to determine lupeol content in BAS collected from three different locations. The amount of lupeol was found to be about 50–100 µg/g of dried powder. The results revealed no statistically significant differences between the two methods. These findings revealed the validated RP-HPLC and TLC-densitometry that were easily applied for routine quality control of BAS for the first time.