TLC-DENSITOMETRIC QUANTIFICATION OF NEGUNDOSIDE, URSOLIC ACID, EUGENOL, LUPEOL, ANDβ-SITOSTEROL USING HPTLC FROMVITEX NEGUNDOLEAVES

Vitex negundo Linn (Verberaceae) is an important drug in Indian, Chinese, and Malaysian Systems of medicine. The leaves are reported to show anti-inflammatory, anti-ulcer, anti-implantation, and analgesic activities. They contain negundoside, nishindaside, agnuside, luteolin, vitexicarpin, ursolic acid, and β-sitosterol. Lupeol, an important marker compound has not been reported from V. negundo until recently. Our preliminary TLC experiments indicated the presence of lupeol in the leaves of V. negundo. Subsequently, lupeol was isolated by preparative TLC. The identity of lupeol was confirmed by comparing the mass spectra [m/z value 427.8 (M+1) and 425.8 (M−1)]; melting point (215°C, using DSC) and co-TLC with the standard compound. A High Performance Thin Layer Chromatography (HPTLC) method for the quantification of five marker compounds, namely, negundoside, ursolic acid, eugenol, lupeol, and β-sitosterol from the leaves of V. negundo was developed and validated as per ICH guidelines. The amounts of negundoside, ursolic acid, eugenol, lupeol, and β-sitosterol were found to be 1.048, 0.013, 0.115, 0.116, and 0.043% w/w, respectively. The developed methods were found to be accurate, precise, and reproducible. They can be used for routine quality control of herbal material and formulations containing these compounds.