n-Hexane fraction (100 mg) of V. negundo leaves was dissolved in 40 mL of n-hexane and the volume was made up to 50 mL in a volumetric flask. This fraction was used for the quantification of lupeol and β-sitosterol. Fifteen microliters of this dilution was applied in triplicate on a TLC plate along with standards, lupeol, and β-sitosterol. The plate was developed as mentioned above and scanned at 525 nm. The peak areas were recorded and the amount of lupeol and β-sitosterol was calculated using the respective calibration curves.