An extraction and analytical protocol for saponins were established for Verbascum thapsus L., a medicinal plant. Four different kinds of plant sample were analyzed; field-grown, in vitro cultured, commercially obtained leaves and field-grown capsules. A cleanup procedure with Octadecyl (C18) Solid Phase Extraction Column was used prior to HPLC analysis. Ilwensisaponin A was used as an external standard and digitoxin as an internal standard. C18 reverse phase column and gradient elutions (Acetonitrile with 0.1 % Orthophosphoric acid and Water with 0.1 % Orthophosphoric acid) were used for HPLC analysis. Commercially obtained leaves had a higher concentration of saponin (0.215 mg/g tissue) than other leaves (0.081-0.198 mg/g tissue) and capsule sample (0.016 mg/g tissue)