Simultaneous estimation of three triterpenoids‑ursolic acid, b‑sitosterol and lupeol from flowers, leaves and formulations of Rhododendron arboreum Smith. using validated HPTLC method

Abstract

Background: This paper enfolds a rapid and sensitive high‑performance thin‑layer chromatographic (HPTLC) method for the simultaneous estimation of three triterpenoids namely ursolic acid, β‑sitosterol and lupeol from the leaves, flowers and herbal formulations of Rhododendron arboreum Smith., an ethnomedicinal Himalayan tree. All the three phytoconstituents have high therapeutic value. Aims and Objectives: The main aim is to separate, resolve and simultaneously quantitate the three markers‑ursolic acid, β‑sitosterol and lupeol from R. arboreum using normal phase HPTLC. Materials and Methods: Separation was performed on TLC aluminium plates precoated with silica 60 F254 followed by detection of ursolic acid, β‑sitosterol and lupeol carried out by derivatizing the plate with 10% methanolic sulphuric acid reagent followed by heating at 110°C for 7 min. Camag TLC scanner 4 equipped with winCATS software was used for densitrometric scanning at 366 nm. The proposed method was further validated in terms of linearity, precision, accuracy and sensitivity as per the International Conference on Harmonisation (ICH) guidelines. Results: A good linear relationship was obtained for the calibration plots with r2 = 0.999, 0.993 and 0.995 for ursolic acid, β‑sitosterol and lupeol, respectively. Accuracy of the method was checked by recovery study conducted at three different levels with the average recovery between 95% and 98% for all the three markers. Conclusion: The developed method can be used for the assessment of the quality of botanicals in terms of bioactive content. Key words: Ethnomedicinal, Rhododendron arboreum, simultaeneous estimation, triterpenoids