HPTLC Quantification of Withaniasomnifera (L.) Dunal

Abstract

At present study a simple, selective, easy, robust, sensitive and precise method isdeveloped to investigate the outcomes of HPTLC for the simultaneous determination of3 key withanolides, namely withaferin A, withanolide A and withanolide B inAshwagandha; Withaniasomnifera plant part samples. The method is analyzedsimultaneously on Camag aluminum per-coated silica gel 60- F254 HPTLC plates, usingtoluene: ethylacetate:formic acid:ethanol (6:3:0.1:0.6 v/v/v/v) as mobile phase. TheQuantitative evaluation of withanolides was performed by densitometry in thereflection-absorption mode at 530 nm. The method was validated for recovery,precision, accuracy, robustness, limit of detection, limit of quantitation, and specificityconferring to International Conference on Harmonization guidelines. Specificity ofquantification was confirmed using retention factor (Rf) values, UV Visual spectralcorrelation, and electrospray ionization mass spectra of marker compounds in sampletracks. This combination of techniques has been applied, for the first time, for theestimation of withaferin-A, withanolide A and withanolide B in fruit, leaf, stem androot parts samples of Withaniasomnifera. Present method may be beneficial in a qualitycontrol setting as it would allow rapid qualitative analysis of herbal material whilesustaining accurate quantification of extract composition and can be used for routineanalysis as well as for quality control of raw materials and herbal formulation