Quantitative Determination of Shikimic Acid and Pinitol in Abies pindrow Aerial Parts Using TLC

Abstract

Kumar and Kumar: TLC Densitometry of Abies pindrow on the Basis of PhenolsAbies pindrow Royle (Himalayan Fir; Pinaceae) has been traditionally used in the treatment of various ailments and reported to contain cyclic polyols as major class of phytoconstituents. But no work has ever been carried out to standardize A. pindrow on the basis of cyclic polyols. Thus, it was considered worthwhile to determine quantitatively contents of main cyclic polyols, i.e., shikimic acid and pinitol in A. pindrow aerial parts using TLC densitometry. Shikimic acid and pinitol were resolved on TLC plates using solvent systems, chloroform:methanol:formic acid (14:6:1) and chloroform:methanol (3:2), respectively. The shikimic acid and pinitol were derivatized with anisaldehyde-sulphuric acid reagent and ammonical silver nitrate reagent respectively. The spots of shikimic acid and pinitol were visualized by heating derivatized plates. The plates were then scanned in TLC scanner at 500 nm. The contents of shikimic acid and pinitol were estimated to be 0.0511% and 0.0987% w/w, respectively, in A. pindrow aerial parts. The developed TLC densitometric methods for estimation of cyclic polyols in A. pindrow aerial parts were found to be precise, accurate and specific on the basis of validation parameters.Key word: Abies pindrow, Pinitol, Shikimic acid, TLC densitometry.Abies pindrow Royle (Silver Fir; family: Pinaceae) is an Indian traditional plant, which is distributed throughout Western Himalayas [1] . The plant has long history of use as anxiolytic, analgesic, antidiabetic and antiinflammatory [2] . The traditional uses of A. pindrow have been validated by scientific studies, but these studies employed crude uncharacterized extracts [3][4][5] . Sporadic phytochemical work has been carried out on the plant. Chalcone glycosides, flavonoids, fatty acids, hydrocarbons and terpenoids have been isolated from A. pindrow [6][7][8][9] . As the case with most plant drugs, A. pindrow has not been standardized on the basis of marker compounds. Marker-based standardization is important for reproducibility of therapeutic efficacy and safety of the plant. High Performance Thin Layer Chromatography (HPTLC) has gained popularity in standardizing plants by estimating the content of, preferably, bioactive constituents. Thus, the present investigation was undertaken to estimate the contents of two main cyclic polyols constituents of A. pindrow using TLC densitometry. Methanol, chloroform (E-Merck, Delhi, India), and formic acid (Ranbaxy Laboratory Chemicals, Mumbai, India), of AR grade, sulphuric acid, glacial acetic acid (E-Merck Ltd., Mumbai), sodium hydroxide, silver nitrate, ammonia solution and anisaldehyde (S.D. Fine Chemicals, Mumbai) were used in present investigations. Shikimic acid and pinitol, isolated from A. pindrow aerial parts, were used as marker compounds. The isolated compounds have been characterized by 1 HNMR and 13 CNMR spectral analysis.The stock solutions of marker compounds were prepared by separately dissolving accurately weighed 5 m...