IDENTIFICATION AND QUANTIZATION OF OLEANOLIC ACID IN STEM AND ROOTS OF ACHYRANTHES ASPERA USING VALIDATED HPTLC METHOD

Objective: Active chemical constituent in plants exhibits pharmacological action. Hence this research work aimed to identify and quantitate oleanolic acid. Methods: A simple, high-performance thin-layer chromatography (HPTLC) is developed and validated to identify and quantify oleanolic acid. The extracts were spotted as bands on aluminum TLC plates which were pre-coated with silica gel. Different compositions of mobile phase tried for high resolution and reproducible results. Mobile phase composition toluene: methanol: ethyl acetate (8.5:1:0.5 V/V) utilized. Plates were derivatized with 10% sulphuric acid and scanned. Oleanolic acid was detected quantitatively at 529nm. Result: In the selected mobile phase, R_f  value for oleanolic acid was found to be 0.35, which indicated a good separation. The Chromatography method was validated with respect to  Linearity range (ng), Intraday and Interday precision, Instrumental Precision, Quantitation Limit i.e., LOQ (ng), Detection Limit, i.e., LOD (ng), Repeatability for oleanolic acid as per ICH guidelines. Oleanolic acid calibration graph in 200-1200 ng range per band was found linear with correlation coefficient value 0.998. All the parameters obeyed acceptable criteria as per the ICH guidelines. In the extract, the oleanolic acid was found 0.29% w/w. Hence the recently developed HPTLC technique was found prompt, cost-effective, precise, simple, accurate, reproducible, and rapid for analysis of quantity and quality of oleanolic acid in stem and roots petroleum ether extract of Achyranthes aspera. For other chromatographic techniques, this validated method of HPTLC may be useful as an economical alternative.