DEVELOPMENT AND VALIDATION OF HPTLC METHOD FOR IDENTIFICATION AND QUANTIFICATION OF URSOLIC ACID IN THE LEAVES OF ALSTONIA SCHOLARIS

Herbs have been used since antiquity for treatment and management of various disease conditions. Their ability to cure or prevent diseases is due to storehouses of various phytoconstituents like alkaloids, flavonoids, glycosides, terpenoids, steroids, tannins, phenols, etc. present in them. Pentacyclic triterpenoids have gained greater attention of researchers due to its wide array of pharmacological actions. Thus, a simple and precise HPTLC method has been developed for identification and quantification of one such ubiquitous pentacyclic triterpenoid- ursolic acid in the leaves of Alstonia scholaris. The optimized mobile phase Toluene: Ethylacetate (7:3v/v) gave a better resolution of bands of ursolic acid at 540 nm after derivatizing plate with ASR. The R_f value for ursolic acid was found to be 0.46. The amount of ursolic acid quantified in ethanol, and chloroform extracts of leaves of Alstonia scholaris were 2.34 and 2.38 %w/w, respectively. The developed method was validated in terms of linearity, range, specificity, precision, accuracy, LOD, and LOQ following ICH guidelines. The calibration curve of ursolic acid was linear between 500-900 ng/band with a correlation coefficient (r²) 0.9990. Inter-day and intra-day precision were assessed in terms of %RSD, which was less than 2% in both the cases, thus indicating good precision of the method.  LOD and LOQ were 56.79 and 172.10 ng/band, respectively. Thus, the developed method was found to be quick, simple, specific, precise, and accurate for identification and quantification of ursolic acid in the leaves of Alstonia scholaris and can be used for routine quality control of the plant.