DEVELOPMENT OF A VALIDATED HPTLC METHOD FOR QUANTIFICATION OF ESCULIN IN DIFFERENT FRACTIONS OF CICHORIUM INTYBUS LEAF EXTRACT

Objective: To develop a simple, rapid, sensitive and validated High performance thin layer chromatography method (HPTLC) for identification and quantification of esculin (6, 7-dihydroxycoumarin 6-glucoside), a glycosidic coumarin in different fractions of Cichorium intybus leaves extract prepared using soxhlet extraction . Method: The separation was achieved on silica gel 60 F254 HPTLC plates using ethyl acetate: methanol: water: glacial acetic acid (20:2:1:4, v/v/v/v) as the mobile phase. Densitometric analysis of esculin was carried out using reflectance/fluorescence mode at 343 nm. Results: Densitometric analysis of different fractions of leaf extract gave compact spots for esculin (Rf 0.36). The linear regression analysis of data for the calibration plots showed a good linear relationship with r=0.9989. The average recovery of esculin was 98.34%, indicating the good reproducibility. The amount of esculin was found to be equal in both water (94.9 µg g-1 dw) and ethanol (93.4µg g-1dw) fraction. Statistical analysis of the data showed that the method is reproducible. Conclusion: The method was found to be simple and convenient for rapid screening and quantification of esculin in the biological samples of C. intybus.