Development and Validation of HPTLC Technique For Simultaneous Estimation of Bioactives of Moringa Oleifera Leaves

Abstract

Moringa Oleifera belonging to family Moringaceae is a significant versatile tree having bunch of nutritional and therapeutic values. MO is also known as the Miracle Tree because many of its components are useful. Leaves contain many phenolics, flavonoids, glucosinolates, alkaloids and isothiocyanates. Leaves are widely used in many market formulations but very few analytical techniques have been reported for characterization of bio actives in crude extract which can be used for analysis of formulation also. Determination of various bio actives in MO leaves was done by developing a simple and precise HPTLC method. In which chlorogenic acid, rutin, quercetin and gallic acid were determined simultaneously and its analysis was done by using silica gel 60F254 TLC plates. Scanning of plates was done using CAMAG TLC scanner 3 using UV densiometrically at wavelength of 254nm. After preliminary screening using mobile system with varied proportion of polar and non-polar solvents, toluene, ethyl acetate, methanol and formic acid in the ratio of 7:6:2:1 was finalized to be used as a mobile phase. Different Rf values observed at 0.72 for quercetin, 0.09 for rutin, 0.55 for gallic acid and 0.15 for chlorogenic acid. The developed method was validated for linearity, accuracy, precision, range, LOD and LOQ. Linearity range was found to be 100 to 600 nanogram per band with correlation coefficient (R2) of 0.9931, 0.9963, 0.9956 and 0.9954 for quercetin, rutin, gallic acid and chlorogenic acid respectively, high accuracy (97-105%), intra- and interday precision (%RSD < 2), with LOD 58.56, 36.05, 27.59 and 12.47 ng/band and LOQ 177.48, 109.26, 83.62 and 37.78 ng/band for quercetin, rutin, gallic acid and chlorogenic acid respectively