FINGERPRINT ANALYSIS FOR VALIDATION AND SIMULTANEOUS QUANTIFICATION OF QUERCETIN AND KAEMPFEROL IN MORINGA OLEIFERA BY RP-HPLC AND HPTLC

To establish authenticity and integrity for usage of herbal drugs quantitative analysis by sensitive analytical instrument is highly desirable. Quercetin and Kaempferol from whole plant of Moringa oleifera was chosen for simultaneous quantification by RP-HPLC and HPTLC method due to their pharmacological activities. HPTLC method of quercetin and kaempferol was developed on HPTLC plates using Toluene: Ethyl acetate: Chloroform: Formic acid (6:2:5:1.5) as mobile phase. Densitometric evaluation of flavonoids was performed at λ = 240 nm in absorbance mode. Furthermore, RP-HPLC method was developed using mobile phase containing acetonitrile and 0.1 M phosphate buffer (pH 2.5 adjusted by orthophosphoric acid) using C₁₈ column in gradient mode. Chromatographic methods were validated as per ICH guidelines. Excellent linear relationships over the concentration range (1 – 40 µg/ml) were observed with the r² value higher than 0.999 for both the phyto-constituents by RP-HPLC. Limit of detection and quantification were found to be in range of 0.2 – 0.5 µg/ml with percent recovery of 98 – 101%; while r² value of quercetin and kaempferol were found to be 0.9764 and 0.9823 by HPTLC. The average recovery values for fruits of Moringa oleifera were within range of 98 – 101% by HPTLC. In conclusion, RP-HPLC method was found to be more precise and sensitive as compared to HPTLC for obtaining acceptable resolution between quercetin and kaempferol. Therefore, HPLC method can be used for the routine analysis of the quality of herbal extracts and formulations of Moringa oleifera.