Simultaneous determination of rutin, isoquercetin, and quercetin flavonoids in Nelumbo nucifera by high‑performance liquid chromatography method

Objective: The present study was investigated to provide a documentary evidence for the determination of rutin, isoquercetin, and quercetin flavonoids from the flora of Nelumbo nucifera by reversed‑phase high‑performance liquid chromatography (RP‑HPLC). Materials and Methods: RP‑HPLC analysis was performed by gradient elution with a low-pressure gradient using 0.5% acetic acid: acetonitrile as a mobile phase with a flow rate of 1.0 ml/min. The separation was done at 26°C using a Kinetex XB-C18 column as stationary phase and the detection wavelength at 356 nm. The proposed method was validated as per International Conference on Harmonisation guidelines with respect to specificity, linearity, precision, accuracy, limit of detection (LOD), and limit of quantification (LOQ). Results: The validated results were within the acceptable limits. In specificity, the retention time of rutin, isoquercetin, and quercetin peak in the sample was matched with the reference standard peak and showed good resolution. An excellent linearity was obtained with correlation coefficient (r) higher than 0.999. In precision, the repeatability and intermediate showed <1.0% of % relative standard deviation of peak area percentage indicating high precision and accurate. The recovery rate for rutin, isoquercetin, and quercetin was between 99.85%–101.37%, 101.90%–103.24%, and 101.74%–106.73%, respectively. The lower LOD and LOQ of rutin, isoquercetin, and quercetin enable the detection and quantitation of these flavonoids in N. nucifera at low concentrations. Conclusion: The developed analytical method is convenient for the determination of flavonoids content in herbal drugs.