Development and validation of a HPLC method for the quantification of three flavonoids in a crude extract of Dimorphandra gardneriana

ABSTRACT

A method for separation and quantification of three flavonoids by reverse-phase high performance liquid chromatography (HPLC) was developed and validated. Flavonoids present in a crude methanolic extract of the inner bark of Dimorphandra gardneriana Tul., Fabaceae, were analyzed. Rutin, isoquercitrin and quercetin were used as calibration standards. The analysis was performed using a Thermo Scientific Hypersil C18 column (250 x 4.0 mm i.d., 5 μm particle size), as stationary phase, with a flow rate of 0.8 mL/min and detection at a wavelength of 356 nm. The proposed method was validated by resolution RE No. 899/2003 of the National Health Surveillance Agency. In this study, an excellent linearity was obtained with r higher than 0.99. Besides, the chromatographic peaks showed good resolution. With other validation data, including precision, specificity, accuracy and robustness, this method demonstrated good reliability and sensitivity, and can be conveniently used for the quantification of rutin, isoquercitrin and quercetin in crude methanolic extract of D. gardneriana pods. Furthermore, there are the advantages of easy sample preparation and short time between each injection.