Development and validation of a high performance liquid chromatography-diode array detection (HPLC-DAD) method for the quantification of rutin and isoquercetin in Morus nigra L. (Moraceae)

Abstract

Morus nigra L. is widely used by the population of the São Francisco Valley (Brazil) for the treatment of several diseases such as hypertension, diabetes, arthritis etc. The objective of this study was to develop and validate an analytical method that is capable of identifing and quantifing flavonoids in the ethanolic crude extract (Mn-EtOH) and its ethyl acetate phase (Mn-AcOEt) obtained from M. nigra leaves using high performance liquid chromatography (HPLC). The plant material was collected and processed to obtain Mn-EtOH, which was subjected to a process of liquid/liquid partition, from which the AcOEt phase was obtained. The method was developed using an reverse phase (RP) C-18 column (250 × 4.6 mm, 5 μm) as the stationary phase. A mixture of acidified water (0.01% trifluoroacetic acid): acetonitrile was used as the mobile phase in a gradient system with a flow rate of 0.8 mL/min at a wavelength of 340 nm for the detection and determination. The method developed was considered to be specific: linear (R2 > 0.99), selective, accurate, and robust.   Key words: Flavonoids, isoquercetin, rutin, morus.