Development and application of high-performance liquid chromatography for the study of two new oxyprenylated anthraquinones produced by Rhamnus species.

Rhamnus spp. is known to contain biologically active anthraquinone secondary metabolites but the presence of oxyprenylated ones is not reported. To this aim, a new simple, and accurate analytical method was developed to reveal chemical fingerprint of these analytes in plant extracts. Plant samples were analysed after extraction with n-hexane (first step) and methanol (second step) using a C(18) column with a mobile phase composed of 35% of water:65% of methanol (both with 1% formic acid, v/v) at 0.7 mL min⁻¹ flow rate in gradient elution mode. For quantitative analyses, selective detection was performed at 435 nm. The limit of quantification (LOQ) was 0.5 μM, with the only exception of 3-geranyloxyemodin for which the LOQ value was 5.0 μM, and external matrix-matched standard curves showed linearity up to 125 μM. The within- and between-batch precision (RSD%) values ranged from 0.2% to 12.9% while within- and between-batch trueness (bias%) values ranged from 12.2% to 12.7%. The method was applied to evaluate for the first time the presence and the quantities of oxyprenylated anthraquinones in Rhamnus spp. barks as well as the anthraquinone profile of Rhamnus pumila Turra. The proposed method could be directly applied to the selective quantification of these analytes in natural sources.
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