Abstract
Anecdotal evidence from Danish folk medicine suggests that the tea of Calluna vulgaris Hull. has a nerve calming effect [1]. Unpublished results from our department show that the methanol extract of C. vulgaris has an inhibitory effect on monoamine oxidase A (MAO-A). For bioassay-guided isolation, dried, ground aerial parts of C. vulgaris (100g) were defatted twice with 1000ml heptane. The plant material was extracted twice with 1000ml methanol. The combined methanol extracts were evaporated to dryness, re-dissolved in 1000ml 80:20 methanol: water and partitioned three times against 300ml heptane. Methanol was removed from the methanol: water fraction by evaporation and the volume was made up with water. The resulting water fraction was partitioned three times against 300ml ethyl acetate. The three fractions (heptane, ethyl acetate and water) were tested for MAO-A inhibitory activity [2]. The active ethyl acetate fraction was fractionated on a VLC column (240g silica gel 60), eluted with heptane, mixures of heptane: ethyl acetate: methanol, mixtures of dichloromethane: methanol and methanol. A total of 34 fractions were spotted on a TLC plate and fractions showing similar patterns were combined. The fractions were tested in the MAO-A assay. The most active fraction was investigated by HPLC (C18) employing a gradient from 0 to 100% acetonitrile over 30min, revealing one major peak in the chromatogram. This peak was active in the MAO-A assay. The compound was re-crystallised from acetone yielding 14.3mg pure compound which was investigated by 1H-NMR. By comparison with 1H-NMR literature data [3, 4] the compound was identified as quercetin. LC-MS analysis revealed a molecular weight of 302.1g/mol confirming the identification. The IC50-value of quercetin was 18±2µM in the MAO-A assay (clorgylin: 0.2±0.02µM).
Published Version
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