This work has been digitalized and published in 2013 by Verlag Zeitschrift für Naturforschung in cooperation with the Max Planck Society for the Advancement of Science under a Creative Commons Attribution 4.0 International License. Dieses Werk wurde im Jahr 2013 vom Verlag Zeitschrift für Naturforschung in Zusammenarbeit mit der Max-Planck-Gesellschaft zur Förderung der Wissenschaften e.V. digitalisiert und unter folgender Lizenz veröffentlicht: Creative Commons Namensnennung 4.0 Lizenz. 5,5',6,6'-Tetrahydroxy-3,3'-biindoIyl from Beetroot {Beta vulgaris) Peel Extract Tytti Kujala*, Karel Klika, Vladimir Ovcharenko, Jyrki Loponen, Maarit Vienola and Kalevi Pihlaja Department of Chemistry, Vatselankatu 2, FIN-20014 University of Turku, Finland Fax: +3582-333 6700. E-mail: tytti.kujala@utu.fi * Author for correspondence and reprint requests Z. Naturforsch. 56c, 714-718 (2001); received May 2/May 31, 2001 Beta vulgaris, Beetroot, Nitrogen Heterocycles, Biindolyls A compound of unusual structure was isolated extract and identified as 5,5',6,6'-tetrahydroxy-3,3' NMR and MS studies. Introduction Recently, plant material extracts containing phe nolic compounds have been screened as potential new sources of natural antioxidants. Beet extracts, especially the extracts from the peel, have shown strong antioxidant activity in these evaluations re sulting in increased attention to the natural pro ducts present in beetroot (Vinson et al., 1998; Kähkönen et al., 1999). As part of our ongoing investigation into the distribution and properties of known phenolics in red beetroot peel (Kähkönen et al., 1999; Kujala et al., 2000), a novel compound, 5,5',6,6'-tetrahydroxy-3,3'-biindolyl (1, Fig. 1), was isolated. It was shown to be a phenolic compound by a modified Folin-Ciocalteu phenol test (Nurmi et al., 1996) and was thus of prime interest to our work despite its highly unstable na ture when isolated. Materials and Methods Extraction and isolation Red beetroots (Beta vulgaris, cultivar “Rubia“) were washed, hand-peeled, and the collected peels cut into small pieces and stored at -25° C until lyophilisation. After the lyophilised plant material was reduced to powder by mortar and pestle, 96 g of the peel was extracted in 8.0 g portions by ho mogenising (Ultra-Turrax T25, Janke & Kunkel, IKA-Labortechnik, Germany) each portion twice for four min with 160 ml of 80% aq. MeOH. Sam ples were centrifuged (10 min, 1500xg) and the combined extracts evaporated to syrup under re from red beetroot (Beta vulgaris) peel -biindolyl based on the combination of duced pressure. The syrups were stirred with MeCN (3 x 50 ml for 30 min) in a planar mixer (Promax 2020, Heidolp, Germany). The combined MeCN extracts of all syrups were taken to dryness under reduced pressure and the residue was ex tracted with EtOAc. The EtOAc was evaporated to dryness under reduced pressure and the water- soluble fraction of the extract was removed to water. A small portion of the sample was ace- tylated with the remainder subjected to semiprep arative purification using a LiChrocart column {Lichrospher 100 RP-18, 250 x 10 mm I. D., 10 /um, Merck, Darmstadt, Germany; elution with MeCN (A) and HCOOH - water (B) (1:99, v/v), 0 - 10 min, 95% B followed by 10-25 min, 5-16% A in B (linear gradient) with detection at 305 nm}. From the combined fractions, a distinct and attrac tive yellow/orange isolate (as an aqueous solution) was obtained which quickly discoloured to a dark brown solution (several hours) followed by the formation of black precipitate (one to several days). A syrup was also prepared as mentioned earlier from 4.0 g of dried peel and dissolved in water (6.0 ml) for HPLC-MS analysis. NMR NMR spectra were acquired on a JEO L Alpha 500 NMR spectrometer operating at 500.16 MHz for :H and 125.78 MHz for 13 C. Spectra were re corded at 25° C in D 2 0 ; both XH and 13C spectra were referenced internally to TMS (0 ppm for both). ID proton and carbon spectra were ac- 0939-5075/2001/0900-0714 $ 06.00 © 2001 Verlag der Zeitschrift für Naturforschung. Tübingen •www.znaturforsch.com • D