Linear Sesquiterpene Lactones from Anthemis auriculata and Their Antibacterial Activity Revecca Theodori, ² Anastasia Karioti, ² Ana Ranc ˇic ´, ‡ and Helen Skaltsa* ,² Department of Pharmacognosy and Chemistry of Natural Products, School of Pharmacy, UniVersity of Athens, Panepistimiopolis, Zografou, GR-157 71, Athens, Greece, and Department of Plant Physiology, Institute for Biological Research, 29 NoVembar 142, 11 000 Belgrade, Serbia and Montenegro ReceiVed July 20, 2005 Three linear sesquiterpene lactones, anthecotulide (1), hydroxyanthecotulide (2), and acetoxyanthecotulide (3), were isolated from the aerial parts of Anthemis auriculata. Five known flavonoids, taraxa-20(30)en-3-ol (4), and methyl vanillate (5) were also isolated. The structures and the relative configuration of the new compounds 2 and 3 were deduced by spectroscopic methods. The in vitro activity of compounds 1-5 against 10 bacterial species and one fungus was tested using the microdilution method. The genus Anthemis comprises about 130 species predominately distributed around the Mediterranean, but species are also found in Southwest Asia and South Africa, several of which are aromatic, herbal medicines, insecticides, and dyes. 1,2 Anthemis auriculata Boiss. (Asteraceae), an annual herb, is an endemic plant of the South Balkan peninsula and Turkey. 1 As a continuation of our research on Greek Anthemis species, 3 we report here on the chemical profile and the antibacterial activity of secondary metabolites isolated from A. auriculata. A lipophilic extract of the aerial parts of A. auriculata was chromatographed on a silica gel column using cyclohexane contain- ing increasing amounts of EtOAc and acetone. Further chroma- tography of the main fractions afforded three linear sesquiterpene lactones, anthecotulide (1), 4-6 2, and 3, as well as taraxast-20(30)- en-3-ol (4) 4,5 and methyl vanillate (5). 7 From a polar extract of the same plant material five known flavonoids, namely, apigenin, 8 luteolin, 8 pectolinaringenin, 9 eriodictyol, 10 and luteolin 7-O--D- glucopyranoside, 11 were isolated. Compounds 2 and 3 are new natural products. The structures of the compounds were established by means of 1D and 2D NMR, MS, and UV spectroscopic analysis. The main constituents of A. auriculata were flavonoids, while sesquiterpene lactones were isolated in smaller amounts. Compound 2 (Figure 1) showed in its mass spectrum a molecular ion [M + H] + at m/z 265.1440, compatible with the molecular formula C 15 H 20 O 4 . The IR spectrum exhibited absorption bands typical of hydroxyl (3400 cm -1 ) and carbonyl groups, 1770 (Cd O, γ-lactone) and 1715 (CdO, ketone). The 13 C NMR spectrum of compound 2 displayed 15 carbons, which were assigned by HSQC, HMBC, and DEPT 135° experiments to the resonances of three tertiary methyls, one olefinic and two aliphatic methylenes, one oxygenated, one nonoxygenated, and two olefinic methines, three quaternary carbons, and two carbonyls. The presence of an R-methylene-γ-lactone moiety was confirmed by the 13 C NMR signals at δ 170.9 (OCO) and 134.3 and 124.6 ppm (CdCH 2 ). Detailed 2D NMR analysis showed that compound 2 is a linear type sesquiterpene lactone. The 1 H spin systems H-3/H-15a/H-15b and H-3/CH 2 -14 were assigned by COSY. HMBC correlations between H-15a, H-15b/C-1, H-14a, H-14b/C-1, and H-3/C-1 suggested that these protons belong to a lactone ring. Further COSY correlations H-3/H-4 and H-4/H-5, as well as HMBC cross-peaks H-13/C-5, H-13/C-6, H-7/C-6, H-7/C-8, H-9/C-8, H-9/C-12, and H-9/C-11, revealed the structure as shown. The trans-configuration of the 5,6-double bond was confirmed by NOESY experiments. NOE interactions between H-3/H-15b and H-4/H-15b indicated that these protons have the same orientation (Figure 2a). From the above observations, compound 2 was assigned as (+)-4-[(E)-1-hydroxy- 3,7-dimethyl-5-oxo-octa-2,6-dienyl]-3-methylenedihydrofuran-2- one [4-hydroxyanthecotulide]. The (3S,4S)-configuration of 4-hydroxyanthecotulide was con- firmed by Mosher’s ester methodology. 12 The (S)- and (R)-MTPA esters of compound 2 were prepared, and chemical shifts of the H-5, H-3, H-14a, H-15a, and H-15b protons were measured for both derivatives. Using ∆ δ ) δ S - δ R methodology for H-5, a positive value for ∆ δ was observed, and for protons H-3, H-14a, * To whom correspondence should be addressed. Tel and fax: + 30107274593. E-mail: skaltsa@pharm.uoa.gr. ² University of Athens. ‡ Institute for Biological Research. Figure 1. Structure of the isolated sesquiterpene lactones. Figure 2. (a) Relative configuration of compound 2. (b) Chemical shift differences, ∆ δ (δ S - δ R ), for the (S)- and (R)-MTPA esters of compound 2. 662 J. Nat. Prod. 2006, 69, 662-664 10.1021/np058084i CCC: $33.50 © 2006 American Chemical Society and American Society of Pharmacognosy Published on Web 03/11/2006