Two New Antiinflammatory Elemanolides from Centaurea Chilensis
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.,...... Joumalof <.:.- ETHNO-- PHARMACOLOGY Journa1 of Ethnopharmacology 40 (1993) 149-153 Two new antiinflammatory elemanolides from Centaurea chilensis Rosa E. Negretea, Nadine Backhousea, Ivan Cajigala, CarIa Delportea, Bruce K. Cassels*b, Eberhard Breitmaierc, Gert EckhardtC apharmacognosy Laboratory, Faculty o/ Chemical and Pharmaceutical Sciences, hDepartment o/ Chemistry, Faculty o/ Sciences, University o/ Chile, Casilla 653, Santiago 1, Chile cOrganic Chemistry lnstitute, Bonn University, Bonn, Germany (Received 19 March 1992; revision received 8 Ju1y 1993; accepted 8 Ju1y 1993) Abstraet Two previously undescribed elemanolide esters, the 2-methylpropanoate and 2-methyl-2-propenoate of 11,13• dehydromelitensin, were iso1ated in the course of a bioassay-guided fractionation from the aeria1 parts of Centaurea chilensis Hook, et Am" used traditionally to treat 'gout and rheumatism', The mixture ofboth substances exhibits anti• inflammatory activity in the carrageenan-induced paw edema assay. Key words: Antiinflammatory; E1emanolides; Centaurea chilensis; Asteraceae l. Introduction (Negrete et al., 1988b), and 11{3-H-II,13-dihydro• desacyl-cynaropicrin 8{3-D-glucoside (Negrete et The genus· Centaurea (Asteraceae, tribe al., 1988a). Cynareae) is a well-known source of sesquiterpene No elemanolides have been reported previously lactones. As part of a study of species growing in from C. chilensis Hook. et Am., but other Cen• Chile, we have now obtained two new elemanolide taurea species have afforded melitensin, dehydro• esters from C. chilensis Hook. et Am. Earlier melitensin, and several ester s of these sesquiter• chemical studies on this plant had afforded {3• pene lactones: melitensin was obtained originally sitosterol and stigmasterol (Negrete et al., 1984), from C. melitensis L. (González et al., 1971) and the flavonoids apigenin 7-0-glucoside, chrysoeriol later from C. aspera varo stenophyl/a (Dufour) 7-0-glucoside, chrysoeriol, luteolin, kaempferol, Nyman (Picher et al., 1984); its structure and quercetin, and hispidulin (Negrete et al., 1988a), as stereochemistry have been confirmed by X-ray well as the guaianolides dehydrocostus lactone analysis (Tortajada et al., 1988); this elemanolide and 8a-hydroxydehydrocostus lactone (Negrete has al so been synthesized via Cope rearrangement et al., 1984), 8a-acetoxydehydrocostus lactone of cnicin to 11,13-dehydro-melitensin (González et al., 1974a), which was first isolated from C. pul/ata • Corresponding author. 0378-8741/93/$06.00 © 1993 Elsevier Scientific Pub1ishers Ireland Ltd. AlI rights reserved. SSDl 0378-8741(93)01055-Q 150 R.E. Negrete et al. / J. Ethnopharmacol. 40 (1993) 149-153 L. (González et al., 1974b); C. melitensis afforded ing, respectively 50, 300, and 400 g of residue after melitensin and 11, 13-dehydromelitensin ¡3-hyd• removing the solvents in vacuo. All 3 extracts were roxybutyrates (González et al., 1975); C. arbuti• administered to guinea pigs (see below) to monitor folia Svent. gave 11,13-dehydromelitensin iso• potential antiinflammatory activities. The chloro• valerate (isoarbutifolin) (González et al., 1981), C. form extract proved to be the most active in the phrygia L. yielded the 8a-(5-hydroxy)-angeloyl carageenan-induced paw edema assay, and was ester (Tsankova and Ognyanov, 1985) and C. therefore chromatographed on a silica gel G cineraria L. ssp. umbrosa (Lacaita) Pign. (syn. C. column, eluting with petroleum ether-CH2Cl2 ucriae Lacaita ssp. ucriae) gave the 2-methyl-3,4• mixtures of increasing polarity as shown in dihydroxybutyrate (Bruno and Herz, 1988). Table 1. C. chilensis has been used in folk medicine for 11,13-Dehydromelitensin 2-methylpropanoate 'gout and rheumatism', although the treatment (1) and 2-methyl-2-propenoate (I1) from fractions advocated in recent years only involves external B, C and D (see Table 1) were purified by further use (Muñoz et al., 1981). Several guaianolides are chromatography on silica gel G with petroleum known to possess anti-inflammatory activity, ether-CH2Cl2 (3:2) increasing the CH2CI2 propor• which has been related to the presence of the tion up to 100%. The major constituent obtained a-methylene-')'-Iactone moiety (Hall et al., 1979, in this way (300 mg) seemed to be homogeneous 1980), and this suggested a possible relationship on TLC (silica gel; light petrol-EtOAc 7:3; between the chemistry of this plant and its tradi• RF = 0.35) and had the appearance of a slightly tional use. In this paper we describe the bioassay• greenish lacquer, insoluble 'in petroleum ether, guided isolation, structure elucidation, and anti• somewhat soluble in CHCI3 and in EtOAc, very inflammatory activity of a mixture of two soluble in MeOH, giving a purple Lieber• previously undescribed elemanolide esters: 11,13• mann-Burchard reaction. In CHCI3, the mixture dehydromelitensin 2-methylpropanoate (1) and 2• apparently polymerizes, giving a white precipitate methyl-2-propenoate (I1), from C. chilensis. which cannot be redissolved. EIMS m/z data were: (relative abundance) 264.1366 (0.26%; calculated 2. Materials and methods ClsH2004, 264.1361), 246.1265 (7.65%; calculated ClsHIS03, 246.1256), 228.1149 (8.43%; calculated Aerial parts of Centaurea chilensis were col• C'SH'60b 228.1149). Complete lH_ and l3C_ lected in the flowering season (December) in Las NMR assignments are shown in Table 2. Tacas, La Serena (near sea level, 300S latitude), Anti-inflammatory activity was tested against Chile. A voucher specimen (L. Arancibia 16078) is carrageenan-induced edema in the guinea pig on deposit in the Botany Laboratory of the Facul• hindpaw. A-Carrageenan (1% in saline, 0.1 mI) was ty of Chemical and Pharmaceutical Sciences, Uni• injected into the left plantar aponeurosis of groups versity of Chile, Santiago (SQF). Melting points are uncorrected and were de ter• mined on a Kofler hot stage. IR spectra (KBr Table 1 discs) were recorded utilizing a Leitz IIIG infrared Fraetionation of the ehloroform extraet of Centaurea chilensis spectrometer. NMR spectra in CDCI3 with TMS as internal standard were recorded at 400 MHz Fraetion 25:75Solvent75:2550:50100%1000;',100'10petroleumCH2Cl2petroleumMeOHeompositionether/CH2Cl2ethernone+Anti-+++++ B ('H) and 100.5 MHzCFED (l3C) on a Bruker WM-400 inf1ammatoryaetivity A spectrometer, using commercially available pulse programs for DEPT and 2D spectra ('H_1H and 'H-l3C COSY, COLOC). The HREIMS was recorded on a Kratos MS-30 instrument. The air-dried, ground plant material (4.0 kg), was extracted successively (at room temperature) with petroleum ether, CHCI3, and MeOH, yield- R.E. Negrete et al. / J. Ethnopharmacol. 40 (1993) /49-153 151 of 10-15 guinea pigs of both sexes (Pirbright) by the paw, immersed up to a mark made weighing 200-300 g, fasted for 24 h, with water ad previously just above the ankle, before injecting libitum, and kept at 18-20°C. The volume of the carrageenan, and 3 h later. Drugs or vehicle (pro• injected paw was determined with a plethysmo• pyleneglycol) were administered orally 1 h before meter by measuring the volume of water displaced injection. The percent inhibition of edema· was Table 2 l3c and IH chemical shifts from the 100 MHz 13C and 400 MHz IH NMR spectra - assignments based on the 100/400 MHz FI• decoupled CH correlation CHCCCH147.87113.28112.97145.59613C46.1570.3751.8180.03173.72139.13119.59171.4053.1342.9535.1966.6819.1119.3119.03 C Atoms CH3CH2CH2CHn147.85113.2546.05167.67139.08137.43126.95119.6380.0070.9651.7453.0342.9218.4119.26 61H 3JHH coupling 153a 17a13b13a12IOb19b14a14b18a11b16b18b19a17blOa49b7b3b6b9a8b8a27a5a5blb1a (ppm) b for compound 11. (ppm) (H) a11a6afor compound16al. 6.03; 5.58; 5.54 1.96; 6.14; 5.69 5.83 2.49; 5.00; 4.90; 1.71; 1.16 5.83 2.51; 5.00; 4.90; 1.68; 1.19 a2.49; 4.05; 3.94; 5.40 b 2.51; 4.05; 3.94; 5.40 2.90; 4.46; 6.03 3.05; 4.48; 6.03 6.14; 1.96 6./4; 5.69 1.96 6.03; 5.54 3.05 6.03; 5.58 2.90 5.40; 5.00 4.05; 3.94; 2.49 5.40; 5.00 4.05; 3.94; 2.51 a5.00; 4.90 b5.02; 5.02 4.46 2.90; 2.49 4.48 3.05; 2.51 5.27 3.05; 1.88; 1.68 5.22 2.90; 1.88; 1.71 4.05; 3.94 5.40; 5.00 2.90 5.22; 6.03; 5.58 3.05 4.48; 5.27; 6.03; 5.54 2.49 4.46; 1.16; 1.88; 5.00; 5.40 2.51 4.48; 1.19; 1.88; 5.00; 5.40 1.88; 1.71 1.16 1.88; 1.68 1.19 2.49; 5.00; 4.90; 1.16; 1.88; 1.71 2.51; 5.02; 5.02; 1.19; 1.88; 1.68 2.61 1.16 2.49; -1.71 1.19 2.51; 1.68 1.17 1.19 1.19 1.17 1.96 6.14; 5.69 152 R.E. Negrete et al. /1. Ethnopharmacol. 40 (1993) 149-153 calculated for each group with respect to its made on the basis of DEPT, homo- and hetero• vehicle-treated control group, and analyzed nuclear COSY, and COLOC spectra, and are sum• statistically using Student's t-test. marized in Table 2. 3. Results 4. Discussion and conclusion A 20% infusion of the aerial parts of this plant, The fact that germacradienolides undergo facile similar to that recommended by folk healers, thermal isomerization to elemadienolides has led administered orally to guinea pigs, exhibited weak to the suggestion that some of these latter products (20%) but significant (P < 0.05) anti-inflamma• may be artefacts (Barrero et al., 1989). The drying tory activity at 4 ml!kg in a carrageenan-induced of the plant material, the extraction and all purifi• paw edema assay (Winter et al., 1962).The residue cation steps relating to compounds 1 and n, of the methanol extract of C.