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Terpenoids and phenolics from Inula ensifolia
Biochemical Systematics and Ecology 38 (2010) 232–235
Contents lists available at ScienceDirect
Biochemical Systematics and Ecology journal homepage: www.elsevier.com/locate/biochemsyseco
Terpenoids and phenolics from Inula ensifolia Anna Stojakowska a, *, Janusz Malarz a, Szymon Zubek b, Katarzyna Turnau c, Wanda Kisiel a a b c
´w, Poland Department of Phytochemistry, Institute of Pharmacology, Polish Academy of Sciences, 12 Sm˛etna Street, 31-343 Krako ´w, Poland Mycology Unit, Institute of Botany, Jagiellonian University, 46 Lubicz Street, 31-512 Krako ´w, Poland Institute of Environmental Sciences, Faculty of Biology and Earth Sciences, Jagiellonian University, 7 Gronostajowa Street, 30-387 Krako
a r t i c l e i n f o Article history: Received 18 May 2009 Accepted 12 December 2009 Keywords: Inula ensifolia Asteraceae Thymol derivatives Norisoprenoids Flavonoids Dicaffeoylquinic acids
1. Subject and source Inula ensifolia L. (Asteraceae, Inuleae) is a herbaceous perennial, about 45 cm high, with grey-green, lance-shaped leaves and anthodia composed of bright-yellow, narrow ray florets surrounding darker disc florets. Whole plants of I. ensifolia were collected from xerothermic grasslands nearby Kalina-Lisiniec (Miecho´w Upland, Poland; coordinates: 50 330 9400 N, 20170 9700 E), in June 2008, and identified by Teresa Anielska M.Sc. from the Institute of Environmental Sciences, Jagiellonian University in Krako´w. A voucher specimen (08/08) has been deposited at the Garden of Medicinal Plants, Institute of Pharmacology, Polish Academy of Sciences, Krako´w.
2. Previous work The species lacks detailed phytochemical investigation. Literature data are sparse. Wollenweber et al. (1997) examined five Inula species (Inula britannica L., Inula germanica L., Inula salicina L., Inula helenium L., I. ensifolia L.) with respect to exudate flavonoid production. However, no exudate flavonoids were found in I. ensifolia aerial parts. Thin-layer chromatography of methanolic extracts from disc and ray flowers of I. ensifolia revealed the presence of phenolic acids (caffeic and chlorogenic) and flavonoids (apigenin and hyperin) (Pe´ter and Do´sa, 2002). Secondary metabolites of plants from the genus Inula, including monoterpenoids, sesquiterpenoids and flavonoids, have been reviewed by Konovalov and Khubieva (1997) and recently by Zhao et al. (2006). Although the plant has no any documented medicinal use, an antiproliferative activity of methanolic extract from I. ensifolia against human cancer cell lines in vitro has been reported (Re´thy et al., 2007).
* Corresponding author. Tel.: þ48 12 6623217; fax: þ48 12 6374500. E-mail address: [email protected] (A. Stojakowska). 0305-1978/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.bse.2009.12.011
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3. Present study The present report deals with the isolation of five thymol derivatives (1–5) from the roots of I. ensifolia, as well as atocopherol (6), two norisoprenoids (7, 8), three quercetin derivatives (9–11) and four caffeoylquinic acids (12–15) from its aerial parts (Fig. 1). The air-dried roots of I. ensifolia (10.8 g) were powdered and exhaustively extracted with CHCl3 at room temperature with shaking. The obtained extract was concentrated under reduced pressure to give 0.162 g of an oily residue. This residue, after fractionation by preparative TLC (Merck, Art. 1.05553; hexane-EtOAc, 9:1) followed by semipreparative HPLC on a Delta-Pak C-18 column (particle size 15 mm, 25 100 mm) coupled to a dual wavelength UV/vis detector operating at 205 and 270 nm, using MeOH–H2O mixture (13:7) at a flow rate of 6 ml min1, yielded: 1 (21.9 mg), 2 (2.1 mg), a mixture (2.8 mg) of 3 and 4 (by 1 H NMR), and a fraction (1.5 mg) containing 5 as the main component (by 1H NMR and ESIMS). The aerial parts (59.6 g) of I. ensifolia were powdered and extracted successively with CHCl3, followed by pure MeOH at room temperature with shaking and the solvents were evaporated under reduced pressure which furnished crude CHCl3 (2.98 g) and MeOH (5.50 g) extracts. The crude CHCl3 extract was fractionated by column chromatography on a silica gel (Merck, Art 7754) using gradients of EtOAc in hexane as a solvent system. Elution with hexane-EtOAc 9:1 followed by preparative TLC in the solvent system of the same composition led to the isolation of 6 (3.3 mg). From the more polar fractions (hexane-EtOAc, 1:1) a mixture (2 mg) of 7 and 8 (by 1H NMR) was isolated. The crude MeOH extract was subjected to column chromatography on Sephadex LH-20 (Pharmacia Biotech) eluted with MeOH/H2O. The eluted fractions were monitored by analytical RP-HPLC using an Agilent 1200 Series HPLC system (Agilent Technologies, USA) equipped with a Rheodyne manual sample injector, quaternary pump, degasser, column oven and a diode array detector. Chromatographic separations were carried out at 25 C, on a Zorbax Eclipse XDB-C18 column (4.6 150 mm, 5 mm particle size; Agilent Technologies, USA) with a mobile phase consisting of H2O/HCOOH/CH3COOH 99/0.9/0.1 (solvent A) and MeCN/MeOH/HCOOH/CH3COOH 89/10/0.9/0.1 (solvent B), at a flow rate of 1 ml/min, using 10 ml injections. The gradient elution conditions described by Spitaler et al. (2006) were used. This procedure yielded: 12 (100 mg), 13 (300 mg), a mixture (160 mg) of 9 and 10 (by 1H NMR and co-HPLC with authentic samples), a mixture (304 mg) of 13, 14 and 15 (by 1H NMR), and 11 (60 mg), in that order. The mixtures were not further separated, as the 1H NMR signals could be readily assigned to the respective compounds. Optical rotations were determined with a PolAAr31 automatic polarimeter (Optical Activity LTD). 1H and 13C NMR spectra were measured in CDCl3 or in CD3OD (phenolic compounds) on a Varian Mercury-VX 300 spectrophotometer operating at 300.08 MHz (1H) and 75.46 MHz (13C). COSY experiments were carried out using the same instrument. Chemical shifts (d in ppm) were referenced to TMS. Mass spectra (ESIMS) were recorded on a Bruker Esquire 3000 mass spectrometer. The isolated compounds were identified based on their physical ([a]D, whenever possible) and spectroscopic data, including NMR and MS, and their comparison with literature data for: 7-isobutyroyloxythymol methyl ether (1, Shtacher and Kasman, 1971; Anthonsen and Kjøsen, 1971), 10-isobutyroyloxy-8,9-epoxythymol isobutyrate (2), 10-(2-methylbutyroyloxy)-8,9-epoxythymol isobutyrate (3), 10-isovaleroyloxy-8,9-epoxythymol isobutyrate (4) (Bohlmann et al., 1969; Zee et al., 1998), 7,10diisobutyroyloxy-8,9-epoxythymol isobutyrate (5, Bohlmann and Zdero, 1977), a-tocopherol (6, Baker and Myers, 1991; Malik et al., 1997), megastigmane aglycone - 3b-hydroxy-5b,6b-epoxy-b-ionone (7, Cha´vez et al., 1997; Xian et al., 2006), loliolide (8, Kisiel, 1992; El Hattab et al., 2008), quercetin-3-O-b-glucopyranoside (isoquercitrin, 9, Shoeb et al., 2007), quercetin-3-O-bgalactopyranoside (hyperin, 10, An et al., 2008), quercetin-3-O-b-(600 -caffeoylgalactopyranoside) (11, Shigematsu et al., 1982), chlorogenic acid (12, Pauli et al., 1999), 1,5-, 3,4- and 3,5-dicaffeoylquinic acids (13–15, Merfort, 1992; Basnet et al., 1996; Islam et al., 2002). 4. Chemotaxonomic significance A total of 15 terpenoid and phenolic compounds have been isolated from the roots and aerial parts of I. ensifolia. Except for compounds 10 and 12, the other isolated constituents are reported for the first time from I. ensifolia. Quinic acid derivatives (12–15) appeared to be major secondary metabolites (over 1% yield) of the aerial parts. The paraphyletic genus Inula L., heterogenous with respect to morphology and chromosome numbers, comprises ca. 100 species native to Eurasia and Africa (Bremer, 1994). Within the genus, a group of resiniferous taxa, including I. helenium – a well known medicinal plant, has been distinguished by Anderberg (1991). Roots of the resiniferous species usually contain essential oils with the eudesmane-type sesquiterpene lactones alantolactone and/or isoalantolactone as major constituents. Separate position of these species has been confirmed by ITS sequence analysis (Eldena¨s et al., 1998; Francisco-Ortega et al., 2001). Within Inula species lacking resin canals, a classification based on morphological data has been proposed (Anderberg, 1991). According to this proposal, I. ensifolia has been included in the I. salicina group together with I. germanica, Inula hirta, Inula helvetica and Inula viscidula. Of these, only I. salicina and I. germanica have been phytochemically investigated (Anthonsen and Kjøsen, 1971; Konovalova et al., 1974; Bohlmann et al., 1978, 1985). Both plant species contained sesquiterpene lactones in their aerial parts (I. germanica – germacranolides, I. salicina – eudesmanolides). The compounds were absent from the plant material under study. This absence of sesquiterpene lactones situates I. ensifolia closer to some representatives of the postulated Inula decurrens group, i.e. Inula bifrons and Inula conyza. Moreover, I. germanica is rich in exudate flavonoids, derivatives of luteolin, scutellarein and quercetagenin, whereas only one derivative of luteolin was found in I. salicina exudate and no flavonoids in an exudate of I. ensifolia were found (Wollenweber et al., 1996). It seems that, neither exudate flavonoids nor esterified thymol derivatives are helpful chemotaxonomical markers within Inula. The flavonoids due
234
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CH2R1
7
O
1
O
O
2
6 5
O
3
4 8
OMe
9
O
1
CH3
CH3
O
CH3
10
R1 H H H OiBu
2 3 4 5
H3C
OR2
R2 iBu MeBu iVal iBu
CH3
CH3
CH3 HO CH3
6
O
12
11 2 3
HO
1 4
7
6
O
5
9
8
11
HO 3
10
2
4 1
O
5 6
O
8 7
13
10
9
7
8 OH OH
HOOC OR
OH
9 10 11
OR4
6
O
HO
1
1
2
4
5
OR3
3
OR1
OR2
R2 H H Caff Caff
R3 H H Caff H
O
R - Glc R - Gal R - (6''-caffeoyl)Gal
12 13 14 15
R1 H Caff H H
R4 Caff Caff H Caff
Fig. 1. Structures of thymol derivatives (1–5), a-tocopherol (6), norisoprenoids (7, 8), flavonoids (9–11) and quinic acid derivatives (12–15), isolated from Inula ensifolia (iBu, isobutyroyl; MeBu, 2-methylbutyroyl; iVal, isovaleroyl; Caff, caffeoyl; Glc, b-glucopyranosyl; Gal, b-galactopyranosyl).
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to limited data available, and the thymol derivatives due to their common occurrence in members of Inuleae and in plants of Helenieae and Eupatorieae tribes. Further phytochemical studies of Inula sp. are needed to support classification efforts, which nowadays are based mainly on morphological traits. References An, R.-B., Sohn, D.-H., Jeong, G.-S., Kim, Y.-C., 2008. Arch. Pharm. Res. 31, 594. Anderberg, A.A., 1991. Plant Syst. Evol. 176, 75. Anthonsen, T., Kjøsen, B., 1971. Acta Chem. Scand. 25, 390. Baker, J.K., Myers, C.W., 1991. Pharm. Res. 8, 763. Basnet, P., Matsushige, K., Hase, K., Kadota, S., Namba, T., 1996. Biol. Pharm. Bull. 19, 1479. Bohlmann, F., Niedballa, U., Schulz, J., 1969. Chem. Ber. 102, 864. Bohlmann, F., Zdero, C., 1977. Phytochemistry 16, 1243. Bohlmann, F., Mahanta, P.K., Jakupovic, J., Rastogi, R.C., Natu, A.A., 1978. Phytochemistry 17, 1165. Bohlmann, F., Baruah, R.N., Jakupovic, J., 1985. Planta Med. 51, 261. Bremer, K., 1994. Asteraceae. Cladistics & Classification. Timber Press, Portland. Cha´vez, J.P., Dos Santos, I.D., Cruz, F.G., David, J.M., Yang, S.-W., Cordell, G.A., 1997. Phytochemistry 46, 967. Eldena¨s, P., Anderberg, A.A., Ka¨llersjo¨, M., 1998. Plant Syst. Evol. 210, 159. El Hattab, M., Culioli, G., Valls, R., Richou, M., Piovetti, L., 2008. Biochem. Syst. Ecol. 36, 447. Francisco-Ortega, J., Park, S.-J., Santos-Guerra, A., Benabid, A., Jansen, R.K., 2001. Biol. J. Linn. Soc. 72, 77. Islam, M.S., Yoshimoto, M., Yahara, S., Okuno, S., Ishiguro, K., Yamakawa, O., 2002. J. Agric. Food Chem. 50, 3718. Kisiel, W., 1992. Pol. J. Chem. 66, 1449. Konovalov, D.A., Khubieva, Sh.I., 1997. Rastit. Resur. 33, 87. Konovalova, O.A., Rybalko, K.S., Sheichenko, V.I., 1974. Chem. Nat. Comp. 10, 591. Malik, M.N., Fenko, M.D., Shiekh, A.M., Wisniewski, H.M., 1997. J. Agric. Food Chem. 45, 817. Merfort, I., 1992. Phytochemistry 31, 2111. Pauli, G.F., Kuczkowiak, U., Nahrstedt, A., 1999. Magn. Reson. Chem. 37, 827. Pe´ter, A., Do´sa, G., 2002. Acta Bot. Hung. 44, 129. Re´thy, B., Csupor-Lo¨ffler, B., Zupko´, I., Hajdu´, Z., Ma´the´, I., Hohmann, J., Re´dei, T., Falkay, G., 2007. Phytother. Res. 21, 1200. Shigematsu, N., Konno, I., Kawano, N., 1982. Phytochemistry 21, 2156. Shoeb, M., Jaspars, M., Mac Manus, S.M., Celik, S., Nahar, L., Kong-Thoo-Lin, P., Sarker, S.D., 2007. J. Nat. Med. 61, 164. Shtacher, G., Kasman, Y., 1971. Tetrahedron 27, 1343. Spitaler, R., Schlorhaufer, P.D., Ellmerer, E.P., Merfort, I., Bortenschlager, S., Stuppner, H., Zidorn, C., 2006. Phytochemistry 67, 409. Wollenweber, E., Do¨rr, M., Fritz, H., Valant-Vetschera, K.M., 1997. Z. Naturforsch. 52c, 137. Xian, Q., Chen, H., Liu, H., Zou, H., Yin, D., 2006. Environ. Sci. Pollut. Res. 13, 233. Zee, O.P., Kim, D.K., Lee, K.R., 1998. Arch. Pharm. Res. 21, 618. Zhao, Y.-M., Zhang, M.-L., Shi, Q.-W., Kiyota, H., 2006. Chem. Biodiv. 3, 371.
Contents lists available at ScienceDirect
Biochemical Systematics and Ecology journal homepage: www.elsevier.com/locate/biochemsyseco
Terpenoids and phenolics from Inula ensifolia Anna Stojakowska a, *, Janusz Malarz a, Szymon Zubek b, Katarzyna Turnau c, Wanda Kisiel a a b c
´w, Poland Department of Phytochemistry, Institute of Pharmacology, Polish Academy of Sciences, 12 Sm˛etna Street, 31-343 Krako ´w, Poland Mycology Unit, Institute of Botany, Jagiellonian University, 46 Lubicz Street, 31-512 Krako ´w, Poland Institute of Environmental Sciences, Faculty of Biology and Earth Sciences, Jagiellonian University, 7 Gronostajowa Street, 30-387 Krako
a r t i c l e i n f o Article history: Received 18 May 2009 Accepted 12 December 2009 Keywords: Inula ensifolia Asteraceae Thymol derivatives Norisoprenoids Flavonoids Dicaffeoylquinic acids
1. Subject and source Inula ensifolia L. (Asteraceae, Inuleae) is a herbaceous perennial, about 45 cm high, with grey-green, lance-shaped leaves and anthodia composed of bright-yellow, narrow ray florets surrounding darker disc florets. Whole plants of I. ensifolia were collected from xerothermic grasslands nearby Kalina-Lisiniec (Miecho´w Upland, Poland; coordinates: 50 330 9400 N, 20170 9700 E), in June 2008, and identified by Teresa Anielska M.Sc. from the Institute of Environmental Sciences, Jagiellonian University in Krako´w. A voucher specimen (08/08) has been deposited at the Garden of Medicinal Plants, Institute of Pharmacology, Polish Academy of Sciences, Krako´w.
2. Previous work The species lacks detailed phytochemical investigation. Literature data are sparse. Wollenweber et al. (1997) examined five Inula species (Inula britannica L., Inula germanica L., Inula salicina L., Inula helenium L., I. ensifolia L.) with respect to exudate flavonoid production. However, no exudate flavonoids were found in I. ensifolia aerial parts. Thin-layer chromatography of methanolic extracts from disc and ray flowers of I. ensifolia revealed the presence of phenolic acids (caffeic and chlorogenic) and flavonoids (apigenin and hyperin) (Pe´ter and Do´sa, 2002). Secondary metabolites of plants from the genus Inula, including monoterpenoids, sesquiterpenoids and flavonoids, have been reviewed by Konovalov and Khubieva (1997) and recently by Zhao et al. (2006). Although the plant has no any documented medicinal use, an antiproliferative activity of methanolic extract from I. ensifolia against human cancer cell lines in vitro has been reported (Re´thy et al., 2007).
* Corresponding author. Tel.: þ48 12 6623217; fax: þ48 12 6374500. E-mail address: [email protected] (A. Stojakowska). 0305-1978/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.bse.2009.12.011
A. Stojakowska et al. / Biochemical Systematics and Ecology 38 (2010) 232–235
233
3. Present study The present report deals with the isolation of five thymol derivatives (1–5) from the roots of I. ensifolia, as well as atocopherol (6), two norisoprenoids (7, 8), three quercetin derivatives (9–11) and four caffeoylquinic acids (12–15) from its aerial parts (Fig. 1). The air-dried roots of I. ensifolia (10.8 g) were powdered and exhaustively extracted with CHCl3 at room temperature with shaking. The obtained extract was concentrated under reduced pressure to give 0.162 g of an oily residue. This residue, after fractionation by preparative TLC (Merck, Art. 1.05553; hexane-EtOAc, 9:1) followed by semipreparative HPLC on a Delta-Pak C-18 column (particle size 15 mm, 25 100 mm) coupled to a dual wavelength UV/vis detector operating at 205 and 270 nm, using MeOH–H2O mixture (13:7) at a flow rate of 6 ml min1, yielded: 1 (21.9 mg), 2 (2.1 mg), a mixture (2.8 mg) of 3 and 4 (by 1 H NMR), and a fraction (1.5 mg) containing 5 as the main component (by 1H NMR and ESIMS). The aerial parts (59.6 g) of I. ensifolia were powdered and extracted successively with CHCl3, followed by pure MeOH at room temperature with shaking and the solvents were evaporated under reduced pressure which furnished crude CHCl3 (2.98 g) and MeOH (5.50 g) extracts. The crude CHCl3 extract was fractionated by column chromatography on a silica gel (Merck, Art 7754) using gradients of EtOAc in hexane as a solvent system. Elution with hexane-EtOAc 9:1 followed by preparative TLC in the solvent system of the same composition led to the isolation of 6 (3.3 mg). From the more polar fractions (hexane-EtOAc, 1:1) a mixture (2 mg) of 7 and 8 (by 1H NMR) was isolated. The crude MeOH extract was subjected to column chromatography on Sephadex LH-20 (Pharmacia Biotech) eluted with MeOH/H2O. The eluted fractions were monitored by analytical RP-HPLC using an Agilent 1200 Series HPLC system (Agilent Technologies, USA) equipped with a Rheodyne manual sample injector, quaternary pump, degasser, column oven and a diode array detector. Chromatographic separations were carried out at 25 C, on a Zorbax Eclipse XDB-C18 column (4.6 150 mm, 5 mm particle size; Agilent Technologies, USA) with a mobile phase consisting of H2O/HCOOH/CH3COOH 99/0.9/0.1 (solvent A) and MeCN/MeOH/HCOOH/CH3COOH 89/10/0.9/0.1 (solvent B), at a flow rate of 1 ml/min, using 10 ml injections. The gradient elution conditions described by Spitaler et al. (2006) were used. This procedure yielded: 12 (100 mg), 13 (300 mg), a mixture (160 mg) of 9 and 10 (by 1H NMR and co-HPLC with authentic samples), a mixture (304 mg) of 13, 14 and 15 (by 1H NMR), and 11 (60 mg), in that order. The mixtures were not further separated, as the 1H NMR signals could be readily assigned to the respective compounds. Optical rotations were determined with a PolAAr31 automatic polarimeter (Optical Activity LTD). 1H and 13C NMR spectra were measured in CDCl3 or in CD3OD (phenolic compounds) on a Varian Mercury-VX 300 spectrophotometer operating at 300.08 MHz (1H) and 75.46 MHz (13C). COSY experiments were carried out using the same instrument. Chemical shifts (d in ppm) were referenced to TMS. Mass spectra (ESIMS) were recorded on a Bruker Esquire 3000 mass spectrometer. The isolated compounds were identified based on their physical ([a]D, whenever possible) and spectroscopic data, including NMR and MS, and their comparison with literature data for: 7-isobutyroyloxythymol methyl ether (1, Shtacher and Kasman, 1971; Anthonsen and Kjøsen, 1971), 10-isobutyroyloxy-8,9-epoxythymol isobutyrate (2), 10-(2-methylbutyroyloxy)-8,9-epoxythymol isobutyrate (3), 10-isovaleroyloxy-8,9-epoxythymol isobutyrate (4) (Bohlmann et al., 1969; Zee et al., 1998), 7,10diisobutyroyloxy-8,9-epoxythymol isobutyrate (5, Bohlmann and Zdero, 1977), a-tocopherol (6, Baker and Myers, 1991; Malik et al., 1997), megastigmane aglycone - 3b-hydroxy-5b,6b-epoxy-b-ionone (7, Cha´vez et al., 1997; Xian et al., 2006), loliolide (8, Kisiel, 1992; El Hattab et al., 2008), quercetin-3-O-b-glucopyranoside (isoquercitrin, 9, Shoeb et al., 2007), quercetin-3-O-bgalactopyranoside (hyperin, 10, An et al., 2008), quercetin-3-O-b-(600 -caffeoylgalactopyranoside) (11, Shigematsu et al., 1982), chlorogenic acid (12, Pauli et al., 1999), 1,5-, 3,4- and 3,5-dicaffeoylquinic acids (13–15, Merfort, 1992; Basnet et al., 1996; Islam et al., 2002). 4. Chemotaxonomic significance A total of 15 terpenoid and phenolic compounds have been isolated from the roots and aerial parts of I. ensifolia. Except for compounds 10 and 12, the other isolated constituents are reported for the first time from I. ensifolia. Quinic acid derivatives (12–15) appeared to be major secondary metabolites (over 1% yield) of the aerial parts. The paraphyletic genus Inula L., heterogenous with respect to morphology and chromosome numbers, comprises ca. 100 species native to Eurasia and Africa (Bremer, 1994). Within the genus, a group of resiniferous taxa, including I. helenium – a well known medicinal plant, has been distinguished by Anderberg (1991). Roots of the resiniferous species usually contain essential oils with the eudesmane-type sesquiterpene lactones alantolactone and/or isoalantolactone as major constituents. Separate position of these species has been confirmed by ITS sequence analysis (Eldena¨s et al., 1998; Francisco-Ortega et al., 2001). Within Inula species lacking resin canals, a classification based on morphological data has been proposed (Anderberg, 1991). According to this proposal, I. ensifolia has been included in the I. salicina group together with I. germanica, Inula hirta, Inula helvetica and Inula viscidula. Of these, only I. salicina and I. germanica have been phytochemically investigated (Anthonsen and Kjøsen, 1971; Konovalova et al., 1974; Bohlmann et al., 1978, 1985). Both plant species contained sesquiterpene lactones in their aerial parts (I. germanica – germacranolides, I. salicina – eudesmanolides). The compounds were absent from the plant material under study. This absence of sesquiterpene lactones situates I. ensifolia closer to some representatives of the postulated Inula decurrens group, i.e. Inula bifrons and Inula conyza. Moreover, I. germanica is rich in exudate flavonoids, derivatives of luteolin, scutellarein and quercetagenin, whereas only one derivative of luteolin was found in I. salicina exudate and no flavonoids in an exudate of I. ensifolia were found (Wollenweber et al., 1996). It seems that, neither exudate flavonoids nor esterified thymol derivatives are helpful chemotaxonomical markers within Inula. The flavonoids due
234
A. Stojakowska et al. / Biochemical Systematics and Ecology 38 (2010) 232–235
CH2R1
7
O
1
O
O
2
6 5
O
3
4 8
OMe
9
O
1
CH3
CH3
O
CH3
10
R1 H H H OiBu
2 3 4 5
H3C
OR2
R2 iBu MeBu iVal iBu
CH3
CH3
CH3 HO CH3
6
O
12
11 2 3
HO
1 4
7
6
O
5
9
8
11
HO 3
10
2
4 1
O
5 6
O
8 7
13
10
9
7
8 OH OH
HOOC OR
OH
9 10 11
OR4
6
O
HO
1
1
2
4
5
OR3
3
OR1
OR2
R2 H H Caff Caff
R3 H H Caff H
O
R - Glc R - Gal R - (6''-caffeoyl)Gal
12 13 14 15
R1 H Caff H H
R4 Caff Caff H Caff
Fig. 1. Structures of thymol derivatives (1–5), a-tocopherol (6), norisoprenoids (7, 8), flavonoids (9–11) and quinic acid derivatives (12–15), isolated from Inula ensifolia (iBu, isobutyroyl; MeBu, 2-methylbutyroyl; iVal, isovaleroyl; Caff, caffeoyl; Glc, b-glucopyranosyl; Gal, b-galactopyranosyl).
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