Biological activity and essential oil composition of two new Tanacetum chiliophyllum (Fisch. & Mey.) Schultz Bip. var. chiliophyllum chemotypes from Turkey

Industrial Crops and Products 39 (2012) 97–105

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Biological activity and essential oil composition of two new Tanacetum chiliophyllum (Fisch. & Mey.) Schultz Bip. var. chiliophyllum chemotypes from Turkey夽 Kaan Polato˘glu a,b,∗ , Betül Demirci c , Fatih Demirci c , Nezhun Gören a , Kemal Hüsnü Can Bas¸er d Yıldız Technical University, Faculty of Science & Letters, Department of Biology, 34210 I˙ stanbul, Turkey Near East University, Faculty of Pharmacy, Department of Analytical Chemistry, 10 Mersin, Turkey c Anadolu University, Faculty of Pharmacy, Department of Pharmacognosy, 26470 Eskis¸ehir, Turkey d King Saud University, Faculty of Science & Letters, Department of Botany and Microbiology, 11451 Riyadh, Saudi Arabia a

b

a r t i c l e

i n f o

Article history: Received 7 October 2011 Received in revised form 29 January 2012 Accepted 2 February 2012 Keywords: Tanacetum Asteraceae Essential oils Chemotypes Camphor Antibacterial activity

a b s t r a c t Water-distilled essential oils from aerial parts of Tanacetum chiliophyllum (Fisch. & Mey.) Schultz Bip. var. chiliophyllum, from two different localities in Turkey were analyzed by GC and GC/MS. The flower and stem oils of T. chiliophyllum var. chiliophyllum collected from Van-Muradiye location were characterized with camphor (32.5%, 36.2%), 1,8-cineole (1.6%, 16.1%), chamazulene (9.2%, 2.9%), for the first sample from this location and 1,8-cineole (12%, 18.4%), terpinene-4-o1 (10.3%, 9%), (E)-sesquilavandullol (5.8%, 1.6%), p-cymene (5.4%, 5.4%), hexadecanoic acid (4.2%, 7.6%) for the second plant sample. The flower and stem oils of the plant collected from Van-Güzeldere location were characterized with 1,8-cineole (22.1%, 28.9%), terpinene-4-ol (6.5%, 5.6%), ␣-pinene (5.3%, 1.5%). Five chemotypes were proposed according to biosynthetic origin of the main components of the investigated oils and previous investigations and they were tested with AHC analysis. Antibacterial activity of the oils were evaluated for five Gram-positive and five Gram-negative bacteria by using a broth microdulition assay. The highest activity was observed on Escherichia coli with the stem oil of the first sample from Muradiye (62.5 ␮g/mL) which gave same MIC with the positive control chloramphenicol. The highest DPPH scavenging activity was observed on the stem oil from the first sample of Muradiye location at 15 mg/mL concentration (79.1%). The oils showed moderate DPPH scavenging activity when compared with the positive control ␣-tocopherol. All of the oils except for flower and stem oils from the second sample of Muradiye location showed toxicity against Vibrio fischeri in the TLC-bioluminescence assay. © 2012 Elsevier B.V. All rights reserved.

1. Introduction Tanacetum chiliophyllum varieties are native plants of south west Asia including Turkey as well as north west of Iran, Azerbeijan and Armenia. This species is represented in Turkey with four varieties which are var. monocephalum, var. oligocephalum, var. heimerlei and var. chiliophyllum (Davis, 1975). Tanacetum chiliophyllum var. chiliophyllum is known with “c¸eren”, “ormadere” and “yavs¸an” local names in Eastern Anatolia (Altundag and Özhatay, 2009; Altundag and Ozturk, 2011).

夽 Partially presented at the International Symposium on “7th Plant Life of South West Asia” 25–29 June 2007 Eskis¸ehir, Turkey. ∗ Corresponding author at: Near East University, Faculty of Pharmacy, Department of Analytical Chemistry, 10 Mersin, Turkey. Tel.: +90 392 680 2000; fax: +90 392 680 2038. E-mail addresses: [email protected] (K. Polato˘glu), [email protected] (B. Demirci), [email protected] (F. Demirci), [email protected] (N. Gören), [email protected] (K.H.C. Bas¸er). 0926-6690/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.indcrop.2012.02.005

Decoction of the flowers heads of T. chiliophyllum var. chiliophyllum is used against pulmonic disorders, kidney stones and as antipyretic in traditional medicine (Altundag and Ozturk, 2011). It was worthy to note that in our field studies villagers reported snake deterrent properties of this plant however, to the best of our knowledge there is no report on the plant regarding such activity. Various biological activities due to sesquiterpene lactone content of Tanacetum species are well known (Gören et al., 2002) however, to the best of our knowledge only report on the biological activity of the extracts of T. chiliophyllum varieties was on insecticidal activity and DPPH scavenging activity of T. chiliophyllum var. monocephalum extracts (Polato˘glu et al., 2011c). Previous reports indicate the isolation of sesquiterpene lactone tamirin (Matsakanyan and Revazova, 1974), flavonoids ether, 6-hydroxyluteolin-6,3 4 scutellarein-6,7,4 -trimethyl trimethyl ether, 6-hydroxyluteolin-6,3 -dimethyl ether, scutellarein-6,7-dimethyl ether and flavonols quercetagetin3,6,4 -trimethyl ether, quercetagetin-3,6,7-trimethyl ether (Wollenweber et al., 1989) from T. chiliophyllum var. chiliophyllum.

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Also new sesquiterpene lactone type compounds chiliophyllin, heimerlein and known spiciformin, deacetyllauerenbiolide, 1␣hydroperoxy-1-desoxo chrysanolide, tabulin, tanachin, tamirin, dentatin, were isolated from T. chiliophyllum var. heimerlei (Gören and Tahtasakal, 1993, 1994) and 1-epi-chiliophyllin from T. chiliophyllum var. monocephalum (Polato˘glu et al., 2011c). Flavonoids triterpenes 4 ,5,7-trihydroxy-3 ,8-dimethoxyflavone, and  4 ,5,7-trihydroxy-8-methoxyflavone, neolupenyl acetate (lup12-ene-3␤-acetate) were also reported from T. chiliophyllum var. monocephalum (Polato˘glu et al., 2011c). Biological activities and composition of the essential oil from T. chiliophyllum var. chiliophyllum growing in Erzurum and Elazı˘g locations were reported in the literature. The main essential oil components were reported as camphor 28.5%, 1,8-cineole 17.1%, camphene 7.1%, isobornyl propionate 5.4%, carveol 4.5%, 3-cyclohexen-l-ol 3.5% for the plant from Elazı˘g and camphor 17.9%, 1,8-cineole 16.6%, bomeol 15.4%, dihydro-␣-cyclogeranyl hexanoate 10.1% and dihydro-␣-cyclogeranyl pentanoate 3% for the plant from Erzurum (Bagci et al., 2008; Salamci et al., 2007). Also the essential oil composition of this plant from Bayburt location reported as camphor 16.8%, cis-chrysanthenyl acetate 16.3%, ␣-thujone 12.5% and nonadecane 3.6% (Bas¸er et al., 2001). Previous reports show variation in the main components of T. chiliophyllum var. chiliophyllum essential oils. There are many examples of chemo variation in the essential oils at species level such as the examples for T. vulgare, T. nubigenum species (Chanotiya et al., 2005; Chanotiya and Mathela, 2007; Collin et al., 1993; Dev et al., 2001; Hendriks et al., 1990; Judzentiene and Mockute, 2004, 2005; Keskitalo et al., 2001; Mathela et al., 2008; Ognyanov et al., 1992; Rohloff et al., 2004). In addition variation of the oils obtained from subspecies (Judzentiene and Mockute, 2005; Polato˘glu et al., 2009a) and varieties (Bagci et al., 2008; Salamci et al., 2007) of genus Tanacetum have been also reported in recent years. In our ongoing research on phytochemical and biological investigation of this genus in Turkey (Polato˘glu et al., 2009a,b, 2010a,b,c, 2011a,b,c) here we report on the antibacterial, cytotoxic, DPPH scavenging activities and essential oil composition of T. chiliophyllum var. chiliophyllum from two different localities in East Anatolia region of Turkey that showed differences in their morphological properties and in their essential oil compositions. 2. Materials and methods 2.1. Plant materials Plant materials were collected during the flowering period in 22–23 July 2006 from two different locations provinces VanMuradiye at 2494 m above sea level and Van-Güzeldere at 2805 m above sea level. Two different sample groups were collected from the province Van-Muradiye since there were two groups of plants that distinctively showed morphological variations from each other and another sample group was collected from the province Van-Güzeldere. Voucher specimens have been deposited at the Herbarium of the Faculty of Science, Istanbul University (Voucher no. ISTE 83756, ISTE 85431 and ISTE 85430 respectively), Turkey. Plant materials were identified by Dr. Kerim Alpınar. 2.2. Isolation of the essential oils Flowers and stems (100 g each) of the plant samples from province Van-Muradiye (A, B, C, and D) and Van-Güzeldere (E and F) locations were separately subjected to hydrodistillation for 4 h using a Clevenger-type apparatus to produce the oils. Second sample (C and D) from Muradiye with 68 g flowers and 100 g stems were subjected to same procedure. Blue colored oils were obtained

from the first plant sample of Muradiye with 0.1% (A), 0.2% (B) (v/w) yields for flowers and stems respectively. Yellow colored oils were obtained from the second sample of Muradiye with 0.06% (C), 0.06% (D) (v/w) yields for flowers and stems respectively. Yellow colored oils were obtained from the sample of Güzeldere with 0.16% (E), 0.1% (F) (v/w) yields for flowers and stems respectively.

2.3. Gas chromatography–mass spectrometry analysis The essential oil analysis were done simultaneously by gas chromatography (GC) and gas chromatography–mass spectrometry (GC–MS) systems. The GC–MS analysis were done with an Agilent 5975 GC-MSD system with Innowax FSC column (60 m × 0.25 mm, 0.25 ␮m film thickness) and helium as carrier gas (0.8 mL/min). Oven temperature was programmed to 60 ◦ C for 10 min and raised to 220 ◦ C at rate of 4 ◦ C/min. Temperature kept constant at 220 ◦ C for 10 min and than raised to 240 ◦ C at a rate of 1 ◦ C/min. The injector temperature was set at 250 ◦ C. Split flow was adjusted at 50:1. Mass spectra were recorded at 70 eV with the mass range m/z 35 to 450. The GC analyses were done with Agilent 6890N GC system. FID detector temperature was set to 300 ◦ C and same operational conditions applied to a duplicate of the same column used in GC–MS analyses. Simultaneous auto injection was done to obtain the same retention times. Relative percentage amounts of the separated compounds were calculated from integration of the peaks in FID chromatograms (Table 1). Identification of essential oil components were done by comparison of their retention times with authentic samples or by comparison of their relative retention index (RRI) to series of nalkanes. Computer matching against commercial (Wiley GC/MS Library, Adams Library, MassFinder 2.1 Library) (Bas¸er and Demirci, 2007) and in-house “Bas¸er Library of Essential Oil Constituents” built up by genuine compounds and components of known oils, as well as MS literature data (Jennings and Shibamoto, 1980; Joulain and König, 1998) was used for identification.

2.4. Antibacterial activity assay Five Gram-positive bacteria (Staphylococcus aureus ATCC 6538, Staphylococcus epidermis ATCC 12228, Bacillus cereus NRRL B-3711, Bacillus subtilis NRRL B-4378, Meticillin resistant S. aureus (Clinical isolate)) and five Gram-negative bacteria (Escherichia coli NRRL B3008, Pseudomonas aeruginosa ATCC 27853, Enterobacter aerogenes NRRL 3567, Proteus vulgaris NRRL B-123, Salmonella typhimurium ATCC 13311) were used in this study. The minimum inhibitory concentration (MIC) values were determined for all of the oils, on each organism by using microplate dillution method (Iscan et al., 2002). Stock solutions of the oils (2 mg/mL) and standard antibacterial compound chloramphenicol (2 mg/mL) were prepared. Liquid medium was diluted by adding 25% DMSO or CH3 OH. Serial dilution was done on 96-well microlitre plates. Bacteria were standardized according to McFarland No: 0.5 after incubation 24 h at 37 ◦ C on MHB. Cultures were mixed with essential oils and were incubated 24 h at 37 ◦ C. Minimum inhibitory concentrations (MIC: ␮g/mL) were detected at the minimum concentration where bacterial growth was inhibited. 1% 2,3,5-Triphenyltetrazolium chloride (TTC, Aldrich, St. Louis, MO, USA) was used as an indicator of bacterial growth. Essential oil free solutions were used as blank controls and chloramphenicol was used as a positive control. All the experiments were performed in triplicate and means of results were given for the MIC values of the oils (Table 2).

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Table 1 Composition (%) of Flower and Stem Oils of T. chiliophyllum var. chiliophllum from Güzeldere and Muradiye locations. (Continued overleaf). Compound

RRI

A

B

C

D

E

F

Tricyclene ␣-Pinene ␣-Thujene Santolinatriene Camphene Hexanal ␤-Pinene Sabinene Thuja-2,4(10)-diene ␦-3-Carene Myrcene ␣-Phellandrene ␣-Terpinene Dehydro 1,8-cineole Isoamyl isobutyrate Limonene 1,8-Cineole p-Mentha-1,3,6-triene cis-Anhydrolinalool oxide (Z)-3-Hexenal Isochrysanthenone 2-Pentyl furan trans-Anhydrolinalool oxide ␥-Terpinene 2-Methylbutyl butyrate p-Cymene Isoamyl isovalerate Terpinolene 2-Methyl butyl isovalerate 6-Methyl-5-heptene-2-one 1,2,3 Trimethylbenzene 1-Hexanol Nonanal Rosefuran ␣-Thujone ␥-Campholene aldehyde ␣,p-Dimethyl styrene Filifolene trans-Linalool oxide (Furanoid) ␤-Thujone 1-Octen-3-ol 2,6-Dimethyl-1,3(E),5(E),7-octatetraene trans-Sabinene hydrate Linalool-7-oxide-3-one Longipinene* Cyclosativene ␣-Copaene ␣-Campholene aldehyde Decanal Chrysanthenone Camphor trans-Chrysanthenyl acetate Benzaldehyde Dihydro achillene (E)-2-Nonanal Linalool Italicene cis-Sabinene hydrate 1-Nonene-3-ol 1-Methyl-4-acetylcyclohex-1-ene trans-p-Menth-2-ene-1-ol cis-Chrysanthenyl acetate Pinocarvone Bornyl acetate Chrysanthenyl propionate* Nopinone Terpinen-4-ol Hotrienol 4-Terpinenyl acetate cis-p-Menth-2-ene-1-ol trans-p-Mentha-2,8-diene-1-ol Thuj-3-ene-10-al Dehydro sabina ketone Myrtenal Sabina ketone Bornyl isobutyrate

1014 1032 1035 1043 1076 1093 1118 1132 1135 1159 1174 1176 1188 1195 1195 1203 1213 1215 1220 1225 1234 1244 1253 1255 1275 1280 1285 1290 1299 1348 1355 1360 1400 1413 1437 1439 1443 1445 1450 1451 1452 1460 1474 1479 1482 1492 1497 1499 1506 1522 1532 1538 1541 1547 1548 1553 1553 1556 1556 1568 1571 1582 1586 1590 1599 1601 1611 1616 1630 1638 1639 1642 1643 1648 1651 1651

– tr – – 0.1 – tr – – – – – tr – – – 1.6 – tr – – – – tr – 0.1 tr – tr – – – 0.1 – 0.2 tr tr 0.1 tr tr – 0.1 – – – – – 0.4 – 0.6 32.5 0.4 – 0.2 – 0.6 – – 0.1 – 0.3 – 3.2 0.2 0.1 – 1.4 2.7 – – 1 – – 0.5 – –

0.1 0.8 tr 0.1 2.2 tr 0.2 tr tr – – tr 0.3 0.1 tr 0.1 16.1 tr – tr tr tr tr 0.5 – 1.9 tr 0.1 tr – 0.1 tr 0.1 tr tr 0.1 tr 0.1 – – – – 0.7 tr – – tr 0.2 – 0.8 36.2 0.4 – 0.1 – 0.1 – 0.6 – – 0.4 – 2.4 0.2 tr – 2.2 0.3 tr – 0.6 tr tr 0.3 tr 0.2

– 0.9 – – – – 0.5 – – – – – – – – – 12 – – – – – – – – 5.4 – – – 0.3 – – – – 3 – 0.1 – – 0.5 – – – – 0.4 – – 0.3 – – 0.3 3.5 – – – 0.9 – – – – 0.6 – 1.2 – – – 10.3 – – 0.5 – – – 0.3 – –

– 0.5 – – – – 0.4 tr – – – – – – – – 18.4 – – – – – – – – 5.4 – – – – – – – – 1.2 – – – – 0.3 – – – – 0.3 – – 0.2 – – tr 2.8 – – – 0.3 tr 2.3 – – 1.1 – 2.1 – 0.4 – 9 – 1 0.8 – – – 0.5 – –

– 5.3 0.1 – 0.4 – 1.9 0.4 – – – – 0.2 – – 0.2 22.1 – – – – 0.1 – 0.5 – 4.2 – – – – – – – – 0.2 – 0.1 – – – 0.1 – 0.1 – 0.4 – – 0.3 – – 0.8 3.7 – – – 0.9 – – – – 0.3 1.4 1.8 – 0.4 – 6.5 – 0.2 0.2 – – – 0.2 – –

– 1.5 tr – 0.1 tr 1 0.1 tr tr tr – 0.3 – – 0.1 28.9 – – tr – 0.1 – 0.9 tr 3.3 – 0.2 tr 0.2 – tr 0.1 0.1 0.2 0.1 tr – – 0.3 0.1 – 2 – 0.2 tr tr 0.3 tr 0.1 0.9 2 tr – tr 0.2 – 1.5 – 0.2 0.6 – 2.8 – 0.4 0.1 5.6 – 0.3 0.4 – – – 0.4 – –

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Table 1 (Continued) Compound

RRI

A

B

C

D

E

F

Chrysanthenyl isobutyrate Umbellulone cis-Verbenol trans-Pinocarveol cis-p-Mentha-2,8-diene-1-ol ␦-Terpineol trans-Verbenol trans-Piperitol (=trans-p-Menth-1-en-3-ol) p-Mentha-1,8-diene-4-ol (=limonene-4-ol) ␣-Terpineol Borneol Cabreuva oxide II Verbenone Germacrene D ␤-Bisabolene ␤-Selinene Chrysanthenyl isovalerate I Carvone cis-Piperitol Chrysanthenyl isovalerate II Naphthalene Cabreuva oxide IV Isobornyl isovalerate ar-Curcumene p-Methyl acetophenone Cumin aldehyde Myrtenol trans-p-Mentha-1(7),8-diene-2-ol p-Mentha-1,5-diene-7-ol (E,E)-2,4-Decadienal 2-Phenyl ethyl acetate (E)-␤-Damascenone trans-Carveol Geraniol p-Cymene-8-ol 10-epi-Italicene ether (E)-Geranyl acetone cis-Carveol Italicene ether Geranyl isovalerate cis-p-Mentha-1(7),8-diene-2-ol Nonadecane trans-Jasmone (E)-␤-Ionone cis-Jasmone Piperitone oxide 2-Phenylethyl-2-methylbutyrate 2-Phenylethyl isovalerate o-Cresol Caryophyllene oxide trans-␤-Ionone-5,6-epoxide Amyl phenyl acetate Salvial-4(14)-ene-1-one Pentadecanal (E)-Nerolidol p-Mentha-1,4-diene-7-ol Caryophylla-2(12),6(13)-diene-5-on Octanoic acid (E)-Sesquilavandulyl acetate Cumin alcohol cis-Bejarol Hexahydrofarnesyl acetone Spathulenol ␣-Bisabolol oxide B 3,4-Dimethyl-5-pentyliden-2(5H)-furanone Nor-copaanone 1-Tetradecanol ␥-Decalactone (E)-Sesquilavandulol Nonanoic acid Thymol Docosane Eremoligenol ar-Turmerol ␣-Bisabolol Carvacrol trans-␣-Bergamotol

1656 1657 1663 1670 1678 1682 1683 1689 1700 1706 1719 1722 1725 1726 1741 1742 1743 1751 1758 1760 1763 1768 1770 1786 1797 1802 1804 1811 1814 1827 1838 1838 1845 1857 1864 1867 1868 1882 1892 1893 1896 1900 1948 1958 1969 1983 1988 1992 2004 2008 2009 2012 2037 2041 2050 2073 2074 2084 2100 2113 2122 2131 2144 2156 2179 2179 2179 2183 2183 2192 2198 2200 2204 2214 2232 2239 2247

0.9 – – 2.5 0.7 tr 0.3 – – 0.4 2.7 – 0.1 – – 0.1 2 0.1 0.1 2.1 – – 0.1 – – – 0.5 0.2 – 0.2 – 0.1 0.8 tr 0.1 – – 0.1 – – 0.3 – – – – 0.1 0.1 – – 0.5 – – 0.2 0.1 – 0.1 0.3 0.1 – 0.2 – 0.4 0.7 – – 0.3 – 0.1 0.3 – – – 0.1 – – – –

2.2 – – 1.2 0.2 0.1 – – – – 2.8 – 0.1 tr – 0.1 3 0.1 0.1 2.8 – – 0.2 – – tr 0.2 0.2 – – – – 0.3 – 0.1 – – – – – 0.3 – – – – – – 0.1 – 0.1 – – – – – 0.1 0.1 – – 0.2 – 0.2 0.5 – 0.1 – 0.1 tr 0.1 – – – – – – tr tr

– – – 1.2 – 0.4 – 0.5 – 1.8 0.8 0.5 – – 0.2 – – 0.2 0.4 – – 0.3 – 0.2 0.3 – 0.4 – – tr – – 0.5 – – tr – – 0.1 – – – 0.1 – 0.4 – – – – – – – – – – – – – 1.1 – tr 0.2 0.8 0.5 – – – – 5.8 – – tr – 0.2 2.5 – –

– – – 1.7 – 0.5 1 0.5 – 0.9 0.5 tr – – 0.1 – – 0.5 0.5 – – – – 0.1 – – 0.6 – – – – – 0.4 – 0.5 – 0.2 – – – – – – 0.3 0.2 – – – – – – – – – – – – – 0.5 0.5 – 0.4 0.9 0.2 tr – – – 1.6 tr – – – 0.3 2.1 – –

– 2.4 – 1.5 – 0.3 0.2 – – 2.1 0.3 0.2 – – – – – 0.1 0.1 – 0.2 0.1 – – – tr 0.4 – – – – – 0.3 0.1 0.4 – – – – – – – – – 0.4 – – – tr 0.3 – – – – – – – – – – – – – – – – – – 3.6 – – – – 0.2 0.4 – –

– 2.6 0.2 1.7 – 0.6 1.1 0.2 tr 1.1 0.4 – – – 0.1 – – 0.2 0.1 – 0.4 – – 0.1 – 0.1 0.5 0.1 0.1 tr 0.1 0.1 0.4 tr 0.2 – 0.1 – – 0.1 tr tr 0.1 0.2 0.2 – tr – 0.2 tr tr 0.1 – – 0.1 0.1 – – – 0.3 – 0.6 0.3 0.1 0.3 – – – – 0.1 1.9 – – 0.2 1.3 – –

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Table 1 (Continued) Compound

RRI

A

B

C

D

E

F

␣-Eudesmol ␤-Eudesmol Intermedol Guaia-6,10(14)-diene-4␤-ol (2E,6E)-Farnesyl acetate 4-Oxo-␣-ylangene 1,4-Dimethyl azulene Decanoic acid Tricosane Caryophylladienol I Caryophylladienol II Eudesm-4(15),7-dien-4␤-ol Farnesyl acetone Caryophyllenol I Caryophyllenol II Tetracosane Chamazulene Kaur-16-ene Pentacosane Dodecanoic acid 1-Octadecanol Phytol Tetradecanoic acid Heptacosane Pentadecanoic acid Nonacosane Hexadecanoic acid Monoterpene hydrocarbons Oxygenated monoterpenes Sesquiterpene hydrocarbons Oxygenated sesquiterpenes Others Total

2250 2257 2264 2269 2271 2289 2291 2298 2300 2316 2324 2369 2384 2389 2392 2400 2430 2438 2500 2503 2607 2622 2670 2700 2822 2900 2931

– 4.7 – – – – 0.6 tr 0.5 0.2 0.6 0.1 – 0.2 0.2 – 9.2 – 0.5 – 0.1 – tr 0.5 – tr 1.6 1.6 65.8 1 14.6 0.8 83.5

– 1.1 – – – tr 0.2 – 0.1 0.1 0.3 tr – tr 0.1 – 2.9 – 0.1 – 0.2 0.4 0.2 0.1 – – 1.1 3.2 59.3 0.9 16.2 0.2 91.1

3.4 0.8 1 2 – – – tr 1.6 – – – tr – – 0.6 – – 1.5 tr – – 1.3 0.8 tr tr 4.2 – 15.8 3.5 34.7 7.3 77.6

1.4 0.8 0.7 – 0.2 – – tr 0.4 – – – – – – – – – 0.7 tr – 0.5 1.1 0.9 – 0.2 7.6 6.5 80.1 – 0.4 1.6 76.5

0.5 0.4 – – 0.2 – – 0.9 0.6 – 0.2 – – – 0.3 0.3 – – 0.6 – – – – 0.6 – – 1.7 3.4 76.9 0.2 3 0.5 72.9

0.1 0.4 0.3 0.1 0.1 – – tr 0.1 – 0.1 – – – – 0.1 – tr 0.2 – 0.3 0.3 0.8 0.3 – 0.5 2.3 – 8.3 4.8 16 25.3 78.3

RRI, relative retention indices; tr, trace (<0.1%); A, T. chiliophyllum var. chiliophyllum from Muradiye sample 1 – flower oil; B, T. chiliophyllum var. chiliophyllum from Muradiye sample 1 – stem oil; C, T. chiliophyllum var. chiliophyllum from Muradiye sample 2 – flower oil; D, T. chiliophyllum var. chiliophyllum from Muradiye sample 2 – stem oil; E, T. chiliophyllum var. chiliophyllum from Güzeldere sample – flower oil; F, T. chiliophyllum var. chiliophyllum from Güzeldere sample – stem oil. * Correct isomer not identified.

2.5. Vibrio fischeri cytotoxicity assay 5 ␮L of 2 mg/mL ethanol solutions of the essential oils were applied on HPTLC plates (Merck Darmstadt, Germany) by the help of Automatic TLC Sampler 4 (Camag Muttenz, Switzerland). Freezedried, luminescent Vibrio fischeri microorganisms obtained from the kit were inoculated on the medium provided by the kit (ChromadexTM Irvine, CA, USA). Culture of the microorganism was incubated for 24–30 h at 28 ◦ C. Previously prepared HPTLC plates were dipped into the freshly grown luminescent culture with an automatic immersion device (Camag Muttenz, Switzerland) and excess of the culture removed from the plates with a squeegee. Plates were photographed at −30 ◦ C with CCD camera of BioLuminizer (Camag Muttenz, Switzerland). Cytotoxicity of the oils were detected as black spots on the photographs (Verbitski et al., 2008) (Table 2). 2.6. DPPH scavenging activity assay DPPH scavenging activity of the oils were determined with DPPH radical protocol (Yamaguchi et al., 1998). A modified protocol for HPTLC-DPPH (Shikov et al., 2007) was used. Stock solutions of the oils from the first sample of Muradiye, and Güzeldere samples (10 and 15 mg/mL), positive control ␣-tocopherol (Aldrich, St. Louis, MO, USA) (10 and 15 mg/mL) and DPPH (0.1 mM) (Aldrich, St. Louis, MO, USA) were prepared with CH3 OH. 200 ␮L of the oil solutions were mixed with 1000 ␮L of DPPH solution as well as positive controls and essential oil free blank controls in 1.5 mL Eppendorf tubes and vortexed for 2 min. After incubating all the samples and controls for one hour in dark at room temperature, 2 ␮L of them were applied on an aluminium 60 F254 TLC

Plate (Merck Darmstadt, Germany) with 5 mm band length by the help of Linomat 5 TLC applicator system (Camag Muttenz, Switzerland). After preparing samples and controls on the TLC; plates were scanned at 517 nm with a TLC Scanner 3 (Camag Muttenz, Switzerland) and absorbance of the bands were detected. Percent of DPPH scavenging activity was calculated according to % DPPH Scav. Prop. = [(AControl − ASample )/AControl ] × 100 formula. The data obtained from this formula were analyzed with Tukey’s honest significance test (Table 3). 2.7. Statistical analysis Essential oil composition data of T. chiliophyllum var. chiliophyllum reported in the literature (Bagci et al., 2008; Bas¸er et al., 2001; Salamci et al., 2007) and from the present work was used in the agglomerative hierarchical cluster (AHC) analysis. Data set was prepared based on the qualitative and quantative properties of the oils reported in the literature and from the GC/MS analysis performed in this research. AHC analysis were performed using XLSTAT – Pro 2011 program trial version (Addinsoft, New York, USA). AHC analysis were done by pearson’s dissimilarity method and unweighted pair-group average as aggregation criterion. A dendrogram was obtained showing dissimilarity of the analysed oils within the range 0–1 (Fig. 1). 3. Results and discussion 3.1. Plant samples Three plant samples collected from two locations three different populations showed morphological differences such as height

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Fig. 1. Dendrogram obtained from AHC analysis of T. chiliophyllum var. chiliophyllum essential oils based on the chemical composition of the three samples and those from the literature.

of the stems and number of capitula per stem. The second plant sample from Muradiye location exhibited same morphological properties with the Güzeldere sample. However, the first plant sample exhibited different number of capitula per stem and stem height than the other plant samples. Differences observed in the morphology of the plants were within the range of the systematic identification key given in the flora records (Davis, 1975).

3.2. Essential oil compositions In the oils of first plant sample from Muradiye location, 87 and 106 compounds were detected representing 83.5% (A), 91.1% (B) for flower and stem oils, respectively. 61 compounds were identified in the oils of the second plant sample from Muradiye representing 77.6% (C), 76.5% (D) flower and stem oils, respectively. 61 and 114 compounds were identified in the oils from Güzeldere, representing 72.9% (E), 78.3% (F) of the flower and stem oils respectively. The first sample from Muradiye was dominated by camphor (32.5% and 36.2%), 1,8-cineole (1.6% and 16.1%), chamazulene (9.2% and 2.9%), ␤-eudesmol (4.7% and 1.1%), pinocarvone (3.2% and 2.4%) for flower (A) and stem (B) oils respectively. The second sample from Muradiye was dominated by 1,8-cineole (12% and 18.4%), terpinene-4-ol (10.3% and 9%), (E)-sesquilavandulol (5.8% and 1.6%), p-cymene (5.4% and 5.4%), hexadecanoic acid (4.2% and 7.6%), trans-chrysanthenyl acetate (3.5% and 2.8%) for flower (C) and stem (D) oils respectively. Güzeldere sample was dominated with 1,8-cineole (22.1% and 28.9%), terpinene-4-ol (6.5% and 5.6%), ␣-pinene (5.3% and 1.5%), p-cymene (4.2% and 3.3%) and transchrysanthenyl acetate (3.7% and 2%) for flower (E) and stem (F) oils respectively.

Some of the food poisoning cases are associated with B. cereus. Contamination of food resulting from this bacteria results in production of enterotoxin causing food poisoning. Enterotoxins causes severe nausea, vomiting and diarrhea (Ehling-Schulz et al., 2004; Kotiranta et al., 2000). Similarly some strains of E. coli can also cause food related infections such as gastroenteritis and urinary tract infections. Previous research also indicated high activity of T. chiliophyllum var. chiliophyllum oils against E. coli (Bagci et al., 2008; Salamci et al., 2007). However, there is no report on the activity of T. chiliophyllum var. chiliophyllum oils against Enterobacter aerogenes, P. vulgaris, S. epidermis, B. cereus. Additional studies should be done in order to find the active constituents in the oil B. The cytotoxicty assay employed in the research was used to evaluate possible general toxicity of the essential oil samples as an initial indicator. All of the oils except for the C, flower oil of the second sample from Muradiye showed toxicity to V. fischeri. The toxicity was observed at low concentrations when compared to vitamin C. This confirmed the possible occurrence of a substance or a mixture of substances which inhibited the growth of V. fischeri (see Table 2). 3.4. DPPH scavenging activity Highest DPPH radical scavenging activity was observed on the samples with 15 mg/mL concentration. Oils A, B (B: 79.1%, A: 78.1%) showed higher DPPH scavenging activity than oils E, F (F: 65.6%, E: 55.2%). In the oils of samples from both locations stem oils showed higher DPPH scavenging activity than the flower oils. DPPH scavenging activity of the oils were concentration dependent. None of the oils showed high DPPH scavenging activity when compared with the positive control ␣-tocopherol (15 mg/mL: 94.6%) at the same concentration (see Table 3).

3.3. Antibacterial and cytotoxic activity 3.5. Agglomerative hierarchical cluster analysis Highest antibacterial activity was observed for the oil B against E. coli (62.5 ␮g/mL) which was the same MIC of the positive control chloramphenicol. Also stem oil from the first plant sample of Muradiye (125 ␮g/mL) and flower oil E from Güzeldere sample (125 ␮g/mL) showed the same MIC of the positive control against B. cereus.

The variation in the essential oils of T. chilliophyllum var. chilliophyllum samples presented in this research proved existence of different chemotypes in this variety. In order to verify these chemotypes a dendrogram was obtained from agglomerative hierarchical clustering of the oil compositions given in this research and the

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103

Table 2 Antibacterial (MIC: ␮g/mL) and cytotoxic activity of T. chiliophyllum oils from Güzeldere and Muradiye locations.* Microorganism

A

B

C

D

E

F

Chloramphenicol

Staphylococcus aureus Meticillin resistant S. aureus Staphylococcus epidermis Bacillus cereus Bacillus subtilis Escherichia coli Pseudomonas aeruginosa Enterobacter aerogenes Proteus vulgaris Salmonella typhimurium Vibrio fischeri

250 500 250 250 500 ≥500 500 500 250 500 Toxic

500 125 250 125 125 62.5 250 500 250 250 Toxic

150 750 150 150 375 150> 150 150> 150> 150> –

500 500 500 500 500 500 500 500> 500> 500 Toxic

250 250 125 125 250 250 250 500 250 500 Toxic

≥500 250 500 500> 500 ≥500 500 500> 500> 500> Toxic

62.5 62.5 31.2 125 62.5 62.5 31.2 62.5 62.5 125 NA

NA, not available; MIC, MIC values are given as ␮g/mL. Results are given as the mean values of three parallel experiments.

*

data from the previous reports. As expected very low dissimilarity was observed between the oils C, D and E, F. These oils formed a group together with 0.58 dissimilarity where dissimilarity ranged 0–1. The small difference observed in AHC analysis points out minor compounds such as ␣-pinene and ␤-pinene which exists in E, F oils with higher amounts. Similarly ␣-thujone which existed with high amounts in C, D oils while it was absent in E, F oils. Instead of ␣thujone E, F oils contained umbellulone. Oils C, D and E, F were separated from the rest of the oils with 0.86 dissimilarity in the dendrogram. In the dendrogram oil B and the previously reported oil from Elazı˘g location (Bagci et al., 2008) formed a group which showed a 0.53 dissimilarity. Higher similarity of the stem oil B with the oil from the previous report (Bagci et al., 2008) and its higher dissimilarity with A was observed since the flower oil contained lower amounts of 1,8-cineole and higher amounts of chamazulene than B. The previous report on the oil obtained from the same plant from Elazı˘g indicate high amounts of 1,8-cineole (Bagci et al., 2008) similarly to B. However, the reported oil does not contain chamazulene instead it contains azulene in very small amounts (0.1%) (Bagci et al., 2008). Other minor compounds such as camphene, isobornylpropionate, carveol and 3-cyclohexen-1-ol were not observed at all or observed in very small quantities in A, B. The oils reported from Erzurum (Salamci et al., 2007) and Bayburt (Bas¸er et al., 2001) locations were seperated from the Elazı˘g (Bagci et al., 2008) and A, B group with 0.65 and 0.74 dissimilarities.

3.6. Defined chemotypes The oils which were obtained from three samples originating from two different locations showed differences in their compositions. The oil compositions of A, B contains high amount of camp hor (A, 32.5%; B, 36.2%) and chamazulene (A, 9.2%; B, 2.9%) unlike the other samples. Due to the occurrence of chamazulene color of these oils were blue. Oils C, D and E, F contained low camphor content (<1%); these oils did not contained chamazulene. All the samples contained high amounts of 1,8-cineole (12–28.9%) except for the oil A (1.6%); in addition to these differences oils A, B contained ␤eudesmol (A, 4.7%; B, 1.1%) higher than the other plant samples (<1%). Second plant sample from Muradiye and Güzeldere sample contained same main and minor components with similar amounts.

The difference in the essential oil compositions of Muradiye samples which were collected from the same location and time presented differences that cannot simply explained by environmental conditions. The compounds such as chamazulene, ␤-eudesmol were present in the first sample from Muradiye in high quantity and completely missing or in very low quantity in the other samples. Additionally C, D and E, F did not contained the rearrangement products of bornyl cation such as camphene, camphor and borneol or these compounds exists in very low quantities in these oils unlike A, B and the previous literature (Bagci et al., 2008; Salamci et al., 2007; Bas¸er et al., 2001). The C, D and E, F are mainly characterized with 1,8-cineole, terpinen-4-ol and other by products of ␣-terpinyl cation such as thujones and pinenes but except the products of bornyl cation. Bornyl cation and pinyl cation are produced in plants by folding of the ␣-terpinyl cation by an enzyme mediated mechanism that is reported to be responsible for formation of these bicyclic compounds. The nature of the compound whether bornyl type or pinyl type is decided on the which end of the double bond was involved in the reaction (Dewick, 2001). It is fair to point out that until now only in one previous report existance of pinane type bicyclic monoterpene was reported in high quantity for T. chilliophyllum var. chilliophyllum from Bayburt location which also contained bornane type monoterpenes (Bas¸er et al., 2001). This previously reported plant from Bayburt should be defined as cis-chrysanthenyl acetate/camphor chemotype. On the other hand until now chamazulene is not reported from T. chilliophyllum var. chilliophyllum essential oils. Chamazulene is actually an artifact which is obtained from degradation of the heat sensitive guaianolide type compound matricin (Dewick, 2001). The existence of chamazulene only in A, B oils suggests the guanolide content of this plant; therefore it could be defined as chamazulene/camphor chemotype. The essential oil composition of the oils C, D and E, F which were poor on bornane type compounds suggests that these two are 1,8-cineole/terpinen-4ol chemotypes. Another chemotype is previously reported from Elazı˘g location which contained camphene in high quantity unlike other plants together with camphor and 1,8-cineole in its oil (Bas¸er et al., 2001). This previously reported plant from Elazı˘g location should be defined as camphor/1,8-cineole/camphene chemotype. Finally another article on this plant from Erzurum location reports an unusual compound dihydro-␣-cyclogeranyl derrivatives in

Table 3 DPPH scavenging activity (%) of the essential oils.* Concentration

A

B

E

F

␣-Tocopherol

15 mg/mL 10 mg/mL

78.1 ± 0.96b** 71.7 ± 0.56c

79.1 ± 1.2b 74.7 ± 0.42b

55.2 ± 1.97d 31.9 ± 0.77e

65.6 ± 2.61c 44.3 ± 0.56d

94.6 ± 0.96a 94.5 ± 0.79a

* **

Results are given in means of three parallel experiments. Different letters show the statistical differences of the values at p < 0.05 probability level.

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high amounts together with 1,8-cineole, camphor and borneol (Salamci et al., 2007). This composition suggests this plant from Erzurum location could be dihydro-␣-cyclogeranyl hexanoate/1,8cineole/campor/borneol chemotype. With this report we presented 2 new chemotypes (chamazulene/camphor – Muradiye sample 1 oils A, B; 1,8-cineole/terpinen-4-ol – Muradiye sample 2 oils C, D and Güzeldere sample oils E, F) of T. chilliophyllum var. chilliophyllum. The results obtained from AHC analysis matched with the proposed chemotypes except for the results observed for the oils A, B and the previous report (Bagci et al., 2008). However, the facts that plant parts studied in Elazı˘g sample contains aerial parts unlike our study and also essential oils of A (flower) and B (stem) oils presented differences in their composition, clarifies this mismatch. Results of biological activity of the oils also matched the similarities observed in the oils. Except for a couple of microorganisms similar activity was observed for C, E and D, F. 4. Conclusion The present study points out differences for the essential oil composition of T. chiliophyllum var. chiliophyllum from different locations. Chemotypes of this plant were established depending on the biosynthetic origin of the major compounds in the oils studied and the previous reports on the oils from the same plant. AHC analysis of the essential oil compositions obtained from this research and the previous reports clearly indicated the differences of proposed chemotypes as expected. However, additional essential oil studies on T. chiliophyllum var. chiliophyllum from different locations together with DNA isolation and comparison studies are still necessary in order to define the exact position of defined chemotypes and to identify other chemotypes from this species. Antibacterial activities together with moderate DPPH scavenging activity of the oil B showed that essential oils of this chemotype may be considered as beneficial oil which could have potential uses. However, in vivo toxicity tests, activity guided isolation studies are required to find the active principles and potential beneficial uses for this oil. Chemotypes of the plant should be well defined in order to locate the chemotype with the highest activity. To the best of our knowledge this is the first report on DPPH scavenging activity, V. fischeri cytotoxic activity and antibacterial activity of T. chiliophyllum var. chiliophyllum chemotypes. Acknowledgements This research was supported by the Scientific and Technological Research Council of Turkey (TUBITAK-TBAG 104T306) and State Planning Organization (Project no. 27-DPT-01-07-01). Also infrastructure of Anadolu University Research Funding (Project no. BAP-060301) was used. We would like to thank Ms. Gamze C¸ayırdere who assisted the antimicrobial evaluations. References Altundag, E., Ozturk, M., 2011. Ethnomedicinal studies on the plant resources of east Anatolia, Turkey. Proc. Soc. Behav. Sci. 19, 756–777. Altundag, E., Özhatay, N., 2009. Local names of some useful plants from I˘gdir province (East Anatolia). J. Fac. Pharm. I˙ stanbul 40, 101–115. Bagci, E., Kursat, M., Kocak, A., Gur, S., 2008. Composition and antimicrobial activity of the essential oils of Tanacetum balsamita L. subsp. balsamita and T. chiliophyllum (Fisch. et Mey.) Schultz Bip. var. chiliophyllum (Asteraceae) from Turkey. J. Essent. Oil Bear. Pl. 11, 479–484. Bas¸er, K.H.C., Demirci, B., Tabanca, N., Özek, T., Gören, N., 2001. Composition of the essential oils of Tanacetum armenum (DC.) Schultz Bip., T. balsamita L., T. chiliophyllum (Fisch. & Mey.) Schultz Bip. var. chiliophyllum and T. haradjani (Rech. fil.) Grierson and the enantiomeric distribution of camphor and carvone. Flavour Fragr. J. 16, 195–200.

Bas¸er, K.H.C., Demirci, F., 2007. Chemistry of essential oils. In: Berger, R.G. (Ed.), Flavours and Fragrances, Chemistry, Bioprocessing and Sustainability. SpringerVerlag, Berlin, pp. 43–86. Chanotiya, C.S., Sammal, S.S., Mathela, C.S., 2005. Composition of a new chemotype of Tanacetum nubigenum. Indian J. Chem. B 44, 1922–1926. Chanotiya, C.S., Mathela, C.S., 2007. Two distinct essential oil bearing races of Tanacetum nubigenum Wallich ex DC from Kumaon Himalaya. Nat. Prod. Commun. 2, 785–788. Collin, G.J., Deslauriers, H., Pageau, N., Gagnon, M., 1993. Essential oil of Tansy (Tanacetum vulgare L.) of Canadian origin. J. Essent. Oil Res. 5, 625–627. Davis, P.H., 1975. Flora of Turkey and The East Aegean Islands, vol. 5. University Press, Edinburgh. Dev, V., Beuchamp, P., Kashyap, T., Melkani, A., Mathela, C., Bottini, A.T., 2001. Composition of the essential oil of Tanacetum nubigenum Wallich ex De. J. Essent. Oil Res. 13, 319–323. Dewick, P.M., 2001. Medicinal Natural Products ‘A Biosynthetic Approach’, 2nd ed. John Wiley & Sons Ltd., Chichester. Ehling-Schulz, M., Fricker, M., Scherer, S., 2004. Bacillus cereus, the causative agent of an emetic type of food-borne illness. Mol. Nutr. Food Res. 48, 479–487. Gören, N., Tahtasakal, E., 1993. Sesquiterpenes of Tanacetum chiliophyllum var. heimerlei. Phytochemistry 34 (4), 1071–1073. Gören, N., Tahtasakal, E., 1994. A further investigation on Tanacetum chiliophyllum var. heimerlei. Turk. J. Chem. 18, 296–300. Gören, N., Arda, N., C¸alıs¸kan, Z., 2002. Chemical characterization and biological activities of the genus Tanacetum (Compositae). In: Rahman, Atta-ur (Ed.), Studies in Natural Product Chemistry, vol. 27. Elsevier Science Press, pp. 547–658. Hendriks, H., Van Der Elst, D.J.D., Van Putten, F.M.S., Bos, R., 1990. The essential oil of Dutch Tansy (Tanacetum vulgare L.). J. Essent. Oil Res. 2, 155–160. Iscan, G., Kirimer, N., Kürkcüoglu, M., Baser, K.H.C., Demirci, F., 2002. Antimicrobial screening: Mentha piperita essential oil. J. Agric. Food Chem. 50, 3943–3946. Jennings, W.G., Shibamoto, T., 1980. Quantitative Analysis of Flavor and Fragrance Volatiles by Glass Capillary GC. Academic Press, New York. Joulain, D., König, W.A., 1998. The Atlas of Spectra Data of Sesquiterpene Hydrocarbons. EB-Verlag, Hamburg. Judzentiene, A., Mockute, D., 2004. Composition of the essential oils of Tanacetum vulgare L. growing wild in Vilnius District (Lithuania). J. Essent. Oil Res. 16, 550–553. Judzentiene, A., Mockute, D., 2005. The injlorescence and leaf essential oils of Tanacetum vulgare L. var. vulgare growing wild in Lithuania. Biochem. Syst. Ecol. 33 (5), 487–498. Keskitalo, M., Pehu, E., Simon, J.E., 2001. Variation in volatile compounds from tansy (Tanacetum vulgare L.) related to genetic and morphological differences of genotypes. Biochem. Syst. Ecol. 29, 267–285. Kotiranta, A., Lounatmaa, K., Haapsalo, M., 2000. Epidemiology and pathogenesis of Bacillus cereus infections. Microbes Infect. 2, 189–198. Mathela, C.S., Padalia, R.C., Joshi, R.K., 2008. Variability in fragrance constituents of Himalayan Tanacetum species: commercial potential. J. Essent. Oil Bear. Pl. 11, 503–513. Matsakanyan, V.A., Revazova, L.V., 1974. Tamirin from Tanacetum chiliophyllum. Chem. Nat. Compd. 3, 396–397. Ognyanov, I., Min, F.T.B., Todorova, M., Kuleva, L., 1992. Chemotypes in some bulgarian populations of Chrysanthemum vulgare (L.) Bernh. Comptes rendus de l’Acadaemie Bulgare des Sciences 45 (4), 29–31. Polatoglu, K., Gören, N., Bas¸er, K.H.C., Demirci, B., 2009a. The variation in the essential oil composition of Tanacetum cadmeum (Boiss.) Heywood ssp. orientale Grierson from Turkey. J. Essent. Oil Res. 21, 97–100. Polato˘glu, K., Gören, N., Bas¸er, K.H.C., Demirci, B., 2009b. The essential oil composition of Tanacetum densum (Labill.) Heywood ssp. sivasicum Hub.-Mor. & Grierson from Turkey. J. Essent. Oil Res. 21, 1–3. Polato˘glu, K., Demirci, F., Demirci, B., Gören, N., Baser, K.H.C., 2010a. Essential oil composition and antibacterial activity of Tanacetum argenteum (Lam.) Willd. ssp. argenteum and T. densum (Lab.) Schultz Bip. ssp. amani Heywood from Turkey. J. Oleo Sci. 59 (7), 361–367. Polato˘glu, K., Demirci, F., Demirci, B., Gören, N., Baser, K.H.C., 2010b. Antimicrobial activity and essential oil composition of a new T. argyrophyllum (C. Koch) Tvzel var. argyrophyllum chemotype. J. Oleo Sci. 59 (6), 307–313. Polato˘glu, K., Baser, K.H.C., Gören, N., Demirci, B., Demirci, F., 2010c. Antibacterial activity and the variation of Tanacetum parthenium (L.) Schultz Bip. essential oils from Turkey. J. Oleo Sci. 59 (4), 177–184. Polato˘glu, K., Demirci, B., Gören, N., Bas¸er, K.H.C., 2011a. Essential oil composition of endemic Tanacetum zahlbruckneri (Náb.) and Tanacetum tabrisianum (Boiss.) Sosn. and Takht. from Turkey. Nat. Prod. Res. 25 (6), 576–584. Polato˘glu, K., Demirci, B., Gören, N., Bas¸er, K.H.C., 2011b. Essential oil composition of Tanacetum kotschyi from Turkey. Chem. Nat. Compd. 47 (2), 297–299. Polato˘glu, K., Karakoc¸, Ö.C., Gökc¸e, A., Gören, N., 2011c. Insecticidal activity of Tanacetum chiliophyllum (Fisch. & Mey.) var. monocephalum grierson extracts and a new sesquiterpene lactone. Phytochem. Lett. 4, 4432–4435. Rohloff, J., Mordal, R., Dragland, S., 2004. Chemotypical variation of Tansy (Tanacetum vulgare L.) from 40 different locations in Norway. J. Agric. Food Chem. 52, 1742–1748. Salamci, E., Kordali, S., Kotan, R., Cakir, A., Kaya, Y., 2007. Chemical compositions antimicrobial and herbicidal effects of essential oils isolated from Turkish Tanacetum aucheranum and Tanacetum chiliophyllum var. chiliophyllum. Biochem. Syst. Ecol. 35, 569–581.

K. Polato˘glu et al. / Industrial Crops and Products 39 (2012) 97–105 Shikov, A.N., Pozharitskaya, O.N., Ivanova, S.A., Makarov, V.G., 2007. Separation and evaluation of free radical-scavenging activity of phenol components of Emblica officinalis extract by using an HPTLC-DPPH method. J. Sep. Sci. 30, 1250–1254. Verbitski, S.M., Gourdin, G.T., Ikenouye, L.M., McChesney, J.D., 2008. Detection of Actaea racemosa adulteration by thin-layer chromatography combined thinlayer chromatography bioluminescence. J. AOAC Int. 91, 268–275.

105

Wollenweber, E., Mann, K., Vetschera, K.M.V., 1989. External flavonoid aglycones in Artemisia and some further Anthemidae (Asteraceae). Fitoterapia 60 (5), 460–463. Yamaguchi, T., Takamura, H., Matoba, T., Terao, J., 1998. HPLC method for evaluation of the free radical-scavenging activity of foods by using 1,1-diphenyl-2picrylhydrazyl. Biosci. Biotechnol. Biochem. 62, 1201–1204.