J. Agric. Food Chem. 2003, 51, 7115−7121
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Seasonal and Phenological Variations of the Essential Oil from the Narrow Endemic Species Artemisia molinieri and Its Biological Activities VEÄ RONIQUE MASOTTI,*,† FABIEN JUTEAU,† JEAN MARIE BESSIEÅ RE,§ JOSETTE VIANO†
AND
Laboratoire Dynamique et Ressources du Ve´ge´tal, E.A. 2202-Biodiversite´, Universite´ de Provence, UFR DENTES et SVTE, case 17, 3 place Victor Hugo, F-13331 Marseille Cedex 3, France, and Ecole Nationale Supe´rieure de Chimie de Montpellier, 8 rue Ecole Normale, F-34296 Montpellier, France
The volatile components of the aerial parts of Artemisia molinieri, an endemic wormwood of southern France, were analyzed by GC and GC-MS. Among the 69 compounds identified, major components were ascaridole (19-76%), R-terpinene (traces-36%), p-cymene (1-17%), 1,8-cineole (0.3-8%), and germacrene D (0.6-15%). Quantitative variations have been characterized following the season, the phenological cycle, and the aging of the plants. Bioassays have been performed on a sample of essential oil, which has shown a strong inhibition of the growth of both tested yeasts (Candida albicans and Saccharomyces cerevisiae var. chevalieri) and minor activity on both tested Gram-negative bacteria (Escherichia coli and Enterococcus hirae). The oils have shown interesting antioxidant activities on the basis of R-tocopherol as reference compound, up to 400-1200%. KEYWORDS: Artemisia molinieri; Asteraceae; essential oils; chemical composition; ascaridole; antibacterial and antifungal activities; antioxidant; chemiluminescence; GC; GC-MS
INTRODUCTION
The genus Artemisia is one of the largest in the Asteraceae family, consisting of more than 800 species that are widespread all over the world. In this genus, some species are consumed as spices (tarragon) or alcoholic drinks (black and common wormwoods), and many of them have been used since ancient times as folk remedies and credited with a long list of medicinal uses, including antimalarial, antiviral, antitumor, spasmolytic, and others (1). Most of these interests can be related to the high amounts of volatile terpenic compounds that can be found in the essential oil, giving to the wormwoods their aromatic and medicinal properties (2-4). Artemisia molinieri [discovered by Que´zel et al. (5)] is a restricted endemic species, located on two temporary freshwater marshes in southeastern France. This species has been registered in the Red Book of French flora as a plant threatened by extinction (6) and subjected to regional protection since 1994 in France. Both marshes are included in the LIFE international program and belong to the NATURA 2000 European network. As for many endangered species, it is important to determine highly valuable components in A. molinieri to assess protection status. Previous analyses on A. molinieri have utilized diethyl ether extracts, which contain mainly ascaridole and two bisabolol * Corresponding author [telephone (33) 4 91 10 62 61; fax (33) 4 91 10 62 58; e-mail address:
[email protected]]. † Universite ´ de Provence. § Ecole Nationale Supe ´ rieure de Chimie de Montpellier.
oxide derivative (7). Some of the major compounds of its essential oil have been identified as 1,4-cineole and p-cymene (8) or R-terpinene, ascaridole, and p-cymene (9). The only exhaustive study of the essential oil of A. molinieri has been published by Carnat et al. (10), but these analyses deal with only one population, at flowering stage. These authors mentioned ascaridole, p-cymene, R-terpinene, 1,8-cineole, and germacrene D as major compounds. A. molinieri antimicrobial activity has been tested on the flavonoid content of this species (11). These authors have reported antifungal activity equivalent to that of nystatine against Candida albicans. It is important to note that the antioxidant activity of mugworts has not often been studied: the methanolic extract of Artemisia maritima has shown a weak antioxidant effect (12), but chlorogenic acid of Artemisia iwayamogi has shown the same activity as ascorbic acid (1), and the antioxidant power of Artemisia judaica essential oil was comparable to that of butylated hydroxytoluene (BHT). MATERIALS AND METHODS Plant Material. Aerial parts of A. molinieri Que´zel, Barbero et R. Loisel, were harvested in southeastern France (departement of Var) in both known populations, near Besse-sur-Issole (Lac Gavoti) and near Flassans-sur-Issole (Lac Redon), at two different stages of development in 1999 (buds and flowering) and at four different stages of development in 2000 (vegetative, buds, flowering, and seeding). The aerial parts were harvested early in the morning, from numerous representative plants, randomly chosen. Plant material was taken immediately to the
10.1021/jf034621y CCC: $25.00 © 2003 American Chemical Society Published on Web 10/22/2003
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Table 1. Qualitative and Quantitative Composition (Peak Area Percent) of Seasonally Harvested A. molinieria location:
Lac Gavoti Aug 1999
physiological status: yield (%): sample:
B 0.8 1
F 0.7 2
V 0.9 3
V 0.8 4
B 0.9 5
V 0.5 6
S 0.5 7
− 0.1 0.2 0.2 − tr 0.1 tr 10.6 9.6 2.8 0.2 − 0.6 0.1 0.2 0.1 0.2 0.2 0.3 0.1 0.1 0.3 0.2 0.1 0.4 0.1 0.1 0.2 0.2 0.2 0.1 − 0.2 1.0 − − 0.5 0.1 tr 0.6 0.1 36.8 0.2 − 1.2 0.6 2.3 0.4 0.1 − 0.6 0.2 2.6 1.1 0.2 0.4 tr 0.3 tr 0.4 0.2 0.4 0.1 0.9 − 4.1 0.2
− − − tr − − − − 0.5 6.5 1.1 − − 0.2 − 0.1 − 0.1 0.2 0.1 − 0.1 0.2 − − 0.1 0.1 − − − − − − − 0.5 − − − − − 3.6 0.1 74.7 tr − 0.5 0.7 2.6 0.6 0.1 − 1.7 0.2 1.5 0.7 − 0.1 − 0.1 − 0.1 − − − 0.1 − 0.6 −
0.1 0.2 0.5 0.7 − 0.1 0.1 0.1 34.2 11.4 3.1 0.7 0.1 1.3 − 0.2 0.2 0.2 0.4 0.3 0.1 0.1 0.4 0.2 − 0.3 0.2 − 0.1 − − 0.1 0.1 0.2 0.9 0.2 − 0.1 0.1 0.2 2.9 − 20.0 0.3 0.1 0.8 0.5 1.0 1.8 − − 0.3 − 1.1 0.5 − 0.1 − tr − 0.2 tr 0.2 0.4 0.7 0.3 3.5 −
− − − − − − − − 0.7 2.5 0.7 − − 0.2 − 0.1 tr 0.1 0.1 0.1 tr − 0.3 tr 0.1 0.3 − 0.1 − − 0.1 0.1 − 0.1 1.0 tr tr 0.5 − 0.1 4.6 0.8 64.9 0.3 0.2 1.0 0.7 − 0.6 − − 5.5 0.3 1.6 1.0 0.1 0.1 tr 0.1 tr 0.1 0.1 0.2 0.1 1.0 0.1 3.0 0.2
− − − 0.1 − − 0.1 − 6.9 9.7 2.4 − − 0.3 − 0.2 0.1 0.2 0.2 0.1 tr − 0.3 0.1 0.1 0.3 − 0.1 − tr − 0.1 − 0.2 0.9 tr tr 0.5 0.1 0.1 5.0 − 53.5 0.3 0.2 0.9 0.8 tr 0.8 − − 0.6 0.2 2.0 1.0 0.1 0.1 0.1 0.2 − tr 0.2 0.1 0.1 1.1 0.3 3.8 0.3
− − − − − − − − tr 1.1 0.4 − − 0.1 − 0.3 − 0.1 0.3 0.1 − − 0.5 − tr 0.4 − − − tr 0.1 0.1 − 0.2 1.4 0.1 0.1 0.4 0.1 0.1 4.7 − 44.9 0.3 − 0.5 0.9 7.6 1.8 − − 1.2 0.3 1.4 1.6 tr tr 0.1 0.3 tr 0.1 0.2 − tr 2.6 − 15.1 0.3
− − − 0.1 − − − − 0.1 16.9 7.8 − − 0.2 − 0.2 − 0.1 0.2 0.1 − 0.1 0.4 0.1 0.2 0.4 − 0.1 − 0.1 0.2 0.1 − 0.2 1.2 0.1 tr 0.5 0.1 0.1 4.1 − 39.8 0.4 − 1.0 1.1 7.5 1.2 − − 0.6 0.4 1.1 0.8 − − − 0.2 tr 0.1 0.1 − tr 1.3 − 7.3 0.1
compound 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 67 68
R-thujene R-pinene R-fenchene sabinene 1-octen-3-ol β-pinene myrcene dehydro-1,8-cineole R-terpinene p-cymene 1,8-cineole (Z)-β-ocimene (E)-β-ocimene γ-terpinene NI cis-sabinene hydrate terpinolene p-cymenene trans-sabinene hydrate 2-methylbutyl 2-methylbutyrate amyl isovalerate p-mentha-1,3,8-triene cis-p-menth-2-enol p-mentha-1,5,8-triene trans-pinocarveol trans-p-menth-2-enol NI sabina ketone NI pinocarvone trans-β-terpineol δ-terpineol rosefurane epoxide NI terpinen-4-ol 4-methylacetophenone cryptone p-cymen-8-ol cis-piperitol trans-piperitol cis-isoascaridole NI ascaridole cumin aldehyde NI cis-piperitone oxide trans-piperitone oxide cis-carvenone oxide* trans-carvenone oxide* NI NI thymol NI trans-isoascaridole carvacrol cumin alcohol eugenol NI R-copaene β-bourbonene (Z)-jasmone β-caryophyllene (E)-β-farnesene dehydrosesquicineole linalyl 2-methylbutyrate γ-curcumene germacrene D β-selinene
identification abc abc abc abc abc abc abc abc abc abc abc abc abc abc abc abc abc abc ab ab abc abc a abc abc abc abc abc abc abc abc ab ab abc abc abc ab abc abc ab ab a a abc ab abc abc abc abc abc abc abc abc abc ab abc abc abc
May 2000
Lac Redon
harvest date:
Aug 2000
Oct 2000
RI 928 935 947 972 973 975 990 990 1022 1028 1032 1038 1048 1059 1065 1067 1088 1088 1098 1103 1108 1112 1122 1135 1136 1141 1148 1153 1154 1161 1162 1166 1167 1168 1180 1182 1183 1190 1198 1208 1240 1244 1248 1252 1253 1254 1258 1262 1272 1283 1285 1286 1290 1295 1296 1348 1360 1368 1382 1390 1399 1425 1460 1460 1461 1482 1486 1489
Aug 1999 B 0.9 8
F 0.8 9
May 2000
Aug 2000
Oct 2000
V 0.7 10
V 0.5 11
B 0.9 12
F 1.3 13
V 0.6 14
S 0.6 15
tr − 0.4 0.7 0.1 − 0.2 − 36.4 12.4 3.5 1.3 0.1 1.3 − 0.1 0.2 0.2 0.2 0.3 tr 0.1 0.3 0.2 tr 0.2 tr − tr − − − tr 0.3 0.8 − − 0.1 tr 0.1 1.2 0.1 19.0 0.2 0.1 0.8 0.2 1.2 0.1 tr tr 0.2 0.2 0.9 0.5 0.1 0.2 − − 0.1 0.4 0.2 0.2 0.6 0.6 0.5 4.9 −
− − − 0.1 0.1 − − − 0.6 4.5 3.7 − − 0.2 tr 0.2 tr 0.1 0.2 0.1 − − 0.4 0.1 tr 0.2 − − 0.1 − tr 0.1 tr 0.1 1.4 − tr 0.4 0.2 0.1 2.6 0.3 61.4 0.3 0.1 0.9 0.7 3.2 0.8 0.2 0.2 0.6 0.4 2.2 1.2 0.1 0.1 tr 0.2 − 0.1 0.1 0.1 0.1 0.8 − 6.5 0.1
− − − 0.1 − − − − 6.3 7.5 3.3 0.1 − 0.5 0.1 0.1 0.1 0.1 0.1 0.1 − tr 0.3 0.1 0.1 0.2 − tr tr tr 0.1 0.1 tr 0.1 0.9 − tr 0.3 tr tr 1.6 0.3 55.0 0.3 − 1.1 0.9 4.5 0.6 0.2 0.2 0.5 0.4 2.3 1.1 0.1 0.1 − 0.2 tr 0.2 0.1 0.1 tr 0.5 − 5.5 0.1
− − − − − − − − 0.2 1.7 0.3 − − 0.1 tr tr − tr 0.1 0.1 − − 0.2 − 0.1 0.1 − tr tr tr tr tr tr tr 0.4 − − 0.2 − − 5.6 − 76.1 tr − 1.0 0.3 1.8 0.4 0.2 0.1 0.2 0.2 3.0 1.4 − 0.1 − 0.1 − 0.2 0.1 tr − 0.5 − 1.9 0.1
− − − − − − − − tr 0.7 0.4 − − 0.1 0.1 0.1 − − 0.4 − − − 0.3 − 0.1 0.3 − 0.1 − 0.1 0.1 0.1 − 0.1 0.9 0.1 − 0.3 0.1 0.1 4.2 − 54.7 0.2 − 0.9 0.7 6.4 1.5 0.3 0.2 0.7 0.3 1.6 1.1 − − 0.1 0.1 − 0.1 0.1 tr − 0.6 − 7.4 0.1
− − − − − − − − tr 1.1 0.7 tr tr 0.2 0.2 0.1 tr 0.2 0.2 0.2 − − 0.3 tr 0.1 0.3 − tr tr tr 0.1 0.1 0.1 0.1 0.8 tr − 0.4 tr 0.1 4.6 − 74.0 0.2 − 1.2 0.9 5.0 0.5 0.2 0.1 0.5 0.3 2.6 0.7 − 0.1 − 0.1 0.1 0.2 0.1 − − 0.2 − 0.9 0.1
peak area % − − − tr − − − − 0.2 13.1 0.7 − − 0.3 tr − − 0.1 0.1 tr − − 0.3 0.1 − 0.1 − − − − − − − − 0.8 − − 0.2 − tr 4.0 0.1 66.8 tr − 0.8 0.6 3.4 − 0.1 tr 1.3 0.2 1.7 0.9 tr 0.1 tr − − 0.2 − − − 0.4 0.1 1.9 −
− − − − − − − − 0.8 15.9 1.2 − − 0.1 − − − − − − − − 0.1 tr − 0.1 − − − − − − − − 0.4 − − 0.1 − − 3.0 0.1 67.7 tr − 0.5 0.3 2.3 − − − 1.2 0.3 1.2 0.7 − 0.1 − − − 0.1 − − − 0.1 0.1 2.6 −
Phenological Variations of the Essential Oil from Artemisia molinieri
J. Agric. Food Chem., Vol. 51, No. 24, 2003
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Table 1. (Continued) location:
Lac Gavoti
harvest date:
Aug 1999
physiological status: yield (%): sample: compound 69 70 71 72 73 74 75 76 77 78 79 80 81 82 83 84 85 86 87
isoamyl phenylacetate bicyclogermacrene linalyl valerate δ-cadinene (E)-nerolidol NI spathulenol salvialenone NI linalyl 3-methylhexanoate R-bisabolol oxide B R-bisabolol NI R-bisabolol oxide A NI NI NI NI NI total identified (%)
B 0.8 1 identification a abc ab abc abc abc ab ab abc abc abc
F 0.7 2
May 2000 V 0.9 3
Lac Redon
Aug 2000 V 0.8 4
B 0.9 5
Oct 2000 V 0.5 6
RI 1490 1499 1510 1524 1559 1573 1574 1608 1637 1654 1656 1681 1730 1746 1765 1806 1832 1847 1860
S 0.5 7
Aug 1999 B 0.9 8
F 0.8 9
May 2000
Aug 2000
Oct 2000
V 0.7 10
V 0.5 11
B 0.9 12
F 1.3 13
V 0.6 14
S 0.6 15
0.3 0.3 − tr − 0.1 − − 0.3 0.1 0.5 0.3 − 0.4 − − 2.8 0.1 2.8
0.1 0.2 0.1 − 0.1 0.1 0.3 tr − 0.1 0.7 − − 0.1 − 0.3 1.1 0.1 0.5
0.2 0.2 0.1 tr 0.1 0.1 0.2 0.1 0.1 0.1 0.4 0.1 − 0.3 − 0.1 0.8 0.2 0.4
tr 0.2 tr − − − 0.1 − 0.1 0.1 0.4 0.2 − 0.4 − 0.1 0.8 0.3 0.5
0.2 0.3 tr − − 0.2 0.2 − 0.3 0.1 1.9 0.1 tr 0.1 0.1 0.9 3.3 0.2 6.3
tr tr tr − − 0.1 0.1 0.1 0.1 − 0.7 0.1 − 0.1 − 0.1 0.5 0.2 0.1
93.2
96.5 97.0 97.7 87.6 98.1
peak area % − 0.4 0.3 0.1 tr 0.2 0.1 − − 0.2 2.5 0.6 0.3 1.4 0.3 0.1 4.7 1.0 4.4
tr 0.1 − − − − 0.1 − − − 0.1 0.1 0.1 0.6 0.1 0.1 0.4 0.3 0.2
88.0 98.3 93.7
− 0.2 0.2 tr − 0.1 0.2 0.2 − 0.2 0.7 0.3 tr 0.3 tr 0.1 1.3 0.2 1.2
0.3 0.2 tr tr − tr − 0.3 − tr 0.6 0.4 tr 0.6 − − 2.9 0.1 2.7
0.2 0.2 0.2 tr − − 0.3 0.2 − 0.1 1.1 0.3 − 0.3 − 0.2 1.5 0.2 1.4
0.2 0.5 0.2 tr − 0.2 0.4 0.4 0.2 0.3 1.3 0.6 0.1 0.6 0.1 0.2 2.2 0.3 2.4
95.3
96.4 93.7 97.6
0.1 0.3 0.1 tr − 0.1 0.1 0.1 − 0.2 0.6 − − tr − 0.3 0.6 0.2 0.6
0.1 0.1 − − − − 0.1 tr − tr 0.2 0.1 0.1 0.2 tr 0.1 0.3 0.1 0.1
tr 0.1 − − − − − − − − 0.1 0.1 − 0.1 − − 0.2 0.1 0.3
99.0 99.0
RI, retention indices relative to C8−C22 n-alkanes on DB5 column; V, vegetative plant; B, budding plant; F, flowering plant; S, seeding plant; 1, a ) mass spectra, b ) retention index, c ) comparison with standard; tr, traces (
