Aromas From Quebec. II. Composition of the Essential Oil of the Rhizomes and Roots of Asarum canadense L.

Transcription

1 Collin et al. Aromas From Quebec. II. Composition of the Essential Oil of the Rhizomes and Roots of Asarum canadense L. André Bélanger, Centre de recherche et de développement en horticulture 430, boul. Gouin, Saint-Jean-sur-Richelieu (Québec), Canada J3B 3E6 Guy Collin* and François-Xavier Garneau, Corporation LASEVE, Université du Québec à Chicoutimi, Saguenay (Québec), Canada G7H 2B1 Hélène Gagnon and André Pichette, Laboratoire LASEVE, Université du Québec à Chicoutimi, Saguenay (Québec), Canada G7H 2B1 Abstract The fresh rhizomes and roots of Asarum canadense were steam-distilled to yield oils in 1.30% to 1.96%, respectively. The oils were analyzed by a combination of GC-FID and GC/MS techniques. In the case of the rhizome oil, the total amount of phenyl ethers including methyl eugenol, elemicins, and asaricin was about 58%. In the root oil, (E)-isoelemicin was the major product (20%) accompanied by elemicin (4.9%). From retention indices on both nonpolar and polar columns and MS considerations, it may be inferred that methoxy-elemicin and both (E)- and (Z)- methoxy-isoelemicin as well as the new hydroxyl derivatives : hydroxyl-elemicin and (E)- and (Z)-hydroxy-isoelemicins were characterized in the oils. Key Word Index Asarum canadense, Aristolochiaceae, essential oil composition, methyl eugenol, (E)-isoelemicin, linalool, junenol, methoxylated elemicin, hydroxy elemicin isomers. Introduction The essential oil of Asarum canadense L., Aristolochiaceae, is also known as Canadian Snakeroot oil. The chemical composition of the oil obtained from the steam-distilled ground root material of Asarum canadense was described some 20 years ago (1). Chalcone glycosides and flavonol glycosides (2) as well as aristolochic acid (3) were identified in the organic solvent extracts of this plant. Experimental Plant material: Roots and rhizomes of A. canadense were collected from plants cultivated in the La Pocatière region, Quebec Province on an experimental plot of a forest of maples. Isolation of the essential oil: The rhizomes (3.31 g) of the plant were steam-distilled in a 1 L glass flask for 3 h giving g of oil in a yield of 1.30% (w/w). The roots (6.60 g) of the plant were subjected to steam distillation in a 500 ml glass flask for 3 h giving 0.13 g of oil in a yield of 1.96% (w/w). GC-FID and GC/MS analyses: GC-FID and GC/MS analyses were carried out on a Hewlett-Packard 5890 gas chromatograph fitted with a non-polar DB-5 capillary column and a polar Supelcowax 10 column (30 m 0.25 mm; film thickness 0.25 mm), split mode, split ratio: 1:50. GC/MS analyses were performed on HP 5972 mass spectrometer at 70 ev coupled to an HP 5890 GC equipped with a DB-5 column (same as above) scan time 1.6/s; acquisition mass range: Temperature program for both the GC and GC/MS was 40 C for 2 min, then 2 C/min to 210 C and held constant for 33 min. Identification of the components was done by comparison of their retention indices with standards, and by comparison of their mass spectra with literature values (4 6). Quantitative data were obtained electronically from GC-FID peak area percentages without the use of correction factors. Results and Discussion Composition of the essential oil: The composition of the oil obtained from the fresh rhizomes is given in Table I. The major products observed in the fresh rhizomes oil were methyl *Address for correspondence /10/ $14.00/ Allured Business Media Received: February 2008 Revised: April 2008 Accepted: August /Journal of Essential Oil Research Vol. 22, March/April 2010

2 A. canadense eugenol (53.8%) and linalool (12.5%). The total amount of products similar to or belonging to the same family of phenyl ethers including methyleugenol, elemicin, isoelemicin and asaracin was about 58%. Alcohols formed the second most important group with a total of about 25%. Finally, esters amounted to about 5%. This composition was qualitatively similar but quantitatively different to that published by Guenther s books, for example phenol ethers 36.9%, alcohols 13.3%, and esters 27.5% (7). The very low percentage of asarone differentiates the present oil from the Asarum europaeum oil (7). Methyl eugenol, linalyl acetate and linalool have been reported as the main compounds (36%, 28% and 5%, respectively) of Canadian snakeroot oil (1). The composition of the root oil was more complex than the oil from the rhizomes (Table II). In the root oil, (E)-isoelemicin was the major product (20%) and was accompanied by elemicin (4.9%). A sesquiterpene, junenol and an unidentified product (each ~14%) were also major compounds. Alcohols (< 2%) and esters (< 3%) were among the minor constituents. Numerous sesquiterpenes were also present with percentages in the 0 5% range. The most important compounds of this family were guaia-6,9-diene (3.7%), zingiberene (3.1%) and seychellene (2.3%). GC and GC/MS of unknown components: Of particular interest was the presence of unusual compounds. The mass spectra of certain components of the oil showed well-defined and important molecular peaks at 238 and 224 (Figure 1). Moreover, the 208, 238 and 224 partial ion chromatograms showed similar, somewhat symmetric patterns on both nonpolar (Figure 2) and polar columns. The three compounds with a 208 molecular ion peak were identified as elemicin and (Z)- and (E)-isoelemicin. The first 238 molecular ion peak was identified from the 75k NBS Data bank as 1,2,3,4-tetramethoxy- 5-(2-propenyl)benzene (methoxyelemicin). Considering the great similarity of the structure of the molecules, one would expect that the substitution of an H atom located on the benzene ring by a methoxy group will produce a similar effect on the Kovat s indices (K.I.) of the elemicin and isoelemicin isomers. The D values [d = K.I.(R-O-CH 3 ) - K.I.(R- H)] shown in Table III confirm this situation. One can observe a small decrease on the polar Kovat s values (D = -26 ± 8) and a small increase on the non-polar column (D = +28 ± 14). The same observation can be made upon substitution by an OH group: the D values increase by 69 ± 10 and 220 ± 10 units on the non-polar and polar columns, respectively. From the additive rules of the Kovat s system (8), it may be inferred that the two other 238 molecular ion peaks belong to (Z)- and (E)-methoxy-isoelemicin: (Z) and (E)-5-(1-propenyl)-1,2,3,4- tetramethoxybenzene. Moreover, one can see that the differences in the K.I. observed between the polar and the non-polar columns (the D values in Table III) vary from 624 to 663 upon the addition of a methoxy radical. In contrast, this difference is in the range in the alkyl methyl ether on the non-polar column (9). The same type of discrepancies, resulting from the addition of an OH radical, between the three elemicin isomers and the alkane series may be observed. These D values are in the and ranges for the n-1-alcohol and 2-n-alcohol series, respectively (10). The same values reported in Table III Vol. 22, March/April 2010 are in the range. The structure and polarity of the elemicin derivatives are quite different than those of alkane derivatives and the effects of an OH or a CH 3 O radicals on the K.I. increments are different in both groups of compounds (See Figure 3). The three methoxy elemicin and isoelemicins are known. They were reported in certain varieties of parsley (8). Methoxyelemicin was reported in Asarum canadense (11) and Asiasarum heterotropoides (12) and in various Asarum species found in Japan (13,14). The two methoxy-isoelemicin isomers were reported in the oil of Pituranthos scoparius from Algeria (15). A biosynthetic route of these molecules was proposed (14). It would appear that the present work is the first report on the hydroxyl elemicin and isoelemicins although the 6-(2-propenyl)-2,3,6-trimethoxyphenol was observed in an ethanol acetate extract of Ocotea cymbarum (16). (E)- and (Z)-3-(1-propenyl)-2,5,6-trimethoxyphenols were also observed in the roots of Smirnowia iranica (17). Table I. Percentage composition of the essential oil of the fresh rhizomes of Asarum canadense L. Component K. I. Peak DB-5 S-wax area % myrcene a-terpinene limonene b-phellandrene (Z)-b-ocimene (E)-b-ocimene g-terpinene terpinolene linalool terpinen-4-ol a-terpineol nerol geraniol linalyl acetate bornyl acetate neryl acetate geranyl acetate methyl eugenol seychellene guaia-6,9-diene methyl isoeugenol allo-aromadendrene germacrene D ar-curcumene asaricin unidentified elemicin unidentified junenol unidentified (E)-isoelemicin selin-11-en-4-a-ol Total ( % ) Although both Adams data bank and MassFinder 3 give good fits for the furopelargone A mass spectrum, the DB-5 R.I. values do not agree : 1540 (5) and 1517 (6); 2 105(100), 43 (99), 161(95), 81(84); 121(72), 204(155), 189(67), 133(63), 91(58), 93(57); 55(52), 41(50), 80(48), ; 3 43(100), 81(90), 161(59), 105(52), 204(47), 93(47), 121(47), 41(41), 92(40), 135(32), 189(32), 136(31), 95(29), Mw = 222?; Also see lower right MS on Figure 1. Journal of Essential Oil Research/165

3 Collin et al. methoxyelemicin (E)-methoxyisoelemicin hydroxyelemicin (Z)-hydroxyisoelemicin (E)-hydroxyisoelemicin unknown + (Z)-methoxyisoelemicin Figure 1. Recorded 70 ev- cracking Mass spectra of elemicin and isoelemicin derivatives obtained from the essential oil of the roots of Asarum canadense. On the lower right spectrum, one can see the 238 and 223 peaks attributed to the (Z)- methoxyisoelemicin molecule. MS recorded in the tail of an unidentified compound (R.I. DB-5: 1617). Acknowledgments The authors thank France Boudreau for her technical assistance. References 1. a. M.G. Motto and N.J. Secord, Composition of the essential oil from Asarum canadense (Aristolochiaceae). J. Agric. Food Chem., 33, (1985). b. B.M. Lawrence, Progress in Essential Oils Snakeroot oil. Perfum. Flavor., 11(6), (Dec. 1986/Jan. 1987). 2. T. Iwashina and J. Kitajima, Chalcone and flavonol glycosides from Asarum canadense (Aristolochiaceae). Phytochemistry, 55, (2000). 3. B.T. Schaneberg, W.L. Applequist and I.A. Khan, Determination of aristolochic acid I and II in North American species of Asarum and Aristolochia. Pharmazie, 57, (2002). 4. National Bureau of Standards. 75k NBS data bank, Washington, DC (2003). 5. R.P. Adams, Identification of Essential Oil Components by Gas Chromatography/Mass Spectrometry. 4 th Edn., Allured Publ. Corp., Carol Stream, IL (2007). 6. D.H. Hochmuth, MassFinder 3/Terpenoids Library. ( com), Hamburg, Germany (2004). 166/Journal of Essential Oil Research Vol. 22, March/April 2010

4 A. canadense Figure 2. Partial 208, 238, and 224 ions chromatogram (DB-5 column) of the essential oil of the rhizomes of Asarum canadense elemicin methoxyelemicin (E)-methoxyisoelemicin (Z)-isoelemicin (E)-isoelemicin (Z)-methoxyisoelemicin hydroxyelemicin (Z)-hydroxyisoelemicin (E)-hydroxyisoelemicin Vol. 22, March/April 2010 Journal of Essential Oil Research/167

5 Collin et al. Table II. Percentage composition of the essential oil of the roots of Asarum canadense L. Component K. I. Peak DB-5 S-wax area % a-phellandrene p-cymene Tr. limonene Tr. b-phellandrene ,8-cineole Tr. terpinolene linalool terpinen-4-ol a-terpineol methyl thymol linalyl acetate bornyl acetate geranyl acetate bicyclo-4(15)-oppositene methyl eugenol cascarilladiene opposita-4(15),7-diene a-guaiene trans-a-bergamotene seychellene guaia-6,9-diene (Z)-methyl isoeugenol a-humulene a-patchoulene (E)-b-farnesene a-amorphene germacrene D ar-curcumene g-muurolene d-selinene zingiberene (E)-methyl isoeugenol Component K. I. Peak DB-5 S-wax area % a-bulnesene d-amorphene germacrene A 1512 Tr. b-bisabolene a-bulnesene b-sesquiphellandrene elemicin germacrene B ,2,3,4-tetramethoxy-5-(2- propenyl)benzene b : see text unidentified c (Z)-isoelemicin trans-dracunculifoliol ,3,4-trimethoxy-6-(2- propenyl)phenol d : see text junenol epi-junenol unidentified (see note 1, Table I) (Z)-1,2,3,4-tetramethoxy-5-(1- propenyl)benzene e ~ (E)-isoelemicin patchoulol selin-11-en-4-a-ol a-asarone 1669 Tr. (Z)-2,3,4-trimethoxy-6-(1- propenyl)phenol f : see text Tr. (E)-1,2,3,4-tetramethoxy-5-(1- propenyl)benzene g juniper camphor (E)-2,3,4-trimethoxy-6-(1- propenyl)phenol h : see text Total (%) 84.8 Tr. = trace (> 0.01); b methoxy-elemicin; c see footnote 2 in Table I; d hydroxyelemicin; e (Z)-methoxy-isoelemicin; f (Z)-hydroxy-isoelemicin; g (E)- methoxy-isoelemicin; h (E)-hydroxy-isoelemicin. Table III. The effect of methoxy or hydroxy substitution on the K.I. values of the three elemicin isomers Column elemicin (Z)-isoelemicin (E)-isoelemicin Mean d or K.I. d K.I. d K.I. d D value Swax DB D ± 30 methoxy addition Swax ± 10 DB ± 14 D ± 16 hydroxy addition Swax ± 10 DB ± 10 D ± 10 D (Kovat s units) = Swax-10 Kovat s index DB-5 Kovat s index; d (Kovat s units) = observed Kovat s index - Kovat s index of the parent elemicin isomer on the same column. 7. E. Guenther, The Essential Oils. Vol. 6, pp , Krieger Publ. Co., Malabar, FL (1952). 8. E. Stahl and H. Jork, Chemische rassan bei arzneipflanzen I. Mitt.: Untersuchung der kulturvarietäten europäischer petersilienherkünfte. Arch. Pharm., 297, (1964). 9. W. Jennings and T. Shibamoto, Qualitative Analysis of Flavor and Fragrance Volatiles by Glass Capillary Gas Chromatography. Academic Press, New York, NY (1980). 10. H. Gagnon, F.-I. Jean and G. Collin, La chromatographie gazeuse sur colonnes capillaires (CG) Usages des indices de Kovats (I.K.). 3rd 168/Journal of Essential Oil Research Vol. 22, March/April 2010

6 A. canadense Figure 3. Molecular structures of elemicin, (Z)- and (E)-isoelemicins and their proposed monomethoxylated and monohydroxylated derivatives Colloquium on Natural Products from Vegetal Origin. pp , Saint- Jean-sur Richelieu 1995, Oct , Univ. Quebec. Chicoutimi, Canada (1996). 11. L. Bauer, C.L. Bell, J.E. Gearien and H. Takeda, Constituents of the rhizome of Asarum canadense. J. Pharm. Sci., 56, (1967). 12. Y. Saiki, Y. Akahori, M. Tadataka, K. Morinaga, T. Taira, S. Fukushima and T. Harada, Gas chromatography of natural volatile oils II. Gas chromatography of the volatile oils of plants belonging to Asiasarum and Asarum genera. Yakugaku Zasshi 87, (1967). 13. N. Hayashi, M. Solichin, T. Sakao, Y. Yamamura and H. Komae, An approach to chemotaxanomy of the Asarum subgenus heterotropa. Biochem. Syst. Ecol., 8, (1980). 14. N. Hayashi, J. Ding, Z. Ding, Z. Chen, Y. Yi and H. Komae, Volatile constituents of the essential oils of four Chinese species in the genus Asarum (Aristolochiaceae). Z. Naturforsch., 45c, (1990). 15. G. Vernin, C. Lageot, C. Ghiglione, M. Dahia and C. Parkanyi, GC/MS analysis of the volatile constituents of the essential oils of Pituranthos scoparius (Coss et Dur.) Benth. et Hook. from Algeria. J. Essent. Oil Res., 11, (1999). 16. C.C. Andrei, R. Braz-Filho and O.R. Gottlieb, Allylphenols from Ocotea cymbarum. Phytochemistry, 27, (1988). 17. M. Lambert, D. Staerk, S.H. Hansen, M. Sairafianpour and J.W. Jaroszewski, Rapid extract dereplication using HPLC-SPE-NMR: analysis of isoflavonoids from Smirnowia iranica. J. Nat. Prod., 68, (2005). Vol. 22, March/April 2010 Journal of Essential Oil Research/169