A Study of the Mediterranean Oregano Populations. Chemical Composition of Essential Oils of Origanum ehrenbergii Boiss. from Two Populations in Lebanon
Abstract
Seedlings obtained from seeds of three populations of oregano (Origanum ehrenbergii) harvested in Lebanon and sown at a research station in France were planted in three replicates in a Fischer block design. oils produced by water distillation were subjected to analysis by GC and GC/MS. The oils of one of the populations were uniform and carvacrol-rich. The other population displayed a variable chemical composition, including thymol, thymol-carvacrol and p-cymene/y-terpinene chemotypes.
Key Word Index
Origanum ehrenbergii, Lamiaceae, essential oil composition, thymol, carvacrol, γ-terpinene.
Introduction
Origanum ehrenbergii belongs to the Prolaticorolla section of Group C in the classification of letswaart (1). Although many species belonging to the genus Origanum have been studied by Baser and Duman (2-6), Arnold et al. (7), Valentini et al. (8), Skoula et al. (9) and Melegari et al. (10), no work has been done on the chemical composition of O. ehrenbergii. However, O. compactum Benth. and O. laevigatum Boiss. from the same group have been studied by Charai et al. (11), Baser et al. (12) and Tucker and Maciarello (13), respectively. Origanum compactum was found to contain carvacrol (56%), γ-terpinene (15%) andp-cymene (9%), while O. laevigatum was found to be rich in sesquiterpenes, principally bicyclogennacrene (25-38%), germacrene D (20-21%) and β-caryophyllene (17-14%).
Origanum ehrenbergii is endemic in Lebanon where it grows at an altitude of up to 1500 m. It occurs on dry limestonebased stony scrubland and in open pinewood understories in acid soils. It is used as a condiment, although the less strongly flavored O. syriacum L. is preferred.
Origanum ehrenbergii as a crop is similar to the ordinary oregano found in France. It grows rapidly, but its yield of dry matter is moderate. It is a prostrate plant that sends out numerous floral scapes and blooms abundantly.
The work reported here was carried out on batches of seeds from three distinct populations located in Lebanon, grown at the CNPPMAI facility at Milly-La-Forêt in 1999, 2000 and 2001.
Experimental
Plant material: The seeds collected in 1998 from three sample populations designated 98-011, 98-012 and 98-034 located near Aaquoura, Choueir and Bolonia in Lebanon were grown first in 1999 and disposed in separate plots at CNPP-MAI, with three repeats each. Plants were harvested in 1999 and 2000. New plantings were carried out in 2000 for further studies in 2000 and 2001. The 1999 harvest comprised only enough material for a pooled study of each population, but the 2000 and 2001 harvests allowed a plant-by-plant study.
The plant was harvested in full flower, dried away from light and isolated by use of a Clevenger-type water distillation unit for 3 h. Specimens of each were kept at CNPPMAI.
GC: The gas phase chromatography analysis was carried out on a Delsi DI 200 instrument equipped with a flame ionization detector and a DBS column (25 m x 0.25 mm, df: 0.25 µm) with a split flow rate of 60 mL/inin, nitrogen as carrier gas and temperature programming (5 min at 50°C and 3°C/min up to 220°C), injector temperature 220°C and detector temperature 235°C. Quantitative data were obtained from FID area percents without the use of correction factors.
GC/MS: The oils were analyzed on a Hewlett-Packard gas chromatograph Model 6890 coupled to a Hewlett-Packard MS Model 5873 equipped with an HP5 column (30 m x 0.25 mm, df: 0.25 µm) programmed from 50°C (5 min) to 300°C at 5°C/ min, and 5 min hold. The carrier gas was helium (1 mL/min), injection in split mode (1/10); injector and detector temperatures 250°C and 280°C, respectively. The MS ran in electron impact mode at 70 eV, electron multiplier 2200 V, ion source temperature 230°C. Mass spectral data were acquired in the scan mode in the m/z range 33-450.
Identification was carried out by calculating retention indices and comparing mass spectra with those in data banks; personal, Adams (14) and McLafferty and Stauffer (15).
Results and Discussion
The yields of dry matter were similar for all three batches, at around 15 g/plant. The essential oil contents were about 3.6 mL/100g of dry matter, giving an average oil yield of 0.47 mL/plant. GC and GC/MS analysis identified 63 constituents, of which 28 were quantified, the others being present only in trace amounts (
Sample 98-011 was carvacrol-rich (71.5-78.5%) for the three replicates, which were thus fairly uniform. Some γ-terpinene (2.6-6%) and p-cymene (5-7%) were also found. Thymol was present in only trace amounts (0.9-1.3%). The second and third samples (98-012 and 93-034) differed from the first and were highly heterogeneous. Two repeats (98-012) were rich in thymol/carvacrol (35.2-38.3% / 21.2-22.5%) accompanied byyterpinene (14.5-2.1%) and p-cymene (10.6-8.9%), three repeats (one 98-012, two 98-034) were thymol-rich (52.4-59.2%) with very small amounts of carvacrol 1.6-4.9%, and γ-terpinene and p-cymene comprising 11,1-14.7% and 5.5-10.6%. The last 93034 repeat stood out by its high thymol content (28.5%), usual γ-terpinene content (10.6%) and particularly large amounts of p-cymene (33.1%). This population study of O. ehrenbergii revealed four chemotypes illustrated by histograms shown in Figure 1. These results compared with those obtained forother species in the same group and section differed markedly from those obtained for O. laevigatum, which were sesquiterpene-rich. The oil from the carvacrol chemotype in 98-011 was chemically similar to that analyzed by Charai et al. (11).
The overall compositions obtained on repeats for each of the three samples were pooled averages, and may have hidden more specific compositions. We therefore isolated oils from the most prolific individual plants. We studied 34 plants in 2000, six of 98-011 and three of 98-012 in 2001, 23 plants of 98-011 and two new individuals planted in 2000. The analytical results are given in Table II.
The 23 individuals of 98-011, planted in 1999, survived two winters and proved hardy. Most of the oils had the same composition, with carvacrol as the main constituent (78.2-87.6%). Five other individual plants differed notably in their linalool contents (7.9-16.4%), and contained less carvacrol (7.9-16.4%). One plant had an unusual thymol content (13.5%). Linalool was present only in traces in the pooled samples, where linalool-rich individuals were presumably few in number (Figure 1).
The few individuals studied in 2000 and planted in 1999 (98-012) or 2000 (98-068) showed that these samples were especially sensitive to harsh climate. There was no 98-034 material. The five individuals studied were all of different composition (Barchart 5); four (98-012) were all thymol-rich to different extents (40.9-72.9%), with correspondingly ranging complementary carvacrol (2.9-24.6%) and p-cymene contents (16.7-15.5%), γ-terpinene content ranging from 1.1-11.5%. The last individual displayed a more marked predominance of carvacrol (55.1%) with thymol at 13.1%, p-cymene at 9.5% and γ-terpinene at 2.8%.
A carvacrol chemotype (Figure 1) was thus clearly defined in both the pooled study of repeats and in individual plants. Thymol-rich, thymol/carvacrol andthymol/carvacrol/p-cymene/ γ-terpinene chemotypes (Figure 1) were represented by only a few individuals. The most abundant (carvacrol) chemotype is thus readily available, but the others would require further selective breeding before they could be exploited.
References
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