Abstract
The chemical compositions of the essential oils of Seseli peucedanoides (M.Bieb.) Koso-Pol. inflorescences and leaves, isolated by hydrodistillation, and headspace volatiles, obtained by the static headspace method, were analyzed in detail by gas chromatography (GC) and GC/mass spectrometry (MS). In total, 74 constituents were identified, representing more than 98% of the observed GC peaks. The number of identified essential oil components obtained from the inflorescences was 63 while for the leaf essential oil it was 46. A much smaller number of compounds, 26 for inflorescences and 21 for leaves, were detected in the headspace samples. In both essential oils the most abundant compounds were the same, (E)-caryophyllene and germacrene D, only in different proportions. The main components in the headspace specimens were α-pinene and (E)-β-ocimene with (E)-caryophyllene and (Z)-3-hexen-1-ol also determined in significant percentages. The major class of compounds identified in the investigated essential oils was hydrocarbon sesquiterpenes with a share of over 80%, while the most dominant class of the headspace volatiles was hydrocarbon monoterpenes, which contribute slightly less than 80% of the total.
The Apiaceae Lindley (Umbeliferae Juss.) is a well-known family of flowering plants, containing about 450 genera and 3700 species throughout the world and distributed primarily in temperate regions of the northern hemisphere. 1 One of the largest genera in this family is the genus Seseli L. which comprises up to 140 species. 2 Seseli is an old Greek name that was used by Hippocrates for certain members of the Apiaceae family and several species of the genus are reported in ancient literature for various healing effects. 3 Genus Seseli is widely distributed in the Irano-Turanian, Euro-Siberian, and East Mediterranean geographical region and contributes 10 species to the flora of Serbia. 4 One of them is Seseli peucedanoides (M.Bieb.) Koso-Pol. This is a perennial green or gray-green plant with large spindle-shaped roots and vertical stems branched in the form of a shield, up to 70 cm high. Leaves are linear, twice pinnate shared with petiole expanded in leaf sheath. Flowers are small and yellow, arranged in an umbelliform inflorescence.
On the basis of previously conducted chemical studies, it is known that Seseli species are a source of coumarins, sesquiterpenes, and phenyl propanoides. 5-9 Previous analyses of the essential oils from various Seseli species demonstrated that the main components are quite different from species to species: sabinene, α-pinene, and β-phellandrene in S. globiferum aerial parts 10 ; α-pinene in S. rigidum Waldst. & Kit. aerial parts and fruits 11 as well as in S. pallasi stems and fruits while n-nonane and (Z)-β-ocimene were the main compounds in the roots 12 ; germacrene D and sabinene in S. annuum and S. gummiferum Boiss. & Heldr. subsp. corymbosum aerial parts 13,14 ; β-pinene and germacrene D in S. montanum subsp. tommasinii. 15 But, to date, there is only one report regarding S. peucedanoides (M.Bieb.) Koso-Pol. essential oil with respect to its compositional analysis (and the oil obtained only from the air-dried aerial parts of the plant). 16
To the best of the authors’ knowledge, this study is the first report on the chemical composition of the essential oils and volatiles obtained from fresh vegetative parts (inflorescences and leaves) of S. peucedanoides, accomplished by gas chromatography (GC) and GC/mass spectrometry (MS) analysis of the essential oils and headspace analysis of the corresponding plant parts volatiles.
Table 1 lists the identified volatile constituents, their retention indices, and relative percentages in the oils and headspace specimens of the investigated taxa. The analyses allowed the identification of 74 compounds in total: 63 for the inflorescences (OI) (amounting 98.9% of the total oil), 46 for the leaves (OL) (98.7% of the oil), and much less components for the headspace samples of S. peucedanoides, 26 for inflorescences (HSI) and 21 for leaves (HSL) (99.3% and 99.7% of the total components detected, respectively). In both essential oils the main compounds were the same, (E)-caryophyllene and germacrene D, but in different proportions, 33.3% and 24.2% in inflorescences versus 12.5% and 49.6% in the leaf oil, respectively. Other abundant components (with the yield over 4.0%) from OI were δ-cadinene (9.9%) and (E)-β-ocimene (4.4%), while from OL δ-cadinene (6.3%), (E,E)-α-farnesene (5.9%), and bicyclogermacrene (5.4%) were determined in higher percentage. The major class of compounds identified in oils was hydrocarbon sesquiterpenes, with a share of 84.6% (OI) and 87.1% (OL). On the other hand, the most abundant components in the headspace samples were α-pinene (40.3% and 47.3%) and (E)-β-ocimene (17.3% and 15.1%) in the inflorescences and leaves, respectively. Other compounds present in significant amounts in HSI were (E)-caryophyllene (16.3%) while in HSL they were (Z)-3-hexen-1-ol (14.7%), myrcene (4.1%), and (Z)-β-ocimene (4.0%). As could be expected, due to the applied technique (static headspace), the predominant class of compounds in both HSI and HSL were hydrocarbon monoterpenes (lower boiling points) with the share of 79.7% and 78.0%, respectively.
Chemical Composition (%) of Seseli peucedanoides Essential Oils and HS Volatiles.
Compounds listed in order of elution on a HP-5 MS column. RI, experimentally determined retention indices on the mentioned column by co-injection of a homologous series of n-alkanes C8-C32; AI, Adams retention indices; *, identified by comparison of those mass spectra with spectra in the NIST library as well as with retention indices obtained on HP-5 MS column; tr, trace (< 0.05%); -, not detected. Essential oil samples: OI, inflorescences; OL, leaves. Headspace samples: HSI, inflorescences; HSL, leaves.
Comparing the main constituents of the essential oils from the aerial parts of S. peucedanoides with other previously investigated species of Seseli genus, it can be noticed that there are significant differences in the main constituent composition between different species.
It seems that the basic similarity between the essential oils from the aerial part obtained from various taxa of the Seseli genus is the occurrence of α-pinene as main constituent, while the presence of other constituents varies between different species. α-Pinene (35.9%), sabinene (8.8%), (E)-sesquilavandulol (8.4%), and β-pinene (7.0%) are the main constituents of herbal parts of S. tortuosum grown in Turkey. 17 In the Balkan endemic species S. rigidum the volatile oil from aerial parts and fruit also contains α-pinene as the predominant component (57.4% and 23.3%, respectively). In the aerial parts of the same species, limonene, camphene, and sabinene are present in high amounts, while in the fruit oil, β-phellandrene and sabinene were found in significant quantities. 18 The main components of the essential oil from S. campestre aerial parts are α-pinene (38.6%), β-pinene (17.5%), and (E)-sesquilavandulol (10.3%). 19 On the other hand, there are many reports regarding some other species of Seseli genus that have different main constituents of the essential oils from the aerial parts. Marongui reported that the major compounds in the essential oil from leaves of S. bocconi obtained from different areas of Sardinia were himachalol, sabinene, β-phellandrene, α-humulene, and γ-himachalene. 20 The essential oil from the fruit of S. petraeum from Turkey contained a high level of carotol (20.7%), γ-terpinene (11.3%), sabinene (9.5%), and germacrene D (7.8%) while dominant components of the essential oil from the fruit of S. andronakii were carotol (52.7%) and germacrene D (8.7%). 21 Therefore, it can be concluded that the results obtained on the composition of the essential oil from S. peucedanoides inflorescences and leaves do not have any similarities with the previously published results both in terms of the main components and the prevalent classes of compounds.
The most confusing fact is the complete difference in the essential oil composition of S. peucedanoides in this study from the previous report. 16 Bulatović et al. found that major class of compounds in air-dried S. peucedanoides aboveground parts oil was hydrocarbon monoterpenes (~90%), while dominant components were α-pinene (69.4%), β-pinene (4.9%), and limonene (4.6%). These components were not even detected in the leaves oil while in the inflorescences oil they were present only as minor contributors (1.2%, 0.1%, and 0.8%, respectively). Furthermore, the most abundant compounds from both OI and OL, (E)-caryophyllene and germacrene D, in the previous survey were found only in small amounts or were not detected at all. Possible explanations for the above-mentioned differences are the influence of plant drying, environmental factors (climate and soil type), genetic variability of the investigated populations, and/or the interaction between genotype and environment.
Experimental
Plant Material
Seseli peucedanoides (M.Bieb.) Koso-Pol. was harvested in June 2017, Radan Mt, Serbia (43.025864, 21.550523), altitude 748 m, in full flowering phenophase. The plant material was collected and identified by Bojan Zlatković and the voucher specimen was deposited in the Herbarium Moesiacum Niš (HMN), Department of Biology and Ecology, Faculty of Science and Mathematics, University of Niš, under the acquisition number 13315. The plant was carefully divided into parts: inflorescences and leaves.
Essential Oil Isolation and HS Sample Preparation
The fresh samples were hydrodistilled for 2.5 hours using a Clevenger-type apparatus to produce a small amount (<0.05%, w/w) of oil, which was trapped in n-hexane. The oils were stored at 4°C in the dark until analyzed. For static headspace experiments, 500 mg of milled fresh inflorescences and leaves were put separately into 20 mL HS vials then each soaked with 2 mL of distilled water. The samples were heated at 80°C for 20 minutes with the next mixing program: shaking for 5 seconds, pause for 2 seconds; 500 µL of vapor generated from the specimens was drawn out from the vial using a gas-tight syringe (90°C) and injected directly into the chromatographic column.
Gas chromatography and GC/MS Analyses
The GC/MS analyses of the volatiles were carried out (three repetitions for each sample) in an Agilent Technologies 7890A gas chromatograph equipped with a fused silica capillary column HP-5 MS (5% phenyl methyl siloxane, 250 µm × 25 m, film thickness 0.25 µm, Agilent Technologies, Santa Clara, CA, USA) and coupled with a 7000 MS/MS triple quadrupole system, operating in the MS1 scan mode, of the same company. The GC was operated under the following conditions: injector and interface temperature was 230°C and 280°C, respectively; oven temperature was programmed from isothermal at 50°C for 2.25 minutes, raised to 290°C at a rate of 4°C/min. As a carrier gas, helium at 1.0 mL/min was used, at constant flow mode. Exact 1 µL of the oil solution in n-hexane (1:100) was injected at split ratio 40:1. Electron impact mass detector was operated at the ionization energy of 70 eV, in the 40 to 440 amu range, with scanning speed of 0.32 seconds. For GC (flame ionization detection; FID) analyses, the same column and chromatographic conditions were applied as described for GC/MS. FID detector temperature was 300°C. The percentage amounts of the separated compounds were calculated from the GC peak areas using the normalization method without correction factors. The data are reported as mean values of three sample injections.
Identification of Volatile Compounds
Qualitative analysis of the constituents was based on the comparison of their linear retention indices (relative to retention times of C8-C32 n-alkanes on a HP-5 MS column) with literature values and their mass spectra with those from Adams, Wiley 6, NIST 11, Essential oils libraries of known essential oils, applied on Agilent Mass Hunter Workstation (B.06.00) and AMDIS (2.1, DTRA/NIST, 2011).
Footnotes
Acknowledgments
The authors owe a special gratitude to Bojan Zlatković, Department of Biology and Ecology, Faculty of Science and Mathematics, University of Niš.
Declaration of Conflicting Interests
The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
Funding
The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: This study was supported by Ministry of Education, Science and Technological Development of Serbia (Project No. 172047).