Comparison of Microwave-Assisted Hydrodistillation and Hydrodistillation Methods for the Fruit Essential Oils of Foeniculum vulgare
Microwave-assisted hydrodistillation (MWHD) and hydrodistillation (HD) were carried out for the analysis of volatile components in whole and ground fruits of Foeniculum vulgare Miller (fennel). Fruits were distilled using a microwave oven modified to fit a Clevenger-type apparatus. The effect of microwave energy on the yield and composition of the essential oil was investigated against the classical hydrodistillation. All the essential oils were analyzed by GC-FID and GC/MS. (E)-anethole was found as the main compound in the oils of both whole and ground materials (82.2-86.8%) using die two methods. Methyl chavicol (4.0-4.9%) and limonene (2.2-4.9%) were also found in fennel oils obtained by HD and MWHD. The amounts of identified components in the oils obtained from whole and ground fennel fruits were not affected significantly by microwave energy except for limonene.
Key Word Index
Foeniculum vulgare, Apiaceae, fennel, microwave-assisted hydrodistillation, hydrodistillation, essential oil composition, (E)-anethole.
The fruits oiFoeniculum vulgare Miller (Fennel) (Apiaceae) are used as a spice in Europe, Northern Africa, West, Central and South Asia, as well as Turkey. Fennel fruits are used as appetizers, digestive, sedative and colic (1) and added in breads, fishes, salads and cheeses as a spice (2). Two varieties of fennel are known and reported in the European Pharmacopoeia. Foeniculum vulgare Miller subsp. vulgare var. vulgare known as bitter fennel contains (E)-anetiiole (60%) and fenchone (15%) and F. vulgare Miller subsp. vulgare var. dulce (Miller) Thellung known as sweet fennel contains (E)-anethole (80%) as die main component. The oil of fennel fruits is used as an ingredient of cosmetic and pharmaceutical products for its balsamic, cardiotonic, digestive, lactagogue and tonic properties. The composition of die oil of fennel has been reported. (E)-Anediole, fenchone (present in bitter varieties), estragóle (metìiyl chavicol), limonene, a- pinene, and a- phellandrene were found as the main components (2-5). The amount of methyl chavicol and the ratio of (E)-anethole and methyl chavicol vary between the two types. The carcinogenic effect of methyl chavicol has been reported. Scientific committee on food of European Commission reported its opinions on methyl chavicol in 2001 (6). In this report, chemical characterization, exposure assessment, metabolism, and toxicity of mediyl chavicol were given. According to this report, limited widi respect to the standard long-term bio-assays, have shown diat mediyl chavicol is a weak inducer of hepatocarcinogenicity in mice treated orally, by i.p. or s.c. injection. The induction of liver tumours seems to depend on formation of l’-hydroxymetabolites. Metabolic studies indicate that in the high dose range of carcinogenicity studies (150-600 mg/kg bw) the production of G-hydroxy-methyl chavicol, expressed as percentage of die dose, is about 5-10 times higher dian that at lower doses (0.05-50 mg/kg bw). l’-Hydroxyestragole has been found also in the urine of men dosed with 100 µg estragole/day for six months. Mediyl chavicol occurs naturally in a variety of foods including tarragon (60-75%), sweet basil (20-43%), sweet fennel (5-20%), green anise (1%), and star anise (5-6%). There are 28 food categories identified by industry to which mediyl chavicol can be added. For these food categories, a concentration of 10 mg methyl chavicol/kg food was assumed for food in general and a concentration of 50 mg/kg for food containing herbs and spices. For alcoholic beverages, canned fish and fats and oils, die following concentrations were applied as specified by die Council of Europe (2001): alcoholic beverages 100 mg/kg; canned fish 50 mg/kg; fats and oils 250 mg/kg (6).
Fennel oils are obtained by distillation of dried fruits . There are several reports on their composition (2-5). Conventional hydrodistillation (HD) using modified Clevenger-type apparatus is widely used to obtain the essential oil from plants for scientific investigations and for oil yield determinations. It involves distillation of die plant material in water for 3 h. Microwave-assisted mediods have been used increasingly in die last few years especially for extraction (7-12). Microwave-assisted hydrodistillation (MWHD) metiiod is amore recent technique used to recover volatile components (13-17). In this method, plant material placed in a Clevenger-type apparatus is heated inside a microwave oven for a short period of time to extract the essential oil. Heat is produced by microwave energy. The sample reaches its boiling point very rapidly, leading to a very short extraction or distillation time (7-17). Widi the microwave distillation technique it is possible to achieve distillation with the indigenous water of the fresh plant material.
The aim of diis study was to provide a rigorous comparison between microwave-assisted hydrodistillation and classical hydrodistiUation techniques to obtain essential oil from fennel fruits. Both whole and ground fruits were evaluated from the point of view for the effect of microwave energy on oil composition.
Plant material and reagents: The ripe fruits of Foeniculum vulgare Miller were obtained from Türer Ltd. Sti., (Izmir, Turkey). Distilled water was used for the distillation of plant materials.
HydrodistiUation (HD): 100 gplant material and 1000 mL distilled water were placed in a 2000 mL round-bottom flask and connected to a Clevenger-type apparatus. HydrodistiUation was performed for 3 h after boiling. Oil yields obtained from die experiments were calculated on moisture free basis.
Microwave-assisted hydrodistiUation (MWHD): The oils were obtained from whole and ground fennel fruits by hydrodistillation for 60 min using a Clevenger-type apparatus placed in a modified microwave oven (Milestone ETHOS E Microwave Labstation). During distillation, time, temperature, pressure and power were monitored and controlled with the “easy-CONTROL” software package of the system. Microwave power applied to die plant material was controlled by a shielded thermocouple inserted directly into the flask. The oven was operated for 10 min at 800 Watts up to 1000C, and dien kept at 1000C for 50 min at 500 Watts followed by 5 min of ventilation. Oil yields obtained from the experiments were calculated on moisture free basis.
Gas Chromatography (GC): The oils were analysed by GC using a Hewlett Packard HP6890 system with Innowax FSC column (60 m × 0.25 mm ∅, with 0.25 µm film thickness ). Nitrogen (from 1.2 to 0.9 mL/min ramp flow) was used as carrier gas. The GC oven temperature was kept at 6O0C for 10 min and programmed to 2200C at a rate of 4°C /min and then kept constant at 2200C for 10 min and programmed to 240°C at a rate of 1 0C /min. Split flow was adjusted at 12 mL/min widi 10:1 split ratio. The injector and detector temperatures were adjusted at 250°C. The relative percentage amounts of die separated compounds were calculated from FID chromatograms.
Gas Chromatography/Mass Spectrometry (GC/MS): The oils were analyzed using a Hewlett Packard G1800A GCD system widi same column and analysis parameters of GC to obtain same chromatographic separation. Helium (0.8 mL/min) was used as carrier gas. The mass range was recorded from m/z 35 to 425. The split ratio and the injector temperature were adjusted as 50:1 and 250°C respectively. MS were recorded at 70 eV. Alkanes were used as reference points in the calculation of relative retention indices (RM).
Identification of compounds: The components of die ods were identified by comparison of their mass spectra with tiiose of Baser Library of Essential Oil Constituents, Wiley GC/MS Library, Adams Library, Mass Finder 3 Library and confirmed by comparison of tiieir retention indices.
Results and Discussion
The whole and ground fruits of fennel were distilled both by HD and MWHD methods. All the samples were distilled using a Clevenger-type apparatus for 3 h in heating mande; and for 60 min in the modified microwave oven, resp. The yields of oils calculated on moisture free basis are shown in Table I. The oil yields of whole and ground fruits were found to be similar in HD. However, microwave energy appeared to increase (30%) die yield of oil in ground fennel samples (Table I). In microwave-assisted hydrodistiUation, distillation time was shorter dian classical hydrodistiUation and also the sample reached boiling stage more rapidly. This is an advantage of MWHD when it is compared to classical HD.
The oils were analyzed by GC-FID and GC/MS. Relative percentages of die characterized compounds calculated by FID integrator are shown in Table I. (E)-Anetiiole, estragóle, limonene were found as main compounds in fennel fruit oils. A small amount of a-fenchone was detected in all the oils. According to European Pharmacopoeia, the amount of (E)-anethole has to be more than 80% in oils of die sweet variety (2). The percentages (Table I) of volatile secondary compounds from one experiments in fennel oils were obtained by HD and MWHD varied slightiy by grinding the material. The only exception was limonene (Table I). The percentage of limonene in the oil from ground fruits increased almost twice using microwave energy. Same results were also obtained from other limonene containing plants in our previous researches by MWHD (18,19). The effect of grinding on die isolation of limonene within the Laser trilobum oil was investigated previously by our group (20). Limonene contents were found higher in crushed fruits than in whole fruits (20). This can be explained by the fact that the oxygenated compounds of the oil in whole fruit, which are generally less volatile but have more affinity for water, appear earlier in the distillate tìian do the hydrocarbons which, although more volatile, have less affinity for water. Furthermore, hot water softens and penetrates die fruits. The oxygenated components, which have more affinity for water, diffuse to die surface of the material during distillation and are removed by the passing steam. The diffusion proceeds at die same diminishing rate as the od containing oxygenated components inside die fruit declines (20). In this study, similar results were obtained for limonene (Table I): it can be concluded that microwave energy was not enough to heat the material long enough for extracting limonene from the whole fruits. On die contrary, in HD, die amount of limonene was not affected by grinding, because the distillation time was three times longer than the MWHD.
Accordingto pharmacopoeias, die amounts of (E)-anetìiole, mediyl chavicol and α-fenchone and their ratio are important for the quality assessment of fennel fruits. The percentages of these components were almost die same in the fennel oils obtained by both techniques. But, HD takes 3 h to complete while 1 h is enough for a complete distillation by the microwave technique. The results have shown that diere is slight differ- enee in oil yields by both techniques and similar oil profiles are obtained. MWHD appeared to yield more oxygenated components than terpenic hydrocarbons. This is an advantage in die production of olfactively more appealing oils. MWHD is still in its infancy since there are only a few papers published. More research may prove it to be an advantageous technique over conventional distillation also for industrial applications.
Authors are grateful to AUBIBAM for oil analysis support.
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Müberra Kosar,* Temei Özek, Mine Kürkcüoglu and K. Hüsnü Can Baser,
Faculty of Pharmacy, Department of Pharmacognosy, Anadolu University, 26470 Eskisehir, Turkey
“Address for correspondence
Received: March 2006
Revised: June 2006
Accepted: July 2006
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