Messrs. Christos HASSIOTIS */ & Pavlos EFTHYMIOU M**/
*/ Department of Forestry and Environmental Management, Dimokreitian University of Trace, ORESTIADA
**/ Department of Forestry & Natural Environment. Aristotle University, THESSALONIKI
(Not available in French & Russian)
Across the Mediterranean region aromatic plant species growing naturally. These species are producing essential oils and comprise an important economical indicator for the population. All aromatic plants emit volatile substances into the environment either during their life or during the decomposition process. These volatile constituents affect the top soil microflora and form specific environmental conditions for the recycle process. The aim of this study was to investigate the essential oil degradation and the release rate of the main compounds of the oil of Laurus nobilis during decomposition process.
Key words: Laurus nobilis, Essential oil degradation, Mediterranean region, Decomposition process
One of the major plant categories growing in the Mediterranean region is that of aromatic plants which contain essential oils. It is obvious that essential oils do not survive forever in plant material and the fate of the oil following leaf fall is a question of interest.
Almost all plants, and mainly the aromatic ones, emit volatile substances. The oil represents 1–4% of the dry weight, and there are several ways that these secondary metabolites escape into the environment. According to Margaris and Vokou (1986) the terpenoid emissions participate in photochemical reactions leading to aerosol production. Plant essential oils are complex mixtures of different compounds, and their constituents are insoluble or almost insoluble in water. The oil constituents are influenced by the geographical location of the plant (Rhyu, 1979).
Essential oil decomposition can be occurred even when the plant put into a dark place. As Sombrero (1992) stated, the percentage of oil in the leaves showed a gradual decrease the longer the plant remained in the dark losing approximately 40% of oil in a period of twenty-four days. He also reported that not only the total amount of the oil but also the different oil compounds presented a decrease whereas some others presented an increase (e.g. a-pinene).
The decomposition of the essential oil is achieved by the presence of microorganisms which need to have the enzymatic capacity to break down the organic compounds of the litter. The litter and its components constitute source of carbon and nutrients (Melillo & Aber, 1984). The terpenoids which are characterized by their lability (Knobloch et al., 1989) have been found to interfere with enzymatic reactions of energy metabolism.
The most prominent among the organisms known to attack hydrophobic residues are members of the genus Pseudomonas and Nocardia (Gunsalus and Marshall 1974). It was also found (Stevenson 1967) that Arthrobacter spp. are able to utilize a great number of aromatic hydrocarbons as their sole carbon source. There are a number of reports dealing with the ability of Athrobacter in degrading aromatic structures (Audus & Symonds 1955, Gunderson & Jensen 1956). Similarly a number of Arthrobacter have been shown to produce complex polysaccharides or enzymes involved in the transformation or cleavage of these structures (Cadmus et al. 1963). One hundred and thirty Arthrobacter isolates were tested for their ability to utilize aromatic hydrocarbons as their sole carbon source. Of these organisms, 77% were able to grow on at least two aromatic substrates and many were capable of growing on a wide range of these compounds (Stevenson 1967). The fact that the members of the genus Arthrobacter occur as a major component of the autochthonous8 flora of most soils coupled with the ability to utilize aromatics, suggests that this activity may be the major role of this group of organisms in soil.
Not only the oil but also the organic chemical constituents can affect the litter decay. The constituents of the litter can be divided into three broad groups which begin their net mass loss at different stages of decomposition (Berg et al 1982). The slight water solubility of terpenes would allow some leaching loss of these components. Cineole inhibits respiration (Muller et al. 1969) and alters the anatomy of the roots and seedling cells. As Halligan (1975) reported, camphor and 1,8-cineole were the two most toxic components and contribute to toxicity in the field.
Materials and methods
The target of this study was to investigate the essential oil degradation during decomposition process of Sweet Bay (Laurus nobilis) and how the major compounds are released or decomposed.
The litterbag technique was used to evaluate the litter decay and the fade of the essential oil content and its components. The plant species used for this project was Sweet Bay (Laurus nobilis). Approximately 15g of fresh leaves were added to the plastic bags and sealed with synthetic thread. Three hundred bags were used and each bag had a code number. The bags were put on the ground, by removing the surface vegetation, so that the bags were slightly touching the soil.
The area of the investigation was in the North part of Greece at Chalkidiki peninsula with typical Mediterranean characteristics. Twenty replicates, of fifteen bags were installed in the experimental area. The first sampling was on the 1st May and then periodically, every month. Approximately 1g dw from the litterbags, containing the aromatic plant material, ground in a mill, is put into the capsule and covered by a cotton top. The extraction medium was diethyl ether with 10–4 molar (M) of n-tetradecane to act as an internal standard for any losses. Then after the essential oil analyzed in a Gas Chromatograph. The chromatograph was injected with 2.5 μl of extract for each run. Three replications of each sample were analyzed. Traces and area integration was recorded on an automatic Pye Unicam PU 4810 computing integrator. It was programmed to record and calculate the percentage area of each peak, although ignoring the solvent peak. The extracts were analyzed by GC using the following pure compounds as markers; α-pinene, 1.8 cineole, limonene, fencone, linalol, camphor and eugenol. All data have been tested for statistically important differences with analysis of variance test (ANOVA).
Results and discussion
During the period of the investigation the essential oil content (table 1) of buried Laurus nobilis present a remarkable decrease. A significant fade also observed in the main compounds of the oil (table 2).
8 Autochtonous is an organism type which lives naturally in a certain area.
Table 1. The Percentage of the oil derived from the buried plant material is shown. The plant material was buried from April of 94 to November of 95.
|Date||Oil %||Fade %|
Table 2. The percentage of the main compounds found in the of buried Laurus nobilis is shown. The plant material was buried from April of 94 to November of 95.
|DATE||Compounds (%) in the oil concentration|
|a-pinene||sabinene||myrcene||limonene||1–8, cineole||Cam phor||Lina lool||a-terpineol||a-terpinyla catate||Eugenole|
Table 1 shows that there is a great loss of the oil content through the period of investigation. After eighteen months the buried plant material of aromatic Laurus nobilis lost about 76 % of the initial oil content. All the major compounds of the oil of Laurus nobilis decreased during the study and the differences were observed between the dates of samplings were statistically very important.
Figures 1–3. Representation of a chromatograph. In the charts the ten most important compounds found in the oil of Laurus nobilis are presented. In the charts the compounds as well as the percentage values are also reported. Each chart gives the percentage values of different dates from buried plant material.
The major compound is 1–8 cineole.
|Source of Variation||SS||df||MS||Fcritical||F99|
|Compounds between dates of samplings||1242.0||14||88.7||4.9||2.2|
As is seen in the chromatographs after six months the percentage of cineole dropped from 49.9% to 17.8%, of linallol from 14.2% to 3.3%, of ⇋-terpinyl acetate from 12.1% to 0%. These compounds are catabolised easily from the buried leaves. Another important conclusion could be drawn is that the minor compounds or the subproducts of the major compounds are increasing during the study.
Figure 4. Essential oil and major compounds degradation of Laurus nobilis during the period of eighteen months.
The essential oils have been extensively reported to affect the microorganism populations and moreover the decomposition rates. Even 1,8 cineole, which is very toxic to microorganisms and major compound of sweet bay oil can be degradated by microorganisms. It is reported that 1,8 cineole and -terpinyl acetate were actively metabolised in mature leaves with the help of 14CO2. However (personal results) among different plants Laurus nobilis oil had the highest inhibition against a series of microorganisms. 1,8 cineole was the most effective compound against Rhyzopertha dominica, and Oryzaepfilus surinamensis. Similar results were obtained by Kivanc & Akgul (1986). They found that the most toxic oil against Staphylococus aureus and Proteus vulgaris were the oils from Cumin, Laurel and Oregano.
The magnitude of the ability of microorganisms to decompose plant secondary metabolites is reported by Vokou and Margaris (1988). Under favorable climatic conditions (sufficient moisture) soil microorganisms have the capacity to decompose natural products, such as secondary metabolites, at a rate of at least 1.7 g.m-2 .d -1. Such a rate could hold in the field for a limited period of time, excluding that of the summer drought. It is also been suggested that monoterpenes may serve as carbon and energy resources particularly during periods of carbon stress. The secondary compounds emanate either from the live plant or from the leaf litter (Vokou et al 1984), and reach the soil, where they have a significant effect on microorganisms and mineralization of N, P, etc.
The view that the bacterial populations are able to use the volatile oils as a carbon source is supported by many researchers, experimenting with specific isoprenoid compounds such as b-pinene and camphor (Gibbon et al., 1972; Gunsalus and Marshall 1974). Muller and de Moral (1966) reported that if the volatile oils were to exert any effect, they should be primarily absorbed by or adsorbed on the soil particles or soil colloids.
Since the litter of aromatic plants contains large quantities of volatile oils it might be expected that the decomposition process in Mediterranean ecosystems is mediated by allelopathic interactions, causing shifting of the population balance in the soil fungi and bacteria, particularly in the surface horizons. This activation of bacteria by volatile oils might be considered as an adaptive mechanism where these systems develop.
Audus, L.J. & K.V. Symonds, 1955. Further studies on the breakdown of 2,4-dichlorophenoxyacetic acid by a soil bacterium. Ann. Appl.Biol. 42, pp.174–182.
Berg, B., K. Hannus, T. Popoff & O. Theander, 1982. Changes in organic chemical components on needle litter during decomposition. Long-term decomposition in a Scots pine forest, I. Can. J. Bot. 60, pp.1310–1319.
Cadmus, M.C., H. Gasdorf, A.A. Anderson & R.W. Jackson, 1963. New bacteria polysaccharide from Arthrobacter. Appl. Microbiol. 11, pp.488–492.
Gibbon, G.H., N.F. Millis & S.J. Pirt 1972. Degradation of a-pinene by bacteria. Proccedings IVth International Fermentation Symposium: Fermentation Technology Today, pp.609–612.
Gunderson, K. & H.L. Jensen, 1956. A soil bacterium decomposing organic nitro-compounds. Acta Agr. Scand. 6, pp.100–114.
Gunsalus, I.C. & V.P. Marshall, 1974. Genetic regulation and machanisms of p-450 oxygenase action. In: Survival and toxic environments. Academic Press, New York, San Francisco and London, pp.295–313.
Halligan, J.P., 1975. Toxic terpenes from Artemisia californica. Ecology 56, pp.999–1003.
Kivanc,M. & A.Akgul, 1986. Antibacterial activities of essential oils from Turkish spices and citrus. Flavour and Fragrance Journal 1, pp. 175–179.
Knobloch, K., A. Pauli, B. Iberl, H. Weigand & N. Weis, 1989. Antibacterial and antifungal properties of essential oil components. Journal of essential oil research 1, pp.119–128.
Margaris, N.S. & D. Vokou, 1986. Microbial decomposition of smog organic pollutants. Bull. Environ. Contam. Toxicol. 36, pp. 15–22.
Melillo J.M. & J.D. Aber, 1984. Nutrient immobilization in decaying litter: an exampl of carbon-nutrient interactions. In: Trends in ecological research for the 1980s (J.H. Cooley and F.B. Golley, Eds), pp.193–215. Plenum Press, New York.
Muller, C.H. & R. del Moral, 1966. Soil toxicity induced by terpenes from Salvia leucophylla. Bull. Torey Bot. Club 93, pp. 130–137.
Muller, W.H., P.Lorber, and B. Haley, 1969. Volatile growth inhibitors produced by Salvia leucophylla: Effects on oxygen uptake by mitochondrial suspensions. Torrey Bot. Club 96, pp.89–96.
Rhyu, H.Y., 1979. Gas chromatographic characterization os sages of various geographic origins. Journal of Food Sciences 44, pp.758–762.
Sombrero, C., 1992. Environmental control of oil production in Mediterranean plants. PhD thesis, University of Reading, U.K..
Stevenson, I.L., 1967. Utilization of aromatic hydrocarbons by Arthrobacter spp. Can. J. Microbiol. 13, pp.205–211.
Vokou, D., N.S. Margaris & J.M. Lynch, 1984. Effects of volitile oils from aromatic shrubs on soil microorganisms. Soil Biol. Biochem. 16, pp.509–513.
Vokou, D. & N.S. Margaris, 1988. Decomposition of terpenes by soil microorganisms. Pedobiologia 31, pp.413–419.