Octanoic acid

Abstract

The essential oil of eaglewood tree (Aquilaria agallocha Roxb.) has been considered to be a pathological product. An investigation was carried out to study the difference in composition of oils obtained from healthy, naturally infected and artificially inoculated eaglewood using GC and GC/MS analyses. This investigation showed a marked difference in the oil compositions among the treatments with regards to their quality. Valerianol (3.0%) and tetradecanioc acid (7.1%) contents were recorded higher in the oils of naturally infected plants than in that of healthy ones (0.1% and 6.9%, respectively). Pentadecenoic acid was totally absent in the oils of healthy, whereas it was found in a greater amount (6.8%) in the oil of naturally infected plants. In contrast, dodecanoic acid (3.1%), pentadecanoic acid (6.2%), hexadecanoic acid (31.5%) and octadecanoic acid (4.1%) were found in a higher amount in the oils of healthy plants, while the oils obtained from naturally infected plants contained lower amounts of these components (2.5%, 4.8%, 20.0% and 1.0%, respectively). The oils obtained from the inoculated plants showed almost similar distribution of the components with healthy plants.

Key Word Index

Aquilaria agallocha, Chaetomium gtohosum, Fusarium oxysporum, Thymelaeaceae, essential oil composition, hexadecanoic acid.

Introduction

The eaglewood tree Aquilaria agallocha Roxb. (syn. Acquilaria malaccensis Lamk., family: Thymelaeaceae) is a precious floral wealth of India (1). The resinous patches of fragrant wood of the plant known as 'agar' is used as incense in Egypt, Arabia, and throughout the northeast part of India where it can be found. The oil obtained from agar is described as a stimulant, cardiatonic and carminative. It is also used in the cosmetic and pharmaceutical industries. Agar is considered to be a pathological product produced by fungal invasion of the host (2). Since 1938, few workers have been studying about agar formation and reported the agar zones to be associated with mold and decay fungi (3-11). Association of mycoflora in the seed, rhizosphere and phyllosphere were studied by Tainuli et al. (12-14). Among different fungal species reported to be associated with agar zones, few could exhibit pathogenesis with the development of disease symptoms while others seem to be of saprophytic nature in different eco-geographical conditions. Studies on the oil of infected A. agallocha were made by various workers (15-26). Maheshwari et al. (15) isolated three newsesquiterpenic furanoids of the selinane group from aganvood oil, obtained from the fungus infected plant and their structures and absolute configurations determined by degradative studies and physical measurements. Varma et al. (16) examined that degradative studies and physical measurements supported by an unambiguous synthesis of the derived ketone have led to the assignment of a novel spiro-skeleton to agarospirol, a sesquiterpene alcohol isolated from the essential oil of infected aganvood. Paknikar and Dhavlikar (17) and Paknikar and Naik (18) reported that on hydrogenation of α-agarofuran and β-agarofuran the same dihydroagarofuran was obtained. Thomas and Ozainne (19) used a combination of hydrogenation, ^sup 1^H-NMR, ^sup 13^C-NMR and some naturally occurring dihydroagarofuran and isodihydroagarofuran to unequivocally show that the dihydroagarofuran found was indeed dihydro-β-agarofuran and isodihydroagarofuran was isodihydro-β-agarofuran; two separate compounds.

Pant and Rastogi (20) and Bhandari et al. (21) isolated a new sesquiterpene, agarol and a couinarinolignan, aquillochin, respectively, from the oil of Aquilaria agallocha. Nakanishi et al. (22) showed that a benzene extract of aganvood collected in Indonesia contained a new tricyclic sesquiterpene alcohol as a major constituent. Nagashima et al. (23) further characterized the presence of two more sesquiterpene alcohols, jinkohol II and jinkoh-eremol, from the same Indonesia agarwood oil. Nakanishi et al. (24) again reported that a benzene extract of an Indonesian sample of 'Jinkoh' agarwood was found to contain α-agarofuran, 10-epi-γ-eudesmol and oxo-agarospirol. Ishihara et al. (25) characterized seven new sesquiterpenes based on the guaiane skeleton in a sample of agarwood oil. Five new eudesmane sesquiterpenes and three other compounds further characterized by Ishihara et al. (26) in a sample of agarwood extract produced in the laboratory from A. agallocha of Vietnamese origin.

Vesicular-arbuscular myccorhizal association in the tree species and changes in amino acid composition due to pathogenesis were also studied by Tamuli et al. (27,28). So far the qualitative study of the oils of healthy and infected eaglewood has yet to be investigated. The present investigation was, therefore, undertaken to study the qualitative differences in the oils obtained from healthy, naturally infected and artificially inoculated eaglewood.

Experimental

Artificial inoculation of fungal isolates: The most frequently isolated fungi from infected agarwood (e.g. Chaetomium globosum and Fusarium oxysporum) were inoculated to the healthy plants by artificial boring on to the plants (9). Inoculation was made with two different fungi alone and in their combination. Observations were made at an interval of 30 days after inoculation.

Oil isolation: The materials used for isolating oils were the wood chips of healthy (H), naturally infected (NI) and artificially inoculated plants by two fungal isolates, i.e. Chaetomium glohosum (CG) and Fusarium oxysporum (FO). The materials were hydrodistilled using Clevenger-type apparatus (29). Distillation of the materials was run for 70 h. The oil collected was then dried over anhydrous sodium sulfate and their physical characters like color and odor were recorded. The oils were then stored in sealed container under refrigeration prior to analysis.

GC: A Shimadzu GC-17 A gas chromatograph equipped with a FID detector and a HP-fused silica column (30 m x 0.32 mm, 0.25 µm film thickness) was used. The samples were injected in the split mode, using pressure-controlled helium as carrier gas at a linear velocity of 30 cm/s (at 60°C). Injector and detector temperatures were maintained at 280°C and 300°C, respectively. The column oven temperature was programmed from 50°C (after 3 min) to 300°C at 4.5°C/min. The final temperature was held for 20 min. Peak areas and retention times were measured by electronic integration without the use of correction factors. The relative amounts of individual components are based on the peak areas obtained without FID response factor correction. Temperature programmed (linear) retention indices of the compounds were determined relative to n-alkanes.

GC/MS: Analyses were carried out on a Shimadzu GC-17 A/GC MS- QP 5000 system. A 25m x 0.20 mm fused silica HP-1 column, with a film thickness of 0.33 µm, was employed. The column oven temperature was programmed from 60°C (after 30 min) to 300°C at 5°C/mm. The injector and GC/MS interface temperatures were maintained at 280°C and 300°C, respectively. Helium carrier gas was pressure controlled to give a linear gas velocity of 44 cm/s (at 60°C). Electron ionization mass spectra (70 eV) were acquired over the mass range 10-400 Da at a rate of 2/s.

Component identification: The compounds were identified by matching their retention indices on various columns with those of authentic samples scanned underidentical conditions. Identities of many compounds were further verified by GC/M S where peaks were compared with reference compounds and by matching their 70 eV El mass spectra with those of library search data (30-37).

Results and Discussion

The oil of healthy naturally infected and artificially inoculated plants were analyzed by GC and GC/M S. The data were cited in Table I. Investigation showed a marked difference between the oils obtained from naturally infected and healthy plants with regards to their quality.

A trace of heptanoic acid was found in the oil of healthy plant whereas it was 2.4% in the oil of a naturally infected plant. Octanoic acid and 10-epi-γ-eudesmol were 2.1% and 1.5% in the oils obtained from naturally infected plants but these components were totally absent in the oil of healthy plants. Again, valerianol (3.0%) and tetradecanoic acid (7.1%) contents were recorded higher in the oils of naturally infected plants than in that of healthy ones (0.1% and 6.9%, respectively). Pentadecenoic acid was totally absent in the oils of healthy whereas it was found in a greater amount (6.8%) in the oil of naturally infected plants.

In contrast, some of the components were recorded in a higher amount in the oils of healthy plants than in naturally infected. Dodecanoic acid (3.1%), pentadecanoic acid (6.2%), hexadecanoic acid (31.5%) and octadecanoic acid (4.1%) were found in a higher amount in the oils of healthy plants whereas the oils obtained from naturally infected plants contained lower amounts of these components (2.5%, 4.8%, 20.0% and 1.0%, respectively). Tridecanoic acid and linoleic acid were completely absent in the oils of naturally infected plants while these two components were found in a higher amount (3.2% and 3.4%, respectively) in the oil of healthy plants.

The oils obtained from the inoculated plants showed almost similar distribution of the components. But some of the components were found in the oils of artificially inoculated plants including naturally infected whereas those are totally absent in the oil of healthy plants. 10-epi-γ-Eudesmol and γ-eudesmol were totally absent in the oil of healthy plants while traces of theses components were found in the oils of artificially inoculated plants.

It was observed that the characteristic components of agarwood oil were found to be lower in the oils obtained from healthy samples. The oils obtained from artificially inoculated agarwood have no such differences with the oils of healthy wood though little changes were observed. This may indicate that naturally infected type of agarwood would not be achieved by artificially inoculation of fungal isolates.

The observations made by us showed that the microflorais of great importance in production of specialized type of agarwood for best quality agar oil. However, there may exist variants or eco-types within the agarwood plant species. If natural variant or type exists within the plant species, the fungal pathogens might be host type specific or variant specific. If it is so, there may exist specific host variant pathogen/host type-pathogen relationship which determines the success of artificial inoculation. Therefore, identification of natural variant or eco-type and the specific host-pathogen relationship under different ecological conditions is expected to give clue for unraveling the secret of agar formation. Then only artificial supplement of inoculum to the specific host might give positive result for induction of disease in the plant.

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Phatik Tamuli,* Paran Boruah and Subhan C. Nath

Division of Plant Sciences and Ecology, Regional Research Laboratory, Jorhat - 785 006, Assam, India

Piet Leclercq

Laboratory of Instrumental Analysis, Eindhoven, University of Technology, P.O. Box 513, 5600 MB Eindhoven, The Netherlands

* Address for correspondence

Received: June 2003

Revised: September 2003

Accepted: November 2003

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