Chemical Composition of Essential Oil of Baeckea frutescens L

The volatile components of the leaf oil of Baeckea frutescens L. (Myrtaceae) from the Hatinh Province, Vietnam, were analysed by capillary gas chromatography (GC) and gas chromatography/mass spectrometry (GC/MS) and co-elution techniques. The identified compounds constituted more than 99.5% of the oil contents. Forty-nine compounds have been characterized Original Research Article Dai et al.; IRJPAC, 8(1): 26-32, 2015; Article no.IRJPAC.2015.071 27 among which -humulene (19.2%), β-caryophyllene (17.3%), baeckeol (13.8%), thujene (8.8%), linalool (5.6%) and 1, 8-cineole (5.6%) were the major constituents. This result may represent another chemotype of the oil of B. frutescens. Aims: Vietnam is a country blessed with many plants whose chemical compounds have not been previously examined. The aim of this research is to investigate the chemical constituents of essential oil of Baeckea frutescens. Study Design: Extraction of essential oil from the air-dried leaf samples of B. frutescens and investigation of its chemical constituents. Place and Duration of Study: Mature leaves of B. frutescens were collected from Hatinh Province, Vietnam in October 2013. Methodology: Air-dried and pulverized leaves were subjected to hydrodistillation in accordance with Vietnamese Pharmacopoeia specification to obtained essential oil. The components of the oil were analyzed by gas chromatography (GC) and gas chromatography/mass spectrometry (GC/MS) and co-elution techniques. Results: Forty-nine compounds have been characterized among which -humulene (19.2%), βcaryophyllene (17.3%), baeckeol (13.8%), -thujene (8.8%), linalool (5.6%) and 1, 8-cineole (5.6%) were the major constituents. Conclusion: The present oil compositions were found to be different from the results obtained previously from the essential oils of B. frutescens grown in other parts of the world. The present result may represent another chemotype of the oil of B. frutescens.


INTRODUCTION
Baeckea frutescens L. is a shrub or small tree of the family Myrtaceae. The leaves are opposite, strip or strips tapered, 5-8 mm long, while the solitary flower is bisexual, yellow and white, about 2-3 mm in diameter. The fruits are small, about 1 mm while the seeds have horns. Flowering takes place between July and October [1]. Extracts from the leaves of B. frutescens have displayed anti-inflammatory and antioxidant [2] and cytotoxicity [3] activities. Flavonoids, chromones, sterols [4] and flavonol glycoside [5] were previously isolated from the plant. A biflavonoid compound, 3-O-α-lrhamnopyranosylmyricetinyl-(I-2″,II-2″)-3-O-α-lrhamnopyranosylmyricetin, present in B. frutescens was found to be useful in the prevention of arteriosclerosis [6]. The antioxidant and cytoprotective activities of the biflavonoid baeckein E characterized from the plant has been reported [7], while baeckein I displayed potent anti-inflammatory effect [8]. Other known compounds include baeckein A-D [9,10]. Two phloroglucinols with strong cytotoxicity activity against leukaemia cells (L 1210) were isolated from the dried leaves of B. frutescens [11]. 5-Hydroxy-2-isopropyl-7-methoxychromone isolated from the aerial parts of B. frutescens exhibited toxicity to the brine shrimp Artemia salina [12].
In this paper, the chemical compounds of essential oil obtained from B. frutescens are being reported, as part of an extensive research on the chemical analysis of poorly studied species of Vietnamese flora [21].

Plant Collection
Leaves of B. frutescens were collected in October 2013, in Nghi Xuan District (12.55 N, 109.07 E), Ha Tinh Province, Vietnam. Botanical identification of the plant was performed by Dr. D.N. Dai. The soil at the point of collection was saline and acidic in nature while the temperature was 10ºC. A voucher specimen (DND 127) was deposited at the Botany Museum, Vinh University, Vietnam. 600 g of the plant was collected initially prior to air-drying. Plant samples were air-dried prior to extraction.

Extraction of the Volatile Oil
0.5 kg of air dried sample was shredded and their oils were obtained by hydrodistillation for 3 h at normal pressure, according to the Vietnamese Pharmacopoeia [22]. The yield of essential oil was 0.52% (v/w, light yellow), calculated on a dry weight basis.

Gas Chromatography (GC) Analysis of the Oils
Gas chromatography (GC) analysis was performed on an Agilent Technologies HP 6890 Plus Gas chromatograph equipped with a FID and fitted with HP-5MS column (30 m x 0.25 mm, film thickness 0.25 m, Agilent Technology). The analytical conditions were: carrier gas H 2 (1 mL/min), injector temperature (PTV) 250ºC, detector temperature 260ºC, column temperature programmed from 40ºC (2 min hold) to 220ºC (10 min hold) at 4ºC/min. Samples were injected by splitting and the split ratio was 10:1. The volume injected was 1.0 L. Inlet pressure was 6.1 kPa. The sample was analyzed thrice.

Gas Chromatography -Mass Spectrometry (GC-MS) Analysis
An Agilent Technologies HP 6890N Plus Chromatograph fitted with a fused silica capillary HP-5 MS column (30 m x 0.25 mm, film thickness 0.25 m) and interfaced with a mass spectrometer HP 5973 MSD was used for the GC/MS analysis, under the same conditions as those used for GC analysis. The conditions were the same as described above with He (1 mL/min) as carrier gas. The MS conditions were as follows: ionization voltage 70 eV, emission current 40 mA, acquisitions scan mass range of 35-350 amu at a sampling rate of 1.0 scan/s.

Identification of the Constituents
The identification of constituents was performed on the basis of retention indices (RI) determined by co-injection with reference to a homologous series of n-alkanes, under identical experimental conditions. Further identification was performed by comparison of their mass spectra with those from NIST (Database 69) and Wiley 9 th Version and the home-made MS library built up from pure substances and components of known essential oils, as well as by comparison of their retention indices with literature values [23,24].
When compared with previous studies, it could be seen that only -2-carene and trans-bisabolene were the only compounds that could not be detected in the present study. However, the chemical combination of -humulene/-caryophyllene/baeckeol could be described as a new chemical composition of essential oil of B. frutescens. It may therefore represent a new chemotype of essential oil of B. frutescens. The chemical variations in the oil composition of B. frutescens by different researchers may be due some factors such as collection time, age of the plant, chemotypes, handling conditions, mode of extraction as well differences in geographical and climatic factors between these countries.
The essential oil of B. Frutescens was shown to possess anti-fungal activity against the mycelial growth of five pathogenic fungi [29]. The biological activities of an essential oil may be due to the major compounds or synergy between the major and some minor compounds present in the oil [30]. Referring to literature, -caryophyllene is known to exhibit both antimicrobial and insecticidal activities [30]. Essential oils with moderate to high contents of -humulene and βcaryophyllene have been used to treat itching and other skin problems [31]. Baeckeol and its several analogues have demonstrated potent antioxidant, anti-inflamatory and cytotoxicity activities [7]. Therefore, considering the information on the biological potentials of the compounds present in therein, it may be postulated that the essential oil of B. frutescens may be useful as phytopharmaceuticals.