Chemical Composition of Essential Oils and Antimicrobial Activity of Amomum cinnamomeum from Vietnam

Published in Khimiya Prirodnykh Soedinenii, No. 3, May–June, 2021, pp. 487–489.

The lack of information on the chemical constituents and biological activity of the volatile and non-volatile extracts of Amomum cinnamomeum Skornick., Luu & H.D.Tran, sp. nov. (Zingiberaceae) attracted our interest, hence we report herein the results of a study on the chemical constituents and antimicrobial activity of the essential oil from the rhizome of A. cinnamomeum.

The compositions and biological activities of essential oils from Amomum plants grown in Vietnam [[1]–[4]] and other parts of the world [[5]–[8]] have been reported previously. The results also indicated that monoterpenes and sesquiterpenes were the predominant compounds in the essential oils of a majority of these Amomum plants. In addition, essential oils from these Amomum plants exhibited biological activities, such as antimicrobial [[1], [5]–[8]] and larvicidal activity [[3]], among others. The aim of the present study is to examine the chemical constituents and antimicrobial activity of essential oils from the leaves and rhizomes of A. cinnamomeum grown in Vietnam for the first time and to determine its potential uses. Mature leaves and rhizomes of A. cinnamomeum were collected from Quang Ngai, Nghe An Province, Vietnam, in July 2019. The sample was identified by Dr. D. N. Dai. A voucher specimen, LNS 804, was deposited at the Botany Museum, Nghe An College of Economics, Vietnam. A total of 1 kg of the pulverized sample was used. The essential oil was obtained by hydrodistillation, which was carried out in a Clevenger-type distillation unit designed according to an established specification [[9]].

Gas chromatography (GC) was performed on an Agilent Technologies HP 7890 Plus Gas chromatograph equipped with an FID and fitted with an HP-5MS column (30 m × 0.25 mm, film thickness 0.25 μm, Agilent Technology). The analytical conditions were as described previously [[2]–[9]]. An Agilent Technologies HP 7890N Plus Chromatograph fitted with capillary an HP-5 MS column (30 m × 0.25 mm, film thickness 0.25 μm) and interfaced with a mass spectrometer HP 5973 MSD was used for gas chromatography-mass spectrometry (GC-MS) under the same conditions as those used for gas chromatography as described above. 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. The identification of constituents from the GC/MS spectra of A. cinnamomeum was performed based on retention indices (RI) determined with reference to a homologous series of n-alkanes (C4–C40), under identical experimental conditions. The mass spectral (MS) fragmentation patterns were compared with those of other essential oils of known composition [[10]].

The minimum inhibitory concentration (MIC) and median inhibitory concentration (IC50) values were measured by the microdilution broth susceptibility assay [[11]]. Stock solutions of the oil were prepared in dimethylsulfoxide (DMSO). Dilution series (214, 213, 212, 211, 210, 29, 27, 25, 23, and 21 μg/mL) were prepared in sterile distilled water inside the micro-test tubes, from which they were transferred separately to 96-well microtiter plates. Bacteria grown in double-strength Mueller–Hinton broth or double-strength tryptic soy broth, and fungi sustained in double-strength Sabouraud dextrose broth, were standardized to 5 × 105 and 1 × 103 CFU/mL, respectively. DMSO was used as a negative control. Streptomycin was used as the antibacterial standard, while nystatin and cycloheximide were used as antifungal standards. All experiments were performed in triplicate. After incubation at 37°C for 24 h, the MIC values were determined as the lowest concentration of essential oils of A. cinnamomeum that completely inhibited the growth of the microorganisms. The IC50 values were determined as the percentage inhibition of growth of microorganisms based on the turbidity measurement data of an EPOCH2C spectrophotometer (BioTeK Instruments, Inc Highland Park Winooski, VT, USA) and Rawdata computer software (Belgium).

The average yields of the essential oils were 0.12% and 0.19% (v/w, ± 0.01), respectively, for the leaf and rhizome, calculated on a dry weight basis. Fifty-nine compounds each were identified from both oil samples. The major classes of compounds were monoterpene hydrocarbons (46.9 and 31.9%), oxygenated monoterpenes (14.5 and 32.6%), sesquiterpene hydrocarbons (15.3 and 6.6%), and oxygenated sesquiterpene (16.4 and 25.7%); see Table 1.

TABLE 1. Constituents of Essential Oils from the Leaves and Rhizomes of Amomum cinnamomeum

aElution order on HP-5MS column; b Retention indices on HP-5MS column; – not identified.

TABLE 2. Antimicrobial Activity of the Leaf and Rhizome Essential Oils of A. cinnamomeum

Both the leaf and rhizome oils exhibited no antimicrobial action against Escherichia coli ATCC25922, Pseudomonas aeruginosa ATCC27853, and Salmonella enterica ATCC13076.

The main constituents of the leaves oil were β-pinene (35.8%), (E)-cinnamaldehyde (11.5%), and caryophyllene oxide (5.9%), while the rhizome oil was dominated by fenchyl acetate (13.7%), camphene (12.3%), bornyl acetate (9.7%), β-phellandrene (7.4%), and eudesma-4(15),7-dien-1β-ol (7.3%). This is the first report on the volatile constituents of A. cinnamomeum. Terpene compounds predominate in the essential oils, as was reported for other Amomum oil samples grown in Vietnam, such as A. rubidium [[1]] A. gagnepainii [[12]], A. repoense [[12]], A. longiligulare [[3]], A. villosum [[4]], A. aculeatum [[4]], A. maximum [[13]], and A. microcarpum [[9]]. However, the identities of these terpene compounds differed from one species to another, thus exhibiting chemical variability in their compositional pattern [[9]].

The leaves oil of A. cinnamomeum displayed antimicrobial activity towards Enterococcus faecalis ATCC 299212 (MIC 16.0 μg/mL), Staphylococcus aureus ATCC 25923 (MIC 16.0 μg/mL), and Bacillus cereus ATCC 14579 (MIC 16.0 μg/mL). The oil also inhibited the growth of Candida albicans ATCC 10231, with MIC of 64.0 μg/mL. The median inhibitory concentrations (IC50) against the tested microbes were evaluated as 4.98, 3.78, 5.78, and 28.79 μg/mL, respectively. However, the rhizome oil exhibited antimicrobial activity against the four microorganisms with MIC value of 32.0 μg/mL, while the IC50 values were estimated to be 10.34, 15.98, 9.78, and 9.79 μg/mL, respectively. The MIC and IC50 provided evidence that the leaf and rhizome oils of A. cinnamomeum showed potent antimicrobial activity against E. faecalis, S. aureus, B. cereus, and C. albicans. Both the leaf and stem oils exhibited no antimicrobial action against Escherichia coli ATCC25922, Pseudomonas aeruginosa ATCC27853, and Salmonella enterica ATCC13076. Streptomycin displayed antimicrobial activity with MIC values in the range 0.28 to 3.20 μg/mL, while nystatin had an MIC value of 8.0 μg/mL, with cycloheximide showing activity at MIC of 3.20 μg/mL. This is the first report on the antimicrobial activity of essential oil of space A. cinnamomeum. The results in this study are comparable with data obtained on the antimicrobial action of other Amomum essential oil reported in the literature such as A. rubidium [[1]], A. subulatum [[5]], A. cannicarpum [[14]], A. uliginosum [[15]], A. tsao-ko [[7]], and A. kravanh [[16]]. The antimicrobial activities of the essential oil of A. cinnamomeum can be related to its main compounds or some synergy between the major and minor compounds. The present essential oil constituents, such as β-pinene [[6]], caryophyllene oxide [[17]], (E)-cinnamaldehyde [[18]], fenchyl acetate [[19]], and camphene [[19]], were previously reported to have significant broad-spectrum activity.

This work was supported by the Project "Investigate the current status and value of "ma-gang" medicinal resources in Quang Ngai Province as a basis for management, use, conservation, and sustainable development" funded by Quang Ngai Provincial Department of Science and Technology (Contract No. 06/2018/HD-DTKHCN, dated 25 July 2018).