Abstract
Objective/Background
The essential oils (EOs) of Curcuma species contain various volatile compounds with bioactivity. However, the phytochemical profile of Curcuma thorelii and the antimicrobial activities of Curcuma rhabdota, and Curcuma petiolata have received limited attention. This study aims to characterize and compare the major compounds and the antimicrobial activities of EOs extracted from the rhizomes of C. rhabdota, C. thorelii, and C. petiolata collected in Vietnam.
Methods
EOs were obtained by hydrodistillation of the rhizomes of three Curcuma species. The chemical profiles were determined using gas chromatography-mass spectrometry (GC-MS). The antimicrobial activities against bacteria and a pathogenic fungus were determined through the broth dilution method.
Results
The volatile profiles of C. rhabdota, C. thorelii, and C. petiolata EOs included 63 (97.1%), 47 (98.0%), and 50 (95.6%) compounds, respectively. The major compound in the EO of C. rhabdota rhizomes was 3-carene (16.6%), followed by camphene (9.8%), α-copaene (7.4%), γ-terpinene (7.3%), camphor (5.9%), and β-curcumene (5.7%). The predominant compounds of the EO extracted from C. thorelii rhizome were xanthorrhizol (40.7%), β-curcumene (20.7%), and α-curcumene (8.9%), while camphene (17.0%), (E)-β-elemenone (16.8%), (E)-β-farnesene (13.6%), germacrone (8.9%), 1,8-cineole (7.2%), and camphor (6.0%) were the most abundant components in C. petiolata rhizomes. Except for Pseudomonas aeruginosa, which was less susceptible to the EOs with a minimum inhibitory concentration (MIC) value of 128 μg/mL, the three oil samples exhibited potent antimicrobial activities against all investigated strains with MIC values in the range of 2-32 μg/mL. Especially, the EO of C. thorelii rhizomes showed intense activity against Enterococcus faecalis, Staphylococcus aureus, Bacillus cereus, Escherichia coli, and Candida albicans with a MIC value of 2 µg/mL.
Conclusion
The results showed the chemical variability of EOs from three Curcuma species and the prepared EO samples showed potent antimicrobial activities against several microbial strains, indicating a high potential application as a food preservative and in the pharmaceutical industry.
Introduction
Curcuma is a large genus of the Zingiberaceae family with approximately 130 species growing in South and Southeast Asia. 1 Among them, there are currently more than 27 species of Curcuma widely distributed from the northern to southern provinces in Vietnam.2,3 In the traditional medicine of many countries, Curcuma species have long been used medicinally for treating pneumonia, hormonal disorders, bronchial complaints, leucorrhea, diarrhea, dysentery, and parasitosis, among other conditions.4–6 Curcuma species have been considered a rich source of essential oils (EOs), which are responsible for their pleasant aroma, as well as a valuable source of medicinal substances, such as curcumin, xanthorhizol, curdione, curcuzedoalide, isoprocurcumenol, and turmeronols,7,8 which contribute to their various pharmacological properties. 9 Several studies on Curcuma EOs identified sesquiterpenoids and monoterpenoids as the major components. 10 Various EOs from Curcuma species have been reported to possess strong antioxidant activity.11–13 Other bioactivities of these EOs also have been described such as antibacterial, 11 anti-inflammatory, 14 antimicrobial, 15 antitumor, 16 insecticidal,17,18 antifungal, 19 and cytotoxic activities. 20 Furthermore, curcuminoid compounds are the major constituents of Curcuma species, which are generally regarded as the most active constituents, with a diverse range of bioactivities such as antitumor, anti-inflammatory, and neuro-protective, for treatment of a wide range of ailments, 21 and efficient inhibition of the enzyme tyrosinase. 22 Although studies on the phytochemistry and bioactivities of Curcuma EOs are abundant in the literature, there is no report describing the antimicrobial activity of Curcuma rhabdota Sirirugsa & M.F.Newman and Curcuma petiolata Roxb. Besides, there has been no phytochemical and pharmacological information on Curcuma thorelii Gagnep. Therefore, the present study was conducted to clarify the chemical compositions of EOs prepared from C. thorelii, C. rhabdota, and C. petiolata rhizomes and their antimicrobial activities.
Results and Discussion
EO Composition
Hydrodistillation of the rhizomes of the three Curcuma species all produced pale-yellow oils. The yields of the obtained EOs from C. rhabdota, C. thorelii, and C. petiolata were 0.19%, 0.22%, and 0.17% (w/w), respectively, calculated on a fresh weight basis. The chemical compositions of these EOs are given in Table 1.
Chemical Compositions of EOs From the Rhizomes of Three Curcuma Species.
Abbreviations. RI(Exp.): retention indices on HP-5MS ultra inert column; RI(Lit.): retention indices in literature (NIST 17 and Adams 23 ); content (%) in “Bold” denotes major compounds (> 5%); EO: essential oil.
In general, the C. rhabdota rhizome EO was rich in monoterpene (50.4%) and sesquiterpene (29.3%) hydrocarbons, the C. thorelii rhizome EO was rich in oxygenated sesquiterpenes (44.0%) and sesquiterpene hydrocarbons (37.8%), while the rhizome EO of C. petiolata showed no significant difference between the compound groups.
The analytical results showed the identification of 63, 47, and 50 principal constituents, making a total of 97.1%, 98.0%, and 95.6% of the EOs of C. rhabdota, C. thorelii, and C. petiolata, respectively (Table 1). 3-Carene (16.6%), camphene (9.8%), α-copaene (7.4%), γ-terpinene (7.3%), camphor (5.9%), and β-curcumene (5.7%) were the main components of C. rhabdota rhizome EO. Xanthorrhizol (40.7%), β-curcumene (20.7%), and α-curcumene (8.9%) were the principal compounds of the EO of C. thorelii rhizomes, and camphene (17.0%), (E)-β-elemenone (16.8%), (E)-β-farnesene (13.6%), germacrone (8.9%), 1,8-cineole (7.2%), and camphor (6.0%) were identified as the major constituents of the EO of C. petiolata rhizome. Remarkably, the EOs of C. rhabdota and C. petiolata had camphene and camphor as the same major components, while β-curcumene was found as the major component in both C. rhabdota and C. thorelii EOs.
A comparison of these results with those in the literature showed that there are some differences among the C. rhabdota EOs that have been studied. Specifically, three main constituents of the EO of this plant from Bangkok, Thailand were germacrone (24.4%), butanoic acid butyl ester (14.2%), and butanoic acid-1-methylpropyl ester (8.8%). 24 In another study, 3-carene and copaene were identified as two major compounds in the rhizome EO of this plant from Ubon Ratchathani, Thailand. 25 For C. petiolata, previous results showed marked differences in the quality and quantity of chemical compositions in which β-farnesene (74.8%) and 2-methyl-5-pentanol (84.0%) were revealed to be dominant.26,27 In particular, xanthorrhizol in C. xanthorrhiza rhizomes 28 was found in C. thorelii rhizomes, but was absent from the rhizomes of C. rhabdota and C. petiolata. Additionally, α-curcumene and β-curcumene were found in C. rhabdota and C. thorelii, which is similar to the reported data for other Curcuma species (eg Curcuma amada and Curcuma aromatica29,30). However, these two compounds were not found in the EO of C. petiolata. These results showed that the variability in the chemical constituents of the EOs of Curcuma species depends on different species, different geographical locations, and different extraction methods leading to the variability of EO analytical results.
Antimicrobial Activity Evaluation
The results of the study of the antimicrobial activity of the EOs of the rhizomes of C. rhabdota, C. thorelii, and C. petiolata are presented in Table 2.
Antimicrobial Activity of EOs From the Rhizomes of Three Curcuma Species.
Abbreviation. EO: essential oil.
In general, the results showed that C. thorelii rhizome EO showed higher potential antimicrobial activities than those of C. petiolata and C. rhabdota EO against all investigated strains, except for Pseudomonas aeruginosa. Specifically, the rhizome EO of C. thorelii showed strong antibacterial activities against Gram-positive bacteria (Enterococcus faecalis, Staphylococcus aureus, and Bacillus cereus) with MIC values of 2 μg/mL and Gram-negative bacteria (Escherichia coli and Salmonella enterica), with MIC values of 2-8 μg/mL. The potential antibacterial activity of C. petiolata rhizome EO against E. faecalis was observed with a MIC value of 8 µg/mL, while the values against B. cereus, S. aureus, S. enterica, and E. coli were only 16 µg/mL. Similarly, C. rhabdota rhizome EO was only shown to possess moderate activity against the same bacterial strains with MIC values in the range of 16-32 μg/mL. Compared with the inhibitory effects of the positive controls on those bacteria, C. thorelii showed significantly stronger activity than kanamycin (MIC values of 4-128 μg/mL) and tetracycline (MIC values of 4-64 μg/mL), while C. petiolata and C. rhabdota possessed comparable antimicrobial activities to these antibiotics. Considering P. aeruginosa, all EOs tested exhibited weak antibacterial activity with a MIC value of 128 μg/mL, whereas kanamycin still effectively inhibited this strain, with a MIC of 64 µg/mL. Compared with some previous studies, the antimicrobial activities of C. thorelii, C. petiolata, and C. rhabdota were markedly pronounced. The MIC value of the EO from Euphorbia helioscopia against S. aureus, E. faecalis, and E. coli was 31.25 μg/mL. 31 In another study, the EO from Meistera sudae Šída f. & Škorničk leaf, a Vietnamese Zingiberaceae species, inhibited Bacillus subtilis and S. aureus, with the same MIC value of 25 µg/mL. 32 In recent research, EOs from the leaves of Fokienia hodginsii and Amentotaxus argotaenia strongly inhibited E. faecalis and B. cereus, with MIC values ranging from 32 to 64 µg/mL. 33
For anti-yeast activity, the EO samples also showed moderate to strong effects against Candida albicans. Specifically, the strongest yeast inhibitory effect was found for C. thorelii rhizome EO (MIC = 2 µg/mL), followed by C. petiolata and C. rhabdota rhizome EOs, with a MIC value of 16 μg/mL.
This finding might be valuable because many EOs are known to inhibit significantly the cell wall of only Gram-positive bacteria. 34 Akarchariya et al demonstrated that EOs from other Curcuma rhizomes, such as Curcuma aeruginosa, Curcuma glans, and Curcuma cf. xanthorrhiza, also exhibited good antibacterial activities. 35 The antimicrobial and other bioactivities of the Curcuma genus have been summarized recently. 9
In a previous study, β-curcumene was identified as a characteristic chemical component of the EOs of the Curcuma genus, along with other sesquiterpenes, such as ar-turmerone, α-curcumene, and xanthorrhizol. 9 To date, there have been limited studies on the bioactivities of β-curcumene. However, the EO of C. thorelii rhizomes displayed potential antimicrobial activities and β-curcumene formed 20.7% of the total content, which suggested that the olefinic sesquiterpene might possess promising antimicrobial activities. Xanthorrhizol, a bisabolene-type aromatic sesquiterpene, is the most abundant compound in the EO of C. xanthorrhiza. Recently, xanthorrhizol has become a compound of interest in the pharmacological field, in terms of anti-cancer and antimicrobial activities. The aromatic sesquiterpene exhibited strong antimicrobial effects against S. aureus, E. coli, and Propionibacterium acnes, which explained the strong antimicrobial effects of C. thorelii rhizome EO, which contained more than 40% of xanthorrhizol. 36 Xanthorrhizol was also shown to have potential anticancer properties with inhibitory effects on various cancer cells. Xanthorrhizol also exhibited synergistic effects with curcumin and tamoxifen on cancer cell growth inhibition.37,38 3-Carene is a bicyclic monoterpene, which can be commonly found in Pinus and pepper volatile oils. The compound had strong antibacterial activity against Brochothrix thermosphacta and Pseudomonas fluorescens by damaging cellular membranes, disrupting DNA structure, and interfering with cellular functions. 39 Camphene is a volatile compound commonly found in various aromatic plants, including Thymus, Origanum, and Salvia genera. Camphene was found to exhibit antimicrobial activities against various bacterial and fungal strains. This bicyclic monoterpene was also studied for other pharmacological activities, including antiviral, anti-leishmanial, anti-inflammatory, anti-diabetic, hypolipidemic, and anti-cancer activities. 40 In particular, previous studies demonstrated that 1,8-cineole has antinociceptive, vasodilator, bronchodilator, anti-inflammatory, hepatoprotective, gastroprotective, antibacterial, antimycotic, and antitumorigenic activities. 41 Camphor can be a promising agent for potential antibacterial, antifungal, analgesic, anti-inflammatory, and antioxidant activities. 42 Currently, further studies are needed on these herbs because of their antimicrobial and other biological effects. Based on that scientific basis, the orientation continues for the rational exploitation and use as well as the development of new functional foods and new drugs.
Materials and Methods
Plant Materials
The fresh rhizomes of C. rhabdota Sirirugsa & M.F.Newman and C. petiolata Roxb were collected from Phuoc Vinh Forest, Chau Thanh District, Tay Ninh Province, Vietnam in July 2021, while the fresh rhizomes of C. thorelii Gagnep were taken from Suoi Kiet Commune, Tanh Linh District, Binh Thuan Province, Vietnam in August 2022. The plants were identified by Dr Dang Van-Son (Institute of Tropical Biology, Vietnam Academy of Science and Technology) and Dr Nguyen Danh-Duc (Institute of Applied Technology, Thu Dau Mot University). Voucher specimens HC-008 (C. rhabdota), HC-009 (C. petiolata), and NDD-236 (C. thorelii) were deposited at the herbarium of the Institute of Applied Technology, Thu Dau Mot University, Binh Duong Province, Vietnam.
Isolation of the EOs
Five hundred grams of each sample was washed, pulverized, and distilled using a Clevenger apparatus until the amount of obtained EO was constant. After that, anhydrous sodium sulfate (Na2SO4) was used to completely remove all traces of water. Experiments were performed in triplicate. Finally, the EOs were stored in a refrigerator at 4°C for later analysis of chemical composition and biological activity.
Chemical Characterization of the EOs
GC-MS analysis was performed on an Agilent 7890B GC System equipped with a 5977B MSD model. The GC column was an HP-5MS Ultral Inert (30 m × 0.25 mm, thin film of 0.25 μm). The injection volume (dissolved with a ratio of 1/100, v/v, n-hexane, Merck) was 1 μL using a splitting mode (1:25). The carrier gas was helium with a flow rate of 2.0 mL/min and a column head pressure of 8.2 psi. The inlet-F temperature was 300°C, the MS Quad temperature 50°C, the Aux-2 temperature 300°C, and the MS source 230°C. The GC oven temperature was kept at 60°C for 1 min and increased to 240°C at a rate of 4°C/min and kept constant at 240°C for 4 min. The MS scanned were obtained by electronic impact at 70 eV with automatic scanning in the range of 50-550 amu at 2 scans per second. The volatile constituents were identified by comparison of their mass spectra and retention indices (RI) with the literature (NIST 17 and Adams). 23 MassHunter Workstation Software was used to handle mass spectra and chromatography analysis. Finally, the content of each compound was quantified by dividing their respective peak areas by the sum of all the EO components’ peak areas and multiplying the result by 100.
Antimicrobial Activity Assay
The rhizome EOs of C. rhabdota, C. thorelii, and C. petiolata were analyzed for antimicrobial activity using Gram-positive bacteria namely, E. faecalis ATCC29212, S. aureus ATCC2592, and B. cereus ATCC14579; Gram-negative bacteria, namely E. coli ATCC25922, P. aeruginosa ATCC27853, and S. enterica ATCC13076; and a yeast strain, namely C. albicans ATCC10231. All of them were purchased from the National Institute for Food Control (Hanoi, Vietnam).
The rhizome EOs of C. rhabdota, C. thorelii, and C. petiolata were dissolved in 10% DMSO (dimethyl sulfoxide) in a decreasing concentration range (μg/mL): 256, 128, 64, 32, 16, 8, 4, and 2; the experiment was conducted in triplicate. Fifty microliter of microbial culture, standardized at a concentration of 2 × 105 CFU/mL, was shaken at 120 r/min, and then incubated at 37 °C for 24 h. MIC values were determined as the lowest concentration of each of the EOs which completely inhibited (97%-100%) the growth of the microorganisms and were accurately determined based on turbidity measurement using a BioTek Epoch spectrophotometer (USA) and RawData software. Kanamycin, tetracycline, and cycloheximide were used as the antibacterial and antifungal standards. 43
Conclusions
For the first time, the antimicrobial activity of rhizome EOs of C. rhabdota and C. petiolata, as well as the chemical constituents and antimicrobial activity of the rhizome EO extracted from C. thorelii were reported. The analysis results showed that C. rhabdota consisted mainly of monoterpenes while C. thorelii and C. petiolata were composed of sesquiterpenes. All three EOs showed strong antimicrobial activity against the investigated strains. Therefore, the results of this investigation may hold promise for food, pharmaceutical, and other industrial applications. Finally, the isolation and identification of nonvolatile compounds and their bioactivities from these species could be carried out in the near future.
Supplemental Material
sj-docx-1-npx-10.1177_1934578X231167229 - Supplemental material for Phytochemical Analysis of the Essential Oils From the Rhizomes of Three Vietnamese Curcuma Species and Their Antimicrobial Activity
Supplemental material, sj-docx-1-npx-10.1177_1934578X231167229 for Phytochemical Analysis of the Essential Oils From the Rhizomes of Three Vietnamese Curcuma Species and Their Antimicrobial Activity by Hieu Tran-Trung, Xuan Duc Dau, Thi Chung Nguyen, Hien Nguyen-Thi-Thu, Hieu Nguyen-Ngoc, Thi Giang An Nguyen, Van Trung Hoang, Dang-Khoa Nguyen, Danh Duc Nguyen, Chen Tran Van and Le Duc Giang in Natural Product Communications
Footnotes
Declaration of Conflicting Interests
The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.
Funding
The author(s) received no financial support for the research, authorship, and/or publication of this article.
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References
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