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Chemical composition and antimicrobial activity of the essential oil from the aerial part of Dictamnus dasycarpus Turcz.
Industrial Crops & Products 140 (2019) 111713
Contents lists available at ScienceDirect
Industrial Crops & Products journal homepage: www.elsevier.com/locate/indcrop
Short communication
Chemical composition and antimicrobial activity of the essential oil from the aerial part of Dictamnus dasycarpus Turcz.
T
Haiping Tiana, Hepeng Zhaoa,b, Hongli Zhoua,b, , Yang Zhangc, ⁎
⁎⁎
a
School of Chemistry and Pharmaceutical Engineering, Jilin Institute of Chemical Technology, Jilin 132022, China Engineering Research Center for Agricultural Resources and Comprehensive Utilization of Jilin Provence, Jilin Institute of Chemical Technology, Jilin 132022, China c School of Biology and Food Engineering, Changshu Institute of Technology, Changshu 215500, China b
ARTICLE INFO
ABSTRACT
Keywords: Dictamnus dasycarpus Turcz. Aerial part Chemical composition Antimicrobial activity
Dictamnus dasycarpus Turcz. is an economic plant widely cultivated in China and its dried root bark is commonly prescribed as Cortex Dictamni for the treatment of various inflammatory diseases. Thus, more attention was devoted to keep the high-quality of the root bark of D. dasycarpus, but leaving the aerial part on ground as waste. In this context, we focused on the chemical composition and antimicrobial activity of the essential oil from the aerial part of D. dasycarpus (EOAPDD) for the first time. The extraction yield of EOAPDD was 0.14% by hydrodistillation, and thirty-nine compounds were identified by gas chromatography-mass spectrometry with the highest proportion of sesquiterpenoids (37.3%), followed by monoterpenes (29.4%). The dominant active constituents in EOAPDD are germacrene D (18.0%), terpinolene (13.0%), (Z)-β-ocimene (10.4%), β-caryophyllene (7.74%), phytol (5.20%), and (+)-viridiflorol (3.16%). Essential oil from the aerial part of D. dasycarpus shows pronounced inhibitory activity against Candida albicans with minimum inhibitory concentration of 3.23 mg/mL (4-fold lower than that of chloramphenicol), which may be largely contributed by the terpene compounds (37.3% of sesquiterpenoids and 29.4% of monoterpenes) in EOAPDD. Results obtained from present investigation provided strong evidence that the aerial part of D. dasycarpus also deserves attention, cherishment and deep exploitation due to the promising potential as a nature-based antifungal agent.
1. Introduction Essential oils (EOs), produced by plants as secondary metabolites are usually composed of different compounds with relatively small molecular weights and normally possess remarkable aromatic odor (Zhang et al., 2018). Based on chemical classification, EOs can be mainly divided into aliphatics, aromatics, and terpenoids, and are characterized by low boiling point and high volatility in most cases (Raut and Karuppayil, 2014; Bajer et al., 2018). Essential oils elicit diverse bioactivities including antimicrobial, antioxidant, antidiabetic, and anticancer properties (Edris, 2007), particularly their antimicrobial activities have gained enormous attentions in the past decade due to the promising potential for the substitute of chemically obtained antibiotics (Omonijo et al., 2017). Species of the genus Dictamnus are perennial herbs widely distributed in Europe and north Asia. There are approximately five species that are placed taxonomically in the family Rutaceae including D. dasycarpus, D. angustifolius, D. caucasicus, D. albus, and D. gymnostylis
⁎
(China Flora Editorial Board of China Science Academy, 2008), two of which i.e. D. dasycarpus and D. angustifolius are the species that populate in China (Gao et al., 2011). The dried root bark of D. dasycarpus, also called Cortex Dictamni, is a traditional herbal medicine prescribed for the relief of skin diseases, cough, jaundice, and rheumatism in China, Japan, and Korea (Kim et al., 2013). Various bioactive constituents have been isolated from Cortex Dictamni, such as sesquiterpenes, alkaloids, flavonoids, limonoids, and glycosides. These beneficial ingredients attribute to the pharmacological functions of neuroprotection, immunosuppression, antifungal and antiallergic properties (Han et al., 2015). In addition, Lei et al. prepared the EOs from Cortex Dictamni (EOCD) by hydrodistillation and observed that syn-7-hydroxy-7-anisylnorbornene (29.4%) is the dominant compound and EOCD exerts strong inhibitory effects on Staphylococcus aureus and methicillin-resistant S. aureus (Lei et al., 2008). In China, D. dasycarpus is mainly cultivated on hill slopes and grassy open forests in northeast China, and over 70% of the total output were dedicated by Jilin, Liaoning, and Heilongjiang provinces. Due to the
Corresponding author at: School of Chemistry and Pharmaceutical Engineering, Jilin Institute of Chemical Technology, Jilin 132022, China. Corresponding author at: School of Biology and Food Engineering, Changshu Institute of Technology, Changshu 215500, China. E-mail addresses: [email protected] (H. Tian), [email protected] (H. Zhao), [email protected] (H. Zhou), [email protected] (Y. Zhang).
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https://doi.org/10.1016/j.indcrop.2019.111713 Received 11 April 2019; Received in revised form 19 August 2019; Accepted 20 August 2019 Available online 28 August 2019 0926-6690/ © 2019 Elsevier B.V. All rights reserved.
Industrial Crops & Products 140 (2019) 111713
H. Tian, et al.
particular planting environment, the cultivation of D. dasycarpus is featured by high cost and high difficulty of planting, leading to an increase in the market price of Cortex Dictamni and an enlargement of planting areas for D. dasycarpus year by year (China Flora Editorial Board of China Science Academy, 2008; Li et al., 2018). The economic value of D. dasycarpus depends on its root bark, which can be processed into Cortex Dictamni, thus at the time of harvesting, substantial leaves and stems of D. dasycarpus were optionally discarded as waste on ground, resulting in dissipation of natural resource and environmental pollution. It is therefore important to investigate the bioactive components derived from the leaves and stems of D. dasycarpus to advance the comprehensive and value-added utilization of this precious medicinal herb. Herein, EOs from the aerial part (leaves and stems) of D. dasycarpus (EOAPDD) was prepared by hydrodistillation, after characterization, the antimicrobial activities against several microbial strains were evaluated. To the best of our knowledge, it is the first time to focus on the EOs from the aerial part of D. dasycarpus, which will underlie the development of EOAPDD as a new antimicrobial agent in the future.
2.4. Antimicrobial activity The minimum inhibitory concentration (MIC), defined as the lowest concentration of EOAPDD to inhibit visible microbial growth, was detected using a broth microdilution assay (Jones, 1984) with some modifications. The microbial strains used here were Staphylococcus aureus ATCC 49775, Candida albicans ATCC 10231, Escherichia coli ATCC 33456, Bacillus pumilus ATCC 7065 and Bacillus subtilis ATCC 6633. Essential oil from the aerial part of D. dasycarpus was dissolved in 20% dimethyl sulfoxide (DMSO)-80% distilled water for an initial concentration of 200 mg/mL, 50 μL of which was first added to a 96well plate, then serial dilutions (100 ˜ 0.78 mg/mL) were obtained by 2fold dilution method and added to each well. Twenty five microliters of suspension containing each microbial strain (106 ˜ 108 CFU/mL) was mixed with 100 mL of liquid medium, respectively, then 150 μL of diluted microbial strain suspension was added to each well. Chloramphenicol in a concentration range from 200 to 0.78 mg/mL was used as positive control and 20% DMSO-80% distilled water without EOAPDD was served as negative control. All assays were repeated in triplicate (Cui et al., 2018).
2. Materials and methods
3. Results and discussion
2.1. Plant material
3.1. Essential oil characterization
The discarded leaves and stems of D. dasycarpus were collected on 26 August 2018 from Zhongxin Farmer Specialized Cooperative Society for the Cultivation of Chinese Medicinal Materials (42°4′52″ N, 125°38′8″ E), Liuhe, Jilin, China, and authenticated by Prof. Guangshu Wang, School of Pharmaceutical Sciences, Jilin University, Changchun, Jilin, China. A voucher specimen with herbarium code DTL0810 was deposited at Jilin Engineering Research Center for Agricultural Resources and Comprehensive Utilization, Jilin Institute of Chemical Technology, Jilin, Jilin, China.
Hydrodistillation of the fresh aerial part of D. dasycarpus gave 0.14% (w/w) essential oil, which was faint yellow in color with aromatic odor. Approximately 39 volatile compounds were identified by GC–MS analysis, corresponding to 94.3% of the total constituents. The chemical profiles of EOAPDD were listed in Table 1. It can be seen that EOAPDD was rich in sesquiterpenoids (37.3%) and monoterpenes (29.4%). Germacrene D was the major volatile compound identified in EOAPDD, accounting for 18.0% followed by terpinolene (13.0%), (Z)-βocimene (10.4%), β-caryophyllene (7.74%), phytol (5.20%), and (+)-viridiflorol (3.16%). The major constituents in essential oil from the root bark of D. dasycarpus (Cortex Dictamni) have been identified to be syn-7-hydroxy-7-anisylnorbornene (29.4%), pregeijerene (15.5%) and geijerene (11.4%) (Lei et al., 2008), which indicated a significant divergence on the chemical compositions of essential oil from the root bark of D. dasycarpus compared with those of the aerial part. This may be due to the facts that active constituents in plant usually accumulate in a tissue-specific manner, and different plant organs possess different enzymes that modulate the biosynthesis of secondary metabolites (Ji et al., 2016). It is common that there are variations in chemical compositions of essential oils in different plant organs for the same species, such as Ruta chalepensis (Tounsi et al., 2011), Artemisia persica (Mirjalili et al., 2006), and Blumea balsamifera (Yuan et al., 2016). Meanwhile, the results also suggested that EOAPDD would be considered as a new natural source of sesquiterpenoids-rich essential oil.
2.2. Essential oil extraction Essential oil from the aerial part of D. dasycarpus was extracted by hydrodistillation in a Clevenger-type apparatus (Xi'an Tefic Biotech Co., Limited, Shaanxi, China). A hundred grams of the fresh leaves and stems of D. dasycarpus were weighed and immersed in distilled water. After 5 h of extraction starting from water boiling, EOAPDD was obtained and then stored at 0℃ in sealed vials before gas chromatography-mass spectrometry (GC–MS) analysis (Deng et al., 2016). 2.3. Chemical composition of essential oil Gas chromatography-mass spectrometry analysis was carried out on a GCMS-QP2010 instrument (Shimadzu, Kyoto, Japan) with a Rxi-5Sil MS column (30 m, 0.25 mm, film thickness 0.25 μm) and interfaced with an ion trap detector. The analytical conditions were as follows: transfer line at 250℃; ion trap at 230℃; helium was used as carrier gas at flow rate of 1.0 mL/min; column temperature was programmed at 5℃/min from 40℃ (isothermal for 4 min) to 60℃, held at 60℃ for 2 min, then from 60℃ to 110℃ at 15℃/min, held at 110℃ for 4 min, raised at 5℃/min up to 180℃, held at 180℃ for 5 min, from 180℃ to 280℃ at 25℃/min, held at 280℃ for 5 min in the end. Then 1.0 μL of diethyl ether solution containing 12 mg/mL of EOAPDD was automatically injected into the vaporizer at a split ratio of 1: 30, under an electron ionization energy of 70 eV and within the scan range of 35 ˜ 500 amu. The identification of each compound was assigned by comparing the retention index in relation to standards of n-alkane mixtures (C8-C40) and by comparing its mass spectrum with either reference data from the equipment database (NIST 05) or from the literatures (Elkady and Ayoub, 2018).
3.2. Antimicrobial activity The antimicrobial activities of EOAPDD were evaluated by the inhibition of C. albicans, S. aureus, B. pumilus, B. subtilis and E. coli, respectively. The MIC values of EOAPDD against the above-mentioned microbial strains were listed in Table 2. It can be seen that EOAPDD exhibits certain inhibitory activities against S. aureus, C. albicans, and E. coli with MICs of 1.56 mg/mL, 3.23 mg/mL, and 10 mg/mL, respectively. Particularly the MIC against C. albicans was almost 4-fold lower than that of chloramphenicol, a chemically obtained antimicrobial agent available on the market. Candida albicans is one of the most common pathogenic fungus in nature, which has caused serials of infective diseases, becoming a serious threat to human health. Meanwhile, the abuse of antibiotics aggravated the harmfulness of C. albicans, more and more diseases related to C. albicans infection have emerged and gained enormous attentions (Delcour, 2009). The results 2
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Table 1 Chemical profiles of EOAPDD. No.
Compound
Molecular formula
RI
RI*
Chemical classification
Content/%
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15.
α-Thujene β-Phellandren Myrcene γ- Terpinene α-Terpinene (Z)-β-Ocimene Terpinolene Linalool 1-Nonanal 3,4,4-Trimethyl-2-cyclohexen-1-one 1-(1,4-Dimethyl-3-cyclohexen-1-yl)-ethanone α-Bulnesene β-Caryophyllene δ-Cadinene 4-(1,5-Dimethyl-1,4-hexadienyl)-1-methyl -cyclohexene Germacrene D (Z)-α-Bisabolene α-Caryophyllene α-Muurolene Nerolidol Spathulenol Ledol Cedrol (+)-Viridiflorol N, N-Dimethyldodecanamide 14-Pentadecenoic acid δ-Dodecanolactone Pentadecanal Hexadecanal Cinoxate Cetyl vinyl ether Linolenyl alcohol 1,3,12-Nonadecatriene Phytol Octadecylacetate (9Z,12Z,15Z)-Ethyl octadeca-9,12,15-trienoate E, E, Z-1,3,12-Nonadecatriene-5,14-diol Geranylgeranyl acetate Dotriacontane Total Monoterpene hydrocarbons Sesquiterpene hydrocarbons Oxygen-containing monoterpenes Oxygen-containing sesquiterpenes Others
C10H16 C10H16 C10H16 C10H16 C10H16 C10H16 C10H16 C10H18O C10H18O C9H14O C10H16O C15H24 C15H24 C15H24 C15H24
902 964 975 998 1018 1049 1052 1082 1104 1097 1241 1490 1494 1525 1518
894 964 966 1003 1031 1053 1034 1100 1104 1110 1235 1499 1504 1519 1509
Alkene Alkene Alkene Alkene Alkene Alkene Alkene Alcohol Aldehyde Ketone Ketone Alkene Alkene Alkene Alkene
1.62 0.61 0.44 2.69 0.61 10.4 13.0 0.36 0.30 0.48 0.63 0.65 7.74 2.10 1.94
C15H24 C15H24 C15H24 C15H24 C15H26O C15H24O C15H26O C15H26O C15H24 C15H16O3 C15H28O2 C16H30O2 C16H32O C16H32O C18H26O3 C18H36O C18H32O C19H34 C20H40O C20H40O2 C20H34O2 C19H34 C22H36O2 C32H66
1515 1518 1519 1499 1524 1536 1530 1543 1530 1614 1579 1623 1718 1700 1830 1876 1901 1916 2045 2177 2201 2241 2310 3202
1514 1515 1516 1516 1525 1532 1533 1538 1567 1597 1599 1610 1711 1705 1802 1812 1844 2047 2072 2214 2205 2243 2285 3194
Alkene Alkene Alkene Alkene Alcohol Alcohol Alcohol Alcohol Alcohol Acylamide Acid Ester Aldehyde Aldehyde Ester Alkane Alcohol Alkene Alcohol Ester Ester Alcohol Acid Alkane
18.0 2.42 0.76 0.53 1.15 2.20 0.52 2.44 3.16 0.49 0.33 3.03 0.87 0.44 0.36 0.47 2.28 0.38 5.20 0.34 1.64 2.43 0.53 0.76 94.3 29.4 37.3 0.63 2.20 24.8
16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. 33. 34. 35. 36. 37. 38. 39.
RI refers to the retention index identified by database NIST 05; RI*refers to the retention index calculated from the retention time relative to that of C8 – C40 n-alkanes; EOAPDD refers to the essential oil from the aerial part of D. dasycarpus.
verified by scanning electronic microscopy and transmission electronic microscopy (Martínez et al., 2014). In this work, there are 37.3% of sesquiterpenoids and 29.4% of monoterpenes present in EOAPDD. These terpenes account for 66.7% and may largely contribute to the pronounced inhibitory activity against C. albicans.
Table 2 MIC values of EOAPDD against several microbial strains.
EOAPDD (mg/mL) Chloramphenicol (mg/ mL)
S. aureus
C. albicans
E. coli
B. pumilus
B. subtilis
1.56 < 0.78
3.23 12.5
10 3.12
100 < 0.78
100 < 0.78
4. Conclusions
MIC refers to the minimum inhibitory concentration; EOAPDD refers to the essential oil from the aerial part of D. dasycarpus.
With the aim to promote the comprehensive utilization of D. dasycarpus, the chemical composition and antimicrobial activity of the essential oil derived from the wasted aerial part of D. dasycarpus (EOAPDD) were reported for the first time. There are differences in chemical components of the essential oils between the aerial part and the root bark of D. dasycarpus. Essential oil from the aerial part of D. dasycarpus is rich in sesquiterpenes (germacrene D, β-caryophyllene, and (+)-viridiflorol, etc.) and monoterpenes (terpinolene and β-ocimene, etc.), and exerts obvious inhibitory effects on C. albicans, which reveals the potential of EOAPDD to be used as a novel nature-based antifungal agent.
showed that EOAPDD exhibits more inhibitory effects against C. albicans than those of other microbial strains, indicating a promising potential to be developed as nature-based antifungal agent. Some volatile components in EOAPDD, especially major constituents may contribute to the antifungal activity. Terpenes are natural products that play distinct roles in different organisms where they are presented. Most of terpenes are volatile compounds, which are abundant in essential oils and share a wide range of structural diversity (Gonzalez-Burgos and Gomez-Serranillos, 2012). Terpenes exhibit significant inhibitory capacity against C. albicans via damaging cell wall, which has been 3
Industrial Crops & Products 140 (2019) 111713
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Declaration of Competing Interest
Han, H.Y., Ryu, M.H., Lee, G., Cheon, W.J., Lee, C., An, W.G., Kim, H., Cho, S.I., 2015. Effects of Dictamnus dasycarpus Turcz., root bark on ICAM-1 expression and chemokine productions in vivo and vitro study. J. Ethnopharmacol. 159, 245–252. https:// doi.org/10.1016/j.jep.2014.11.020. Ji, A.J., Luo, H.M., Xu, Z.C., Zhang, X., Zhu, Y.J., Liao, B.S., Yao, H., Song, J.Y., Chen, S.L., 2016. Genome-wide identification of the AP2/ERF gene family involved in active constituent biosynthesis in Salvia miltiorrhiza. Plant Genome 9, 1–11. https://doi. org/10.3835/plantgenome2015.08.0077. Jones, R.N., 1984. NCCLS guidelines: revised performance standards for antimicrobial disk susceptibility tests. Antimicrob. Newsl. 1, 64–65. https://doi.org/10.1016/07381751(84)90027-3. Kim, H., Kim, M., Kim, H., Lee, G.S., An, W.G., Cho, S.I., 2013. Anti-inflammatory activities of Dictamnus dasycarpus Turcz., root bark on allergic contact dermatitis induced by dinitrofluorobenzene in mice. J. Ethnopharmacol. 149, 471–477. https:// doi.org/10.1016/j.jep.2013.06.055. Li, J.S., Nie, L.J., Ji, Y.H., Li, X.L., Chang, Y.F., Li, Z.J., 2018. Establishment of quality evaluation system of standard decoction of Dictamni Cortex pieces. Hebei J. Ind. Sci. Technol. 35, 370–376. https://doi.org/10.7535/hbgykj.2018yx05011. Lei, J., Yu, J., Yu, H., Liao, Z., 2008. Composition, cytotoxicity and antimicrobial activity of essential oil from Dictamnus dasycarpus. Food Chem. 107, 1205–1209. https://doi. org/10.1016/j.foodchem.2007.09.050. Martínez, A., Rojas, N., García, L., González, F., Domínguez, M., Catalán, A., 2014. In vitro activity of terpenes against Candida albicans and ultrastructural alterations. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. 118, 553–559. https://doi.org/10.1016/j. oooo.2014.07.009. Mirjalili, B.F., Meybody, M.H.H., Ardakani, M.M., Rustaiyan, A., Ameri, N., Masoudi, S., Bamoniri, A., 2006. Chemical composition of the essential oil from aerial parts, leaves, flowers and roots of Artemisia persica Boiss. From Iran. J. Essent. Oil Res. 18, 544–547. https://doi.org/10.1080/10412905.2006.9699162. Omonijo, F.A., Ni, L., Gong, J., Wang, Q., Lahaye, L., Yang, C., 2017. Essential oils as alternatives to antibiotics in swine production. Anim. Nutr. Feed Technol. 4, 126–136. https://doi.org/10.1016/j.aninu.2017.09.001. Raut, J.S., Karuppayil, S.M., 2014. Bioprospecting of plant essential oils for medicinal uses. In: Fulekar, M., Pathak, B., Kale, R. (Eds.), Environment and Sustainable Development. Springer, New Delhi, pp. 59–76. https://doi.org/10.1007/978-81-3221166-2_5. Tounsi, M.S., Wannes, W.A., Ouerghemmi, I., Msaada, K., Smaoui, A., Marzouk, B., 2011. Variation in essential oil and fatty acid composition in different organs of cultivated and growing wild Ruta chalepensis L. Ind. Crop. Prod. 33, 617–623. https://doi.org/ 10.1016/j.indcrop.2010.12.028. Yuan, Y., Huang, M., Pang, Y.X., Yu, F.L., Chen, C., Liu, L.W., Chen, Z.X., Zhang, Y.B., Chen, X.L., Hu, X., 2016. Variations in essential oil yield, composition, and antioxidant activity of different plant organs from Blumea balsamifera (L.) DC. At different growth times. Molecules 21, E1024. https://doi.org/10.3390/molecules21081024. Zhang, J., Huang, R.Z., Cao, H.J., Cheng, A.W., Jiang, C.S., Liao, Z.X., Liu, C., Sun, J.Y., 2018. Chemical composition, in vitro anti-tumor activities and related mechanisms of the essential oil from the roots of Potentilla discolor. Ind. Crop. Prod. 113, 19–27. https://doi.org/10.1016/j.indcrop.2017.12.071.
None. Acknowledgments This investigation was supported by the programs of Jilin Province Development and Reform Commission (Grant No. 2019C045-4), the Science and Technology Bureau of Jilin City (Grant No. 201851869), the Science and Technology Department of Jilin province (Grant No. 20190304102YY), and 2019 Team-Guided Graduation Thesis of Changshu Institute of Technology. References Bajer, T., Surmová, S., Eisner, A., Ventura, K., Bajerová, P., 2018. Use of simultaneous distillation-extraction, supercritical fluid extraction and solid-phase microextraction for characterisation of the volatile profile of Dipteryx odorata (Aubl.) willd. Ind. Crop. Prod. 119, 313–321. https://doi.org/10.1016/j.indcrop.2018.01.055. Cui, H., Pan, H.W., Wang, P.H., Yang, X.D., Zhai, W.C., Dong, Y., Zhou, H.L., 2018. Essential oils from Carex meyeriana Kunth: optimization of hydrodistillation extraction by response surface methodology and evaluation of its antioxidant and antimicrobial activities. Ind. Crop. Prod. 124, 669–676. https://doi.org/10.1016/j. indcrop.2018.08.041. China Flora Editorial Board of China Science Academy, 2008. Dictamnus Linnaeus. In: Zhang, D.X., Thomas, G.H. (Eds.), Flora of China. Science Press, Beijing, pp. 74–75. Deng, J.D., He, B., He, D.H., Chen, Z.F., 2016. A potential biopreservative: chemical composition, antibacterial andhemolytic activities of leaves essential oil from Alpinia guinanensis. Ind. Crop. Prod 94, 281–287. https://doi.org/10.1016/j.indcrop.2016. 09.004. Delcour, A.H., 2009. Outer membrane permeability and antibiotic resistance. Biochim. Biophys. Acta 1794, 808–816. https://doi.org/10.1016/j.bbapap.2008.11.005. Elkady, W.M., Ayoub, I.M., 2018. Chemical profiling and antiproliferative effect of essential oils of two Araucaria species cultivated in Egypt. Ind. Crop. Prod. 118, 188–195. https://doi.org/10.1016/j.indcrop.2018.03.051. Edris, A.E., 2007. Pharmaceutical and therapeutic potentials of essential oils and their individual volatile constituents: a review. Phytother. Res. 21, 308–323. https://doi. org/10.1002/ptr.2072. Gao, X., Zhao, P.H., Hu, J.F., 2011. Chemical constituents of plants from the genus Dictamnus. Chem. Biodivers. 8, 1234–1244. https://doi.org/10.1002/cbdv. 201000132. Gonzalez-Burgos, E., Gomez-Serranillos, M.P., 2012. Terpene compounds in nature: a review of their potential antioxidant activity. Curr. Med. Chem. 19, 5319–5341. https://doi.org/10.2174/092986712803833335.
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Contents lists available at ScienceDirect
Industrial Crops & Products journal homepage: www.elsevier.com/locate/indcrop
Short communication
Chemical composition and antimicrobial activity of the essential oil from the aerial part of Dictamnus dasycarpus Turcz.
T
Haiping Tiana, Hepeng Zhaoa,b, Hongli Zhoua,b, , Yang Zhangc, ⁎
⁎⁎
a
School of Chemistry and Pharmaceutical Engineering, Jilin Institute of Chemical Technology, Jilin 132022, China Engineering Research Center for Agricultural Resources and Comprehensive Utilization of Jilin Provence, Jilin Institute of Chemical Technology, Jilin 132022, China c School of Biology and Food Engineering, Changshu Institute of Technology, Changshu 215500, China b
ARTICLE INFO
ABSTRACT
Keywords: Dictamnus dasycarpus Turcz. Aerial part Chemical composition Antimicrobial activity
Dictamnus dasycarpus Turcz. is an economic plant widely cultivated in China and its dried root bark is commonly prescribed as Cortex Dictamni for the treatment of various inflammatory diseases. Thus, more attention was devoted to keep the high-quality of the root bark of D. dasycarpus, but leaving the aerial part on ground as waste. In this context, we focused on the chemical composition and antimicrobial activity of the essential oil from the aerial part of D. dasycarpus (EOAPDD) for the first time. The extraction yield of EOAPDD was 0.14% by hydrodistillation, and thirty-nine compounds were identified by gas chromatography-mass spectrometry with the highest proportion of sesquiterpenoids (37.3%), followed by monoterpenes (29.4%). The dominant active constituents in EOAPDD are germacrene D (18.0%), terpinolene (13.0%), (Z)-β-ocimene (10.4%), β-caryophyllene (7.74%), phytol (5.20%), and (+)-viridiflorol (3.16%). Essential oil from the aerial part of D. dasycarpus shows pronounced inhibitory activity against Candida albicans with minimum inhibitory concentration of 3.23 mg/mL (4-fold lower than that of chloramphenicol), which may be largely contributed by the terpene compounds (37.3% of sesquiterpenoids and 29.4% of monoterpenes) in EOAPDD. Results obtained from present investigation provided strong evidence that the aerial part of D. dasycarpus also deserves attention, cherishment and deep exploitation due to the promising potential as a nature-based antifungal agent.
1. Introduction Essential oils (EOs), produced by plants as secondary metabolites are usually composed of different compounds with relatively small molecular weights and normally possess remarkable aromatic odor (Zhang et al., 2018). Based on chemical classification, EOs can be mainly divided into aliphatics, aromatics, and terpenoids, and are characterized by low boiling point and high volatility in most cases (Raut and Karuppayil, 2014; Bajer et al., 2018). Essential oils elicit diverse bioactivities including antimicrobial, antioxidant, antidiabetic, and anticancer properties (Edris, 2007), particularly their antimicrobial activities have gained enormous attentions in the past decade due to the promising potential for the substitute of chemically obtained antibiotics (Omonijo et al., 2017). Species of the genus Dictamnus are perennial herbs widely distributed in Europe and north Asia. There are approximately five species that are placed taxonomically in the family Rutaceae including D. dasycarpus, D. angustifolius, D. caucasicus, D. albus, and D. gymnostylis
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(China Flora Editorial Board of China Science Academy, 2008), two of which i.e. D. dasycarpus and D. angustifolius are the species that populate in China (Gao et al., 2011). The dried root bark of D. dasycarpus, also called Cortex Dictamni, is a traditional herbal medicine prescribed for the relief of skin diseases, cough, jaundice, and rheumatism in China, Japan, and Korea (Kim et al., 2013). Various bioactive constituents have been isolated from Cortex Dictamni, such as sesquiterpenes, alkaloids, flavonoids, limonoids, and glycosides. These beneficial ingredients attribute to the pharmacological functions of neuroprotection, immunosuppression, antifungal and antiallergic properties (Han et al., 2015). In addition, Lei et al. prepared the EOs from Cortex Dictamni (EOCD) by hydrodistillation and observed that syn-7-hydroxy-7-anisylnorbornene (29.4%) is the dominant compound and EOCD exerts strong inhibitory effects on Staphylococcus aureus and methicillin-resistant S. aureus (Lei et al., 2008). In China, D. dasycarpus is mainly cultivated on hill slopes and grassy open forests in northeast China, and over 70% of the total output were dedicated by Jilin, Liaoning, and Heilongjiang provinces. Due to the
Corresponding author at: School of Chemistry and Pharmaceutical Engineering, Jilin Institute of Chemical Technology, Jilin 132022, China. Corresponding author at: School of Biology and Food Engineering, Changshu Institute of Technology, Changshu 215500, China. E-mail addresses: [email protected] (H. Tian), [email protected] (H. Zhao), [email protected] (H. Zhou), [email protected] (Y. Zhang).
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https://doi.org/10.1016/j.indcrop.2019.111713 Received 11 April 2019; Received in revised form 19 August 2019; Accepted 20 August 2019 Available online 28 August 2019 0926-6690/ © 2019 Elsevier B.V. All rights reserved.
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particular planting environment, the cultivation of D. dasycarpus is featured by high cost and high difficulty of planting, leading to an increase in the market price of Cortex Dictamni and an enlargement of planting areas for D. dasycarpus year by year (China Flora Editorial Board of China Science Academy, 2008; Li et al., 2018). The economic value of D. dasycarpus depends on its root bark, which can be processed into Cortex Dictamni, thus at the time of harvesting, substantial leaves and stems of D. dasycarpus were optionally discarded as waste on ground, resulting in dissipation of natural resource and environmental pollution. It is therefore important to investigate the bioactive components derived from the leaves and stems of D. dasycarpus to advance the comprehensive and value-added utilization of this precious medicinal herb. Herein, EOs from the aerial part (leaves and stems) of D. dasycarpus (EOAPDD) was prepared by hydrodistillation, after characterization, the antimicrobial activities against several microbial strains were evaluated. To the best of our knowledge, it is the first time to focus on the EOs from the aerial part of D. dasycarpus, which will underlie the development of EOAPDD as a new antimicrobial agent in the future.
2.4. Antimicrobial activity The minimum inhibitory concentration (MIC), defined as the lowest concentration of EOAPDD to inhibit visible microbial growth, was detected using a broth microdilution assay (Jones, 1984) with some modifications. The microbial strains used here were Staphylococcus aureus ATCC 49775, Candida albicans ATCC 10231, Escherichia coli ATCC 33456, Bacillus pumilus ATCC 7065 and Bacillus subtilis ATCC 6633. Essential oil from the aerial part of D. dasycarpus was dissolved in 20% dimethyl sulfoxide (DMSO)-80% distilled water for an initial concentration of 200 mg/mL, 50 μL of which was first added to a 96well plate, then serial dilutions (100 ˜ 0.78 mg/mL) were obtained by 2fold dilution method and added to each well. Twenty five microliters of suspension containing each microbial strain (106 ˜ 108 CFU/mL) was mixed with 100 mL of liquid medium, respectively, then 150 μL of diluted microbial strain suspension was added to each well. Chloramphenicol in a concentration range from 200 to 0.78 mg/mL was used as positive control and 20% DMSO-80% distilled water without EOAPDD was served as negative control. All assays were repeated in triplicate (Cui et al., 2018).
2. Materials and methods
3. Results and discussion
2.1. Plant material
3.1. Essential oil characterization
The discarded leaves and stems of D. dasycarpus were collected on 26 August 2018 from Zhongxin Farmer Specialized Cooperative Society for the Cultivation of Chinese Medicinal Materials (42°4′52″ N, 125°38′8″ E), Liuhe, Jilin, China, and authenticated by Prof. Guangshu Wang, School of Pharmaceutical Sciences, Jilin University, Changchun, Jilin, China. A voucher specimen with herbarium code DTL0810 was deposited at Jilin Engineering Research Center for Agricultural Resources and Comprehensive Utilization, Jilin Institute of Chemical Technology, Jilin, Jilin, China.
Hydrodistillation of the fresh aerial part of D. dasycarpus gave 0.14% (w/w) essential oil, which was faint yellow in color with aromatic odor. Approximately 39 volatile compounds were identified by GC–MS analysis, corresponding to 94.3% of the total constituents. The chemical profiles of EOAPDD were listed in Table 1. It can be seen that EOAPDD was rich in sesquiterpenoids (37.3%) and monoterpenes (29.4%). Germacrene D was the major volatile compound identified in EOAPDD, accounting for 18.0% followed by terpinolene (13.0%), (Z)-βocimene (10.4%), β-caryophyllene (7.74%), phytol (5.20%), and (+)-viridiflorol (3.16%). The major constituents in essential oil from the root bark of D. dasycarpus (Cortex Dictamni) have been identified to be syn-7-hydroxy-7-anisylnorbornene (29.4%), pregeijerene (15.5%) and geijerene (11.4%) (Lei et al., 2008), which indicated a significant divergence on the chemical compositions of essential oil from the root bark of D. dasycarpus compared with those of the aerial part. This may be due to the facts that active constituents in plant usually accumulate in a tissue-specific manner, and different plant organs possess different enzymes that modulate the biosynthesis of secondary metabolites (Ji et al., 2016). It is common that there are variations in chemical compositions of essential oils in different plant organs for the same species, such as Ruta chalepensis (Tounsi et al., 2011), Artemisia persica (Mirjalili et al., 2006), and Blumea balsamifera (Yuan et al., 2016). Meanwhile, the results also suggested that EOAPDD would be considered as a new natural source of sesquiterpenoids-rich essential oil.
2.2. Essential oil extraction Essential oil from the aerial part of D. dasycarpus was extracted by hydrodistillation in a Clevenger-type apparatus (Xi'an Tefic Biotech Co., Limited, Shaanxi, China). A hundred grams of the fresh leaves and stems of D. dasycarpus were weighed and immersed in distilled water. After 5 h of extraction starting from water boiling, EOAPDD was obtained and then stored at 0℃ in sealed vials before gas chromatography-mass spectrometry (GC–MS) analysis (Deng et al., 2016). 2.3. Chemical composition of essential oil Gas chromatography-mass spectrometry analysis was carried out on a GCMS-QP2010 instrument (Shimadzu, Kyoto, Japan) with a Rxi-5Sil MS column (30 m, 0.25 mm, film thickness 0.25 μm) and interfaced with an ion trap detector. The analytical conditions were as follows: transfer line at 250℃; ion trap at 230℃; helium was used as carrier gas at flow rate of 1.0 mL/min; column temperature was programmed at 5℃/min from 40℃ (isothermal for 4 min) to 60℃, held at 60℃ for 2 min, then from 60℃ to 110℃ at 15℃/min, held at 110℃ for 4 min, raised at 5℃/min up to 180℃, held at 180℃ for 5 min, from 180℃ to 280℃ at 25℃/min, held at 280℃ for 5 min in the end. Then 1.0 μL of diethyl ether solution containing 12 mg/mL of EOAPDD was automatically injected into the vaporizer at a split ratio of 1: 30, under an electron ionization energy of 70 eV and within the scan range of 35 ˜ 500 amu. The identification of each compound was assigned by comparing the retention index in relation to standards of n-alkane mixtures (C8-C40) and by comparing its mass spectrum with either reference data from the equipment database (NIST 05) or from the literatures (Elkady and Ayoub, 2018).
3.2. Antimicrobial activity The antimicrobial activities of EOAPDD were evaluated by the inhibition of C. albicans, S. aureus, B. pumilus, B. subtilis and E. coli, respectively. The MIC values of EOAPDD against the above-mentioned microbial strains were listed in Table 2. It can be seen that EOAPDD exhibits certain inhibitory activities against S. aureus, C. albicans, and E. coli with MICs of 1.56 mg/mL, 3.23 mg/mL, and 10 mg/mL, respectively. Particularly the MIC against C. albicans was almost 4-fold lower than that of chloramphenicol, a chemically obtained antimicrobial agent available on the market. Candida albicans is one of the most common pathogenic fungus in nature, which has caused serials of infective diseases, becoming a serious threat to human health. Meanwhile, the abuse of antibiotics aggravated the harmfulness of C. albicans, more and more diseases related to C. albicans infection have emerged and gained enormous attentions (Delcour, 2009). The results 2
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Table 1 Chemical profiles of EOAPDD. No.
Compound
Molecular formula
RI
RI*
Chemical classification
Content/%
1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15.
α-Thujene β-Phellandren Myrcene γ- Terpinene α-Terpinene (Z)-β-Ocimene Terpinolene Linalool 1-Nonanal 3,4,4-Trimethyl-2-cyclohexen-1-one 1-(1,4-Dimethyl-3-cyclohexen-1-yl)-ethanone α-Bulnesene β-Caryophyllene δ-Cadinene 4-(1,5-Dimethyl-1,4-hexadienyl)-1-methyl -cyclohexene Germacrene D (Z)-α-Bisabolene α-Caryophyllene α-Muurolene Nerolidol Spathulenol Ledol Cedrol (+)-Viridiflorol N, N-Dimethyldodecanamide 14-Pentadecenoic acid δ-Dodecanolactone Pentadecanal Hexadecanal Cinoxate Cetyl vinyl ether Linolenyl alcohol 1,3,12-Nonadecatriene Phytol Octadecylacetate (9Z,12Z,15Z)-Ethyl octadeca-9,12,15-trienoate E, E, Z-1,3,12-Nonadecatriene-5,14-diol Geranylgeranyl acetate Dotriacontane Total Monoterpene hydrocarbons Sesquiterpene hydrocarbons Oxygen-containing monoterpenes Oxygen-containing sesquiterpenes Others
C10H16 C10H16 C10H16 C10H16 C10H16 C10H16 C10H16 C10H18O C10H18O C9H14O C10H16O C15H24 C15H24 C15H24 C15H24
902 964 975 998 1018 1049 1052 1082 1104 1097 1241 1490 1494 1525 1518
894 964 966 1003 1031 1053 1034 1100 1104 1110 1235 1499 1504 1519 1509
Alkene Alkene Alkene Alkene Alkene Alkene Alkene Alcohol Aldehyde Ketone Ketone Alkene Alkene Alkene Alkene
1.62 0.61 0.44 2.69 0.61 10.4 13.0 0.36 0.30 0.48 0.63 0.65 7.74 2.10 1.94
C15H24 C15H24 C15H24 C15H24 C15H26O C15H24O C15H26O C15H26O C15H24 C15H16O3 C15H28O2 C16H30O2 C16H32O C16H32O C18H26O3 C18H36O C18H32O C19H34 C20H40O C20H40O2 C20H34O2 C19H34 C22H36O2 C32H66
1515 1518 1519 1499 1524 1536 1530 1543 1530 1614 1579 1623 1718 1700 1830 1876 1901 1916 2045 2177 2201 2241 2310 3202
1514 1515 1516 1516 1525 1532 1533 1538 1567 1597 1599 1610 1711 1705 1802 1812 1844 2047 2072 2214 2205 2243 2285 3194
Alkene Alkene Alkene Alkene Alcohol Alcohol Alcohol Alcohol Alcohol Acylamide Acid Ester Aldehyde Aldehyde Ester Alkane Alcohol Alkene Alcohol Ester Ester Alcohol Acid Alkane
18.0 2.42 0.76 0.53 1.15 2.20 0.52 2.44 3.16 0.49 0.33 3.03 0.87 0.44 0.36 0.47 2.28 0.38 5.20 0.34 1.64 2.43 0.53 0.76 94.3 29.4 37.3 0.63 2.20 24.8
16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. 33. 34. 35. 36. 37. 38. 39.
RI refers to the retention index identified by database NIST 05; RI*refers to the retention index calculated from the retention time relative to that of C8 – C40 n-alkanes; EOAPDD refers to the essential oil from the aerial part of D. dasycarpus.
verified by scanning electronic microscopy and transmission electronic microscopy (Martínez et al., 2014). In this work, there are 37.3% of sesquiterpenoids and 29.4% of monoterpenes present in EOAPDD. These terpenes account for 66.7% and may largely contribute to the pronounced inhibitory activity against C. albicans.
Table 2 MIC values of EOAPDD against several microbial strains.
EOAPDD (mg/mL) Chloramphenicol (mg/ mL)
S. aureus
C. albicans
E. coli
B. pumilus
B. subtilis
1.56 < 0.78
3.23 12.5
10 3.12
100 < 0.78
100 < 0.78
4. Conclusions
MIC refers to the minimum inhibitory concentration; EOAPDD refers to the essential oil from the aerial part of D. dasycarpus.
With the aim to promote the comprehensive utilization of D. dasycarpus, the chemical composition and antimicrobial activity of the essential oil derived from the wasted aerial part of D. dasycarpus (EOAPDD) were reported for the first time. There are differences in chemical components of the essential oils between the aerial part and the root bark of D. dasycarpus. Essential oil from the aerial part of D. dasycarpus is rich in sesquiterpenes (germacrene D, β-caryophyllene, and (+)-viridiflorol, etc.) and monoterpenes (terpinolene and β-ocimene, etc.), and exerts obvious inhibitory effects on C. albicans, which reveals the potential of EOAPDD to be used as a novel nature-based antifungal agent.
showed that EOAPDD exhibits more inhibitory effects against C. albicans than those of other microbial strains, indicating a promising potential to be developed as nature-based antifungal agent. Some volatile components in EOAPDD, especially major constituents may contribute to the antifungal activity. Terpenes are natural products that play distinct roles in different organisms where they are presented. Most of terpenes are volatile compounds, which are abundant in essential oils and share a wide range of structural diversity (Gonzalez-Burgos and Gomez-Serranillos, 2012). Terpenes exhibit significant inhibitory capacity against C. albicans via damaging cell wall, which has been 3
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Declaration of Competing Interest
Han, H.Y., Ryu, M.H., Lee, G., Cheon, W.J., Lee, C., An, W.G., Kim, H., Cho, S.I., 2015. Effects of Dictamnus dasycarpus Turcz., root bark on ICAM-1 expression and chemokine productions in vivo and vitro study. J. Ethnopharmacol. 159, 245–252. https:// doi.org/10.1016/j.jep.2014.11.020. Ji, A.J., Luo, H.M., Xu, Z.C., Zhang, X., Zhu, Y.J., Liao, B.S., Yao, H., Song, J.Y., Chen, S.L., 2016. Genome-wide identification of the AP2/ERF gene family involved in active constituent biosynthesis in Salvia miltiorrhiza. Plant Genome 9, 1–11. https://doi. org/10.3835/plantgenome2015.08.0077. Jones, R.N., 1984. NCCLS guidelines: revised performance standards for antimicrobial disk susceptibility tests. Antimicrob. Newsl. 1, 64–65. https://doi.org/10.1016/07381751(84)90027-3. Kim, H., Kim, M., Kim, H., Lee, G.S., An, W.G., Cho, S.I., 2013. Anti-inflammatory activities of Dictamnus dasycarpus Turcz., root bark on allergic contact dermatitis induced by dinitrofluorobenzene in mice. J. Ethnopharmacol. 149, 471–477. https:// doi.org/10.1016/j.jep.2013.06.055. Li, J.S., Nie, L.J., Ji, Y.H., Li, X.L., Chang, Y.F., Li, Z.J., 2018. Establishment of quality evaluation system of standard decoction of Dictamni Cortex pieces. Hebei J. Ind. Sci. Technol. 35, 370–376. https://doi.org/10.7535/hbgykj.2018yx05011. Lei, J., Yu, J., Yu, H., Liao, Z., 2008. Composition, cytotoxicity and antimicrobial activity of essential oil from Dictamnus dasycarpus. Food Chem. 107, 1205–1209. https://doi. org/10.1016/j.foodchem.2007.09.050. Martínez, A., Rojas, N., García, L., González, F., Domínguez, M., Catalán, A., 2014. In vitro activity of terpenes against Candida albicans and ultrastructural alterations. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. 118, 553–559. https://doi.org/10.1016/j. oooo.2014.07.009. Mirjalili, B.F., Meybody, M.H.H., Ardakani, M.M., Rustaiyan, A., Ameri, N., Masoudi, S., Bamoniri, A., 2006. Chemical composition of the essential oil from aerial parts, leaves, flowers and roots of Artemisia persica Boiss. From Iran. J. Essent. Oil Res. 18, 544–547. https://doi.org/10.1080/10412905.2006.9699162. Omonijo, F.A., Ni, L., Gong, J., Wang, Q., Lahaye, L., Yang, C., 2017. Essential oils as alternatives to antibiotics in swine production. Anim. Nutr. Feed Technol. 4, 126–136. https://doi.org/10.1016/j.aninu.2017.09.001. Raut, J.S., Karuppayil, S.M., 2014. Bioprospecting of plant essential oils for medicinal uses. In: Fulekar, M., Pathak, B., Kale, R. (Eds.), Environment and Sustainable Development. Springer, New Delhi, pp. 59–76. https://doi.org/10.1007/978-81-3221166-2_5. Tounsi, M.S., Wannes, W.A., Ouerghemmi, I., Msaada, K., Smaoui, A., Marzouk, B., 2011. Variation in essential oil and fatty acid composition in different organs of cultivated and growing wild Ruta chalepensis L. Ind. Crop. Prod. 33, 617–623. https://doi.org/ 10.1016/j.indcrop.2010.12.028. Yuan, Y., Huang, M., Pang, Y.X., Yu, F.L., Chen, C., Liu, L.W., Chen, Z.X., Zhang, Y.B., Chen, X.L., Hu, X., 2016. Variations in essential oil yield, composition, and antioxidant activity of different plant organs from Blumea balsamifera (L.) DC. At different growth times. Molecules 21, E1024. https://doi.org/10.3390/molecules21081024. Zhang, J., Huang, R.Z., Cao, H.J., Cheng, A.W., Jiang, C.S., Liao, Z.X., Liu, C., Sun, J.Y., 2018. Chemical composition, in vitro anti-tumor activities and related mechanisms of the essential oil from the roots of Potentilla discolor. Ind. Crop. Prod. 113, 19–27. https://doi.org/10.1016/j.indcrop.2017.12.071.
None. Acknowledgments This investigation was supported by the programs of Jilin Province Development and Reform Commission (Grant No. 2019C045-4), the Science and Technology Bureau of Jilin City (Grant No. 201851869), the Science and Technology Department of Jilin province (Grant No. 20190304102YY), and 2019 Team-Guided Graduation Thesis of Changshu Institute of Technology. References Bajer, T., Surmová, S., Eisner, A., Ventura, K., Bajerová, P., 2018. Use of simultaneous distillation-extraction, supercritical fluid extraction and solid-phase microextraction for characterisation of the volatile profile of Dipteryx odorata (Aubl.) willd. Ind. Crop. Prod. 119, 313–321. https://doi.org/10.1016/j.indcrop.2018.01.055. Cui, H., Pan, H.W., Wang, P.H., Yang, X.D., Zhai, W.C., Dong, Y., Zhou, H.L., 2018. Essential oils from Carex meyeriana Kunth: optimization of hydrodistillation extraction by response surface methodology and evaluation of its antioxidant and antimicrobial activities. Ind. Crop. Prod. 124, 669–676. https://doi.org/10.1016/j. indcrop.2018.08.041. China Flora Editorial Board of China Science Academy, 2008. Dictamnus Linnaeus. In: Zhang, D.X., Thomas, G.H. (Eds.), Flora of China. Science Press, Beijing, pp. 74–75. Deng, J.D., He, B., He, D.H., Chen, Z.F., 2016. A potential biopreservative: chemical composition, antibacterial andhemolytic activities of leaves essential oil from Alpinia guinanensis. Ind. Crop. Prod 94, 281–287. https://doi.org/10.1016/j.indcrop.2016. 09.004. Delcour, A.H., 2009. Outer membrane permeability and antibiotic resistance. Biochim. Biophys. Acta 1794, 808–816. https://doi.org/10.1016/j.bbapap.2008.11.005. Elkady, W.M., Ayoub, I.M., 2018. Chemical profiling and antiproliferative effect of essential oils of two Araucaria species cultivated in Egypt. Ind. Crop. Prod. 118, 188–195. https://doi.org/10.1016/j.indcrop.2018.03.051. Edris, A.E., 2007. Pharmaceutical and therapeutic potentials of essential oils and their individual volatile constituents: a review. Phytother. Res. 21, 308–323. https://doi. org/10.1002/ptr.2072. Gao, X., Zhao, P.H., Hu, J.F., 2011. Chemical constituents of plants from the genus Dictamnus. Chem. Biodivers. 8, 1234–1244. https://doi.org/10.1002/cbdv. 201000132. Gonzalez-Burgos, E., Gomez-Serranillos, M.P., 2012. Terpene compounds in nature: a review of their potential antioxidant activity. Curr. Med. Chem. 19, 5319–5341. https://doi.org/10.2174/092986712803833335.
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