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ORIGINAL ARTICLE
Year : 2020  |  Volume : 16  |  Issue : 71  |  Page : 625-629  

A special aromatic Chrysanthemum breed with high content of thujone


1 College of Pharmacy, Key Laboratory of Chinese Medicine Resources and Compound Prescription, Ministry of Education and Hubei University of Chinese Medicine, Wuhan, Hubei Province, China
2 Hubei Bencaohui Agricultural Development Co. Ltd., Xianning City, Hubei Province, China

Date of Submission22-Aug-2019
Date of Decision05-Nov-2019
Date of Acceptance06-May-2020
Date of Web Publication20-Oct-2020

Correspondence Address:
Keli Chen
Key Laboratory of Chinese Medicine Resources and Compound Prescription, Ministry of Education and Hubei University of Chinese Medicine, Wuhan 430065
China
Yimei Liu
Key Laboratory of Chinese Medicine Resources and Compound Prescription, Ministry of Education and Hubei University of Chinese Medicine, Wuhan
China
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/pm.pm_382_19

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   Abstract 


Background: A cultivar of Chrysanthemum, whose whole plant has strong and special fragrance, was found in Jiugong Mountain area, Xianning City, Hubei Province. Objective: The objective of the study was to determine taxonomic status of the cultivar in genus Dendranthema. Materials and Methods: The differences of morphological characteristics and volatile oil components between this cultivar and Dendranthema indicum, D. indicum var. aromaticum, and Dendranthema molifolium cv. “Jinju” were compared. Results: The cultivar was found to be a Chrysanthemum breed with unisexual flowers. The volatile oil content was 0.25% in its stem and leave of the breed and about 1.81% in its inflorescence which was much higher than that of other 3 species. There are 17 common components among volatile oils in inflorescence of the four Chrysanthemum species, but the thujone content of this cultivar is much higher than other three species. Conclusions: The cultivar is a new, cultivated Chrysanthemum breed, rich in volatile oil and thujone content, and named as Dendranthema morifolium (Ramat.) Tzvel. cv. “Jiugong Xiangju.”

Keywords: Cultivated breed, Dendranthema molifolium,tujone, volatile oil


How to cite this article:
Zhang Y, Cheng R, Xu Z, She Y, Chen K, Liu Y. A special aromatic Chrysanthemum breed with high content of thujone. Phcog Mag 2020;16:625-9

How to cite this URL:
Zhang Y, Cheng R, Xu Z, She Y, Chen K, Liu Y. A special aromatic Chrysanthemum breed with high content of thujone. Phcog Mag [serial online] 2020 [cited 2020 Dec 5];16:625-9. Available from: http://www.phcog.com/text.asp?2020/16/71/625/298650



SUMMARY

  • A cultivar of Chrysanthemum which was found in Jiugong Mountain area, Xianning City, Hubei Province can be identified by comparing morphological characteristics and volatile oil components with Dendranthema indicum, D. indicum var. aromaticum, and Dendranthema molifolium cv.“Jinju”. The cultivar is a new cultivated Chrysanthemum breed, rich in volatile oil and thujone content,and named as Dendranthema morifolium (Ramat.) Tzvel. cv. “Jiugong Xiangju”




Abbreviations used: G: Dendranthema morifolium (Ramat.) Tzvel. cv. “Jiugong Xiangju”; J: Dendranthema molifolium (Ramat.) Tzvel. cv. “Jinju”; Y: Dendranthema indicum (L.) Des Moul; S: Dendranthema indicum var. aromaticum Q. H. Liu et S. F. Zhang.


   Introduction Top


Medicinal Chrysanthemum is the capitulum of Dendranthema molifolium (Ramat.) Tzvel. (Family Compositae) and it is a famous medicinal and edible plant material with a long history of cultivation in China. In Xianning City of Hubei Province in China, a Chrysanthemum cultivar was found, which stems, leaves, and flowers have strong fragrance. The cultivar is cultivated in the geographical and ecological area around the Jiugong Mountains which is located in Hubei and Jiangxi province and has long application history as both edible and medicinal Chrysanthemum in Xianning City of Hubei Province and Jiujiang City of Jiangxi Province. It is believed that the cultivar has a good effect on relieving eye fatigue and improving eye diseases. Due to its aromatic smell of stems and leaves and growing in the Jiugong Mountains, it is named as Jiugong Xiangju (JGXJ).

In Dendranthema genus, plants with fragrance throughout the whole plant are relatively rare; two species of them are found in Hubei Province. They are Dendranthema indicum var. aromaticum Liuet al.[1] and D. molifolium (Ramat.) Tzvel. cv. “Jinju.”[2] However, according to our survey in Jiangling county, Jingzhou city, the stems and leaves of the gold Chrysanthemum have no aroma. Therefore, in this article, morphological characteristics of these species and contents of the volatile oils in their inflorescences were compared, and the compounds of volatile oil in the stems, leaves, and inflorescences of the cultivar, JGXJ, were analyzed to determine its location in Dendranthema genus in plant taxonomy.


   Materials and Methods Top


Materials

Ten samples of inflorescences, come from JGXJ, D. indicum, D. indicum var. aromaticum, and D. molifolium cv. “Jinju,” were separately collected from Jiujiang City, Jiangxi Province, and Xianning City, Tongshan or Badong country, Hubei Province and identified by Professor Keli Chen of the Hubei University of Chinese Medicine [Table 1]. Samples G5 and G6 come from the stems and leaves of JGXJ, collected from as G1 and G3, respectively.
Table 1: Similarity analysis of 10 samples compared with common pattern

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Reagents and instruments

Reagents used were anhydrous ether (analytically pure, Tianjin Deen Chemical Reagent Co., Ltd.) and anhydrous sodium sulfate (analytically pure, Sinopharm Chemical Reagent Co., Ltd.).

Instruments used were pulverizer, high-frequency desktop LCD ultrasonic cleaner; MS204S one ten-thousandth analytical balance (METTLER TOLEDO); electronic thermostat electric heating jacket (Tianjin Taisite Instrument Co., Ltd.); and Thermo Scientific™ TRACE 1310 ISQ LT Single Quadrupole GC-MS (Thermo Fisher, USA), Thermo TG-1701MS capillary column (0.25 μm × 0.25 mm mm30 m).

Preparation of plant extracts

Each sample powder (50 g) was soaked with distilled water (600 mL) for 1.5 h, sonicated for 40 min, and then placed into a 2 L round-bottomed flask and extracted by steam distillation for 8 h, and then, the distillate was collected and extracted with ether. After the ether layer was dried with anhydrous sodium sulfate, the ether was evaporated to obtain pure volatile oil. It was placed in a sealed glass bottle and stored at 4°C until analysis.

Sample solution

Before injection, 100 μl of volatile oil of each sample was dissolved in 1 ml of chromatographically pure ether, filtered through a 0.45 μm microporous membrane, and for use.

Gas chromatography–mass spectrometry analysis conditions

The GC column was Thermo TG-1701MS capillary column (0.25 μm × 0.25 mm 30 m), injection volume was 1.0 μL, split ratio was 50:1, carrier gas was high purity helium, flow rate was 1.0 mL·min−1, and inlet temperature was 250°C. Programmed temperature included initial temperature at 50°C held for 5 min and heated to 250°C at 2°C ·min−1 for 5 min.

Mass spectrometry (MS) transmission line temperature was 250°C, ionization mode was electron ionization source, electron energy: 70 eV, ion source temperature: 200°C, ion scanning range: 50–500 (m/z), and solvent delay time: 3 min.

Qualitative and quantitative analysis

The mass spectral data of the obtained chromatographic peaks were qualitatively searched by component analysis Xcalibur software and Mainlib database, and the relative percentage of each component in the volatile oil was calculated according to the peak area normalization method.


   Results Top


Morphological characteristics

According to our observation, the morphological characteristics of JGXJ are as follows: the plant height is 40–50 cm. Stem is erect, sparsely pilose, and densely in the inflorescence branches. Leaf blade is ovate to broadly ovate, fissure to mid-crack, lobes apex slightly acute, base slightly cuneate (not truncate), and both surfaces dark green and sparsely pubescent. Involucral bracts are two-layered, the outer layer is ovate or ovate-triangular, outside tomentose, and bracts margin brownish broadly membranous. Capitula is 2.5–4 cm in diameter. In the outermost layer, there are ray florets, female, white, and 11–12 mm wide. The others are tubular florets, all female, in the outer florets yellow-white or white, yellow in the middle, all white when fully matured, and apex 5 denticulate. The flowering period is from November to December.

The morphological characteristics of four species, including JGXJ, D. indicum var. aromaticum, D. indicum, and D. molifolium cv. “Jinju” were compared, as shown in [Table 2].
Table 2: The morphological characteristics of four Chrysanthemum species

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Volatile oil yield

The volatile oils of 12 Chrysanthemum samples were extracted. The volatile oil in stem and leaf of JGXJ is yellowish brown with a yield of 0.25%. The yield and color of volatile oil of 10 inflorescences samples are shown in [Table 1]. (The volatile oil yield [%] = constant weight of volatile oil [g]/sample mass [g].)

[Table 1] shows that there were some differences in the volatile oil content and color of different samples. The yield of volatile oil of JGXJ inflorescences was significantly higher than that of the other three Chrysanthemum s species and also higher than that of its stems and leaves.

Comparative analysis of compounds

GC-MS analysis of the volatile oils 10 inflorescences samples showed that the volatile oils contained 17 same compounds in the inflorescences of all the four species of chrysanthemum. However, it is noticed that, among them, the volatile oil of D. molifolium cv. “Jinju” does not contain thujone [Table 3].
Table 3: Chemical constituents and relative contents of volatile oils in 10 samples

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Similarity analysis of gas chromatography spectra

Four batches of GC data from the volatile oil of JGXJ (G1–G4) [Figure 1] were introduced into the software of Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System (2004 A Edition) to generate a common pattern map of JGXJ [Figure 2]. Then, the similarity analysis was carried out on the GC maps of the other six samples of other three Chrysanthemum species and the common pattern map® [Figure 3]; the results are shown in [Table 1].
Figure 1: Gas chromatography–mass spectrometry superposition map of volatile oil of Jiugong Xiangju

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Figure 2: Gas chromatography–mass spectrometry standard map of volatile oil of Jiugong Xiangju

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Figure 3: Gas chromatography–mass spectrometry stack map of volatile oil of four Chrysanthemum species

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Gas chromatography–mass spectrometry analysis of volatile oil from Jiugong Xiangju

The volatile oils of six JGXJ samples (G1–G6) were analyzed by GC-MS to obtain the total ion chromatogram, and then, qualitative and quantitative analysis was carried out according to the above method, as shown in [Table 4].
Table 4: Composition and relative content of volatile oil in inflorescence, stem, and leaf of JGXJ

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As shown in [Table 4], 27 chemical constituents were identified in the volatile oils of stems, leaves, and inflorescences of JGXJ, and terpene compounds were the main components. The oil content and composition of stems and leaves were different from that of the inflorescences, and three compounds are found only in leaves and stems and can be used to differentiate the volatile oil of inflorescences from that of leaves and stems. Moreover, there are 24 same components which are contained in stems, leaves, and inflorescences, and among them, thujone was the main component in the volatile oil of leaves and stems (>30%) and inflorescences (>45%).


   Discussion and Conclusion Top


According to the results of the analysis from [Table 3], the content of thujone in the volatile oil of inflorescences of JGXJ is as high as 45% or more. It is the highest content of thujone found in plants of genus Dendranthema so far, so JGXJ can be used as a new source plant for extracting thujone. Thujone, a kind of monoterpene ketones, has been proved to promote the proliferation of spleen cells, thymocytes, and production of cytokines (such as interleukin-2 and interferon-gamma),[3] and to have anti-tumor ability.[4],[5]

According to the reports, thujone has been found in volatile oils of D. indicum[6],[7],[8],[9] and its variety D. indicum var. aromaticum[10],[11],[12] but not found in that of species series of D. molifolium. Hence, it is supposed that JGXJ has a closer genetic relationship with D. indicum than D. molifolium. However, the content of thujone in volatile oils of JGXJ inflorescences was higher than 45%, much higher than that from D. indicum and D. indicum var. aromaticum. Moreover, there are 21 same components in the volatile oils of inflorescences of JGXJ and two species of D. indicum, but the similarity of the volatile oil components between JGXJ and the two species were only 0.080%–0.110%. Hence, in chemical composition, the breed of JGXJ is obvious different from D. indicum and D. indicum var. aromaticum.

According to Flora of China,[13]D. indicum is a wild species, propagates mainly by seeds, its ray florets with only pistil flowers, but its tubular florets with both pistil and stamen and all yellow, while D. molifolium is pluralistic in source and may come from cross-breeding between species and propagates mainly by stem or rhizome. According to the morphological observation for JGXJ, all flowers in its inflorescences are unisexual no matter the ray or tubular florets, and without seeds its propagation depends on asexual reproduction. The average diameter of its inflorescences is 2.5–4 cm, larger than that of D. indicum. All ray florets are white and the tubular flowers are white when completely mature, while all the tubular florets of D. indicum are yellow. Hence, the characteristics of the reproductive organ flowers of JGXJ are significantly different from those of D. indicum in structure and morphology but significantly compatible with those of D. molifolium. Therefore, even though JGXJ is only 40–50 cm tall and contains thujone, it can still and only be identified as a cultivated breed of D. molifolium, namely here as Dendranthema morifolium (Ramat.) Tzvel. “Jiugong Xiangju” cv. nov. Of course, JGXJ maybe is a hybrid of D. indicum (or its variants) and a breed of D. molifolium for containing thujone and with strong aromas in stems and leaves. To have an insight into the breed of D. molifolium and JGXJ, further work is needed in genetic structure and chemical components including flavonoids and organic acids.

Acknowledgements

Thanks to the platform of the Chinese Medicine Resource Center for providing the experimental condition platform, and Dr. Ling Xiao for guiding the experiment.

Financial support and sponsorship

The project was supported by the National Key Research and Development Plan (Grant No. 2017YFC1700704).

Conflicts of interest

There are no conflicts of interest.



 
   References Top

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Li QH, Jia WJ, Yang DH. The investigation on geographical distribution, ecological, habit and storage quantity on a new resource plant of Hubei, Dendranthema indicum(L.) Des Monl. var. Aromaticum. J Wuhan Bot Res 1983;2:239-45, 338.  Back to cited text no. 1
    
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Siveen KS, Kuttan G. Augmentation of humoral and cell mediated immune responses by Thujone. Int Immunopharmacol 2011;11:1967-75.  Back to cited text no. 3
    
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Siveen KS, Kuttan G. Thujone inhibits lung metastasis induced by B16F-10 melanoma cells in C57BL/6 mice. Can J Physiol Pharmacol 2011;89:691-703.  Back to cited text no. 4
    
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Biswas R, Mandal SK, Dutta S, Bhattacharyya SS, Boujedaini N, Khuda-Bukhsh AR. Thujone-rich fraction of Thuja occidentalis demonstrates major anti-cancer potentials: Evidences from In vitro studies on A375 cells. Evid Based Complement Alternat Med 2011;2011:568148.  Back to cited text no. 5
    
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Zhu SY, Yang Y, Yu HD, Ying Y, Zou GL. Chemical composition and antimicrobial activity of the essential oils of Chrysanthemum indicum. J Ethnopharmacol 2005;96:158.  Back to cited text no. 6
    
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He XZ, Jiang JH, Zhao L. Chemical components analysis of volatile oil in Flos Chrysanthemi Indici from Anhui. Applied Chem Ind 2014;43:1536-9, 46.  Back to cited text no. 7
    
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Zhang YM, Huang YF, Tao L, Huang JW. Chemical components of essential oils from Flos Chrysanthemi Indici in different areas. Zhongguo Zhong Yao Za Zhi 2002;27:265-7.  Back to cited text no. 8
    
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Zhou X, Mo BB. Study on chemical components of volatile oil from Chrysanthemum indicum L. growing in Guizhou and Guangdong provinces. West China J Pharm Sci 2001;16:330-3.  Back to cited text no. 9
    
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Fan S, Chang J, Zong Y, Hu G, Jia J. GC-MS analysis of the composition of the essential oil from Dendranthema indicum var. Aromaticumusing three extraction methods and two columns. Molecules 2018;23. pii: E576.  Back to cited text no. 10
    
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Wang CZ, Su Y, Li D, Cai B, Guo YL. Analysis of volatile organic compounds from Dendranthema indicum var. Aromaticum by headspace gas chromatography-mass spectrometry and accurate mass measurement. Analyt Lett 2010;43:2297-310.  Back to cited text no. 11
    
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Jian L, Sun M, Zhang QX. Analysis on aroma compositions in flowers, stems and leaves of Chrysanthemum indicum var. Aromaticum. J Northwest A F Univ 2014;42:87-92.  Back to cited text no. 12
    
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Editorial Committee of Chinese Botany, Chinese Academy of Sciences. Flora of China. Beijing: science Press. 1983. p. 32.  Back to cited text no. 13
    


    Figures

  [Figure 1], [Figure 2], [Figure 3]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]



 

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