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Helichrysum gymnocephalum Essential Oil: Chemical Composition and Cytotoxic, Antimalarial and Antioxidant Activities, Attribution of the Activity Origin by Correlations


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Helichrysum gymnocephalum essential oil (EO) was prepared by hydrodistillation of its leaves and characterized by GC-MS and quantified by GC-FID. Twenty three compounds were identified. 1,8-Cineole (47.4%), bicyclosesquiphellandrene (5.6%), γ-curcumene (5.6%), α-amorphene (5.1%) and bicyclogermacrene (5%) were the main components. Our results confirmed the important chemical variability of H. gymnocephalum. The essential oil was tested in vitro for cytotoxic (on human breast cancer cells MCF-7), antimalarial (Plasmodium falciparum: FcB1-Columbia strain, chloroquine-resistant) and antioxidant (ABTS and DPPH assays) activities. H. gymnocephalum EO was found to be active against MCF-7 cells, with an IC(50) of 16 ± 2 mg/L. The essential oil was active against P. falciparum (IC(50) = 25 ± 1 mg/L). However, the essential oil exhibited a poor antioxidant activity in the DPPH (IC(50) value > 1,000 mg/L) and ABTS (IC(50) value = 1,487.67 ± 47.70 mg/L) assays. We have reviewed the existing results on the anticancer activity of essential oils on MCF-7 cell line and on their antiplasmodial activity against the P. falciparum. The aim was to establish correlations between the identified compounds and their biological activities (antiplasmodial and anticancer). β-Selinene (R² = 0.76), α-terpinolene (R² = 0.88) and aromadendrene (R² = 0.90) presented a higher relationship with the anti-cancer activity. However, only calamenene (R² = 0.70) showed a significant correlation for the antiplasmodial activity.
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Molecules 2011, 16, 8273-8291; doi:10.3390/molecules16108273
ISSN 1420-3049
Helichrysum gymnocephalum Essential Oil: Chemical
Composition and Cytotoxic, Antimalarial and Antioxidant
Activities, Attribution of the Activity Origin by Correlations
Samia Afoulous 1, Hicham Ferhout 2, Emmanuel Guy Raoelison 3, Alexis Valentin 4,
Béatrice Moukarzel 4, François Couderc 1 and Jalloul Bouajila 1,*
1 Laboratoire des Interactions Moléculaires et Réactivité Chimique et Photochimique UMR CNRS 5623,
University of Toulouse, University of Paul-Sabatier, 118 route de Narbonne, F-31062 Toulouse,
2 Nat’Ex Biotech. Bat 7, 55 avenue Louis Breguet, 31400 Toulouse, France
3 Laboratoire de Phytochimie et Standardisation, IMRA, BP: 3833 Antananarivo 101, Madagascar
4 Faculté of Pharmacy, University of Toulouse, UMR 152 IRD-UPS Pharma-DEV, University of
Paul Sabatier Toulouse 3, 35 chemin des maraîchers, 31062 Toulouse Cedex 9, France
* Author to whom correspondence should be addressed; E-Mail:;
Tel.: +33-561558333; Fax: +33-561558155.
Received: 19 September 2011; in revised form: 23 September 2011 / Accepted: 23 September 2011 /
Published: 29 September 2011
Abstract: Helichrysum gymnocephalum essential oil (EO) was prepared by hydrodistillation
of its leaves and characterized by GC-MS and quantified by GC-FID. Twenty three
compounds were identified. 1,8-Cineole (47.4%), bicyclosesquiphellandrene (5.6%),
γ-curcumene (5.6%), α-amorphene (5.1%) and bicyclogermacrene (5%) were the main
components. Our results confirmed the important chemical variability of H. gymnocephalum.
The essential oil was tested in vitro for cytotoxic (on human breast cancer cells MCF-7),
antimalarial (Plasmodium falciparum: FcB1-Columbia strain, chloroquine-resistant) and
antioxidant (ABTS and DPPH assays) activities. H. gymnocephalum EO was found to be
active against MCF-7 cells, with an IC50 of 16 ± 2 mg/L. The essential oil was active
against P. falciparum (IC50 = 25 ± 1 mg/L). However, the essential oil exhibited a poor
antioxidant activity in the DPPH (IC50 value > 1,000 mg/L) and ABTS (IC50 value =
1,487.67 ± 47.70 mg/L) assays. We have reviewed the existing results on the anticancer
activity of essential oils on MCF-7 cell line and on their antiplasmodial activity against the
P. falciparum. The aim was to establish correlations between the identified compounds and
Molecules 2011, 16 8274
their biological activities (antiplasmodial and anticancer). β-Selinene (R² = 0.76),
α-terpinolene (R² = 0.88) and aromadendrene (R² = 0.90) presented a higher relationship
with the anti-cancer activity. However, only calamenene (R² = 0.70) showed a significant
correlation for the antiplasmodial activity.
Keywords: Helichrysum gymnocephalum; GC-MS; antimalarial activity; cytotoxic activity;
antioxidant activity
1. Introduction
The flora of Madagascar is particularly rich, with a specific diversity and more importantly, it
contains endemic species. In Madagascar, more than 400 species described in other parts of the World,
especially in Southern Africa, Western Europe and Australia, and 115 species, all endemic of the large
Island, are currently known [1]. The gender Helichrysum (from the Greek "helios" sun and "chrysos"
gold) which belongs to Asteraceae family is one of the endemic species.
Helichrysum gymnocephalum (Immortal with naked head) is a shrub from 1 to 4 meters in height
whose ultimate branches are covered by a white and very fine tomentum. The leaves are lengthily
attenuated towards the base in short petioles, covered on the two faces by a tomentum and having
three quite visible principal veins in lower part. Flowering lasts from February till May, seeds are also
papillate [2].
Some medicinal properties of Helichrysum species have been reported, and various studies
have demonstrated the antimicrobial and antiseptic properties of many species from the genus
Helichrysum [3-5]. H. gymnocephalum has been traditionally used therapeutically as a tea or syrup
prepared from the leaves to treat gingivitis or buccal ulcers [6]. It is also used as ointment for
rheumatism or gout by mixing crushed leaves with bacon. The leaves and flowers have properties as
diuretics, stimulants and would relieve neuralgia [7]. Analgesic, aphrodisiac, antiseptic, antiscorbutic,
deodorant, tonic and anthelmintic applications have been reported for H. gymnocephalum [8,9].
H. gymnocephalum is also used in the manufacture of toothpaste for the local market. The leaves are
also used to embalm dead bodies.
The essential oil (EO), a complex mixture of compounds, is considered among the most important
antimicrobial agents [3] present in these plants, and may also have antioxidant and anti-inflammatory
activities. These last years, an overflow of information has appeared on the role of the oxidative stress
in the emergence of a certain number of diseases, such as cancers, cardiovascular diseases and
degenerative diseases related to ageing, in parallel the possible therapeutic role of antioxidants in these
diseases was emphasized [10].
More than a third of the World’s population (about two billion people) lives in malaria-endemic
areas. The majority of deaths caused by falciparum malaria occur in sub-Saharan Africa, primarily
among children younger than 5 years and pregnant women living in remote rural areas with limited
access to health services [11]. Nearly all malaria deaths and a large proportion of morbidity are caused
by Plasmodium falciparum. In the last decades, resistance to several antimalarial drugs became widely
disseminated, while the cost of effective treatments was prohibitive for the majority of the populations
Molecules 2011, 16 8275
in the endemic regions. Thus, rapid development of resistance by P. falciparum to the conventional
drugs necessitates search for new antimalarial drugs [11].
Breast cancer is second only to lung cancer as the most common cancer in women. Roughly
180,000 women are diagnosed with this disease each year, of which 44,000 or almost 20% will die [12].
With increased awareness and increased use of routine mammograms, more women are diagnosed in
the earlier stages of this disease, at which time a cure may be possible. The disease is more common in
women after the age of 40. It is also more frequent in women of a higher social-economic class [12].
Cancer diseases are characterized by an uncontrolled proliferation of cells. They constitute the second
cause of mortality behind cardiovascular diseases in developed countries and the third after infectious
and cardiovascular diseases in developing countries [13]. The use of plant extracts and derived
products in the treatment of cancer is of exceptional value in the control of malignancies, due to the
fact that most of the anticancer drugs severely affect the normal cells [13,14].
The aim of the present study was the evaluation of H. gymnocephalum leaves essential oil for its
possible antioxidant and biological activities. In this study, we: (i) examined the chemical components
of the essential oil by GC-MS and GC-FID; (ii) evaluated their cytotoxic, antimalarial and antioxidant
activities and (iii) reviewed researches for essential oils having an activity against P. falciparum and/or
on MCF-7 cell line in order to identify, by correlation, the main active compounds.
2. Results and discussion
2.1. Chemical Composition
The essential oil yield of H. Gymnocephalum obtained from hydrodistillation of leaves was 0.40%.
Two previous works have quantified the yield of this essential oil. They studied the shoot part of the
plant mainly bark and leaves; extraction yields for essential oil were 0.5% and 0.41% according to
Mollenbeck et al. [26] and Cavalli et al. [27], respectively. In the present work, only the leaves were
used and a similar essential oil yield was obtained.
Twenty three components have been identified in H. gymnocephalum essential oil by GC-MS
(Figure 1 and Table 1). Sesquiterpene hydrocarbons and oxygenated monoterpenes were the major
groups of compounds (32.4% and 54.0%, respectively), while oxygenated sesquiterpenes were not
present. The major monoterpenes found were 1,8-cineole (47.4%), p-cymene (4.3%), (E)-β-ocimene
(2.4%), 2,3-dihydro-1,8-cineole (2.1%) and α-terpinolene (1,3%). Among the sesquiterpene hydrocarbons
bicyclosesquiphellandrene (5.6%), γ-curcumene (5.6%), α-amorphene (5.1%) and bicyclogermacrene
(5%) were present in appreciable quantities. On the other hand, this essential oil contained a phenolic
compound (2,3-di-tert-butylphenol) at 0.5%.
The chemical composition of H. gymnocephalum essential oil has been reported elsewhere.
Mollenbeck et al. [26] have shown that 1,8-cineole (66.7%) was the major compound of the essential
oil, a level which was higher than in our work (47.7%). Also, in our work, the major compounds were
β-caryophyllene (3.3%), γ-terpinene (3.1%), β-pinene (2.7%), α-pinene (2.5%) and ρ-cymene (2.3%).
Molecules 2011, 16 8276
Figure 1. Chromatograms GC-MS of H. gymnocephalum essential oil (1: α-thujene;
2: sabinene; 3: β-pinene; 4: 2,3-dihydro-1,8-cineole; 5: α-terpinene; 6: p-cymene;
7: limonene; 8: 1,8-cineole; 9: (E)-β-ocimene; 10: α-terpinolene; 11: α-phellandrene;
12: terpinen-4-ol; 13: α-terpineol; 14: α-copaene; 15: aromadendrene; 16: bicyclo-
sesquiphellandrene; 17: γ-curcumene; 18: β-selinene; 19: bicyclogermacrene;
20: α-amorphene; 21: 2,3-di-tert-butylphenol; 22: calamenene; 23: δ-cadinene).
Table 1. Chemical composition of H. gymnocephalum essential oil.
N RI Compounds (%)
1 928 α-Thujene 1.0
2 967 Sabinene 0.3
3 971 β-Pinene 1.1
4 984 2,3-Dihydro-1,8-cineole 2.1
5 1010 α-Terpinene 1.3
6 1018 p-Cymene 4.3
7 1022 Limonene 0.5
8 1026 1,8-Cineole 47.4
9 1052 (E)-β-Ocimene 2.4
10 1084 α-Terpinolene 1.3
11 1164 α-Phellandrene 0.2
12 1175 Terpinen-4-ol 2.7
13 1187 α-Terpineol 1.8
14 1373 α-Copaene 0.4
15 1438 Aromadendrene 2.0
1 2
4 5
9 10 11
1213 14
Molecules 2011, 16 8277
Table 1. Cont.
N RI Compounds (%)
16 1470 Bicyclosesquiphellandrene 5.6
17 1473 γ-Curcumene 5.6
18 1485 β-Selinene 3.3
19 1494 Bicyclogermacrene 5.0
20 1497 α-Amorphene 5.1
21 1502 2,3-di-tert-butylphenol 0.5
22 1512 Calamenene 1.8
23 1521 δ-Cadinene 3.6
Identified components 99.3
Monoterpene hydrocarbons 8.1
Monoterpenes oxygenated 54.0
Sesquiterpenes hydrocarbons 32.4
Others 4.8
Cavalli et al. [27] have shown that the chemical composition of the essential oil of
H. gymnocephalum was dominated by 1,8-cineole (59.7%), p-cymene (6.3%), α-pinene (4.9%),
β-pinene (3.3%) and terpinen-4-ol (2.6%). These published compositions of H. gymnocephalum
essential oil indicated the presence of oxygenated sesquiterpenes, which were not present in our work.
On the other hand, we found more α-thujene (1%) and δ-cadinene (3.6%) compared to Cavalli et al. [27]
0.5% and 0.5%, respectively. For terpinen-4-ol, we obtained 2.6% which was similar to the result of
Cavalli et al. [27] (2.7%) but higher than the result of Mollenbeck et al. [26] (1.3%). An intermediate
concentration (4.3%) of p-cymene was obtained compared to Cavalli et al. [27] and Mollenbeck et al. [26],
with 6.3% and 2.3%, respectively. On the other hand, β-pinene (1.1%) was obtained in the lowest
concentration compared to these two studies of Cavalli et al. [27] and Mollenbeck et al. [26] (3% and
2.7%, respectively).
We identified some new compounds which were not reported in these two previous studies. The
total yield of these new compounds varied between 2.1 and 5.6% (Figure 2). The presence of
2,3-dihydro-1,8-cineole (2.1%), (E)-β-ocimene (2.4%), β-selinene (3.3%), bicyclogermacrene (5.0%),
α-amorphene (5.1%), bicyclosesquiphellandrene (5.6%) and γ-curcumene (5.6%) was evidenced.
Figure 2. Structures of new abundant compounds identified in H. gymnocephalum
essential oil compared to other studies of same essential oil.
Molecules 2011, 16 8278
Figure 2. Cont.
As shown in Table 1, the differences between our work and previously published results concerned
extraction yield, the number of identified compounds and their respective contents. In our
investigation, H. gymnocephalum plants were harvested in July 2008, and we treated only leaves for
the essential oil extraction. Indeed, H. gymnocephalum was harvested in March 1997 by the team of
Mollenbeck et al. [26] and between November–December 1994 by Cavalli et al. [27].
2.2. Antioxidant Activity
Data presented here is the first bibliographical report on the antioxidant activity of
H. gymnocephalum. Furthermore, our results (Table 2) demonstrated that the essential oil of
H. gymnocephalum has poor antioxidant activity against DPPH (IC50 value > 1,000 mg/L) and ABTS+
(IC50 = 1,487.67 ± 47.70 mg/L). These results may be attributed to a low antioxidant activity (in these
two tests) of the compounds identified in the essential oil of H. gymnocephalum. Four other
Helichrysum species (H. dasyanthum, H. excisum, H. felinum and H. petiolare) were also reported to
exhibit also a low antioxidant activity against test DPPH (IC50 > 100 mg/L) [28].
Table 2. Anticancer, antimalarial and antioxidant activities (IC50 (mg/L)) of
H. gymnocephalum essential oil.
Sample Anticancer
Antioxidant activity
(DPPH assay)
Antioxidant activity
(ABTS assay)
Essential oil 16 ± 2 25 ± 1 >10000 1487.67±47.70
Control 0.218
a ± 0.04 0.10
± 0.09 3.75 c ± 0.01 1.84 c ± 0.03
a Doxorubicin; b Chloroquine; c Ascorbic acid.
Molecules 2011, 16 8279
2.3. Cytotoxic Activity
In recent years, considerable attention has been focused to identifying naturally occurring
substances able to inhibit, delay or reverse the process of multistage carcinogenesis. Plant essential oils
are believed to reduce the risk of cancer when used in prevention [29].
In this work, the cytotoxic activity of H. gymnocephalum leaf essential oil against human breast
cancer cells MCF-7 (IC50 = 16 ± 2 mg/L) was reported for the first time.
Data in Table 3 shows the anti-cancer activity against MCF-7 cells (IC50 mg/L) of several essential
oils reported in the literature and their main components. Essential oils were extracted from
H. gymnocephalum, Schefflera heptaphylla [30], Laurus nobilis, Origanum syriacum, Origanum vulgare,
Salvia triloba [31], Heteropyxis dehniae [32], Schinus molle, Schinus terebenthifolius [33], Salvia
officinalis [34], Melaleuca alternifolia [35], Citrus limon, Citrus medica, Citrus sinensis [36] and
Talauma gloriensis [37].
Based on the bibliographical review, we can consider that the IC50 of our essential oil (16 mg/L ± 2)
is very well positioned among these studied oils in the Table 3. Among the studied essential oils only
five showed higher anti-cancer activities than H. gymnocephalum’s essential oil. Indeed, nine essential
oils had lower activities for the same test (Table 3).
2.4. Antimalarial Activity
Many studies on the antiplasmodial activity of crude essential oils have been reported [38,39]. The
in vitro antiplasmodial activity of H. gymnocephalum essential oil was determined against the FcB1
chloroquine-resistant strain of P. falciparum (Table 2). The antimalarial activity of the essential oil
(IC50 values) was 25 ± 1 mg/L. Since the value of IC50 was found between 5 and 50 mg/L, it can be
considered that H. gymnocephalum essential oil has a good activity against P. falciparum [24]. This
rather high value of IC50 compared to that of chloroquine (IC50 = 0.1 ± 0.09 mg/L) can be explained by
the low concentration of the active compound(s) since the essential oil is a multi-components mixture.
Data in Table 4 summarize the antimalarial activity [IC50 (mg/L)] of all essential oils cited in the
literature and their components. These EO have been obtained from H. gymnocephalum, Xylopia
phloiodora, Pachypodanthium conne, Antidesma laciniatum, Xylopia aethiopica, Hexalobus
crispiorus[40], Salvia stenophylla, Salvia runcinata, Salvia repens [41], Salvia albicaulis, Salvia
dolomitica [42], Lippia multiflora [43], Helichrysum cymosum [44], Artemisia gorgonum Webb [45],
Arnica longifolia, Aster hesperius and Chrysothamnus nauseosus [46]. The results based on this
bibliographical review (Table 4) showed that the IC50 of our essential oil (25 mg/L ± 1) was at an
interesting level, quite well positioned among these studied oils. We found seven essential oils which
have higher antimalarial activity than that of H. gymnocephalum essential oil. In addition, 11 essential
oils had lower activities in similar tests (Table 4).
Molecules 2011, 16 8280
Table 3. Anticancer activity and chemical composition of essential oils (I: Helichrysum gymnocephalum; II: Schefflera heptaphylla [30];
III: Laurus nobilis; IV: Origanum syriacum; V: Origanum vulgare; VI: Salvia triloba [31]; VII: Heteropyxis dehniae [32]; VIII: Schinus
molle; IX: Schinus terebenthifolius [33]; X: Salvia officinalis [34]; XI: Melaleuca alternifolia [35]; XII: Citrus limon; XIII: Citrus medica;
XIV: Citrus sinensis [36]; XV: Talauma gloriensis [37]).
Essential oil I II III IV V VI VII VIII IX
Anticancer activity (IC50 mg/L) 16 ± 2 7.3 101.7 ± 7.9 130 ± 52.2 30.1 ± 1.14 174.3 ± 73.04 150 * 54 ± 10 47 ± 9
α-Thujene 1.0 0.62 0.47 0.54 0.38
Sabinene 0.3 6.92 0.35 5.56 0.24 0.02 0.02
β-Pinene 1.0 22.24 4.55 0.6 1.3 8.89 4.96 3.09
2,3-Dihydro-1,8-cineole 2.0
α-Terpinene 1.2 0.7 0.37 4.46 0.44
p-Cymene 4.2 0.74 30.22 3.83 0.34 1.5 2.49 7.34
Limonene 0.5 3.61 2.1 1.68 0.9
1,8-Cineole 47.4 40.91 0.27 45.16 0.4
(E)-β-Ocimene 2.4
α-Terpinolene 1.3 0.35 0.43
α-Phellandrene 0.2 46.52 34.38
Terpinen-4-ol 2.7 1.55 1.09 0.07 0.03
α-Terpineol 1.8 0.62 3.6 8.38 5.6
α-Copaene 0.4 0.6 0.11 0.19
Aromadendrene 2.0 0.49 0.19
Bicyclosesquiphellandrene 5.6
γ-Curcumene 5.6
β-Selinene 3.3 0.3 1.1
Bicyclogermacrene 5.0 1.05
α-Amorphene 5.1
2,3-Di-tert-butylphenol 0.5
Calamenene 1.8
δ-Cadinene 3.6 2.8 0.27 0.69
Molecules 2011, 16
Table 3. Cont.
Essential oil X XI XII XIII XIV XV
Anticancer activity (IC50 mg/L) 554.4 ± 1.5 310 * 10 1 0.5 14.1
α-Thujene 0.31
Sabinene 0.41 0.1
β-Pinene 2.57 0.91 16.3 3.7
α-Terpinene 0.2 5.76 0.1
Limonene 1.7 98.4 56.6 98.4 0.8
1,8-Cineole 17.52 19.29
(E)-β-Ocimene 0.1
Terpinen-4-ol 1.01 42.62
α-Terpineol 0.27 11.3
α-Copaene 0.1
Bicyclogermacrene 2.1
δ-Cadinene 3.3
Molecules 2011, 16 8282
Table 4. Antipaludic activity and chemical composition of essential oils (I: Helichrysum gymnocephalum; II: Xylopia phloiodora;
III: Pachypodanthium conne; IV: Antidesma laciniatum; V: Xylopia aethiopica; VI: Hexalobus crispiorus [40]; VII: Salvia stenophylla;
VIII: Salvia runcinata; IX: Salvia repens [41]; X: Salvia albicaulis; XI: Salvia dolomitica[42]; XII: Lippia multiflora [43]; XIII: Helichrysum
cymosum [44]; XIV: Artemisia gorgonum Webb [45]; XV: Arnica longifolia; XVI: Aster hesperius; XVII: Chrysothamnus nauseosus [46];
XVIII: Essential oil [39]). * IC50 was calculated from percentage; na: no activity.
Antipalaudic activity (IC50 mg/L) 25 ± 1 17.9 16.6 29.4 17.8 2.0 4.38 ± 1.07 1.23 ± 0.31 1.68 ± 0.26 6.4 ± 2.0 4.8 ± 0.7
α-Thujene 1.0 0.59 0.61
Sabinene 0.3 0.59 0.46 0.1 0.2 0.1
β-Pinene 1.0 0.68 10.07 0.12 0.7 0.8 3.0
2,3-Dihydro-1,8-cineole 2.0
α-Terpinene 1.2 0.48 0.43 0.3 0.2
p-Cymene 4.2 0.35 0.32 1.72 0.2 0.7 2.5 0.2
Limonene 0.5 5.3 0.6 9.8 9.4 0.6
1,8-Cineole 47.4 2.0 9.4
(E)-β-Ocimene 2.4 1.93 1.13 0.8 1.5
α-Terpinolene 1.3
α-Phellandrene 0.2
Terpinen-4-ol 2.7 0.16 0.49 0.16 0.8
α-Terpineol 1.8 4.99 2.7 6.2
α-Copaene 0.4 0.53 7.06 2.2 4.07 13.27 0.1
Aromadendrene 2.0 1.08 2
Bicyclosesquiphellandrene 5.6
γ-Curcumene 5.6
β-Selinene 3.3 0.28 2.2
Bicyclogermacrene 5.0
α-Amorphene 5.1
2,3-Di-tert-butylphenol 0.5
Calamenene 1.8 2.32 0.93 1.09
δ-Cadinene 3.6 15.11 8.06 1.3 4.3 10.07 0.5 0.3
Molecules 2011, 16
Table 4. Cont.
Antipalaudic activity (IC50 mg/L) 2–4 1.25 ± 0.77 5.2 ± 0.77 na na na 40.15 * 205.19 * 10.82 * 72.68 *
α-Thujene 0.8 0.8
Sabinene 0.3 0.2
β-Pinene 3.7 96.32
α-Terpinene 1.2 1.0
p-Cymene 0.2 2.4 0.1 0.1 0.6 93.19
Limonene 0.2 7.2 4.7 92.98
1,8-Cineole 3.1 20.4 0.1 0.1 93.13
(E)-β-Ocimene 3.6 0.3
α-Phellandrene 1.8
Terpinen-4-ol 0.2
α-Terpineol 0.3 2.6 0.2 0.2 0.1 0.3
α-Copaene 0.3 1.2 0.2
Aromadendrene 1.5
γ-Curcumene 0.5
β-Selinene 0.3
Calamenene 0.1 0.2
δ-Cadinene 0.2 0.4 0.8
Molecules 2011, 16 8284
To study the role of the various components of an essential oil in the biological activities obtained,
we performed a complete survey of the activities and the percentage of compounds present in order to
assign the origin of the activity. The aim of the study was to find correlations between each component
present in our literature search of all essential oils of various plants tested for biological activities
(anticancer and antimalarial). Correlations between our essential oil, other oils and their components
are listed in Tables 3 and 4, with the IC50 on the tested biological target (MCF-7 cell line or
P. falciparum).
2.5. Cytotoxic Activity Correlations
We established correlations between compound contents and anti-cancer activity against the MCF-7
cells. Aromadendrene, α-terpinolene and β-selinene showed good correlations, respectively R² = 0.90,
0.88 and 0.76 (chemical structures are presented in Figure 3).
Figure 3. Structures of compounds which have good correlation for anticancer (a–c) and
antimalarial (d) activities.
α-terpinolene (a) aromadendrene (b)
β-selinene (c) calamenene (d)
To our knowledge, no literature has cited any anticancer activity of these three compounds
(β-selinene, aromadendrene and α-terpinolene). We will carry out a deeper study to focus on their
specific biological activity against MCF-7 cell line.
Based on this correlation study, we noticed that six components were present only in our essential
oil, namely bicyclosesquiphellandrene (5.6%), α-amorphene (5.1%), 2,3-dihydro-1,8-cineole (2%),
limonene (0.5%), and 2,3-di-tert-butylphenol (0.5%). In addition, for bicyclogermacrene (5.0%) and
(E)-β-ocimene (2.4%), the correlation has not been established because we did not have enough data
(only two essential oils).
A deep study of the anticancer activity of aromadendrene, α-terpinolene, β-selinene, bicyclo-
sesquiphellandrene, α-amorphene, 2,3-dihydro-1,8-cineole, calamenene, 2,3-di-tert-butylphenol,
bicyclogermacrene and (E)-β-ocimene will have to be carried out to determine the origin of the
anticancer activity of H. gymnocephalum. On the same target, we should evaluate the synergy between
these compounds and their own contribution to the final anticancer activity.
Molecules 2011, 16
2.6. Correlations for Antimalarial Activity
We established correlations between both major and minor components and IC50 (antimalarial
activity). Calamenene showed the highest correlation R2 = 0.7 (Figure 3). Our results are in accordance
with those of Van Zyl et al. [39]; the former showed that p-cymene, limonene and β-pinene had low
antiplasmodial activity (IC50 = 205.20, 72 and 40.16 mg/L, respectively). Indeed, they also showed that
there are significant synergies (the sum of the fractional inhibitory concentrations (FIC) < 0.5)
between p-cymene and other compounds such as E- and Z-(±)-nerolidol, carvacrol and γ-terpinene
(0.09, 0.02 and 0.37, respectively). We noticed that bicyclosesquiphellandrene (5.6%), α-amorphene
(5.1%), bicyclogermacrene (5.0%), 2,3-dihydro-1,8-cineole (2.0%), and 2,3-di-tert-butyl-phenol
(0.5%) were present only in our essential oil. It is possible that these compounds may be involved in
the activity of our essential oil.
3. Experimental
3.1. Extraction of the Essential Oil
The leaves of H. gymnocephalum were collected in Antananarivo, Madagascar (July 2008). Steam-
distillation was used to extract the essential oil according to the European Pharmacopoeia protocol [15].
The essential oil was dried with anhydrous sodium sulphate, filtered and stored in sealed vials at 4 °C
prior to analyses.
3.2. Chemicals
All chemicals used were of analytical reagent grade. All reagents were purchased from Sigma-
Aldrich-Fluka (Saint-Quentin, France).
3.3. Gas Chromatography and Gas Chromatography-Mass Spectrometry
Quantitative and qualitative analysis of the essential oil was carried out by gas chromatography-
flame ionization detection (GC-FID) and gas chromatography-mass spectrometry (GC-MS). Gas
chromatography analyses were carried out on a Varian Star 3400 Cx chromatograph (Les Ulis, France)
fitted with a fused silica capillary DB-5MS column (5% phenylmethylpolysyloxane, 30 m × 0.25 mm,
film thickness 0.25 µm). Chromatographic conditions were 60 °C to 260 °C temperature rise with a
gradient of 5 °C/min and 15 min isotherm at 260 °C. A second gradient was applied to 340 °C at
40 °C/min. Total analysis time was 57 min. For analysis purposes, the essential oil was dissolved in
petroleum ether. One microliter of sample was injected in the split mode ratio of 1:10. Helium (purity
99.999%) was used as carrier gas at 1 mL/min. The injector was operated at 200 °C. The mass
spectrometer (Varian Saturn GC/MS/MS 4D) was adjusted for an emission current of 10 µA and
electron multiplier voltage between 1,400 and 1,500 V. Trap temperature was 150 °C and that of the
transfer line was 170 °C. Mass scanning was from 40 to 650 amu.
Compounds were identified by comparison of their Kovats indices (KI) obtained on a nonpolar
DB-5MS column relative to C5-C24 n-alkanes, with those provided in the literature, by comparison of
their mass spectra with those recorded in NIST 08 (National Institute of Standards and Technology)
Molecules 2011, 16
and reported in published articles and by co-injection of available reference compounds. The samples
were analyzed in duplicate. The percentage composition of the essential oil was computed by the
normalization method from the GC peak areas, assuming identical mass response factors for all
compounds. Results were calculated as mean values of two injections from essential oil, without using
correction factors. All determinations were performed in triplicate and averaged.
3.4. Antioxidant Activity
Free Radical Scavenging Activity: DPPH Test
Antioxidant scavenging activity was studied using the 1,1-diphenyl-2-picrylhydrazyl free radical
(DPPH) assay as described by Blois [16] with some modifications. Various dilutions of EO (1.5 mL)
were mixed with a 0.2 mmol/L methanolic DPPH solution (1.5 mL). After an incubation period of
30 min at 25 °C, the absorbance at 520 nm (the wavelength of maximum absorbance of DPPH) were
recorded as A(sample), using a Helios spectrophotometer (Unicam, Cambridge, UK). A blank experiment
was also carried out applying the same procedure to a solution without the test material and the
absorbance was recorded as A(blank). The free radical-scavenging activity of each solution was then
calculated as percent inhibition according to the following equation:
% inhibition = 100 (A(blank) A(sample)) / A(blank)
Antioxidant activities of test compounds or the essential oil were expressed as IC50 values, defined
as the concentration of the test material required to cause a 50% decrease in initial DPPH
concentration. Ascorbic acid was used as a standard. All measurements were performed in triplicate.
3.5. ABTS Radical-Scavenging Assay
The radical scavenging capacity of the samples for the ABTS (2,2-azinobis-3-ethylbenzothiazoline-
6-sulphonate) radical cation was determined as described by Re et al. [17] with some modifications.
ABTS was generated by mixing a 7 mmol/L solution of ABTS at pH 7.4 (5 mmol/L NaH2PO4,
5 mmol/L Na2HPO4 and 154 mmol/L NaCl) with 2.5 mmol/L potassium persulfate (final
concentration) followed by storage in the dark at room temperature for 16 h before use. The
mixture was diluted with ethanol to give an absorbance of 0.70 ± 0.02 units at 734 nm using a
spectrophotometer (Helios). For samples, solutions of the essential oil in methanol (100 μL) were
allowed to react with fresh ABTS solution (900 μL), and then the absorbance was measured 6 min
after initial mixing. Ascorbic acid was used as a standard and the capacity of free radical scavenging
was expressed by IC50 (mg/L). IC50 values were calculated as the concentration required for
scavenging 50% of ABTS radicals. The capacity of free radical scavenging (IC50) was determined
using the same previously used equation for the DPPH method. All measurements were performed in
triplicate. All data of antioxidant activity were expressed as means ± standard deviations (SD) of the
triplicate measurements. The confidence limits were set at P < 0.05. SD did not exceed 5% for the
majority of the values obtained.
Molecules 2011, 16
3.6. Antiplasmodial Activity
The chloroquine-resistant FcB1-Columbia strain of Plasmodium falciparum (IC50 for chloroquine:
186 nM) was cultured continuously according to Trager and Jensen [18], with modifications [19]. The
IC50 values for chloroquine were checked every 2 months, and we observed no significant variations.
The parasites were maintained in vitro in human red blood cells (O±; EFS; Toulouse, France), diluted
to 4% hematocrit in RPMI 1640 medium (Lonza, Emerainville, France) supplemented with 25 mM
Hepes and 30 M NaHCO3 and complemented with 7% human AB+ serum (EFS).
Parasite cultures were synchronized by combination of magnetic enrichment [20] followed by
D-sorbitol lysis (5% of D-sorbitol in sterile water) as described by Lambros and Vanderberg [21]. The
antimalarial activity of essential oil was evaluated by a radioactive micromethod described elsewhere [22].
Tests were performed in triplicate in 96-well culture plates (TPP) with cultures mostly at ring stages
(synchronisation interval, 16 h) at 0.5–1% parasitemia (hematocrit, 1.5%). Parasite culture was
incubated with each sample for 48 h. Parasite growth was estimated by [3H]-hypoxanthine (Perkin-
Elmer, Courtaboeuf, France) incorporation, which was added to the plates 24 h before freezing. After
48 h incubation, plates were frozen-defrosted and each well was harvested on a glass fiber filter.
Incorporated (3H)-hypoxanthine was then determined with a β-counter (1450-Microbeta Trilux,
Wallac-Perkin Elmer). The control parasite cultures, free from any sample, was referred to 100% growth.
IC50 were determined graphically in concentration versus percent inhibition curves. Chloroquine
diphosphate was used as positive control. The antimalarial activity of sample was expressed by IC50,
representing the concentration of drug that induced a 50% parasitaemia decrease compared to the
positive control culture referred to as 100% parasitaemia [23]. According to the literature concerning
plant antiplasmodial activities a sample is very active if IC50 < 5 mg/L, active if IC50 between 5 and
50 mg/L, weakly active if IC50 between 50 and 100 mg/L and inactive if IC50 > 100 mg/L [24].
3.7. Cytotoxicity Evaluation
Cytotoxicity of sample was estimated on human breast cancer cells (MCF-7). The cells were
cultured in the same conditions as those used for P. falciparum, except for the 10% human serum,
which was replaced by 10% foetal calf serum (Lonza). For the determination of pure compound
activity, cells were distributed in 96-well plates at 3 × 104 cells/well in 100 µL, and then 100 µL of
culture medium containing sample at various concentrations were added. Cell growth was estimated by
(3H)-hypoxanthine incorporation after 48h incubation exactly as for the P. falciparum assay. The
(3H)-hypoxanthine incorporation in the presence of sample was compared with that of control cultures
without sample (positive control being doxorubicin) [25].
3.8. Statistical Analysis
All data were expressed as mean ± standard deviation of triplicate measurements. The confidence
limits were set at P < 0.05. Standard deviations (SD) did not exceed 5% for the majority of the values
Molecules 2011, 16
4. Conclusions
In conclusion, we have identified all volatile constituents of H. gymnocephalum leaf essential oil
and evaluated its anticancer, antimalarial, and antioxidant activities. Our results clearly showed that
this essential oil was active against the tumor cell lines MCF-7 and the FcB1 strain of P. falciparum.
Based on established correlations, compounds such as α-terpinolene, aromadendrene and β-selinene
against MCF-7, could be the best candidates for further analysis. Purification of these compounds is
under development to test them separately. In addition, an in depth study could determine the
mechanisms by which these compounds exert their biological activities. The results presented here
can be considered as the first information on the anticancer and antimalarial properties of
H. gymnocephalum. This may also contribute to our knowledge of the genus Helichrysum. These in
vitro results provide some scientific validation for the widespread use of plants from this genus in
traditional medicine.
1. Humbert, H. Flore de Madagascar et des Comores. 189e Famille, Composées (Compositae);
Firmin-Didot et Cie: Paris, France, 1962; pp. 339-622.
2. Zechmeister, L.; Herz, W.; Grisebach, H.; Kirby, G.W.; Chang, C.W.J.; Flament, I. The Chemistry
of Organic Natural Products; Springer-Verlag: New York, NY, USA, 1979; p. 302.
3. Ramanoelina, A.R.P.; Terrom, G.P.; Bianchini, J.P.; Coulanges, P. Contribution à l’étude de
l’action antibactérienne de quelques huiles essentielles de plantes malgaches. Arch. Inst. Past.
Madag. 1987, 53, 217-226.
4. De Medici, D.; Pieretti, S.; Salvatore, G.; Nicoletti, M.; Rasoanaivo, P. Chemical analysis of
essential oils of malagasy medicinal plants by gas chromatography and NMR spectroscopy.
Flav. Fragr. J. 1992, 7, 275-281.
5. Afolayan, A.J.; Meyer, J.J.M. The antimicrobial activity of 3,5,7-trihydroxyflavone isolated from
the shoots of Helichrysum aureonitens. J. Ethnopharmacol. 1997, 57, 177-181.
6. Boiteau, P.; Allorge-Boiteau, L. Plantes médicinales de Madagascar; ACCT: Paris, France, 1993;
pp. 135-136.
7. Debray, M.; Jacquemin, H.; Razafindrambao, R. Contribution à l’inventaire des plantes
médicinales de Madagascar; ORSTOM: Paris, France, 1971; p. 150.
8. Cabanis, Y.; Chabouis, L.; Chabouis, F. Végétaux et groupements végétaux de Madagascar et des
Mascareignes; BDPA: Tananarive, Madagascar, 1969; Tome I–IV, p. 1342.
9. Boiteau, P. Médecine traditionnelle et pharmacopée, précis de matière médicale malgache;
ACCT: Paris, France, 1986; p. 141.
10. Cavalli, J.F. Caractérisation par CPG/IK, CPG/SM et RMN du Carbone-13 d’huiles essentielles
de Madagascar. Ph.D. Thesis. Faculty of Science and Technology, Pascal Paoli University of
Corsica: Corse, France, 2002.
11. Guerin, J.P.; Olliaro, P.; Nosten, F.; Druilhe, P.; Laxminarayan, R.; Binka, F.; Kilama, W.L.;
Ford, N.; White, N.J. Malaria, current status of control, diagnosis, treatment, and a proposed
agenda for research and development. Lancet Infect. Dis. 2002, 2, 564-573.
Molecules 2011, 16
12. Greenlee, R.T.; Murray, T.; Bolden, S.; Wingo, P.A. Cancer statistics. Cancer J. Clin. 2000, 50,
13. Cordell, G.A.; Beecher, C.W.; Pezzut, J.M. Can ethnopharmacology contribute to development of
new anti-cancer? J. Ethnopharmacol. 1991, 32, 117-133.
14. Popoca, J.; Aguilar, A.; Alonso, D.; Villarreal, M.L. Cytotoxic activity of selected plant used as
antitumorals in Mexican traditional medicine. J. Ethnopharmacol. 1998, 59, 173-177.
15. European Pharmacopoeia; Maissoneuve SA: Sainte Ruffine, France, 1983.
16. Blois, M.S. Antioxidant determination by use of free radical stable. Nature 1958, 181, 1199-1200.
17. Re, R.; Pellegrini, N.; Proteggente, A.; Pannala, A.; Yang, M.; Rice-Evans, C. Antioxidant
activity applying an improved ABTS radical cation decolorization assay. Free Radic. Biol. Med.
1999, 26, 1231-1237.
18. Trager, W.; Jensen, J.B. Human malarial parasites in continuous culture. Science 1976, 193, 673-675.
19. Desoubzdanne, D.; Marcourt, L.; Raux, R.; Chevalley, S.; Dorin, D.; Doerig, C.; Valentin, A.;
Ausseil, F.; Debitus, C. Alisiaquinones and alisiaquinol, dual inhibitors of Plasmodium
falciparum enzyme targets from a New Caledonian deep water sponge. J. Nat. Prod. 2008, 71,
20. Ribaut, C.; Berry, A.; Chevalley, S.; Reybier, K.; Morlais, I.; Parzy, D.; Nepveu, F.; Benoit-Vical, F.;
Valentin, A. Concentration and purification by magnetic separation of the erythrocytic stages of
all human Plasmodium species. Malar. J. 2008, 7, 45.
21. Lambros, C.; Vanderberg, J.P. Synchronization of Plasmodium falciparum erythrocytic stages in
culture. J. Parasitol. 1979, 65, 418-420.
22. Benoit-Vical, F.; Valentin, A.; Mallié, M.; Bastide, J.M.; Bessière, J.M. In vitro antimalarial
activity and cytotoxicity of Cochlospermum tinctorium and C. planchonii leaf extracts and
essential oils. Planta Med. 1999, 65, 378-381.
23. Muñoz, V.; Sauvain, M.; Mollinedo, P.; Callapa, J.; Rojas, I.; Gimenez, A.; Valentin, A.;
Mallié, M. Antimalarial activity and cytotoxicity of () roemrefidine isolated from the stem bark
of Sparattanthelium amazonum. Planta Med. 1999, 65, 448-449.
24. Ouattara, Y.; Sanon, S.; Traoré, Y.; Mahiou, V.; Azas, N.; Sawadogo, L. Antimalarial activity
of swartzia madagascarensis desv. (leguminosae), combretum glutinosum Guill. & Perr.
(combretaceae) and tinospora bakis miers. (menispermaceae) Burkina Fasso medicinal plants.
Afr. J. Trad. Cam. 2006, 3, 75-81.
25. Cachet, N.; Hoakwie, F.; Bertani, S.; Bourdy, G.; Deharo, E.; Stien, D.; Houel, E.; Gornitzka, H.;
Fillaux, J.; Chevalley, S.; Valentin, A.; Jullian, V. Antimalarial activity of simalikalactone E, a
new quassinoid from Quassia amara L. (Simaroubaceae). Antimicrob. Agents Chemother. 2009,
53, 4393-4398.
26. Mollenbeck, S.; Konig, T.; Schreier, P.; Schwab, W.; Rajaonarivony, J.; Ranarivelo, L. Chemical
composition and analyses of enantiomers of essential oils from Madagascar. Flav. Frag. J. 1997,
12, 63-69.
27. Cavalli, J.F.; Ranarivelo, L.; Ratsimbason, M.; Bernardini, A.F.; Casanova, J. Constituents of the
essential oil of six Helichrysum species from Madagascar. Flav. Fragr. J. 2001, 116, 253-256.
Molecules 2011, 16
28. Lourens, A.C.U.; Reddya, D.; Baser, K.H.C.; Viljoena, A.M.; Van Vuuren, S.F. In vitro
biological activity and essential oil composition of four indigenous South African Helichrysum
species. J. Ethnopharmacol. 2004, 95, 253-258.
29. Sharma, P.R.; Dilip, M.; Mondhe, D.M.; Muthiah, S.; Pal, H.C.; Ashok, K.; Shahi, A.K.; Ajit, K.;
Saxena, A.K.; Qazi, G.N. Anticancer activity of an essential oil from Cymbopogon flexuosus.
Chem.-Biol. Inter. 2009, 179, 160-168.
30. Li, Y.L.; Yeung, C.M.; Chiu, L.C.M.; Cen, Y.Z.; Ooi, V.E.C. Chemical composition and
antiproliferative activity of essential oil from the leaves of a medicinal herb, Schefflera
heptaphylla. Phytother. Res. 2009, 23, 140-142.
31. Al-Kalaldeh, J.Z.; Abu-Dahab, R.; Afifi, F.U. Volatile oil composition and antiproliferative
activity of Laurus nobilis, Origanum syriacum, Origanum vulgare, and Salvia triloba against
human breast adenocarcinoma cells. Nutr. Res. 2010, 30, 271-278.
32. Sibanda, S.; Chigwada, G.; Poole, M.; Gwebu, E.T.; Noletto, J.A.; Schmidt, J.M.; Rea, A.I;
Setzer, W.N. Composition and bioactivity of the leaf essential oil of Heteropyxis dehniae from
Zimbabwe. J. Ethnopharmacol. 2004, 92, 107-111.
33. Bendaoud, H.; Romdhane, M.; Souchard, J.P.; Cazaux, S.; Bouajila, J. Chemical composition and
anticancer and antioxidant activities of Schinus Molle L. and Schinus Terebinthifolius Raddi
Berries essential oils. J. Food Sci. 2010, 75, 466-472.
34. El Hadri, A.; Gómez del Río, M.A.; Sanz, J.; González Coloma, A.; Idaomar, M.; Ozonas, B.R.;
González, J.B.; Sánchez Reus, M.I. Cytotoxic activity of α-humulene and trans-caryophyllene
from Salvia officinalis in animal and human tumor cells. An. R. Acad. Nac. Farm. 2010, 76, 343-356.
35. Liu, X.; Zu, Y.; Fu, Y.; Yao, L.; Gu, C.; Wang, W.; Efferth, T. Antimicrobial activity and
cytotoxicity towards cancer cells of Melaleuca alternifolia (tea tree) oil. Eur. Food Res. Technol.
2009, 229, 247-253.
36. Monajemi, R.; Oryan, S.; Haeri-Roohani, A.; Ghannadi, A.; Jafarian, A. Cytotoxic effects of
essential oils of some Iranian citrus peels. Iranian J. Pharm. Res. 2005, 3, 183-187.
37. Haber, W.A.; Agius, B.R.; Stokes, S.L.; Setzer, W.N. Bioactivity and chemical composition of the
leaf essential oil of Talauma gloriensis Pittier (Magnoliaceae) from Monteverde, Costa Rica.
Rec. Nat. Prod. 2008, 2, 1-5.
38. Lopes, N.P.; Kato, M.J.; De A Andrade, E.H.; Maiaa, J.G.S.; Yoshida, M.; Planchart, A.R.;
Katzin, A.M. Antimalarial use of volatile oil from leaves of Virola surinamensis (Rol.) Warb. by
Waiǎpi Amazon Indians. J. Ethnopharmacol. 1999, 67, 313-319.
39. Van Zyl, R.L.; Seatlholo, S.T.; Van Vuuren, S.F.; Viljoen, A.M. The biological activity of
20 nature identical essential oil constituents. J. Essential Oil Res. 2006, 18, 129-133.
40. Boyom, F.F.; Ngouana, V.; Amvam Zollo, P.H.; Menut, C.; Bessiere, J.M.; Gut, J.; Rosenthal, P.J.
Composition and anti-plasmodial activities of essential oils from some Cameroonian medicinal
plants. Photochemistry 2003, 64, 1269-1275.
41. Kamatou, G.P.P.; Viljoen, A.M.; Gono-Bwalya, A.B.; Van Zyl, R.L.; Van Vuuren, S.F.;
Lourens, A.C.U.; Baser, K.H.C.; Demirci, B.; Lindsey, K.L.; Van Staden, J.; Steenkamp, P. The
in vitro pharmacological activities and a chemical investigation of three South African Salvia
species. J. Ethnopharmacol. 2005, 102, 382-390.
Molecules 2011, 16
42. Kamatou, G.P.P.; Viljoen, A.M.; Figueiredo, A.C.; Tilney, P.M.; Van Zyl, R.L.; Barroso, J.G.;
Pedro, L.G.; Van Vuuren, S.F. Trichomes, essential oil composition and biological activities of
Salvia albicaulis Benth. and S. dolomitica Codd, two species from the Cape region of South
Africa. S. Afr. J. Bot. 2007, 73, 102-108.
43. Valentin, A.; Pélissier, Y.; Benoit, F.; Marion, C.; Kone, D.; Mallie, M.; Bastide, J.M.;
Bessiere, J.M. Composition and antimalarial activity in vitro of volatile components of lippia
multiflora. Photochemistry 1995, 40, 1439-1442.
44. Van Vuuren, S.F.; Viljoen, A.M.; Van Zyl, R.L.; Van Heerden, F.R.; Baser, K.H.C. The
antimicrobial, antimalarial and toxicity profiles of helihumulone, leaf essential oil and extracts of
Helichrysum cymosum (L.) D. Don subsp. Cymosum. S. Afr. J. Bot. 2006, 72, 287-290.
45. Ortet, R.; Thomas, O.P.; Regalado, E.L.; Pino, J.A.; Filippi, J.J.; Fernandez, M.D. Composition
and biological properties of the volatile oil of Artemisia gorgonum Webb. Chem. Biodivers. 2010,
7, 1325-1333.
46. Tabanca, N.; Demirci, B.; Crockett, S.L.; Baser, K.H.C.; Wedge, D.E. Chemical composition and
antifungal activity of Arnica longifolia, Aster hesperius, and Chrysothamnus nauseosus essential
oils. J. Agric. Food Chem. 2007, 55, 8430-8435.
Sample Availability: Samples of the compounds are available from the authors.
© 2011 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article
distributed under the terms and conditions of the Creative Commons Attribution license
... However, works focused on the chemical composition of essential oils. Only a few species have been studied in terms of biological activity: H. gymnocephalum (Antiplasmodial and antioxydant activies, cytotoxicity) [21,22], H. faradifani (Insecticidal activity) [12], H. bracteiferum and H. lavanduloides (Antibacterial activities) [22]. ...
... The HIEO ester index (54) is significantly higher than that of Chromolaena odorata (21) and for Pimenta racemosa (33.75) [43], but by far inferior to that of Kaempferia galanga rhizomes EO (189.65) [39] and Cananga odorata flower EO (350.6) ...
Full-text available
The present work aimed to study the composition and antimicrobial properties of the essential oil of Helichrysum ibityense leaves (HIEO). HIEO was extracted by hydrodistillation from fresh leaves with a yield of 1.9%. It had a relative density of 0.9247 at 20℃, a refractive index of 1.4706, an optical rotation of-0°33, an acid index of 2.10 and an ester index of 54. Gas chromatography analysis/flame ionisation detection of HIEO had identified 17 components, representing more than 99.54% of the overall composition of the HIEO including 1.8-cineole (69.46%) the major component, α-terpinene (4.62%), β-caryophyllene (4.39%), α-pinene (4.13%), β-pinene (2.96%), p-cymene (2.39%) and γ-terpinene (2.16%). The HIEO antimicrobial activity was tested against nine pathogenic bacteria including Staphylococcus aureus, Staphylococcus pneumoniae, Streptococcus pyogenes, Clostridium perfringens, Bacillus cereus, Pseudomonas aeruginosa, Enterobacter cloacae, Enterobacter aerogenes, Yersinia enterocolitica, and Candida albicans using the Disc diffusion and the Microdilution assays. HIEO exhibited broad activity spectrum and high microbial activity with inhibition zones (IZ) ranging from 12 to 35 mm. On Bacillus cereus, the most sensitive bacterium, its Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) were 9.37 mg/ml and 13.22 mg/ml respectively. With an MBC/MIC ratio of 1.41, HIEO exerted a bactericidal action on Bacillus cereus. When administered orally to mice at 1000 mg/kg body weight, HIEO caused symptoms of intoxication which disappeared after 24 h. These preliminary results revealed HIEO could be used to treat different infectious diseases if its safety will be confirmed.
... However, a few phytoconstituents of plants extracts do not show anticancer activity as individual compounds; however, when they are combined with other compounds, they exude anticancer activity. For example, β-selinene, aromadendrene and α-terpinolene together show cytotoxicity against MCF-7 cells [24]. ...
... The results were compared with other studies, and a correlation was determined between the content of various terpenes and the cytotoxicity of the essential oils of fifteen different plants. A high correlation of cytotoxic effects on MCF-7 was found for β-selinene (R 2 = 0.76), α-terpinolene (R 2 = 0.88) and aromadendrene (R 2 = 0.90), [29]. ...
Cannabis (Cannabis sativa L.) for medical purposes has been legalized again in many countries in recent years. Currently, only two major cannabinoids (Δ⁹-THC and CBD) are considered in the legislation and medication, which is not sufficient in case of dried plant material or resulting extract. Other substances (mainly terpenes/terpenoids), or their specific combinations, could influence the resulting therapeutic effect for specific oncology diagnosis and specific patients. Six different genotypes (Conspiracy Kush, Jilly Bean, Jack Cleaner 2, Jack Skellington, Nordle and Nurse Jackie) were cultivated indoor at the Czech University of Life Sciences Prague. Ethanol extracts taken from the inflorescences were assayed for their content of main cannabinoids and terpenes/terpenoids. The extracts were used for in vitro cytotoxicity studies on hepatocarcinoma human cell lines Hep-G2 and colorectal carcinoma human cell lines Caco-2 and Ht-29. Healthy lung fibroblast MRC-5 and healthy intestinal cells FHs 74 Int were used to compare selectivity of cytotoxicity. The average content of Δ⁹-THC in extracts was 59.1 ± 2.43%, and of CBD 1.84 ± 0.17%. The content of main cannabinoids in the Nurse Jackie genotype extract was significantly greater than that of the other genotypes. Overall, more than 60 different terpenes/terpenoids were identified in the extracts. The major terpenes/terpenoids detected in most genotypes were limonene, linalool, α-terpineol, β-caryophyllene, trans-α-bergamotene, α-humulene, β-caryophyllene oxide, guaiol, γ-eudesmol, β-eudesmol and α-bisabolol. Differences in the terpene composition of individual genotypes were caused by minor terpenoids, such as β-ocimene, isopulegol acetate, β-elemene, β-selinene and spathulenol. All extracts were highly cytotoxic to Ht-29 colorectal carcinoma cells and showed positive selectivity compared to healthy FHs 74 Int colon cells. The Jack Cleaner 2 extract was cytotoxic to all cell lines tested at the lowest concentrations (8.48 ± 2.4–16.14 ± 0,07 μg/mL), but was positively selective only for colorectal cancer cells, especially Ht-29 and to a lesser extent for Caco-2. Similarly, the Nordle extract showed positive selectivity for Ht-29 and Caco-2 only. Jilly Bean was unique in this study, in that its extract functioned on all cell lines at the highest concentrations (20.13 ± 3.05–49.88 ± 1.5 μg/mL), whilst also being highly positively selective in all carcinoma lines (Ht-29, Caco-2 and Hep-G2 hepatocarcinoma) compared to healthy cell lines (FHs 74 Int and MRC-5). The results suggest that Δ⁹-THC and CBD are responsible for the in vitro cytotoxicity of the extracts, but observed differences in selectivity reveal their synergies with other substances. According to chemical analysis, higher concentrations of myrcene, β-elemene, β-selinene and α-bisabolol oxide found in the Jilly Bean genotype may positively affect the selectivity of cytotoxic activity. It is therefore vital that similar studies are performed on other cell lines, in order to be able to recommend these cannabis genotypes for preclinical and clinical studies, which are still lacking.
... The plant is also known to heal wounds. Its essential oil has been used in aromatherapy for its antimicrobial properties (5)(6)(7). From H. gymnocephalum, Ranarivelo et al. (8) isolated Cardamomin with antiplasmodial activity with an IC 50 of 43 µM against Plasmodium falciparum FCM29, a resistant strain to chloroquine. ...
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Lavender with the scientific name Lavandula stricta Del from the mint family is one of the herbs used in traditional Iranian medicine. The aim of this study was to determine the functional groups and qualitative identification, the amount of phenolic and flavonoid compounds, antioxidant and antimicrobial activity of Lavandula stricta essential oil. Essential factor groups were determined by Fourier transform infrared (FTIR) spectroscopy in the wavelength range of 500–4000 cm⁻¹. Total phenols and flavonoids were measured by Folin-Ciocalteu and aluminum trichloride colorimetry, respectively. Antioxidant activity was also determined by two methods of free radical scavenging DPPH and ABTS. The peaks observed in Lavandula stricta essential oil confirmed the presence of O-H, C-H, C=O, C=C, C-C and C-O functional groups of bioactive compounds. The amount of phenol and flavonoids in total essential oil was 68.60±0.68 mg GAE/g and 19.10±0.52 mg QE/g, respectively. Antioxidant activity was also obtained based on the percentage of free radical scavenging DPPH and ABTS equal to 59.50±0.63 and 67.68±0.53, respectively. The antimicrobial activity of essential oil was evaluated by four methods: disk diffusion, agar well, minimum inhibitory concentration and minimum bactericidal concentration. The results of measuring the antimicrobial activity of essential oil by disk diffusion and agar well showed that Gram-positive bacterium Staphylococcus aureus is the most sensitive strain to Lavandula stricta essential oil. The minimum inhibitory concentration of essential oil for Salmonella typhi, Pseudomonas aeruginosa, Listeria innocua, Staphylococcus aureus and Escherichia coli was equal to 3.125 mg/ml and for Bacillus cereus it was equal to 6.25 mg/ml. The minimum bactericidal concentrations for Listeria innocua, Pseudomonas aeruginosa and Salmonella typhi was equal to 100, 200 and 400 mg/ml, respectively, and for other bacteria (Staphylococcus aureus, Bacillus cereus and Escherichia coli) it was more than 400 mg/ml.
Cancer retains a central place in fatality rates among the wide variety of diseases known world over, and the conventional synthetic medicaments, albeit used until now, produce numerous side effects. As a result, newer, better, and safer alternatives such as natural plant products, are gravely required. Essential oils (EOs) offer a plethora of bioactivities including antibacterial, antiviral, antioxidant, and anticancer properties, therefore, the use of EOs in combination with synthetic drugs or aromatherapy continues to be popular in many settings. In view of the paramount importance of EOs and their potential bioactivities, this review summarizes the current knowledge on the interconnection between EOs and cancer treatment. In particular, the current review presents an updated summary of the chemical composition of EOs, their current applications in cancer treatments based on clinical studies, and the mechanism of action against the cancer cell lines. Similarly, an overview of using EOs in aromatherapy and enhancing immunity during cancer treatment is provided. Further, this review focuses on the recent technological advancements such as the loading of EOs using protein microspheres, ligands, or nanoemulsions/nanoencapsulation, which offer multiple benefits in cancer treatment via site-specific and target-oriented delivery of drugs. The continuing clinical studies of EOs implicate that their pharmacological applications are a rewarding research area.
L’usage des plantes médicinales connaît un regain d’intérêt. Cela est lié à la toxicité des produits chimiques, au coût élevé des médicaments chimiques, à l’éloignement et/ou l’insuffisance des centres de santé surtout en milieu rural. L’objectif général de cette étude était d’évaluer les potentialités thérapeutiques de l’huile essentielle des feuilles de l’espèce Erigeron floribundus utilisées en médecine traditionnelle en Côte-d’Ivoire. Les rendements en huile essentielle des feuilles d’ Erigeron floribundus varient de 0,16 % pour les feuilles fraîches et de 0,31 % pour les feuilles séchées. Les indices physiques et chimiques de ces huiles essentielles sont en adéquation avec les critères de qualité des huiles essentielles selon la norme Afnor. L’effet antifongique des deux types d’huiles essentielles a été très remarquable sur les souches fongiques testées, notamment Trichophyton mentagrophytes et Aspergillus fumigatus , avec un effet moindre sur Candida albicans . Cette étude valide ainsi scientifiquement les usages traditionnels des extraits de la plante Erigeron floribundus en médecine traditionnelle.
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Background Malaria still constitutes a major public health menace, especially in tropical and subtropical countries. Close to half a million people mainly children in Africa, die every year from the disease. With the rising resistance to frontline drugs (artemisinin-based combinations), there is a need to accelerate the discovery and development of newer anti-malarial drugs. A systematic review was conducted to identify the African medicinal plants with significant antiplasmodial and/or anti-malarial activity, toxicity, as wells as assessing the variation in their activity between study designs (in vitro and in vivo). Methods Key health-related databases including Google Scholar, PubMed, PubMed Central, and Science Direct were searched for relevant literature on the antiplasmodial and anti-malarial activities of African medicinal plants. Results In total, 200 research articles were identified, a majority of which were studies conducted in Nigeria. The selected research articles constituted 722 independent experiments evaluating 502 plant species. Of the 722 studies, 81.9%, 12.4%, and 5.5% were in vitro, in vivo , and combined in vitro and in vivo , respectively. The most frequently investigated plant species were Azadirachta indica, Zanthoxylum chalybeum, Picrilima nitida, and Nauclea latifolia meanwhile Fabaceae, Euphorbiaceae, Annonaceae, Rubiaceae, Rutaceae, Meliaceae, and Lamiaceae were the most frequently investigated plant families. Overall, 248 (34.3%), 241 (33.4%), and 233 (32.3%) of the studies reported very good, good, and moderate activity, respectively. Alchornea cordifolia, Flueggea virosa, Cryptolepis sanguinolenta, Zanthoxylum chalybeum, and Maytenus senegalensis gave consistently very good activity across the different studies. In all, only 31 (4.3%) of studies involved pure compounds and these had significantly (p = 0.044) higher antiplasmodial activity relative to crude extracts. Out of the 198 plant species tested for toxicity, 52 (26.3%) demonstrated some degree of toxicity, with toxicity most frequently reported with Azadirachta indica and Vernonia amygdalina . These species were equally the most frequently inactive plants reported. The leaves were the most frequently reported toxic part of plants used. Furthermore, toxicity was observed to decrease with increasing antiplasmodial activity. Conclusions Although there are many indigenous plants with considerable antiplasmodial and anti-malarial activity, the progress in the development of new anti-malarial drugs from African medicinal plants is still slothful, with only one clinical trial with Cochlospermum planchonii ( Bixaceae ) conducted to date. There is, therefore, the need to scale up anti-malarial drug discovery in the African region.
In this study, we compared the effects of terpinolene against the anti-cancer agent cisplatin on proliferation, apoptotic activity and intracellular ROS (reactive oxygen species) production in cancerous (MCF-7) and non-cancerous (HEK-293) cell lines. Cisplatin presented a strong growth-inhibitory effect on both MCF-7 (IC25: 18.77 µM) and HEK-293 (IC25: 15.03 µM) cell lines. The growth-inhibitory effect of terpinolene was found to be weaker than cisplatin (IC25: 291.18 µM for MCF-7 and 345.35 µM for HEK 293 cells). The expression levels of BAX (by 1.86-fold), cleaved-PARP (by 2.23-fold) and pro-caspase-8 (by 1.74-fold) proteins increased in response to terpinolene treatment in MCF-7 cells. Curiously, the increase in the expression levels of these apoptotic markers was less pronounced in terpinolene-treated non-cancerous HEK 293 cells. Terpinolene, promoted a higher rate of apoptotic and necrotic cell death on MCF-7 cells (apoptotic cells: 25.28 %; necrotic cells: 17.70 %) when compared with HEK-293 cell line (apoptotic cells: 15.47 %; necrotic cells: 9.42 %). Interestingly, terpinolene caused a marked increase (2.05-fold) in intracellular ROS production in MCF-7 cells while HEK-293 cells appeared to be resistant to terpinolene or cisplatin treatments. In conclusion, although terpinolene showed a weaker growth-inhibitory effect on MCF-7 cancer cells, it exhibited a better selectivity than cisplatin in inducing cell death and intracellular oxidative stress in MCF7 breast cancer cells.
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The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in China and its spread worldwide has become one of the biggest health problem due to the lack of knowledge about an effective chemotherapy. Based on the current reality of the SARS-CoV-2 pandemic, this study aimed to make a review literature about potential anti-coronavirus natural compounds guided by an in silico study. In the first step, essential oils from native species found in the Brazilian herbal medicine market and Brazilian species that have already shown antiviral potential were used as source for the literature search and compounds selection. Among these compounds, 184 showed high antiviral potential against rhinovirus or picornavirus by quantitative structure–activity relationship analysis. (E)-α-atlantone; 14-hydroxy-α-muurolene; allo-aromadendrene epoxide; amorpha-4,9-dien-2-ol; aristochene; azulenol; germacrene A; guaia-6,9-diene; hedycaryol; humulene epoxide II; α-amorphene; α-cadinene; α-calacorene and α-muurolene showed by a molecular docking study the best result for four target proteins that are essential for SARS-CoV-2 lifecycle. In addition, other parameters obtained for the selected compounds indicated low toxicity and showed good probability to achieve cell permeability and be used as a drug. These results guided the second literature search which included other species in addition to native Brazilian plants. The majority presence of any of these compounds was reported for essential oils from 45 species. In view of the few studies relating essential oils and antiviral activity, this review is important for future assays against the new coronavirus.
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Twenty nature identical essential oil constituents from seven structural groups were tested for antimalarial, antimicrobial, antioxidant and toxicity properties. The compounds displayed antimalarial activity (IC50 values ranging from 0.9 to 1528.8 μM) with varying toxicity (IC50 values ranging from 5.5 to 1358.4 μM). The data indicates that (-)-pulegone, (E- & Z-)-(±)-nerolidol, linalyl acetate and (+)-α-pinene have the most potent antimalaial activity. Carvacrol (MIC values ranging from <1.66 to 13.3 mM) followed by geraniol (MIC values: 19.5-51.9 mM) displayed the broadest spectrum of antimicrobial activity. The least toxic of the tested compounds, eugenol, was the only compound with antioxidant activity comparable to ascorbic acid. The results demonstrate that the 20 compounds individually display variable biological activity, although further research is required to determine how they may interact when combined.
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The chemical composition of Artemisia gorgonumWebb essential oil from Cape Verde was analyzed by GC and GC/MS. A total of 111 volatile compounds, accounting for 94.9% of the essential oil, were identified by GC and GC/MS. The major compounds were camphor (28.7%), chrysanthenone (10.8%), lavandulyl 2-methylbutanoate (9.5%), α-phellandrene (5.5%), lavandulyl propanoate (4.2%), camphene (4.0%), and p-cymene (3.4%). The volatile oil of this endemic plant, which is used in Cape Verdean folk medicine against several ailments, was tested for its antioxidant and antimalarial properties, and was found to exhibit free-radical scavenging on 1,1-diphenyl-2-picrylhydrazyl (DPPH), prevention of lipid peroxidation–in vitro by TBARS (thiobarbituric acid reactive species) assay, and antiplasmodial activity.
Background: The purpose of the present work is two-fold: the fractionation of Salvia officinalis essential oil and the cytotoxic study of this oil with its fractions "in vitro" tumor cell lines. Materials and Methods: S. officinalis essential oil was obtained by hydrodistillation and fractionated with column chromatography; the essential oil and its fractions were analyzed by gas chromatography (GC) coupled to mass spectrometry (MS). The cytotoxic activity was evaluated in cellular lines of breast cancer MCF-7, colon cancer HCT-116, and murine macrophage RAW264.7 cell lines by the MTT assay Results: the sub-subfraction F1.1.1 of S. officinalis essential oil containing alpha-humulene present highest activity on RAW264.7 and HCT-116 with IC(50) values of 41.9 and 77.3 mu g/ml, respectively. The sub-subfraction F1.2.1 of S. officinalis essential oil with trans-caryophyllene showed less activity on RAW246.7 and HCT-116 with IC(50) values of 90.5 and 145.8 mu g/ml. Conclusion: This paper suggests that the a-humulene and trans-caryophyllene extracted from S.officinalis essential oil inhibit tumor cell growth.
A method for the screening of antioxidant activity is reported as a decolorization assay applicable to both lipophilic and hydrophilic antioxidants, including flavonoids, hydroxycinnamates, carotenoids, and plasma antioxidants. The pre-formed radical monocation of 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS*+) is generated by oxidation of ABTS with potassium persulfate and is reduced in the presence of such hydrogen-donating antioxidants. The influences of both the concentration of antioxidant and duration of reaction on the inhibition of the radical cation absorption are taken into account when determining the antioxidant activity. This assay clearly improves the original TEAC assay (the ferryl myoglobin/ABTS assay) for the determination of antioxidant activity in a number of ways. First, the chemistry involves the direct generation of the ABTS radical monocation with no involvement of an intermediary radical. Second, it is a decolorization assay; thus the radical cation is pre-formed prior to addition of antioxidant test systems, rather than the generation of the radical taking place continually in the presence of the antioxidant. Hence the results obtained with the improved system may not always be directly comparable with those obtained using the original TEAC assay. Third, it is applicable to both aqueous and lipophilic systems.
Essential oils from three different Asteraceae obtained by hydrodistillation of aerial parts were analyzed by gas chromatography (GC) and gas chromatography–mass spectrometry (GC/MS). Main compounds obtained from each taxon were found as follows: Arnica longifolia carvacrol 37.3%, R-bisabolol 8.2%; Aster hesperius hexadecanoic acid 29.6%, carvacrol 15.2%; and Chrysothamnus nauseosus var. nauseosus-phellandrene 22.8% and-pinene 19.8%. Essential oils were also evaluated for their antimalarial and antimicrobial activity against human pathogens, and antifungal activities against plant pathogens. No antimalarial and antimicrobial activities against human pathogens were observed. Direct bioautography demonstrated antifungal activity of the essential oils obtained from three Asteraceae taxa and two pure compounds, carvacrol and-bisabolol, to the plant pathogens Colletotrichum acutatum, C. fragariae and C. gloeosporioides. Subsequent evaluation of antifungal compounds using a 96-well micro-dilution broth assay indicated that R-bisabolol showed weak growth inhibition of the plant pathogen Botrytis cinerea after 72 h.
METHODS for measuring antioxidants and appraising antioxidant activity appear to be of two general types. If the chemical nature of the antioxidant is known, one may strive for a test specific for the compound or group of interest; for example, the nitroprusside test for sulphydryl groups. Alternatively one may observe the inhibition of some natural oxidative process such as the β-oxidation of fats, as a function of the added antioxidant.