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Anticancer effect of lemongrass oil and citral on cervical cancer cell lines

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Aims: The aim of the present study was to evaluate the anticancer effect of lemongrass oil and citral emulsion on cervical cancer cell lines (HeLa and ME-180) in vitro. Settings and Design: Citral is a very important component in lemongrass oil. It is proved to have anticancer properties in various human cancer cell lines. Methods and Material: DLS analysis revealed the average size of the lemongrass oil emulsion to be 267 nm and the average size of the citral emulsion to be 270 nm. The anticancer effect of both the emulsions was determined by MTT assay, DCFH-DA method, Rh-123 and AO/EtBr-staining. Statistical analysis used: One-way ANOVA followed by DMRT taking p<0.05 to test the significant difference between groups. Results: The results summarize that lemongrass oil and citral emulsions initiate the cancer cell death by decreasing cell proliferation, increasing intracellular ROS, altering mitochondrial membrane potential, and initiating apoptosis in HeLa and ME-180 cell lines. The present findings of this study clearly demonstrate the involvement of oxidative mechanism for the anti-proliferative effect in HeLa and ME-180 cell lines. ME-180 being chemosensitive showed good results at lower concentrations of citral (IC50 24 h 300 μg/ml), as compared to chemoresistant HeLa cells (citral IC50 24 h 500 μg/ml). Whereas lemongrass oil exhibited better activity in both the cell lines (IC50 24 h 200 μg/ml). Conclusions: All the results suggest lemongrass oil and citral emulsion could be considered as potent candidates for anticancer agents.
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41
Pharmacognosy Communications
Volume 3 | Issue 4 | Oct–Dec 2013 www.phcogcommn.org
Research Article
© Copyright 2013 EManuscript Publishing Services, India
*Correspondence
Kavisa Ghosh
Department of Zoology, Annamalai University, Annamalainagar,
Chidambaram 608002.
Mobile: +91 7502699517
E-mail: Kavisa_9@yahoo.co.in
DOI: 10.5530/pc.2013.4.6
Anticancer effect of lemongrass oil and
citral on cervical cancer cell lines
Kavisa Ghosh*
Department of Zoology, Annamalai University, Annamalainagar, Chidambaram 608002
ABSTRACT: Aims: The aim of the present study was to evaluate the anticancer effect of lemongrass oil and citral
emulsion on cervical cancer cell lines (HeLa and ME-180) in vitro. Settings and Design: Citral is a very important
component in lemongrass oil. It is proved to have anticancer properties in various human cancer cell lines. Methods
and Material: DLS analysis revealed the average size of the lemongrass oil emulsion to be 267 nm and the average
size of the citral emulsion to be 270 nm. The anticancer effect of both the emulsions was determined by MTT assay,
DCFH-DA method, Rh-123 and AO/EtBr-staining. Statistical analysis used: One-way ANOVA followed by DMRT taking
p<0.05 to test the signicant difference between groups. Results: The results summarize that lemongrass oil and
citral emulsions initiate the cancer cell death by decreasing cell proliferation, increasing intracellular ROS, altering
mitochondrial membrane potential, and initiating apoptosis in HeLa and ME-180 cell lines. The present ndings of this
study clearly demonstrate the involvement of oxidative mechanism for the anti-proliferative effect in HeLa and ME-180
cell lines. ME-180 being chemosensitive showed good results at lower concentrations of citral (IC50 24 h 300
µ
g/ml), as
compared to chemoresistant HeLa cells (citral IC50 24 h 500
µ
g/ml). Whereas lemongrass oil exhibited better activity in
both the cell lines (IC50 24 h 200
µ
g/ml). Conclusions: All the results suggest lemongrass oil and citral emulsion could
be considered as potent candidates for anticancer agents.
KEYWORDS: Lemongrass oil, Citral, Emulsion, Anticancer, HeLa, ME-180
KEY MESSAGE: Lemongrass oil and citral emulsion are potent candidates for anticancer ointment based drugs.
INTRODUCTION
Cervical cancer is the third most common cancer in
women, and the seventh overall, with an estimated
530,000 new cases in 2008. More than 85% of the
global burden occurs in developing countries, where
it accounts for 13% of all female cancers. In India,
cervical cancer is the rst threat after breast and ovarian
cancer.[1] The options for treating each patient with
cervical cancer depend on the stage of disease. The
stage of a cancer describes its size, depth of invasion
(how far it has grown into the cervix), and how far it
has spread.[2] Early-stage cancer that is conned to the
cervix, offers an excellent outlook, with a success rate
of over 85%. However, if the cancer has spread to the
vagina, surroundings tissues and pelvic area, or else-
where, the outlook is less positive.[3]
Several natural products are nowadays employed as effec-
tive anticancer agents. In the last two decades the search
for novel anticancer agents from natural sources has wit-
nessed an impressive increase of interest. The genus Cym-
bopogon (family Gramineae) has many species of grasses
that grow in tropical and subtropical regions around the
world from mountains to grasslands to arid zones.[4] These
plants produce essential oils with pleasant aromas in their
leaves. Five species yield the three oils of main commer-
cial importance: lemongrass from Cymbopogon citratus of
Malaysian origin (West Indian lemongrass) and Cymbopogon
exuosus (East Indian lemongrass) from India, Sri Lanka,
Burma, and Thailand; palmarosa oil from Cymbopogon
martinii; citronella oil from Cymbopogon nardus (Sri Lanka),
and Cymbopogon winterianus (Java). Cymbopogon exuosus (also
known as East India or Cochin lemongrass) is a perennial,
Anticancer effect of lemongrass oil and citral on cervical cancer cell lines
42
multicut aromatic grass that yields an essential oil used in
perfumery and pharmaceutical industries and vitamin A.[5]
The plants of lemongrass Cymbopogon citratus and C. exuosus
are medium-sized grasses that are commercially grown
for essential oil distillation in Guatemala (C. citratus) and
Cochin, India (C. exuosus). The fresh grass is cut and
allowed to wilt before it is used for steam-distilled oil
production. Oils from different regions have somewhat
different compositions, Guatemalan West Indian (W.I.)
having slightly different citral content than East Indian
(E.I.) Cochin origin. Lemongrass oil is mobile, pale yellow
in color, with a powerful, fresh, oral-herbal odor that is
quite prominent.[6]
Cymbopogon exuosus oil helps with stress-related disor-
ders, and has shown to have antifungal and antimicro-
bial properties.[7] The chemical composition of the oil has
also been reported.[6] The various constituents (%) pres-
ent in the oil from lemongrass variety of C. exuosus such
as geraniol (20.08), geranyl acetate (12.20), α-bisabolol
(8.42) and isointermedeol (24.97) have been individually
reported for their cancer cell cytotoxicity.[6,8]
The essential oil from a lemongrass variety of Cymbopogon
exuosus (CFO) and its major chemical constituent sesqui-
terpene isointermedeol (ISO) were investigated for their
ability to induce apoptosis in human leukemia HL-60
cells, because deregulation of apoptosis is the hallmark
of cancer cells. CFO and ISO inhibited cell proliferation
with IC50 of ~30 and 20 µg/ml, respectively.[6] Two active
compounds, d-limonene and geraniol, were isolated by
fractionation of lemongrass (C. citratus) oil. These were
tested for their capacity to induce activity of the detoxify-
ing enzyme glutathione-S-transferase (GST) in several tis-
sues of the female A/J mice. d-Limonene increased GST
activity two- to three fold than controls in the mouse liver
and the mucosa of the small and large intestines. Gera-
niol showed high GST-inducing activity in the mucosa of
the small and large intestines, which was about 2.5-fold
greater than controls. Induction of increased GST activity,
which is believed to be a major mechanism for chemical
carcinogen detoxication, has been recognized as one of
the characteristics of the action of anticarcinogens.[9] The
essential oil from C. citratus and its isolated principal citral
have been tested for cytotoxicity against P388 leukemia
cells. The cytotoxicity of citral, IC50 against P388 mouse
leukemia cells was 71 µg/ml. In another experiment, IC50
of C. citratus oil in P388 leukemia cells was found to be
5.7 µg/ml.[10] A study also showed that the oil from Cymbo-
pogon exuosus induced differential in vitro cytotoxicity in 12
human cancer cell lines and in vivo tumor growth inhibiton
in murine, Erlich and S-180 tumor models. The oil also
caused dose dependent increase in apoptosis in HL-cells.[11]
The lemongrass oil used for the study had density of
0.89 g/ml.
Citral (gure 1), or 3, 7-dimethyl-2,6-octadienal (C10H16O),
is a mixture of two isomeric acyclic monoterpene alde-
hydes. The two compounds are double bond isomers. The
trans-citral is known as geranial or citral A. The cis-citral is
known as neral or citral B.[5] Geranial has a strong lemon
odor. Neral has a lemon odor that is less intense and
sweeter than geranial. Citral is an aroma compound used
in perfumery for its citrus effect. Citral is also used for a-
voring and fortifying lemon oil. It also has strong antimi-
crobial qualities[12] and pheromonal effects in insects.[13,14]
Citral is used in the synthesis of vitamin A, ionone, and
methylionone, and to mask the smell of smoke.[5]
Citral has been found to be a potent inducer of
glutathione-s-transferase class of enzymes, which provide
protection to healthy hepatocytes against apoptosis dur-
ing chemotherapy of liver cancers.[15] A good superox-
ide scavenging activity (EC50 = 19 mcg/ml) was reported
in Swiss albino mice in citral treated groups, suggesting
that the antioxidant action could be responsible for the
anti-clastogenic effect of citral against nickel chloride.[16]
Citral, at a concentration comparable to that found in a
cup of tea brewed with 1 gram of lemongrass, was found
to induce apoptosis in cancer cells, without any harm to
normal cells. Apoptosis was accompanied by DNA frag-
mentation and caspase-3 catalytic induction.[17] Citral also
disrupts animal microtubules and inhibits polymerization
of microtubules in vitro.[18] Citral was also tested on cyclo-
oxygenase activity. Citral treatment also caused inhibition of
breast cancer MCF-7 cell growth (IC50 -48 h: 18 × 105 m)
with a cycle arrest in G2/M phase, apoptosis induction and
Figure 1. Citral isomers.
Anticancer effect of lemongrass oil and citral on cervical cancer cell lines
43
also a decrease in prostaglandin E2 synthesis. These ndings
suggested that citral has a potential chemopreventive effect.
[19] The citral used for the study had density of 0.89 g/ml.
The aim of the present study was to evaluate the antican-
cer effect of lemongrass oil and citral emulsion on cervi-
cal cancer cell lines (HeLa and ME-180) in vitro.
SUBJECTS AND METHODS
Chemicals
Citral (0.89 g/ml) was purchased from Sigma chemi-
cal Co., St. Louis, USA. Lemongrass oil was purchased
from Varsha Aromatics, Chennai. Other chemicals for
cell cultures were purchased from Himedia, Mumbai. All
other chemicals and solvents were of analytical grade and
obtained from S.D ne chemical, Mumbai and Fisher
Inorganic and Aromatic Limited, Chennai.
Determining the density of lemongrass oil
Density of the lemongrass oil was measured using spe-
cic gravity bottle. Density of the oil was calculated as
follows: Density = Mass/Volume
Where, Mass = (Substance + Bottle Weight) – (Empty
Bottle Weight)
Volume = 10 ml
Determination of citral content
The citral content in lemongrass oil was determined
by sodium bisulphate method (IS 327: 1991 Oil of
Lemongrass–Specication).
Preparation of emulsion
Emulsion formation method was used as described by
Hart et al., 2000.[20]
Particle size and size distribution
The Lemongrass oil and citral emulsion drop mean size
and distribution were measured with a dynamic light
scattering (DLS) instrument (Zetasizer Nano, Malvern
Instruments Ltd. United Kingdom). Lemongrass oil and
citral were sonicated in ultrapure water before measure-
ment. The analysis was performed at a scattering angle
of 173° at a temperature of 25 °C. The wavelength of
the laser used in the Zetasizer Nano instruments for the
measurement, 632.8 nm ‘red’ laser wavelength.
Cell lines and culture conditions
The cervical cancer cell lines (HeLa and ME-180). were
obtained from National Centre for Cell Science (NCCS),
Pune, India. The cells were grown as monolayer in MEM
medium supplemented with 10% FCS, 1 mM sodium
pyruvate, 10 mM HEPES, 1.5 g/L sodium bicarbonate,
2 mM L-glutamine, and 100 U/ml penicillin-streptomycin
at 37 °C in 5% CO2 atmosphere.
Dose xation study
Cells were treated with lemongrass oil and citral in a
concentration range of 10, 50, 100, 150, 200, 250, 300,
350, 400, 450, 500 µg/ml, and incubated for 24 h. The
cytotoxicity was observed by MTT assay according to the
method of Moshmann.[21] IC50 values were calculated and
optimum dose was used for further study.
Experimental groups
Cervical cancer cells (HeLa and ME-180) were divided
into three groups; in each group six samples were pro-
cessed (Table 1).
Determination of intracellular ROS generation
ROS was measured by using a non-uorescent probe, 2,
7-dichlorodihydrouorescein diacetate (H2-DCFDA) that
can penetrate into the intracellular matrix of cells, by the
method of Bhosle et al., 2005.[22]
Change in mitochondrial transmembrane
potential (Δψm)
The change in ψm in different treatment groups was
observed microscopically and determined using uores-
cent dye Rh-123 by the method of Prasad et al., 2010.[23]
Apoptotic morphological changes
Acridine orange (AO) and ethidium bromide (EtBr) stain-
ing of DNA allowed visualization of the condensed chro-
matin of dead apoptotic cells and was determined by the
method of Lakshmi et al., 2008.[24]
Statistical analysis
Statistical analysis was performed by one-way ANOVA
followed by DMRT taking p < 0.05 to test the signicant
difference between groups.
RESULTS
Density of lemongrass oil
The lemongrass oil used for the study had density of
0.89 g/ml.
Table 1: Experimental design
Group 1:
Control (HeLa) Group 1:
Control (ME-180)
Group 2:
HeLa + Lemongrass oil
(200 µg/ml)
Group 2:
ME-180 + Lemongrass oil
(200 µg/ml)
Group 3:
HeLa + Citral (500 µg/ml) Group 3:
ME-180 + Citral (300 µg/ml)
Anticancer effect of lemongrass oil and citral on cervical cancer cell lines
44
Determination of citral content
78.5% of citral was found to be present in the lemongrass
oil used in the study. The sample used thus conforms to
I.S. 327: 1991 reafrmed 2002 specication.
Particle size distribution and size distribution
DLS results revealed that the average size of the lemongrass
oil emulsion was 267 nm (gure 2A) and the average size
of the citral emulsion was found to be 270 nm (gure 2B).
Dose xation study
Inhibitory concentration 50 (IC50) value for lemongrass
oil and citral was found to be 200 µg/ml and 500 µg/ml
for HeLa cell line, and 200 µg/ml and 300 µg/ml for
ME-180 cells respectively, and it was used for further
experiments (gures 3 A and B). Figure 3 C and D shows
the microscopic images showing the morphological
changes of HeLa and ME-180 cells respectively, during
treatment with IC50 concentrations of lemongrass oil and
citral. At these respective IC50 concentrations, 50% of cell
death was observed in HeLa and ME-180 cell line.
Emulsions of lemongrass oil and citral increased
ROS generation in HeLa and ME-180 cells
Lemongrass oil and citral treatments signicantly increased
ROS level in HeLa and ME-180 cells. Among all the
Figure 2. (A and B): Average size of the lemongrass oil and citral emulsion.
Figure 3. (A and B) Percentage cytotoxicity of lemongrass oil and citral in HeLa and ME-180 cell lines. Inhibitory concentration
50 (IC50) value for lemongrass oil and citral was found to be 200 µg/ml and 500 µg/ml for HeLa cell, and 200 µg/ml and 300 µg/ml
for ME-180 cells respectively, and it was used for further experiments. The values are given as mean ± SD of six experiments in each
group. Figure 3 (C and D) Microscopic images showing the morphological changes of HeLa and ME-180 cells respectively, during
treatment with IC50 concentrations of lemongrass oil and citral.
Anticancer effect of lemongrass oil and citral on cervical cancer cell lines
45
doses tested, 200 µg/ml of lemongrass oil in HeLa cells,
and 200 µg/ml of lemongrass oil ME-180 cells showed
maximum generation of ROS (increased uorescence)
(gures 4 A and B). Fluorescence microscopic images
(gures 4 C and D) conrm the DCF uorescence in
lemongrass and citral treated groups.
Emulsions of lemongrass oil and citral modulate
mitochondrial membrane potential in HeLa and
ME-180 cells
Mitochondrial membrane potential was decreased in
lemongrass oil and citral treated groups compared to
control (gures 5 A and B). Fluorescence microscopic
images (gures 5 C and D) show the accumulation of
Rh-123 dye (red condensed spots) in control and the
accumulation have been found to be decreased in lem-
ongrass oil and citral groups as the membrane potential
decreased.
Effect of lemongrass oil and citral emulsions on
apoptotic morphology
The gure 6 (C and D) shows the photomicrographs
of apoptotic morphological changes in lemongrass oil
and citral treated cells. The % apoptotic cell death was
increased in citral and lemongrass oil (increased orange
colored cells) when compared to control. Figure 6
(A and B) show the quantitative result of apoptosis in dif-
ferent treatment groups. HeLa cells showed 95% apop-
totic cells in lemongrass oil and 80% in citral treatment
groups. ME-180 cells showed 98% apoptotic cells in
lemongrass oil and 90% in citral treatment groups.
DISCUSSION
In this present study, we evaluated the anticancer effect
of lemongrass oil and citral on cervical cancer cell lines
(HeLa and ME-180) in vitro. Lemongrass oil and citral
treatment (24 h incubation) signicantly decreased per-
centage of cell viability in HeLa and ME-180 cells. This
suggested that lemongrass oil and citral treatment was
able to inhibit the growth of cancer cells during incuba-
tion. It was found that (IC50) 200 µg/ml (P > 0.05) of
lemongrass oil could greatly inhibit the cell growth in both
the cell lines. Whereas (IC50) 500 µg/ml and 300 µg/ml
(P > 0.05) concentration of citral was required for good
inhibition of HeLa and ME-180 cells growth respec-
tively. Hence, the result indicates that concentration of
phytochemicals play a vital role for cytotoxicity. Previous
study supports the ndings of this study, that citral inhib-
its P388 mouse leukemia cells[10] and oil from Cymbopogon
exuosus induced differential in vitro cytotoxicity in
12 human cancer cell lines and in vivo tumor growth
inhibiton in murine, Erlich and S-180 tumor models. The
oil also caused dose dependent increase in apoptosis in
HL-cells.[11] Mitochondrial activity of cancer cells may be
inuenced by lemongrass and citral, and this might be
the reason for the increased cytotoxicity observed in lem-
ongrass oil and citral treated cells. This hypothesis was
conrmed by evaluating the alteration in mitochondrial
membrane potential in treated groups. It was observed
that mitochondrial membrane potential was decreased in
lemongrass oil and citral and treated groups compared to
Figure 4. (A and B) Effect of lemongrass oil and citral on ROS in HeLa and ME-180 cells. The values are given as mean ± SD of
six experiments in each group (ANOVA followed DMRT). Bars not sharing the common superscripts differ signicantly at p < 0.05
(DMRT). Figure 4 (C and D) Fluorescence microscopic images showing changes in the levels of ROS generation in lemongrass oil
and citral treated ME-180 cells. Arrow mark (→) represents high DCF uorescence.
Anticancer effect of lemongrass oil and citral on cervical cancer cell lines
46
Figure 5. (A and B) Effect of mitochondrial membrane potential in lemongrass oil and citral treated HeLa and ME-180 cells
respectively. The values are given as mean ± SD of six experiments in each group (ANOVA followed DMRT). Bars not sharing
the common superscripts differ signicantly at p < 0.05 (DMRT). Figure 5 (C and D) Fluorescence microscopic images showing
the alteration of MMP by Rh-123 staining in lemongrass oil, citral and cisplatin treated cells. Arrow marks (→) represents dye
accumulation.
Figure 6. (A and B) Effect of apoptotic morphological changes in lemongrass oil and citral treated HeLa and ME-180 cells
respectively. The values are given as mean ± SD of six experiments in each group (ANOVA followed DMRT). Bars not sharing the
common superscripts differ signicantly at p < 0.05 (DMRT). Figure 6 (C and D): Fluorescence microscopic images showing the
apoptotic morphological changes by dual staining in lemongrass oil and citral treated HeLa and ME-180 cells respectively. Arrow
mark (→) represents orange-colored cells which are late apoptotic cells.
control. The results obtained in this experiment showed
that lemongrass oil had higher activity than citral for both
the cell lines. Further an increase in ROS production had
been observed in lemongrass oil and citral when com-
pared to control groups. ROS production is a mechanism
shared by all non-surgical therapeutic approaches for
cancers, including chemotherapy, radiotherapy and pho-
todynamic therapy, due to their implication in triggering
cell death. Therefore ROS are good tools to kill cancer
cells.[25] Several naturally occurring antioxidants have been
reported to be anti-mutagenic/anticarcinogenic in the lit-
erature such as, reservertrol.[26] It had been reported that
citral, in a dose dependent manner inhibited the oxida-
tive process involved in the formation of free radicals in
nickel chloride treated mouse.[16] Increased ROS levels
are thought to constitute an essential step in cell death
induction by many different cytotoxic drugs. In the pres-
ent work, it was found that lemongrass oil emulsion at a
Anticancer effect of lemongrass oil and citral on cervical cancer cell lines
47
concentration of 200 µg/ml in HeLa cells and ME-180
cells showed maximum generation of ROS (p > 0.05).
The previous experiment showed a decrease in mitochon-
drial potential in treated groups compared to control.
Mitochondrion is one of the most important organelles in
regulating cell death as well as a maker in apoptosis.[27] This
was proved by observing the apoptotic morphology of
treated and non treated groups using dual staining meth-
ods. The percentage apoptotic cell death was increased
in lemongrass oil and citral emulsion when compared to
control. HeLa cells showed 95% apoptotic cells in lemon-
grass oil emulsion and 80% in citral emulsion treatment
groups. ME-180 cells showed 98% apoptotic cells in lem-
ongrass oil emulsion and 90% in citral emulsion treatment
groups. Citral has been found to be a potent inducer of
glutathione-s-transferase class of enzymes, which provide
protection to healthy hepatocytes against apoptosis dur-
ing chemotherapy of liver cancers. A good superoxide
scavenging activity (EC50 = 19 mcg/ml) was reported
in Swiss albino mice in citral treated groups, suggesting
that the antioxidant action could be responsible for the
anti-clastogenic effect of citral against nickel chloride.[16]
Citral, at a concentration comparable to that found in a
cup of tea brewed with 1 gram of lemongrass, was found
to induce apoptosis in cancer cells, without any harm to
normal cells. Apoptosis was accompanied by DNA frag-
mentation and caspase-3 catalytic induction.[17] Another
study also reported that citral disrupts animal microtubules
and inhibits polymerization of microtubules in vitro.[18]
Citral was also tested on cyclo-oxygenase activity. Citral
treatment also caused inhibition of breast cancer MCF-7
cell growth (IC50 -48 h: 18 × 105 m) with a cell cycle arrest
in G2/M phase, apoptosis induction and also a decrease
in prostaglandin E 2 synthesis. These ndings suggested
that citral has a potential chemopreventive effect.[19] Thus,
citral seems to have the ability to turn on the apoptosis
process in cancer cells, causing them to die. It also disrupts
animal microtubules, acts as antioxidant in normal cells
and selectively behaves as pro-oxidant in cancer cells.[16,18]
These characteristics of citral make it a very potent candi-
date as anticancer agents.
The various constituents (%) present in the oil from lem-
ongrass variety of C. exuosus such as geraniol (20.08),
geranyl acetate (12.20), α-bisabolol (8.42) and isointer-
medeol (24.97) have been individually reported for their
cancer cell cytotoxicity.[8,6,29,30] Another report showed that
the essential oil from Cymbopogon exuosus (CFO) and its
major chemical constituent sesquiterpene isointermedeol
(ISO) induced apoptosis with in 48 h IC50 of ~30 and
20 µg/ml, respectively, in human leukemia HL-60 cells.[6]
The activities of essential oils have always been cred-
ited to its synergistic effects of its complex mixtures of
various compounds.[31] Synergistic anticancer effects have
also been reported for crude extracts of Polyalthia evecta,
which induced apoptosis in HepG2 cells.[32] Essential
oils are complex mixtures of chemical constituents, thus
more than one chemical constituent in lemongrass oil can
cause cancer cell death and is thus a very promising candi-
date for anti-cancer therapies. These ndings support that
there may be synergy between the citral and other compo-
nents with anticancer activity in lemongrass oil and also
explain the reason for it to show more potent anticancer
activity than citral alone.
Our results summarize that lemongrass oil and citral
emulsion decreased cell proliferation, increased intracel-
lular ROS, altered mitochondrial membrane potential,
and induced apoptosis in HeLa and ME-180 cell lines.
Hence, lemongrass oil and citral emulsion can be consid-
ered as potent anticancer agents and could be useful in
chemotherapy of cervical cancer in vitro. However, fur-
ther investigation warrants proving their in vitro anticancer
efcacy in in vivo models, and understanding the various
properties of lemongrass oil and citral emulsions for the
production of ointment based anticancer drugs, which
can be used in sit.
ACKNOWLEDGEMENT
None
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... e pharmacological properties of lemongrass were due to the presence of citral which is an acyclic monoterpene. Many studies reported that lemongrass oil possesses many pharmacological properties such as antimicrobial [10] and insecticidal [11]properties; only few studies demonstrated the anticancer properties of lemongrass, for instance, cervical cancer, HeLa and ME-180 cells [12], breast cancer (MCF-7) cells [13], prostate cancer, PC3, and LNCap [14]. However, to date its molecular mechanism in prostate cancer cells has not been elucidated. ...
... e therapeutic use of this oil was due to the presence of monoterpenes and myrcene. Few studies demonstrated the anticancer properties of citral in various cancer cell lines including breast (MDI-MB-231), small-cell lung cancer, colorectal cancer (HCT116 and HT29), and cervical cancer (HeLa and ME-180) [12,[29][30][31]. Although various studies in a preliminary level of citral have been studied against various cancer cell lines, the effect of cis-and trans-citral on aggressive prostate cancer (PC3) has not been elucidated so far. ...
Article
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The isomers of citral (cis-citral and trans-citral) were isolated from the Cymbopogon citratus (DC.) Stapf oil demonstrates many therapeutic properties including anticancer properties. However, the effects of citral on suppressing human prostate cancer and its underlying molecular mechanism have yet to be elucidated. The citral was isolated from lemongrass oil using various spectroscopic analyses, such as electron ionized mass spectrometry (EI-MS) and nuclear magnetic resonance (NMR) spectroscopy respectively. We carried out 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay to evaluate the cell viability of citral in prostate cancer cells (PC-3 and PC3M). Furthermore, to confirm that PC3 undergoes apoptosis by inhibiting lipogenesis, we used several detection methods including flow cytometry, DNA fragmentation, Hoechst staining, PI staining, oil staining, qPCR, and Western blotting. Citral impaired the clonogenic property of the cancer cells and altered the morphology of cancer cells. Molecular interaction studies and the PASS biological program predicted that citral isomers tend to interact with proteins involved in lipogenesis and the apoptosis pathway. Furthermore, citral suppressed lipogenesis of prostate cancer cells through the activation of AMPK phosphorylation and downregulation of fatty acid synthase (FASN), acetyl coA carboxylase (ACC), 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR), and sterol regulatory element-binding protein (SREBP1) and apoptosis of PC3 cells by upregulating BAX and downregulating Bcl-2 expression. In addition, in silico studies such as ADMET predicted that citral can be used as a safe potent drug for the treatment of prostate cancer. Our results indicate that citral may serve as a potential candidate against human prostate cancer and warrants in vivo studies.
... The EO constituents of the genus Cymbopogon are bioactive compounds with several tested biological activities such as antioxidant, antiinflammatory, antifungal, antibacterial, and anticancer properties with potential use in medicine [2]. Different types of cancer are among the most common leading causes of death worldwide [3,4]. Citral has shown anticancer activity in various human cancer cell lines, including cervical [4] and stomach [5] cancer cell lines. ...
... Different types of cancer are among the most common leading causes of death worldwide [3,4]. Citral has shown anticancer activity in various human cancer cell lines, including cervical [4] and stomach [5] cancer cell lines. At the same time, different studies have shown the activity of geraniol against lung, prostate, bowel, liver, colorectal, kidney, and skin cancer [6]. ...
Article
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Lemongrass (Cymbopogon citratus) essential oil (EO) is a major source of bioactive compounds (BC) with anticancer activity such as α-citral, limonene, geraniol, geranyl acetate, and β-caryophyllene. Comparative studies about cytokinin effects on BC profiles in lemongrass are missing. Here, we evaluated four cytokinins (2iP, tZ, BAP, and KIN) in two different osmotic media, MS-N (3% sucrose, 3 g L−1 Gelrite™) and MS-S (5% sucrose, 5 g L−1 Gelrite™). It results in a higher multiplication rate in BAP containing medium compared to tZ, KIN, and 2iP (p ≤ 0.05). While shoots grown on MS-N/BAP, tZ, and KIN exhibited a highly branching morphology, MS-N/2iP produced a less branching architecture. BC profile analysis of established plants in pots revealed that their maxima production depends on the in vitro shoot growth conditions: i.e., highest content (80%) of α-citral in plants that were cultured in MS-S/BAP (p ≤ 0.05), limonene (41%) in MS-N/2iP, or geranyl acetate (25.79%) in MS-S/2iP. These results indicate that it is possible to increase or address the production of BC in lemongrass by manipulating the cytokinin type and osmotic pressure in culture media. The culture protocol described here is currently successfully applied for somatic embryogenesis induction and genetic transformation in lemongrass.
... Lemongrass is a wealth of Indonesian spices contains a substantial group of flavonoids, essential oils, and phenolic compounds. The active compounds in lemongrass possess pharmacological activities such as anti-bacterial [3], anti-fungal [2], antioxidants [4], anti-cancer [5], anti-lipidemic [6], antidandruff [7], anti-inflammatory properties [2] [4] enhancing human health. ...
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Lemongrass is a wealth of Indonesian spices that have many functional benefits. The purpose of the study was to extract lemongrass essential oil and determine the difference of the type of material, extraction method, and plant parts on the phytochemical characteristics of the extract produced. Lemongrass is extracted in fresh and dried form. The selected extraction method is sonication and maceration with parts of stalks and leaves as the material. The plant material was extracted using 96% ethanol solvent. The phytochemical characteristics analyzed were antioxidant capacity, total polyphenols, and flavonoids. The results showed the highest antioxidant capacity and total phenol obtained from the fresh leaves extracted by maceration. Plant part and type of material significantly affect antioxidant capacity, total polyphenols, and total flavonoids in lemongrass. The antioxidant capacity of lemongrass is positively correlated with total polyphenols
... Dudai et al. (2005) have studied and justified the inhibiting effect of citral, a constituent of lemongrass, on growth of hepatic cancer cells during initial stages. Citral has also shown antiproliferative effect in delaying the growth and induction of apoptosis of cancer cells responsible for breast cancer (Ghosh 2013;Philion et al. 2017). Lemongrass has been found to possess anti-hypercholesterolemic and antihyperlipidemic properties (Kumar et al. 2011;Lee et al. 2018). ...
Chapter
Secondary metabolites (SMs) are known to have a wide range of therapeutic values. Large numbers of drugs are derived from these SMs. These naturally occurring SMs known to act as a potent source of antimicrobial, antiviral, anti-inflammatory, anticancer, and insecticidal agents. Aromatic plants are the prime source of variety of easily available SMs. Numerous classes of these SMs also act as powerful natural antioxidants. Antioxidants are the compounds that inhibit or slow down the oxidation of other molecules and help to cure the oxidative stress condition. Oxidative stress is the condition where the amount of free radicals in the body of organism exceeds the homeostatic balance of free radicals and indigenous antioxidant. This excess of free redials leads to various types of chain reactions that damage cells. These free radicals are the cause of more than hundred kinds of diseases in living beings. Cymbopogon is a genus of about 180 species of monocots grasses in a family of Poaceae (Gramineae). The species of genus Cymbopogon are rich source of naturally occurring antioxidants (such as phenolic acids, flavonoids, tannins, hydroquinone, terpenoids and fatty alcohols, etc.), and lemongrass (Cymbopogon citrates) is one of them. Further, the pharmacological applications of lemongrass are also well explored. Hence in the present chapter, we intend to discuss the botanical description, traditional uses, phytochemistry, antioxidant potential, health benefits, and potential economic importance of lemongrass.
... We have shown that citral significantly increased ROS levels in both RD and RH30 cells (Figure 3c). This has also been shown previously in both HeLa and ME180 cells (Ghosh, 2013). Co-incubation of 1,000 μM citral with metformin showed quenching of ROS in both cell lines for all concentrations of metformin. ...
Article
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Rhabdomyosarcoma (RMS) is a rare type of soft tissue sarcoma most commonly found in pediatric patients. Despite progress, new and improved drug regimens are needed to increase survival rates. Citral, a natural product plant oil can induce cell death in cancer cells. Another compound, metformin, isolated originally from French lilac and used by diabetics, has been shown to reduce the incidence of cancer in these patients. Application of citral to RMS cells showed increase in cell death, and RD and RH30 cells showed half maximal inhibitory concentration (IC 50) values as low as 36.28 μM and 62.37 μM, respectively. It was also shown that the citral initiated cell apoptosis through an increase in reactive oxygen species (ROS) and free calcium. In comparison, metformin only showed moderate cell death in RMS cell lines at a very high concentration (1,000 μM). Combinatorial experiments, however, indicated that citral and metformin worked antagonistically when used together. In particular, the ability of metformin to quench the ROS induced by citral could lead to the suppression of activity. These results clearly indicate that while clinical use of citral is a promising anti-tumor therapy, caution should be exercised in patients using metfor-min for diabetes.
... • Anticancer properties: Presence of bioactive citral in Lemongrass helps ight cancer either by apoptosis or boosting immune system [12]. ...
... Fragrant lemongrass is a source of important essential oils that has several benefits. It had activity as antibacterial [1,2], as antidiabetic [3], and anticancer agents [4]. It's leaves also widely used as tea herbal [5]. ...
Article
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The citronella oil refining industry contributes greatly to improving the community’s economy. In the supply chain of citronella oil, the community usually acts as a supplier (fragrant lemongrass farmers), manufacturers (distillers of citronella oil), and distributors (collectors of citronella oil). The main problem in the citronella oil refining industry is the uncertainty of the supply chain. Fragrant lemongrass supply chain mapping is done to determine the capacity and needs of all actors involved from raw material sources to retail consumers. Thus, the continuity of fragrant lemongrass supplies increased production capacity in refineries, and market potential can be estimated more precisely. On the other hand, integration between industries is needed to prevent the impact of environmental damage while increasing profits. The symbiosis model of the citronella refining industry is expected to optimize the potential and existing resources through an industrial system that is efficient, integrated, easy to implement, and environmentally sound. This paper will discuss how the application of supply chain management concepts in the symbiosis of the citronella refining industry so that the objectives of cooperation in the environmentally sound supply chain can be realized.
... It suppresses the NF-kappa B activation in RAW264.7, alters potential of mitochondrial membrane inducing apoptosis in ME-180 and cervical cancer cell lines, decreases proliferation of cell and decreased production of nitrous oxide [93,94]. However, it is unstable in acidic environments [95]. ...
Article
In recent years, SLNs and NLCs are among the popular drug delivery systems studied for delivery of lipophilic drugs. Both systems have demonstrated several beneficial properties as an ideal drug-carrier, optimal drug-loading and good long-term stability. NLCs are getting popular due to their stability advantages and possibility to load various oil components either as an active or as a matrix. This review screens types of oils used till date in combination with solid lipid to form NLCs. These oils are broadly classified in two categories: Natural oils and Essential oils. NLCs offer range advantages in drug delivery due to the formation of imperfect matrix owing to the presence of oil. The type and percentage of oil used determines optimal drug loading and stability. Literature shows that variety of oils is used in NLCs mainly as matrix, which is from natural origin, triglycerides class. On the other hand, essential oils not only serve as a matrix but as an active. In short, oil is the key ingredient in formation of NLCs, hence needs to be selected wisely as per the performance criteria expected.
... It had activity as antibacterial to Salmonella choleraesuis, Pseudomonas aeruginosa, Staphylococcus aureus [3], and Acinetobacter baumannii [4]. It also gave activity as antidiabetic [5], atherosclerosis [6], anticancer agents [7]. It's leaves also widely used as tea herbal [8]. ...
Article
Cancer is considered a multifactorial disease and its development could be associated with several factors, for example, rotenone exposition. Unfortunately, many cancers are resistant to chemotherapy, as cervical cancer. Regarding this, lemongrass is a remarkable natural product that presents antioxidant and anticancer activities, which could show therapeutic action against rotenone and cervical cancer. Thus, this study aimed to investigate the antioxidant and anticancer action of lemongrass. An in vitro study was conducted using VERO (kidney cells) and SiHa cell lines (cervical cancer cells). VERO cells were exposed to rotenone and lemongrass extract for 24 and 72 h. While SiHa cells were exposed to lemongrass isolated and associated to chemotherapy, 5-fluorouracil, during 24 and 48 h. After, levels of viability, proliferation, and oxidative metabolism were determined. The results showed that lemongrass presents antioxidant activity on VERO cells by increasing cell viability and proliferation and decreasing oxidative stress caused by rotenone. Moreover, lemongrass showed anticancer activity by decreasing cell viability and increasing oxidative stress parameters on SiHa. Besides, lemongrass had no alteration in the chemotherapy activity. Therefore, this study revealed that lemongrass presents antioxidant and anticancer activity since it can protect against the cytotoxicity of rotenone and reduce the cell viability of cervical cancer.
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Monoterpenes have been identifi ed as responsible of important therapeutic effects of plant-extracts. In this work, we try to compare the cytotoxic effect of six monoterpenes (carvacrol, thymol, carveol, carvone, eugenol and isopulegol) as well as their molecular mechanisms. The in vitro antitumor activity of the tested products, evaluated against fi ve tumor cell lines, show that the carvacrol is the most cytotoxic monoterpene. The investigation of an eventual synergistic effect of the six natural monoterpenes with two anticancer drugs revealed that there is a signifi cant synergy between them (p<5%). On the other hand, the effect of the tested products on cell cycle progression was examined by fl ow cytometry after DNA staining in order to investigate the molecular mechanism of their cytotoxic activity. The results revealed that carvacrol and carveol stopped the cell cycle progression in S phase; however, thymol and isopulegol stopped it in G0/G1 phase. Regarding carvone and eugenol, no effect on cell cycle was observed.
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Objective and Design: To evaluate potential anti-inflammatory properties of tea tree oil, the essential oil steam distilled from the Australian native plant, Melaleuca alternifolia.¶Material and Methods: The ability of tea tree oil to reduce the production in vitro of tumour necrosis factor-α (TNFα), interleukin (IL)-1β, IL-8, IL-10 and prostaglandin E2 (PGE2) by lipopolysaccharide (LPS)-activated human peripheral blood monocytes was examined.¶Results: Tea tree oil emulsified by sonication in a glass tube into culture medium containing 10% fetal calf serum (FCS) was toxic for monocytes at a concentration of 0.016% v/v. However, the water soluble components of tea tree oil at concentrations equivalent to 0.125% significantly suppressed LPS-induced production of TNFα, IL-1β and IL-10 (by approximately 50%) and PGE2 (by approximately 30%) after 40 h. Gas chromatography/ mass spectrometry identified terpinen-4-ol (42%), α-terpineol (3%) and 1,8-cineole (2%, respectively, of tea tree oil) as the water soluble components of tea tree oil. When these components were examined individually, only terpinen-4-ol suppressed the production after 40 h of TNFα, IL-1β, IL-8, IL-10 and PGE2 by LPS-activated monocytes. Conclusion: The water-soluble components of tea tree oil can suppress pro-inflammatory mediator production by activated human monocytes.
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The essential oils isolated from leaves of Cymbopogon citrates and Ocimum gratissimum have been tested for their cytotoxic activity against P 388 leukemia cells. The IC 50 of the Cymbopogon oil was found to be 5.7 μg/ml while that of Ocimum oil was 10.8 μg/ml. The mixture of the oils (1:1 v/v) showed an IC 50 value of 10.2 μg/ml and there was no synergism in the cytotoxic activity. The oils were standardized by their physico- chemical properties.
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Using Carpoglyphus lactis, body-effect of mites on citral composition was determined by incubating a known amount of a substrate (neral or geranial) in hexane at room temperature with the hexane-washed bodies of mites (0.5 g). Half of the substrate (100 ppm, each) was consumed within 30 min and the products of the incubation consisted of four compounds : isomerized citral to an equilibrium state [neral (40%) and geranial (60%), total 26-43 ppm] and reduction products (total 27-59 ppm) of citral, which were identified as nerol and geraniol. The isomerization rate was affected by the incubation solvent and was observed in the following order: ether>hexane>benzene>acetone>ethanol. Reduction products were detected in the ether benzene- and hexane-incubation. Treatment of washed mites with formalin and trichloroacetic acid retarded the isomerization reaction. Treatment of washed mites with tap water did not affect the isomerization rate, but inhibited the reduction of citral. Both reactions were commonly observed in all species of acarid mites tested. © 1983, JAPANESE SOCIETY OF APPLIED ENTOMOLOGY AND ZOOLOGY. All rights reserved.
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The purpose of this study was to examine the inhibitory effect of essential oils against a broad spectrum of microorganisms including bacteria, yeast, molds, and two bacteriophage. The inhibitory effects of 45 oils on eight bacteria (four Gram positive and four Gram negative), two fungi, and one yeast were examined using the disk assay method. Phage inhibition was measured by mixing the oils with a phage suspension, incubating the mixture at 4°C for 24 h, then plating on a lawn of indicator bacteria and assaying for plaque production. Of the oils tested, all oils exhibited inhibition over activity relative to controls. However, a number exhibited only weak inhibition against several gram positive bacteria. Gram negative bacteria were generally more resistant than Gram positive bacteria to oil treatment with Pseudomonas aeruginosa being the most resistant bacteria. Only cinnamon bark (Cinnamomum zeylanicum) and tea tree (Melaleuca alternifolia) oils showed an inhibitory effect against all the test organisms and phage. Coriander oil (Coriandrum sativum) highly inhibited Gram positive bacteria and fungi. Lemongrass (Cymbopogon flexuosus) and Roman chamomile (Chamaemelum nobile) oils showed a high degree of inhibition against both phage types, while 8 oils showed no inhibition against either phage. Angelica (Angelica archangelicd) and pine (Pinus sylvestris) oils inhibited the bacteria, but had no effect on any fungi. Oils that exhibited high antimicrobial properties and the broadest range of inhibition included cinnamon bark (Cinnamomum zeylanicum), lemongrass (Cymbopogon flexuosus), savory (Satureja montana), Roman chamomile (Cbamaemelum nobile), rosewood (Aniba rosaeodora), spearmint (Mentha spicata) and tea tree (Melaleuca alternifolia).
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An update to the American Cancer Society (ACS) guideline regarding screening for the early detection of cervical neoplasia and cancer, based on recommendations from a formal review and recent workshop, is presented. The new screening recommendations address when to begin screening, when screening may be discontinued, whether to screen women who have had a hysterectomy, appropriate screening intervals, and new screening technologies, including liquid-based cytology and HPV DNA testing.
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The present study investigated the chemopreventive potential of geraniol, an acyclic monoterpene alcohol, by monitoring the tumor incidence and analyzing the status of phase II detoxification agents, lipid peroxidation by products and antioxidants in 7,12-dimethylbenz(a)anthracene (DMBA) induced mouse skin carcinogenesis. Skin tumor was developed by painting DMBA (25 microg in 0.1 ml acetone mouse(-1)) in the shaved back of the mice, twice weekly for 8 weeks. We noticed 100% skin tumor formation in mice treated with DMBA alone. The status of phase II detoxification agents and antioxidants were decreased where as lipid peroxidation by products were increased in tumor bearing mice. Oral administration of geraniol at a dose of 250 mg kg(-1) body weight significantly prevented the tumor formation as well as brought back the status of phase II detoxification agents, lipid peroxidation by products and antioxidants to near normal range in DMBAtreated mice. Present results suggest that geraniol might have inhibited abnormal cell proliferation occurring in skin carcinogenesis by modulating the activities of phase II detoxification agents and through its free radical scavenging potential.