This document summarizes a research study that investigated the effect of Fimbristylis ovata, a plant in the sedge family, on vero and MCF-7 cell lines. The study found that a crude methanol extract of F. ovata was able to inhibit the MCF-7 breast cancer cell line with an IC50 value of 73 and the vero cell line with an IC50 value of 280 based on MTT assays. Previous studies had found that F. ovata contains compounds like phenols, saponins, and flavonoids that are known to have anti-cancer effects. The aim of this study was to determine if F. ovata extracts had cytotoxic or anti-cancer potential against these

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Journal of Chemical and Pharmaceutical Research, 2015, 7(9):284-290
Research Article
ISSN : 0975-7384
CODEN(USA) : JCPRC5
284
Effect of Fimbristylis ovata (Burm. F) J. Kern on vero cell and
MCF-7 cell morphology
Sowmya Thomas and E. Joyce Sudandara Priya*
Department of Botany, Madras Christian College (Autonomous), Tambaram East, Chennai, Tamil Nadu, India
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ABSTRACT
Breast cancer is a complex genetic disease that is caused in about 80% of the women due to various environmental
factors. The allopathic treatments which cause side effects are not appreciated in the recent years. Hence there has
been a long-standing interest to find anticancer agents present in plants to eliminate side effects. Fimbristylis ovata
(Burm.f.)Kern which is being used traditionally for the treatment of wounds and other associated diseases which
could lead to malignancy. Since there has been no scientific data on the effect of Fimbristylis ovata on Vero cell line
and MCF-7 cell line the concept of investigating its cytotoxic and anticancer potential was brought up. The tests
were evaluated using MTT assay and it was noted that the crude methanol extract of Fimbristylis ovata could inhibit
MCF-7 cell line at an IC50 value of 73 and Vero cell line at an IC50 value of 280.
Key words: Fimbristylis ovata, Vero Cell line, MCF-7 cell line, anticancer, methanolic extract.
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INTRODUCTION
Cancer is a potentially fatal disease caused by environmental factors that mutate gene encoding critical cell –
regulatory proteins. The resultant aberrant cell behavior leads to expansive masses of abnormal cells that destroy
surrounding normal tissues and can spread to the vital organs resulting in disseminated disease, commonly a
harbinger of imminent patient death [1]. Cancer begins with mutations in DNA, which instructs the cells, how to
grow and divide. Normal cells have the ability to repair most of the mutations in their DNA, but the mutations which
is not repaired and causing the cell to grow becomes cancerous [2]
Breast cancer is the most common cancer and the most frequent cause of death in women [3]. Breast cancer is the
second most common cancer among the women in India, after cancer of the cervix uteri. Locally advanced breast
cancer constitutes more than 50% to 70% of patients presenting for treatment [4].
Breast cancer is caused by repeated exposure of breast cells to circulating ovarian hormones [5]. Clinical, animal
and epidemiological studies have clearly demonstrated that breast cancer is a hormonally mediated disease and
several factors that influence hormonal status or are markers of change in hormonal status have been shown to be
associated with the risk of breast cancer [6]; [7];[8] A variety of constitutional risk factors have been reported such
as multipolarity, early onset of menarche, delayed first birth, late menopause and decreased parity. These risk factors
point toward endogenous estrogens as likely players in the initiation, progression and promotion of breast cancer [9].
Chemotherapy, being a major treatment modality used for the control of advanced stages of malignancies and as a
prophylactic against possible metastasis, exhibits severe toxicity on normal tissues [10]; [11]. Today despite

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considerable efforts, cancer still remains an aggressive killer worldwide. Moreover, during the last decade, novel
synthetic chemotherapeutic agents currently in use clinically have not succeeded in fulfilling expectations despite
the considerable cost of their development. Therefore there is a constant demand to develop new effective and
affordable anticancer drugs [12]. The search is goal – oriented. The goal is to find new active molecules in plants,
with the ability to stop and prevent growth of cancer cells.[13].
Natural products (specifically plants) have received increasing attentions over the past years for their potential as
novel cancer preventive and therapeutic agents [14][15]. The plant derived compounds are inhibitor of various
stages of tumourogenisis and associated inflammatory processes, underlying the importance of these products in
cancer prevention and therapy [12]. It is well established that there is a long history of the use of different medicinal
plants and natural products as anticancer agents in many parts of the world. A variety of medicinal plants have been
useful sources of clinically relevant antitumor compounds [16] In addition, it is reported that 50% of drugs in
clinical trial for anticancer activity were from natural sources or are related to them [17]. Thus, much research
accomplished today focuses on the investigation of antitumor activity of medicinal plants and the development of
new drugs to treat cancers as well as other incurable diseases [18]; [19]; [20]. Although plants have the potential to
synthesize extremely complex molecular structure extensive screening procedures are required to identify, isolate
and purify these compounds. Examples of plant-derived compounds with anticancer activity used clinically include:
the Vincaalkaloids (which induce metaphase arrest by interfering with microtubule assembly) vinblastine, vincristine
and vincamine derived from Vincarosea, and their synthetic derivatesvindesine and vinorelbine; the topoisomerase
inhibitors campothecin, an alkaloid isolated from Campothecaaccuminata, and etoposide, a semi-synthetic
derivative of podophyllotoxin, an antineoplastic glucoside obtained from Podophyllumpeltatum; the taxoids
paclitaxel (taxol) and the synthetic analogue docetaxel (taxotere), which stabilize microtubules and disrupt mitosis.
The various compounds described above are associated primarily with the treatment of breast cancer. [13]
Many natural dietary agents, including vegetables, fruits, herbs, and spices have been used in traditional medicines,
as non-conventional treatments, for thousands of years, but without sufficient proofs. Effectiveness of such natural
dietary agents might lead to the development of natural and novel drugs with low or no side effects.
The genus FimbristylisVahl., belongs to the family Cyperaceae of Monocotyledons. There are about 250 genera
commonly grown in warm climatic condition. Most of the species belonging to this genus are grown as weeds. The
plant is found distributed in Asian continents, Africa, Australia and in other tropics and subtropics and warm
temperate regions. It has a wide range of medicinal properties.
The species Fimbristylis ovata (Burm.f.)Kern.is a perennial sedge, widely distributed in tropics and subtropics and
low lying grasslands. It is often found in marshes and in seasonally inundated areas at the edges of pools, lake sides
and ditches. [21]. Fimbristylis ovata (Burm.f.) Kern., known by its common name flat spike sedge or “Ya-sae-ma”
in Thai, belongs to the family Cyperaceae[22]
Fimbristylis ovata has been long used for treating adenitis, scrofula, syphilis, cough, bronchitis, and asthma [23];
[24]Fimbristylis ovata has antioxidant activity and inhibits RAGE gene expression in a human lung adenocarcinoma
epithelial cell line. [25]
The methanolic extracts of Fimbristylisovata leaves revealed the presence of tannins, saponins, flavonoids,
anthocyanins, β-cyanins, quinines, cardiac glycosides, phenols and coumarins. [26]. But the underlying mechanism
of its therapeutic activity is unknown. Among the biochemical compounds that are mentioned above few of the
compounds such as phenols, saponins, coumarins and flavonoids (Silymarin) are effective against human breast
cancer. The aim of this study was to investigate the effects of Fimbristylis ovata extracts on Human Breast Cancer
(MCF-7) cell lines.
Plants in the family Cyperaceae consist of several components such as phenolic compounds, flavonoids, alkaloid,
glycoside proteins, amino acids, tannins, and saponins.[27]
As presented above, some researches have shown that certain constituents of the Fimbristylis ovata and its ethanolic
extracts of the plant are effective against RAGE genes [25]. But to our knowledge, no studies exist on the effects of
the methanolic extract of Fimbristylis ovata against the proliferation of breast cancer cells. Therefore, the potential

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of Fimbristylis ovata to induce death of human breast cancer cells has been investigated here in the hope of finding a
natural treatment.
EXPERIMENTAL SECTION
Cell lines:-
The human breast cancer cell line MCF-7 (Michigan Cancer Foundation-7) (King Institute, Chennai), is an epithelial
invasive breast ducal carcinoma cell line, which is estrogen and progesterone receptor positive. Vero cell lines
which were acquired from the kidney epithelial cells extracted from African Green Monkey.
Both MCF-7 and Vero cell lines were cultivated in Dulbecco’s Minimal Essential Medium (DMEM) supplemented
with 10% fetal calf serum (FCS) and stored in the incubator provided with 5% CO2 atmosphere at 37ºC, and 96%
relative humidity.
Determination Of Cell’s Morphology Using Inverted Light Microscopy:-
The Vero and MCF-7 cell lines were suspended in DMEM containing 10% FBS and kept for incubation and
subcultured several times. The degree of subculturing a cell line has undergone, is often expressed as “passage
number”, which can generally be thought of as the number of times cells have been transferred from vessel to vessel.
Cell lines at high passage numbers experience alteration in morphology, response to stimuli, growth rates, and
protein expression and transfection efficiency, compared to lower passage cells. After several passage the cell lines
ready for the test is stored in the incubator provided with 5% CO2 atmosphere at 37ºC, and 96% relative humidity.
The growth of the cells was monitored every 24 hours for 3 days under inverted microscope. The medium was then
discarded and the cells were split and transferred into new medium. To determine the effects of Fimbristylis ovata
whole plant crude methanol extract on the morphology of the cells, both MCF-7 and Vero cell lines
The whole plant ofFimbristylis ovata was obtained locally and they were cleaned, shade dried and powdered using
an electric grinder. The extract was prepared viasoxhlet apparatus. For the cell line studies 10 mg of Fimbristylis
ovata crude methanol extract was weighed and dissolved in 25 µl of DMSO without FCS in MEM media. Several
dilutions of the extract in serum-free medium were prepared [extract:SFM (vol/vol); (5µl of extract in 1000µl of
SFM ), 10µ l, 25µ l, 50µl, 75µl, 100µl, 200µl, 300µl, 400 µl, 500 µl and 600 µl for Vero cell lines and till 300 µl for
MCF-7 and were stored at -80ºC.
Effect Of Methanol Extract Of The Plant On The Cell Lines:-
The 96 well plates seeded with Vero &MCF-7 cell lines were taken out of the incubator and brought to the laminar
air flow. The medium in the 96 well plates were discarded. The cells adherent to the surface was left undisturbed.
These cells were treated with extract stock solution with each concentration of the extract in each well. The plates
were then incubated in 5% CO2 atmosphere, at 37ºC, and 96% relative humidity. The treated cell lines were
monitored for 3 days at an interval of 24 hours.
MTT assay:-
After 72 hours, the medium was aspirated carefully and discarded. 50 µl of MTT solution added was to each well.
The plate was incubated for 4 hours at 37ºC in an incubator with 5% CO2 to allow intracellular reduction of the
soluble yellow MTT to the insoluble purple formazan crystals. The supernatant was removed, 50 µl of propanol was
added, and the plates were gently shaken to solubilize the formed formazan. The suspension was transferred to a
spectrophotometer cuvette and absorbance values were read at 570 nm using DMSO as blank.
RESULTS AND DICUSSION
Fimbristylis ovata belonging to Cyperaceae used by the African tribe for wound healing and other ailments were
selected to study its activity against breast cancer. The crude extract of this plant was selected to test the anticancer
activity based on the chemical constituents such as tannins, saponins, flavonoids, cardiac glycosides, phenols and
coumarins which have been reported in Fimbristylisovata (Sowmya Thomas and ShyamaMukundan, 2012). A crude
methanol extract was prepared by Soxhlet extraction method and its potential to treat cancer was assessed by
performing an MTT assay on breast cancer cell lines MCF-7. This was done by monitoring the viability of the
human breast cancer cell line treated with the methanol crude extract of the plant for every 24 hours for 3 days. To
prove it better a MTT assay was tested on Vero cell lines which are normal cell lines obtained from kidney epithelial

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cells of African green monkey. Both the cell lines were stored, maintained and passaged frequently in King Institute,
Chennai, India.
GRAPH - 1
..
GRAPH - 2
..
The result of the cytotoxicity and antiproliferative activity of the methanolic extract are photographed and scanned
in the Figure 1 and 2 respectively. The plant extract (10mg/ml) started to lyse the Vero cell lines at higher with the
IC50 value 280 whereas the extract inhibited the proliferation of MCF-7 at an IC50 value of 73. The efficacy of cell
0
20
40
60
80
100
120
5 10 25 50 75 100 200 300 400 500 600
%ofToxicityofcrudeextract
Concentration of crude extract
CYTOTOXIC ACTIVITY OF THE METHANOL CRUDE EXTRACT OF
Fimbristylis ovata AGAINST VERO CELL LINES
% OF TOXICITY
0
20
40
60
80
100
120
5 10 25 50 75 100 200
%ofthetoxicitycrudeextract
Concentration of the crude extract
CYTOTOXIC ACTIVITY OF THE METHANOL CRUDE EXTRACT OF Fimbristylis ovata AGAINST
MCF-7 CELL LINES
% OF TOXICITY

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death varied depending upon the concentration of the extract which was serially diluted as 5, 10, 25, 50, 75, 100,
200, 300 µl/ml. The IC50 value which was obtained by plotting a graph indicated the apt concentration to inhibit the
proliferation which is depicted in graph 1 and 2 respectively.
FIGURE - 1
FIGURE - 2

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The MCF-7 cell treated with DMSO retains its normal angular or polygonal shape. Most cells have intact and large
vesicular nuclei with prominent nucleoli. The treated MCF-7 cells with 50% extract showed a significant loss of cell
processes and marked changes in morphology such as shrinkage and irregular shape.
Since, no studies exist on the possible mechanism of the therapeutic effect of Fimbristylisovata the potential of the
methanolic extract of the whole plant was investigated to find a natural remedy against breast cancer. Previous
studies have cited the anti-cancer property of phytochemicals such as tannins, saponins, flavonoids, cardiac
glycosides, phenols and coumarins which have been reported inFimbristylisovata (Sowmya Thomas and
ShyamaMukundan, 2012). Therefore, the anti-cancer activity observed in crude methanol extract of this plant
Fimbristylis ovata may be due to phytochemical constituents or due to any single chemical compound. According to
the results obtained, Fimbristylis ovata appeared to be a potent anti-cancer agent and to the best of our knowledge
this is the first report on anti-cancer activity of Fimbristylis ovata against MCF-7 cell line.
CONCLUSION
The IC50 value of the crude extract on MCF-7 cell lines was lower compared to the IC50 value of the crude extract
treated on Vero cell lines. This indicates that the crude methanol extract is more effective on human breast cancer
cell line than on the normal cell lines. With these results it is ascertained that a major bioactive compound present in
the crude extract has a high anti proliferative activity which can be isolated by one of the recent chromatographic
techniques.
Acknowledgement
The authors are thankful to Mr. Ferroz Ahmed, King Institute, Porur, Chennai for providing infrastructural facilities,
support and guidance throughout this work.
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