International Journal of Pharma Sciences

Vol. 3, No. 2 (2013): 185-188

Research Article
Open Access
ISSN: 2320-6810

In vitro anti-cancer activities of few plant extracts

against MCF-7 and HT-29 cell lines
VSPK Sankara Aditya J*1,2, Naresh Kumar L1,2 and Animisha Mokkapati1,2
1
Translational Research Institute of Molecular Sciences, Pithapuram Colony, Maddilapalem, Visakhapatnam, India
2
Department of Biotechnology, GITAM University, Visakhapatnam, India

* Corresponding author: Sankara Aditya J; e-mail: krishnabios369@gmail.com

Received: 15 February 2013 Accepted: 01 March 2013 Online: 05 March 2013

ABSTRACT
Since many years, plants were known to possess anticancer activities against different cancer cell lines. In this
paper, we report a study based on anticancer properties of three plants Rubia cordifolia, Plumbago zeylanica,
Calophyllum inophyllum respectively. The leaves of plant material were collected, shade dried and extracted with
methanol using soxhlet extraction procedure. Anticancer activities are assayed with standard MTT colorimetric
procedure against MCF-7 and HT-29 cell lines. From the analysis it was found that Rubia cordifolia and Plumbago
zeylanica showed nearly 50 % MCF-7 cell line inhibition at 200µg/ml tested dose, where as three plant species
did not display much anticancer activities against HT-29 colon cancer cell line. Further work is in progress to
evaluate the chemical constituents present in these plants.

Keywords: Rubia cordifolia, Plumbago zeylanica, Calophyllum inophyllum, MTT assay, MCF-7, HT-29

INTRODUCTION cordifolia is reported to be active against a diversified

Plants have been used as remedies and botanical
literature has described the usage of plant extracts.
Cancer is a dreadful disease and combating this disease
is of great importance to public health. There is a
necessity for search of new compounds with cytotoxic
activity as the treatment of cancer with the available
anticancer drugs is often unsatisfactory due to the
problem cytotoxicity to the normal cells. Phytochemical
examination has been making rapid progress and
herbal products are becoming popular as sources of
plausible anticancer compounds [1].
Rubia cordifolia is a flowering plant with red
rhizomatous base and roots, a member of Rubiaceae
family, distributed in hilly tracts of India [2]. It has been
reported in literature that the powdered dried roots
and fruits have been tested for the treatment of skin
diseases and disorders of spleen and its preparations
are prescribed for treatment of major burns, bone
fractures and dysentery and it is considered as tonic,
antitussive and useful in chronic low fevers [3]. This
plant has many pharmacological properties like blood
purifier activity, anticancer, astringent, antidysentric,
antiseptic, antirheumatic, hepatoprotective etc. [4-7]. R.
panel of cancer cell lines, such as P388, L1210, L5178Y,
B16 melanoma, Lewis lung carcinoma, and sarcoma-
180[8]. The roots of Rubia cordifolia L showed strong
cytotoxicity against HT-29 and MCF-7 cell lines, and
DNA topoisomerase I and II inhibitory activities [9].
The plant species Plumbago zeylanica popularly known
as Chitraka, belongs to the family Plumbaginaceae , is
distributed throughout the world. Plumbago species
have been shown to contain compounds with
significant activity and its extract has antiplasmodial,
antihyperglycemic and antiallergic properties [10-12].
Ethanolic extract of Plumbago zeylanica possessed
significant anticancer activity and also reduce
peroxidation levels due to higher content of terpenoids
and flavonoids. [13]. It also stimulates the central
nervous system and is cytotoxic for tumor cell activities
[14]. The root is used as laxative, expectorant,
astringent, abortifacient, and in dysentery [15-17].
Roots of Plumbago zeylanica exhibited activity against
Bacillus subtilis, Escherichia coli, Proteus vulgaris,
Salmonella typhimurium, Pseudomonas aeruginosa and
Staphyloccocus aureus[18]. Plumbagin has been shown
to exert anticarcinogenic, antiatherosclerotic, and
antimicrobial effects. Their components possess

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antiatherogenic, cardiotonic, hepatoprotective, and MTT assay

neuroprotective properties [19-22]. Plumbagin has
exhibited anticancer and antiproliferation properties in
a variety of cell lines and examined its effect on cell
cycle distribution and autophagic cell death in two
human breast cancer cell lines, MDA-MB-231 and MCF-
7. Anti-proliferative and apoptotic activity of plumbagin
were examined in vitro using two human colonic cancer
cell lines, HT29 and HCT15. The mechanism of
anticancer activity of plumbagin was established by
analysing cell cycle regulation and signalling molecules
related to cell survival and apoptosis [23].
Calophyllum consists of 130 species distributed in
tropical areas of the world, belongs to a family
Clusiaceae. The tree is believed to have diuretic
properties. The gum that is extracted from the plant is
emetic and purgative and is used in the treatment of
wounds and ulcers [24]. The plant has anti-
inflammatory and pain relieving properties. An infusion
of gum, bark and leaves is used for sore eyes. Recent
studies show that it possesses anti-HIV activity [24-25].
Some of the uses of this plant indicate that there must
be some antibacterial and analgesic properties lying
with the plant [26].
Taking the above facts into consideration as well as the
properties exhibited by these plants, an attempt has
been made, in this paper, to study the possible anti-
cancer activities of the three plant species (Rubia
Cordifolia, Plumbago Zeylenica, Calophyllum
Inophyllum) against MCF-7(Breast cancer) and HT-
29(colon cancer) cell lines.
Sensitivity of MCF-7 and HT-29 cells to Rubia cordifolia,
Plumbago zeylenica, Calophyllum inophyllum was
determined individually by the MTT colorimetric assay.
Cells were seeded in a flat-bottomed 96-well plate and
incubated for 24 h at 37°C and in 5% CO2. Both cell
lines were exposed to all three plant species mentioned
above. The solvent DMSO treated cells served as
control. Cells were then treated with MTT reagent (20
µl/well) for 4 h at 37°C and then DMSO (200 µl) was
added to each well to dissolve the formazan crystals.
The optical density (OD) was recorded at 492 nm in a
microplate reader. Percentage of residual cell viability
was determined as [1−(OD of treated cells/OD of
control cells)]×100.
RESULTS AND DISCUSSION
Screening of methanolic extracts of Rubia cordifolia,
Plumbago zeylanica, Calophyllum inophyllum resulted in
moderate anticancer activities against MCF-7 and HT-
29 cell lines. The inhibitory properties of these extracts
are compared with standard tamoxifen for MCF-7 cell
line and 5-Fluoro Uracil for HT-29 cell line (Tables 1
and 2) respectively. The Percentage cancer cell
inhibition profiles were found to be concentration
dependent (Figures 1 and 2). The maximum
concentration (µg/ml) used in the study was 200
µg/ml. Following results were obtained when
anticancer activities of three plants were studied.
Table 1. Percent cell inhibition of plant extracts on MCF-7 cell
line

Conc. % cell inhibition

(µg/ml)
MATERIALS AND METHODS Rubia Plumbago Calophyllum Tamoxifen
Collection of plant material cordifolia zeylanica inophyllum
Rubia Cordifolia, Plumbago Zeylenica, Calophyllum 12.5 4.04 4.53 0.69 10.7
inophyllum leaves were collected from GITAM 25 17.05 25.792 1.38 34
University, Visakhapatnam and surrounding areas. The 50 26.46 31.23 19.44 66.4
leaves were shade dried and then dried at 70oC in hot 100 33.64 32.13 22.22 82.1
air oven. The dried leaves were then ground to powder 200 46.19 50.23 31.25 85.4
using mortar and pestle. The powder is stored in
airtight closed bottles. Table 2. Percent cell inhibition of plant extracts on HT-29 cell
line
Preparation of plant extracts
Conc % of cell inhibition
The shade dried (100g) powder of each plant material
(µg/ml)
were filled separately in the thimble and extracted with Rubia Plumbago Calophyllum 5 Fluoro
methanol using a Soxhlet extractor followed by cordifolia zeylanica inophyllum Uracil
distillation and evaporation. The plant powder is placed 12.5 5.36 0.68 4.2 24.2
in a cellulose thimble in an extraction chamber, which 25 6.04 4.76 12.80 34.6
is placed on top of a collecting flask beneath a reflux 50 7.38 21.08 12.80 57.5
condenser. Methanol is added to the flask,and the set 100 12.75 22.44 15.24 84.2
up is heated under reflux. When a certain level of 200 16.77 25.17 22.56 88.6
condensed solvent has accumulated in the thimble, it is
siphoned into the flask beneath.Nearly 15 cycles are MCF-7 cell lines grown in DMEM, when subjected to
repeated for 72 hours for all the plant species. After different concentrations of Rubia cordifolia extract
extraction the solvent is removed by rotary evaporator, resulted in 46.19 % inhibition. Similarly Plumbago
yielding the extracted compound. The non-soluble zeylanica resulted in 50.23% of MCF-7 cell death
portion of the extracted solid remains in the thimble, whereas Calophyllum inophyllum displayed weak
and is usually discarded. inhibition of 31.25%.

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CONCLUSION
From the analysis reported above it can be concluded
that Plumbago zeylanica, Rubia cordifolia possess
anticancer properties against breast cancer cell line
(MCF-7). The screening study revealed that the three
plants were found to be inactive against HT-29 colon
cancer cell line. Further experimental analysis on these
plants would definitely reveal the important chemical
constituents responsible for cancer cell death because
the probable inhibitions with active
principles/chemical constituents would be higher than
the extracts (due to the presence of mixture of varied
Figure 1. Effect of plant extracts on MCF-7cell line
constituents).

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Figure 2. Effect of plant extracts on HT-29 cell line
On the other hand, comparison with Tamoxifen showed
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inhibitions. Separation of chemical constituents by
column chromatography followed by thin layer
chromatography identification would reveal the major
components present in these plants. Work is currently
under progress in this direction.
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