Cytotoxicity of ethanol extract

Melastoma malabathricum L
(senduduk) fruit in HeLa cell line
cultur

Deri Edianto, Aznan Lelo, Syafruddin

Ilyas, Marline Nainggolan, Andrijono,
Putri C Eyanoer
 INTRODUCTION

• The leading causes of death worldwide are shifting

from infectious disease to non infectious disease.
• Cancer is predicted to become the leading cause of
death worldwide
• The world’s population of cancer continues to rised
from 14.1 million in 2012 to 20 million new cases by
2020.
• In developing countries like Indonesia where cervical
cancer screening do not have a good regulation, most
cervical cancer patients come in late stage
 INTRODUCTION

• The failure and systemic toxicity effects of

conventional cancer therapy has led to
research in new materials
• Its expected to prevent or slow down the
growth of cancer, but the human body is more
tolerable.
• Today’s study of phytopharmacy is being
conducted to found new drugs which are
more acceptable with less side effects
 INTRODUCTION

• Many studies report the using of polyphenol

as a component for cancer prevention and
treatment. This compound is easily obtained,
but fast metabolized, with low bioavailability.
• Clinical trials of the effects of polyphenol on
cervical cancer are still very limited, mostly
reporting the use of green tea and curcumin
as cervical cancer treatment
 INTRODUCTION

• Senduduk plants (Melastoma malabathricum L)

are traditionally believed to have many medicinal
benefits including anti cancer.
• In addition to anthocyanin, this plants also
contains secondary metabolites such as tannins,
alkoloid, steroid, flavonoid and terpenoid
• Many research report on the effects of
anthocyanin in cancer cell proliferation both in
vitro and in vivo trials
 MATERIALS AND METHODS

Ethanol extract of senduduk fruit

• The senduduk fruits were taken from vacant land in
Gebang district, Langkat Regency, North Sumatera
Province.
• Ripe senduduk fruit that is still intact with a reddish
color rind, is dried in a drying cabinet with 400C until
brittle, than mashed with a blender and macerated
• The extraction was carried out with 80% ethanol,
concentration were carried out with a rotary
evaporator at 500C until a blackish purple solid form
was formed.
Senduduk plant
 MATERIALS AND METHODS

HeLa cell cytotoxic culture test

• Hela cells in this study used Hela cells cultured in
the Department of Parasitology, Faculty of
Medicine, Universitas Gajah Mada, Jogjakarta
• Cultured in petri dishes with RPMI culture media
containing 10% serum foetal bovine, fungizone
(Ampotericin B) 0.5% and Penstrep 2%
• Cells were cultured in 370C incubators with 5%
CO2 levels. After confluent growth, cells were
harvested and recultured for cytotoxic testing
 MATERIALS AND METHODS
• The cytotoxic test used an MTT assay on a 96 well
plate, each well was filled with 10,000 cells, then
was given a senduduk extract in DMSO solutions
with concentration 500, 250, 125, 62.5, 31.25,
15.625, and 7.8125 ug/mL respectively
• Absorption of living cells was assessed using an
Elisa reader with a wavelength of 595 nm
• The content of anthocyanin in the extract was
spectrophotometrically tested at the Saraswati
Indo Genetech laboratory, Bogor
HeLa Cells
 RESULTS
• Life cell calculation from the MTT assay results
using the formula:

• A = control cell (cell + media)

• B = control media
• C = sample + media + cell
 RESULTS
120

100

80

60
% Cell Viability

40

20

0
500 µg/ml 250 μg/ml 125 μg/ml 62.5 µg/ml 31.25 µg/ml 15.625 µg/ml 7.8125 µg/ml
 RESULTS
• The IC50 senduduk extract in HeLa cell culture
was 956 µg/ml
• Antocyanin content examination conducted by
PT Saraswati Indo Genetech, Bogor Indonesia
through spectrophotometry examination were
115.70 mg/kg for simplo test and 115.82
mg/kg for duplo test.
 DISCUSSION
• Many part from senduduk plants contain active
ingredients, including anthocyanin. These ingredients
can be obtained from leaves, stems, flowers and fruit
• Senduduk fruit also contains many active ingredients
that have pharmacological activities such as flavonoids,
tannins, phenols beside anthocyanins
• This study revealed that senduduk fruit contain
anthocyanin
• The anthocyanin content of the senduduk fruit extract
was reported in other study as 49.9 mg/L, while in this
study the levels of anthocyanin in the extract of
senduduk fruit were 115.70 mg / kg.
 DISCUSSION
• The levels of fruit and plant extracts are varied and have
been tried in the cancer treatment of both in vitro and
invivo
• Invivo experiments are generally used as chemopreventive
materials. Guinea pigs were prior exposed with senduduk
extract before cancer was induced.
• In this experiment, the senduduk extract was exposed to
cancer cell culture to assess its impact on the proliferation
of cancer cells
• Although the concentration needed is up to 965 ug/ ml to
reach IC 50, but this extract is quite safe because it is
obtained from plants that are naturally consumed by
humans and birds.
 DISCUSSION
• Anthocyanin as one of the active ingredients in senduduk fruit, can
also be obtained from other colored fruit.
• Anthocyanin can inhibit cancer growth through several mechanisms
such as inhibiting angiogenesis, invasion, and metastasis.
• Bilusic et al reported that the activity of anthocyanin compounds in
various types of cell culture is still controversial. Cytotoxic effect
can be achieved with high dose
• Wang and Stoner reported that cancer cells can take more
anthocyanin, even up to 100 times than normal cells. And can store
anthocyanin longer, up to 12 hours compared to normal cells which
are only less than 2 hours
• Rugina et al found that elderberries fruit extract containing high
levels of anthocyanin with IC50 concentrations of 164.3 μg/mL in
murine melanoma B16-F10 cell culture.