ANTI-ANGIOGENIC ACTIVITY OF MORINGA OLEIFERA ETHANOLIC LEAF

EXTRACT IN CHRIOALLANTOIC MEMBRANE (CAM) ASSAY

A Research Paper
Submitted to the
Department of Family and Community Medicine
College of Medicine

Medicine Authors:

Achondo, Carlos Jr. D Araya, Ederlyn

09275918117 09455840138
carlosachondo76@gmail.com ederlynaraya@gmail.com

Acttanasiri, Ketsara Arenque, Mary Rolhalane Rose

09175414130 09380768250
ampketsara@hotmail.com lhalanearenque@gmail.com

Agraan, Shane Molyn Arocena, Melsergio

09553190292 09157048026
agraanshanev@yahoo.com melsergioa@gmail.com

Alingayao, Liezel Gupta, Ankit

09164176598 09270881616
Alingayaoliezel16@gmail.com ankitsharangupta@gmail.com

Anzures, Yelena Dey, Soumamitra

09363201332 09666543879
ycanzuresmt@gmail.com soumamitradey@gmail.com

Apalisok, Meggyrose Patel, Arpit

09998826922 09394088910
meggydiokno@gmail.com ap6900208@gmail.com

Grace Villanueva, MD, DFM, FPAFP

Janice D. De Leon, MD, DFM, FPAFP

Chairman of the Department of Family and
Community Medicine
College of Medicine
INTRODUCTION
According to the World Health Organization (WHO) that Cancer and related diseases
ranks second as the leading cause of death globally and is responsible for an estimated 9.6
million deaths in 2018 alone. What is more drastic is that the burden of the disease is more
apparent in the low- and middle-income countries for which our own country belongs. Late stage
diagnosis and treatment are common. In the Philippines, cancer still remains to be a national
health priority with significant implications for individuals, families, communities and the health
system.
With the present status quo stated, it is of essence of the current study to better
understand the pathology of malignancy best exemplified by the process of angiogenesis.
Furthermore, as the gap between diagnosis and treatment on angiogenesis-related diseases i.e
cancer continues to grow especially in the low economic class, an opportunity for better health
care delivery must be offered that are novel and locally available but still producing efficacious
benefits to patients who cannot afford the sophisticated treatment that cancer therapy nowadays
employ and this study intends to determine the possibility of using Moringa oleifera commonly
known locally as “Malunggay”

METHODOLOGY

The research design of this study was experimental, in which researchers were testing for
the anti-angiogenic ability of Moringa oleifera using the Chorioallantoic Membrane Assay
(CAM) technique.
Thirty (30) three-day old fertilized duck embryos were obtained. Researchers used a
randomized control sampling for the research subjects. The eggs were placed in the incubator at
a constant temperature of 37.5°C and at a constant humidity.
The Leaves of Moringa oleifera L. that were used in the present study was collected from
a plantation in Laguna, a supplier of a vegetable vendor in Monumento Market. Its identity were
verified and authenticated by sending a voucher specimen in the University of the Philippines
College of Science in Diliman, Quezon City. About 160 g of pulverized plant material were
subjected for successive soxhlet extraction as previously described procedure. The extraction
was carried out using 650 ml of ethanol for 14 hours and the obtained extract was concentrated
to dryness under reduced pressure using rotary vacuum evaporator and was stored at 4°C until
used. This method was done with the help of Adamson University Technology Research and
Development Foundation Incorporated.
The test plant extract with the concentration of 25%, 50% and 75% Moringa oleifera
ethanolic extract were absorbed on the sterile filter paper discs. Discs containing 80% Ethanol
will be used as negative control and disc treated with 200mg Celecoxib + 200ml distilled water
was as positive control. Six eggs were assigned per experimental concentration and control. The
treated filter paper discs were placed onto the CAM. The treated eggs were sealed with sterile
parafilm tape or surgical tape and were incubated for two days. On the 12th day of incubation,
the CAMs were harvested by removing the hard shell leaving intact the soft membrane covering
the embryo. The shell-less embryo was transferred to a petri dish. The CAMs were photographed
under a dissecting microscope and blood vessels in each CAM were counted and analyzed for
blood vessels in the disc application site and the percentage inhibition were calculated.
RESULTS AND DISCUSSION
Table 1: Blood vessels growth on the set-up groups using the plant extract vs the positive

Negative Positive
Samples 25% 50% 75% Control Control
1 - - - - -
2 - - - - +
3 - - - - +
4 - - - - +
5 - - - + +
6 - - - - -
control

In the table that all the groups applied with the plant extract of Moringga oleifera (25%,
50%, 75%) showed no formation of blood vessels. Four of six eggs applied with the
positive control shows nascent blood vessels formation.

Fig 2: Eggs treated with 75% Moringa oleifera ethanolic extract shows no growth of any
blood vessels.

Legend:
(+) = Blood vessel growth was noted
(-) = No blood vessel growth was
noted

Fig 3: The use of the drug Celecoxib 2% as the positive control contributed to the
attributed formation of blood vessel on the membrane of the eggs.
 Consistent with the findings of the study done by Fitch et. Al (2016) mentioned
that methanolic extract of Moringa who oleifera tends to down regulate various angiogenic
factors (VEDF and KDR) in mice cervices , it was also noted in the preliminary
experiment of the current study on the first batch of experimental eggs that it was
apparent that upon usage of the plant extract all the treatment groups did not show any
formation or attempted to form blood vessel.
CONCLUSION
With different concentrations of 25%, 50% and 75% respectively, the attenuated
nascent blood vessel formation was noted.Attesting the possibility of angiogenic inhibitory
effect of this plant variety of real clinical diseases can be benefiting from it as a
potential novel treatment to mention one which angiogenesis play important role in
clinical symptomatology Cancer Malignancy.
Dissimilar to the expected effect of the positive control on its effect on the
experimental eggs opened for further investigations on the current research on how the
effect of the known drug showed a different response on the test eggs.

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