LARVICIDAL POTENCY OF MADRE DE CACAO (Gliricidia sepium) LEAF EXTRACT

AGAINST MOSQUITO (Aedes aegypti) LARVAE

 IPSF OFFICIAL ABSTRACT FORM

Title: LARVICIDAL POTENCY OF MADRE DE CACAO (Gliricidia sepium) LEAF

EXTRACT AGAINST MOSQUITO (Aedes aegypti) LARVAE
Name: AYALISA MAE S. AYAO
ADDRESS: TIMOG STREET BOLILA, MALITA DAVAO OCCIDENTAL
School: MARIANO PERALTA NATIONAL HIGH SCHOOL
The study was conducted to determine the potential of madre de cacao (Gliricidia) against
mosquito (aedes agypti) larvae. The trial was arranged in Randomized Complete Block Design
(RCBD) with five treatments replicated three times. Treatment #1, #2, #4, and #5 has 10
grams, 20 grams, 40 grams, 70 grams, 90 grams and 200 ml of top water. Treatment 4 was
the control group which has mL of tap water. Each replication has ten mosquito larvae put in
an empty container and was sprayed with the treatments three times. One hour after the
application, the number of dead and survived mosquito larvae were counted and recorded.
Treatment 4 which has the greatest amount of madre de cacao leaves extract and has the
highest mortality rate of 90%. The greater the amount of madre de cacao leaves extract, the
higher the mortality rate of mosquito larvae. Otherwise, the treatment that has only water and
no extract of madre de cacao leaves was not able to kill any of the ten mosquito larvae in each
replication. These findings revealed that Madre de cacao leaf extract is a potential larvicide
against mosquito larvae. Results of the study showed that the mortality rate of mosquito larvae
among five treatments does not differ from one another at 1% level of significance. Therefore
G. sepium leaf extract has potent effect on controlling mosquito larvae which can be explored
further.
As part of this project the student directly handed manipulated or interacted with (check all
that apply).
___Human subjects ___Phathogenic agents ___Recommendation DNA
___Non-human vertebrate animal ___Controlled substances ___Human/Animal issues

Student independently performed all producers as outlined in this abstract. ___Yes ___No
This project was conducted at Registered Research Institute. ___Yes ___No
Is this project a continuation? ___ Yes ___No

I/We hereby certify that the above statements are correct and the information provided in the
abstract is the result of one year’s research. I/We also attest that the above properly reflects my/our
own work.

AYALISA MAE S. AYAO

Signature of Individual Finalist or Team Leader

Date Signed___________________
 ACKNOWLEDGEMENT

With profound gratitude and sincere appreciation, the researcher would like to acknowledge
the following persons who contributed much in the whole duration of The Science Investigatory
Project.

To her parents, Mr. & Mrs. Rudy Ayao for giving her the undying support, morally and
financially.

To Mrs. Perciviranda G. Tindoy, Mrs. Shamae C. Magallanes & to Miss. For there expertise
in English grammar and language and for the suggestions;

To her classmates and friends who shared their knowledge and encouragement in
conducting this study;

To the God’s People youth family, for the motivation and support;

To her spiritual mentors, for the prayers and for the words of wisdom;

To Mr. Ryan C. Montano, the Head Teacher IV, for the trust and kindness;

and

Lastly, To GOD for the health and sound mind.

 DEDICATION

To my parents: words are not expressive enough of how I am very grateful to have you who
made my study complete.

To my friends and classmates for their undying support and encouragements.

To God Almighty, for the grace that abounds, for the protection, love and care.
 TABLE OF CONTENTS

PRELIMINARY PAGES
PAGE

APPROVAL SHEET i
ABSTRACT FORM ii
ACKNOWLEGEMENT iii
DEDICATION iv
TABLE OF CONTENTS v
LIST OF TABLES vi
LIST OF APPENDICES vii

CHAPTER I
INTRODUCTION AND RELATED LITERATURE
INTRODUCTION 1
REVIEW OF RELATED LITERATURE 3

II MATERIALS AND METHODS 4

MATERIALS 5
METHODS 6

III RESULTS AND DISCUSSION 8

IV CONCLUSION AND RECOMMENDATION 9

CONCLUSION 9
RECOMMENDATION 10

LITERATURE CITED 11
APPENDICES 12
DOCUMENTATION 12
CURRICULUM VITAE 13
 LIST OF TABLES

TABLE PAGE

1 Raw Materials of the Study 4

2 Laboratory Equipment 5

3 Mortality Rate of Mosquito Larvae 8

4 Anova-Survival 8
 LIST OF APPENDICES

APPENDIX NUMBER PAGE NUMBER

1 DOCUMENTATION 19
 CHAPTER 1
INTRODUCTION

Aedes aegypti borne diseases currently represents a greater health problem in tropical and
subtropical climates of the world. It transmits many serious diseases like Dengue fever which
continues to be a major public health problem in the countries of Western Pacific and Southeast
Asia. The increasing trend of dengue outbreaks accompanied by DHF is posing a problem of
outmost importance to the public health1.

Mosquito control is essential for the proliferation of mosquito borne diseases, environmental
quality and public health. Synthetic insecticides have been not very successful due to human,
technical, operational, ecological and economic factors. This may result to the development of
resistant strains of mosquitoes. One of the major drawbacks of chemical insecticides is that they are
non-selective and could be harmful to other organisms in the environment 2.So there is an urge to
look for an environment friendly, cost effective, biodegradable and target specific insecticides
against mosquitoes. Many of the organic insecticides are based on single ingredient, but plant
products comprises the number of chemicals which can act on the physiological process of
mosquitoes. Extracts from various part of the plant such as leaves, roots, stem and fruits are shown
mosquitocidal (larvicidal) activities3.

Gliricidia sepium often simply referred to as Gliricidia which is commonly known as Kakawiti,
is a medium size leguminous tree belonging to the family Fabaceae. The active medical compounds
present in Gliricidia are afrormosin, medicarpin, tannin, and some isoflavins. Since gliricidia sepium
leaves contains a compound called tannin, is reported to have antidiarrheic, antidysentric,
antimutagenic, antinephritic, antiviral, bacterial, cancer preventive, hepato-protective, pesticide,
psycotrophic, and viricide activities 4. Which is good for mosquito repellant, therefore it can be
possible to repel mosquitoes. Thus, the researcher made special products using plants or fruits as
an alternative material instead of chemicals that can change or destroy our health. The main
purpose of the propose study was to determine the potential effect of gliricidia sepium as a larvicide.
However, the study aims to help the society to control and can be implement to reduce the number
of mosquitoes. This study would be limited only to the madre de cacao plant.

1
Fulmali PV, Walimbe A, Mahadev PV. (2008). Spread, establishment & prevalence of dengue vector”. Indian J Med Res 127:589-601
2
Lee and Kim, 2001Lee SE, Kim JE and Lee HS (2001). Insecticide resistance in increasing interest. Journal of Applied Biological Chemistry
44(3) 105-112
3
Gbolade AA (2001). Plant-derived insecticides in the control of malaria vector. Journal of Tropical Medicinal Plants 2(1) 91-97
4
Akharay FC, Boboyae B, Adetuyi FC (2012). Anti-bacterial, phytochemical and anti-oxidant activities of the leaf extracts of Gliricidia sepium
and Spathodea campanulata. World Appl Sci J;16:523-30
 REVIEW OF RELATED LITERATURE

A. Mosquitoes/Larvae
The mosquitoes are a family of small, midge-like flies: the Culicidae. Although a few species
are harmless or even useful to humanity, most are nuisance because they consume the blood from
living vertebrates, including humans. In feeding blood, various species of mosquitoes transmit some
of the most harmful to human and livestock diseases. Some authorities argue accordingly that
mosquitoes are the most dangerous animals on Earth.

Below is the scientific classification of a mosquito

Kingdom: Animalia

Phylum: Arthropoda

Class: Insecta

Order: Deptera

Suborder: Nematocera

Infraorder: Culicomorpha

Superfamily: Culicidae

Family: Culicidae

Mosquito larvae have a well-developed head with mouth brushes use for feeding, a large
thorax with no legs, and a segmented abdomen. Larvae breath through spiracles located on the
eight abdominal segment, or through a siphon, and therefore must come to the surface frequently.

The larvae spend most of their time feeding on algae, bacteria, and other microbes in the
surface microlayer. They dive below the surface only when disturbed. Larvae swim either through
propulsion with their mouth brushes, or by jerky movements of their entire bodies, giving them the
common name of “wigglers” or “wrigglers.”

Larvae develop through four stages, or instars, after which they metamorphose into pupae.
At the end of each instar, the larvae molt, shedding their skins to allow farther growth.

B. Madre de Cacao
Madre de Cacao or Kakawete is a perennial mostly herbaceous plant in the Fabaceae
family. The genus is native to Central America and possibly northern South America, but some
species have become naturalized around the world. One species, G. sepium, is considered as a
plant that has the ability to eradicate some variety of insect larvae.

Gliricidia sepium has also been used extensively outside its native range in places which
include the Carribean, the Philippines, India, Sri Lanka and West Africa. These landrance
populations are largely remnants of colonial introductions used to shade plantation crops although
more recently they have been integrated into indigenous farming practices uses for fuelwood, living
fences, animal forage, green manure and soil stabilization.

The active medical compounds present in Gliricidia are afrormosin, medicarpin, tannin, and
some isoflavins. Tannin is reported to have antidiarrheic, antidysentric, antimutagenic, antinephritic,
antiviral, bacterial, cancer preventive, hepato-protective, pesticide, psycotrophic, and viricide
activities.5 The leaves of Gliricidia are used as a mosquito repellent, and they have antifungal and
antibacterial activity. Various phytochemicals such as flavonoids, triterpenoid, coumarin, coumaric
acid, melilotic acid, and stigmastanol glucoside have been identified and isolated from various parts
of this plant. 42 known compounds are found in the leaves and flowers of G. sepium. 6
In India many compounds (Phytochemicals) derived from plants has been recognized as an
important source of insecticide and larvicide and many of them exhibit harmful effect on
mosquitoes.7 Gliricidia sepium is a medium sized tree found in most part of India and it was
observed that it is relatively free from insects and disease problems. The presence of Gliricidia
sepium in the field reduces incidence of some fungal and insect attack.
Since it has an larvicidal effect, it is hypothesized that the leaf extracts can be an effective
larvicide against mosquitoes.

Below is the scientific classification of a mosquito

Kingdom: Plantae

Phylum: Spermatophyta

Subphylum: Angiospermae

Class: Dicotyledon

5
Akharay FC, Boboyae B, Adetuyi FC (2012). Anti-bacterial, phytochemical and anti-oxidant activities of the leaf extracts of Gliricidia sepium
and Spathodea campanulata. World Appl Sci J;16:523-30
6
Joji Reddy L, Beena J (2010). Evaluation of antibacterial activity of the bark, flower and leaf extract of Gliricidia sepium”. Int J Curr Pharm
Res;2:18-20
7
) Kuo PM, Chu FH, Chang ST, Hsiao WF and Wang SY (2007). Insecticidal activity of essential oil from Chamaecyparis formosensis.
Holzforschung 61(5) 595-599.
 CHAPTER II
MATERIALS AND METHODS

Table 1 showed the raw material used as a treatment of the study.

Raw Materials Pictures Functions

Madre de cacao leaves

Used as a treatment of the

study.

Table 2 showed the laboratory equipment’s used in the study with their functions.
Materials Picture Functions

Used to store, mix and heat

Beaker the plant sample.

Used to store and mix the

Erlenmeyer Flask raw materials.

Used to transfer the raw

Funnel materials to empty spray
bottle.
 Stirring Rod

Used to stir the raw

materials.

Cheesecloth Used in separating the extract

from the blended Madre de
cacao leaves.

Digital Weighing Scale Used to measure the

kilogram of the raw
materials.

Stop Watch Used for measuring specific

time intervals.

Used as a container for the

Empty Spray Bottle set up and for the
treatments.
 blender Used for further grinding and
mixing the pounded Madre de
Cacao leaves.

Phase I: Plant Materials

Collection of Madre de Cacao Leaves
Gather all the collected madre de cacao leaves. The collected leaves were only picked at
Timog, Bolila, Malita, Davao Occidental.

Phase II: Plant Preparation

Madre de Cacao Leaves Preparation

Weigh the fresh madre de cacao leaves using a digital scale for the better accuracy and was
measure by the different level of concentration. Put and transfer the crushed leaves of madre de cacao blended
with and the extracts were collected using a cheesecloth.

Preparing Mosquito Larvae                                                                                          

Using a net, carefully get the larvae from the water. Then, transfer the larvae into an empty bottle were the
treatments belong. Lastly, count the number of larvae in each treatment accordingly.

Phase III: Data Collection & Analysis

The trial was arranged in Randomized Complete Block Design (RCBD) with five set-ups
replicated three times. 1 hour after the application, the number of dead and surviving mosquito
larvae was counted and recorded.

The Different Treatments of the Study

The four different treatments considered in this study were the following:
T1: 10 grams of madre de cacao leaves + 200 mL tap water
T2: 40 grams of madre de cacao leaves + 200 mL tap water
T3: 70 grams of madre de cacao leaves + 200 mL tap water
T4: 90 grams of madre de cacao leaves + 200 mL tap water
T5: 200 ml top water
 CHAPTER III

RESULTS

Table 3 showed the effects of 10 g, 40 g, 70 g, and 90 g of madre de cacao leaf extract in

200 ml water per treatments to the health, behavior and death of mosquito larvae after 1 hour.
Results showed that the higher average mortality rate between the five treatments are the treatment
#4 with the total the average of 90 percent.

MORTALITY RATE OF MOSQUITO LARVAE

TREATMENT
TEN (1) HOUR AFTER THE APPLICATION AVERAGE
%
R1 % R2 % R3 % MORTALITY

1 2 20% 3 10% 2 20% 2.33 20%

2 4 40% 4 40% 3 40% 3.66 40%

3 7 70% 7 70% 6 80% 6.66 70%

4 9 90% 9 90% 9 90% 9 90%

5 0 0 0 0 0 0 0 0

Table 3: Mortality rate of mosquito larvae

Test on Hypothesis

Table 4 showed the Analysis of Variance of mortality rate of the mosquito larvae (1) hour
after the application of different treatments.
Since the computed F value of 0.000 is less than the tabular value of 0.001, the null
hypothesis is accepted. This implies that the mortality rate of mosquito larvae among five treatments
does not differ from one another base on the level of significance.

Cases Sum of Squares df Mean Square F p

TREATMENT 147.600 4 36.900 184.5 < .001

Residual 2.000 10 0.200  

Table 4: Anova-Survival
 DISCUSSION

In treatment 1 (10 g Madre de cacao leaf extract added in 200 ml water) only 20 percent is
the mortality rate of the mosquito larvae. In treatment 2 with (40 g Madre de cacao leaf extract
added in 200 ml water) resulted to 40 percent mortality rate of the mosquito larvae. In treatment 3
(70 g Madre de cacao leaf extract added in 200 ml water) obtained 80 percent mortality rate of the
mosquito larvae. The treatment that has the greatest amount of Madre de cacao leaf extract
obtained the highest mortality rate of 90 percent mortality rate of mosquito larvae which is the
treatment 4. The result shows that the Madre de cacao leaf extract in 200 ml water the more are the
larvicidal effect to the mosquito larvae. In treatment 5 (control treatment) resulted to 0 mortality rate.
 CHAPTER IV

CONCLUSION

The data presented on the tables indicates that the Madre de cacao (Gliricidia Sepium) leaf
extract has larvicidal potency and effect in getting rid of mosquito larvae.

RECOMMENDATION

In view of aforementioned results, the following are recommended:

1. Further study using other plant parts of madre de cacao as a potential larvicide against
mosquito larvae.

2. Companies may develop mosquito larvicidal product with the use of madre de cacao as
main component.
LITERATURE CITED

Akharay FC, Boboyae B, Adetuyi FC. (2012). Anti-bacterial, phytochemical and anti-oxidant
activities of the leaf extracts of Gliricidia sepium and Spathodea campanulata. World Appl Sci
J;16:523-30.
Alameda County Mosquito Abatement District. (2011). Biological Notes on Mosquitoes. life cycle
of the mosquito.
Centers for Disease Control and Prevention. (2010). Dengue. Entomology and ecology.
Dengue Virus Net. (2011). Aedes aegypti. Dengue transmission by Aedes aegypti mosquito.
Fulmali PV, Walimbe A, Mahadev PV. (2008). Spread, establishment & prevalence of dengue
vector. Indian J Med Res 127:589-601.
Gbolade AA. (2001). “Plant-derived insecticides in the control of malaria vector”. Journal of Tropical
Medicinal Plants 2(1) 91-97.
Gbolade AA, Oyedele AO, Sosan MB, Adewayin FB and Soyela OL. (2000). Mosquito repellent
activities of essential oils from two Nigerian Ocimum species. Journal of Tropical Medicinal Plants
1(1/2) 146-148.
Glover N. (1989). Gliricidia Production and Use (Nitrogen fixing trees Association, Waimanalo,
Hawaii, USA) 44.
Joji Reddy L, Beena J. (2010). Evaluation of antibacterial activity of the bark, flower and leaf extract
of Gliricidia sepium. Int J Curr Pharm Res;2:18-20.
Kuo PM, Chu FH, Chang ST, Hsiao WF and Wang SY. (2007). Insecticidal activity of essential oil
from Chamaecyparis formosensis. Holzforschung 61(5) 595-599.
Lee and Kim, 2001Lee SE, Kim JE and Lee HS. (2001). Insecticide resistance in increasing
interest. Journal of Applied Biological Chemistry 44(3) 105-112.
Taubes G. (1997). A mosquito bites back. New York Times Magazine 24 40-46.
 APPENDICES
DOCUMENTATION
Phase 1: Plant Extraction
COLLECTION OF MATERIALS WEIGHING PROCESS

Phase 2: Experimentation
MIXING AND SEPARATION OF THE EXTRACT

Control Group
CONTROL
gROUP
 Treatment 1 Treatment 2 Treatment 3

Treatment 4

Phase 3: TESTING OF THE LARVICIDE

 CURRICULUM VITAE

Name: Ayalisa Mae S. Ayao

Date of Birth: September 5, 2003

Civil Status: Single

Address: Timog, Bolila, Malita Davao Occidental

Parents:

Mother: Cecille S.Ayao

Father: Rudy S. Ayao

Educational Background

Elementary: Mariano Calungsod Regis Elem. School

Bolila, Malita Davao Occidental

Secondary: Mariano Peralta National High School

Población, Malita Davao Occidental