1

THE INSECTICIDAL PROPERTY OF PANSIT-PANSITAN (Peperomia

pellucida (Linn.) (HBK)) LEAF CRUDE EXTRACT AGAINST GREEN
LEAFHOPPER (Nephotettix malayanus (Ishihara et Kawase))

A Thesis Presented to the College of Arts and Sciences

University of Perpetual Help System DALTA
Pamplona, Las Piñas City

In Partial Fulfillment of the Requirements for the Degree in

Bachelor of Science in Biology

Gerard Majello B. Dalisay

March 2010
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APPROVAL SHEET

The thesis entitled “The Insecticidal Property of Pansit-pansitan

(Peperomia pellucida (Linn.) (HBK)) Leaf Crude Extract Against Green

Leafhopper (Nephotetix malayanus (Ishihara et Kawase))”, prepared by Mr.

Gerard Majello B. Dalisay in partial fulfillment of the requirements for the

Degree in Bachelor of Science in Biology, is hereby recommended for oral

examination.

MARIA LOURDES C. AGUIRRE, MSc.

Adviser

Approved in completion of the requirements for the Degree of Bachelor of

Science in Biology by the Oral Examination Committee.

MIRIAM B. RAÑOLA, M.A.

Member

RAQUEL MARI-ANN O. PAJE, M.A.

Member

MARY GRACE B. MARIBAO, M. Ed.

Chairman

Accepted in partial fulfillment of the requirements for the Degree in

Bachelor of Science in Biology.

FLORENCIA C. MARFIL,Ph.D.
Dean, College of Arts and Sciences
March 2010
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ACKNOWLEDGEMENT

First of all I would like to express my gratitude to the One God for the many

blessings that He has given. He is always there taking good care of me and always

comforts me whenever I was in trouble, pain or lonely. He guided me in all the

activities of my life including the construction of this study and He never lets me

go both in good times and in the bad.

I would also like to thank the following persons who helped me in making

this study.

To my parents, Mr. Honorio and Victoria Dalisay who greatly supported me

in physical, mental, emotional, spiritual and especially financial terms. I thank

them for the guidance, encouragement and love that they gave to me. To my

uncle, Mr. Teodoro Dalisay for the information about my test organism. Mrs.

Irenea Dalisay for assisting the construction of the insect cages. Mr. Gregorio

Gomez Jr., Mrs. Editha Roldan and the rest of the employees of Dalisay Poultry

Supply and Dalisay Piggery Farm for the construction of the insect cages and

assisting me whenever I go to the site of the study. To all the rice farmers of the

town of Lian, Batangas for looking after my experimental set ups whenever I am

unable to do so.

To the thesis adviser, Mrs. Maria Lourdes Aguirre for her extreme patience,

assistance and support. Truly the unmatched and ultimate “Mother of Perpetual

Help” for always listening to our complaints and problems and comforting us

when we are in need.

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To Ms. Miriam Rañola, for the patience and endurance in dealing with ones

papers and giving suggestions that can contribute to the betterment of the study.

To Mrs. Mary Grace Maribao, for the assistance and life-inspiring words

which has proven to be a valuable asset in achieving ones goal.

To Ms. Raquel Paje, for the wise and practical examples which enables one

to efficiently tackle the problems of life.

To Mr. Eduardo Jimmy Arma, for the humorous statements and antics that

never fails to enlighten anyone. Truly a walking piece of pure joy that makes you

feel happy and refreshed anytime, anywhere.

To Mr. Allan Alvarez, Dr. Vendivil and the rest of the staff of the Botany

Division of the National Museum for the plant identification. To Dr. Sinohin,

Mrs. Fe Alzona and the rest of the staff of the Crop Protection Cluster of the

University of the Philippines Los Baños, Laguna for the identification of test

organism. Dr. Timbang and the rest of the staff of the IHS Laboratory of the

University of Perpetual Help System DALTA for lending us laboratory

apparatuses required for the study. Also to Mr. Arturo Tobias Calizon for the

statistical analysis.

To Mr. Paolo Louis Almario for allowing me to be patient, kind and

enduring, I thank you for teaching me all of these. To Mrs. Kristina Ezra Salceda-

Dumdum for the tremendous amount of humor that she has given me. I thank you

for the huge amount of food that you are giving to us whenever we are at your

house. Ms. Monique Moldez for teaching me how to be strong, to be able to fight

back from other people’s insults whether in a casual or formal manner. I am very
 5

grateful for changing me from my doormat and submissive self to the strong

willed and firm individual that I am today. To Ms. Joanna Marie Valloso for

always reminding me of assignments, quizzes and other academic tasks that are

needed to accomplish. You truly are a genuine secretary in training. Thank you

also for sharing your food with me whenever you are unable to finish it.

To Mr. Gene LemuelAbanto, Olivier Thoenen and the rest of the B.S.

Biology students of the University of Perpetual Help System DALTA. I humbly

thank you all for the comedy, drama, horror, romance and all the emotions that

you let me experience. Truly I never have encountered such people and the

experiences that they gave to me are simply life changing and priceless.

Lastly to the people whom I failed to mention I thank you for your

generosity and effort in aiding me in this study. Once more I gratefully thank you

and God bless us all.

Gerard Majello Bulao Dalisay

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ABSTRACT

This study entittled “The Insecticidal Property of Pansit-pansitan

(Peperomia pellucida (Linn.)(HBK)) Leaf Crude Extract Against Green

Leafhopper (Nephotettix malayanus (Ishihara et Kawase))” aims to confirm the

active components present in Pansit-pansitan leaf crude extract using

phytochemical analysis. The study also aimed to determine the insecticidal

property of Pansit-pansitan based on the mortality rate of Green Leafhopper and

compare the different concentrations of Pansit-pansitan leaf crude extract with

Sevin ® 85 WP (positive control).

The leaf crude extract of the plant material was obtained through decoction

method. Phytochemical analysis was done to confirm the different constituents

present in Pansit-pansitan leaf crude extract.

In each treatment three trials have been carried out. These were Sevin ® 85

WP (positive control) and the varying concentrations of the leaf crude extract of

Pansit-pansitan (50%, 75% and 100%) have been prepared by dissolving 5g, 7.5g

and 10g of leaf crude extract in 10ml of distilled water, respectively. Insecticidal

property was determined based on the mortality rate of Green Leafhopper by

administering 10ml of each treatment and the positive control through spray

method.

Results showed that the highest mortality rate of Green Leafhoppers was

obtained for Sevin ® 85 WP (positive control) followed by 100% concentration of

Pansit-pansitan leaf crude extract and the least mortality rate has been observed at

50% concentration. Statistical analysis showed that the computed F value of

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3.303303 is greater than the critical value of F of 3.24 at 0.05 level of significance

thus, the null hypothesis was rejected. There is a significant difference in the

insecticidal property between Pansit-pansitan leaf crude extract and the positive

control, Sevin ® 85 WP.

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TABLE OF CONTENTS

TITLE i
APPROVAL SHEET ii
ACKNOWLEDGEMENT iii
ABSTRACT vi
TABLE OF CONTENTS viii
LIST OF TABLES x
LIST OF FIGURES xi
LIST OF PLATES xii
LIST OF APPENDICES xiii

CHAPTERS

I THE PROBLEM AND ITS BACKGROUND

Introduction 1
Theoretical Framework 2
Conceptual Framework 3
Statement of the Problem 4
Hypothesis 4
Significance of the Study 5
Scope and Delimitation 5
Definition of Terms 7

II REVIEW OF RELATED LITERATURE

Conceptual Literature 8
Scientific Classification of Pansit-pansitan 8
Botanical Description 9
Chemical Content of Botanical Extract 10
Brief Background about Green Leafhopper 12
Classification 12
Taxonomy 13
Life Cycle 14
Sevin 85 ® WP 15
Research Literature 15

III RESEARCH METHODOLOGY AND PROCEDURE

Research Design 18
Research Setting 18
Research Procedure 19
Collection and Preparation of Plant Specimen 19
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Identification of Plant Material 19

Identification of Test Organism 20
Extraction Procedure 20
Decoction Method 20
Preparation of Different Concentrations
of Leaf Crude Extract 20
Preparation of Sevin ® 85 WP (control) 20
Preparation of the Test Organism 21
Preparation of Green Leafhopper Cage 21
Collection of Green Leafhopper 21
Preparation of Green Leafhopper for Mating 21
Application of Leaf Extract to Green Leafhopper 22
Statistical Treatment 22

IV PRESENTATION, ANALYSIS AND

INTERPRETATION OF DATA 23

V SUMMARY, CONCLUSION AND RECOMMENDATION

Summary 28
Conclusion 29
Recommendation 30

BIBLIOGRAPHY

PLATES

APPENDICES

CURRICULUM VITAE
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CHAPTER 1

THE PROBLEM AND ITS BACKGROUND

Introduction

Rice is the staple food of the Filipino people. This particular crop has been

in the Philippine table for thousands of years from its early inhabitants up to the

modern age. It is an annual plant that is cultivated in warm and wet areas in which

the seeds are cooked and used for food.

The said crop is widely cultivated throughout the country, carefully planted

in well irrigated areas, tended by the most skilled farmers and efficiently

processed before going out to the market. However, rice continues to decrease in

yield due to pest infestations thus plant cultivation and harvesting is severely

affected.

The most effective method to get rid of pest infestations nowadays is to use

commercial insecticides however they may cause unwanted effects such as land

poisoning and may endanger human health if not properly handled. Today’s

farmers are running low on funds to buy insecticides which are too expensive.

Thus the use of botanical extracts have been encouraged as possible substitute for

commercially available insecticides.

It is interesting to note that certain plants have insecticidal properties.

Among these plants that have shown such properties is Pansit-pansitan

(Peperomia pellucida (Linn.) (HBK)) . The said herbaceous annual plant contains

chemical substances such as alkaloids that is proven to cause irritation and death

to most types of insects.

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Theoretical Framework

Pansit-pansitan has long been used by people from all parts of the world in a

variety of ways. In the old days infusion of the leaves and stems are used for gout

and arthritis. The plant has a history of ethnomedicinal use. Anti-inflammatory,

chemotherapeutic, and analgesic properties have been found in crude extracts of

Pansit-pansitan. Data in Bolivia documents the whole plant being crushed, mixed

with water, heated, and then administered to stop hemorrhage (www.drugs.com).

Externally it is used as a facial rinse for complexion problems. Pounded

whole plant used as warm poultice for boils, pustules and pimples. It is also used

for headaches and rheumatic pains (http://www.stuartxchange.org/Pansit.html).In

Southeast Asia, Pansit-pansitan is reported to have analgesic, antibacterial and

antifungal activity. According to a recent study, the plant contains alkaloid which

is a plant-derived compound that is toxic or physiologically active. It has many

uses depending on its type such as quinine which has anti-malarial compound or

tropane alkaloids which have been used throughout history as poisons (Murray G,

Frank M, 1973). Phytochemical studies have also revealed the presence of

alkaloids and tannin in abundant amounts (De Padua LS, Pancho JV. 1983).
 3

Conceptual Framework

Green Leafhoppers are a common pest problem throughout the Philippines.

Millions of pesos are spent to control them every year. Pansit-pansitan has been

used as an alternative to commercial insecticides that can be used to exterminate

such rice pests.

Independent variable Dependent variable

Varying concentrations Insecticidal property of

of Pansit-pansitan leaf the leaf crude extract
crude extract (50%,
Mortality rate of Green
75%, 100%) and Sevin
Leafhoppers
® 85 WP (positive
control)

Figure 1. A schematic diagram showing the insecticidal property of Pansit-

pansitan against Green Leafhopper (N. malayanus (Ishihara et

Kawase))
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Statement of the Problem

The study was conducted in order to determine the insecticidal property of

Pansit-pansitan (Peperomia pellucida (Linn.)(HBK)) leaf crude extract against

Green Leafhopper (Nephotettix malayanus (Ishihara et Kawase)).

Specifically it aimed to:

1. confirm the active components present in Pansit-pansitan (Peperomia

pellucida (Linn.)(HBK)) leaf crude extract using phytochemical analysis;

2. determine the insecticidal property of Pansit-pansitan (Peperomia

pellucida (Linn.)(HBK)) against Green Leafhoppers (Nephotettix

malayanus (Ishihara et Kawase)); and

3. compare the different concentrations of Pansit-pansitan (Peperomia

pellucida (Linn.)(HBK)) leaf crude extract with Sevin ® 85 WP (positive

control) against Green Leafhoppers (Nephotettix malayanus (Ishihara et

Kawase)).

Hypothesis

There is no significant difference in the insecticidal property between

Pansit-pansitan (Peperomia pellucida (Linn.)(HBK)) leaf crude extract and the

positive control, Sevin ® 85 WP against Green Leafhoppers (Nephotettix

malayanus (Ishihara et Kawase)).

Significance of the Study

The main purpose of this study is to determine the insecticidal property of

Pansit-pansitan leaf crude extract against Green Leafhopper. The result of the
 5

study will encourage researchers to formulate more plant based products which

are organic, environmental friendly and cost efficient. It could also be used to

help minimize the occurrence of pests on rice and on other crops, thus pest control

would be stabilized. Ultimately, farmers will be financially benefited because of

the inexpensiveness of the plant extract thus they could focus more on increasing

rice production. Also farmer’s consumption of inorganic insecticides will be

reduced which are harmful to the environment and health. Rice production will

increase and the Philippine economy will eventually improve.

Scope and Delimitation

This study focused on the collection and preparation of the plant material.

The plant extraction procedure done for Pansit-pansitan leaves was through

decoction method by which varying concentrations have been derived such as

50%, 75% and 100%. The extraction method was performed in the IHS

Laboratory, University of Perpetual Help System DALTA located in Pamplona,

Las Piñas City. Phytochemical analysis was also conducted in order to confirm

the chemical components of the Pansit-pansitan leaf.

Three treatments were performed in the study. In each treatment three trials

have been carried out. These were Sevin ® 85 WP (positive control) and the

varying concentrations of Pansit-pansitan leaf crude extract. The varying

concentrations of Pansit-pansitan leaf crude extract was prepared by dissolving

5g, 7.5g and 10g of leaf crude extract in 10ml of distilled water in order to obtain

50%, 75% and 100% concentrations, respectively. The study used adult forms of
 6

Green Leafhopper which were about 11 to 13 days upon hatching. The insects

have been cultured in a natural habitat by placing a fine net on a site in the rice

field to ensure that the specimen will not escape and were allowed to reproduce.

Green Leafhopper eggs were then collected after 6 days upon laying by gently

brushing the rice plants located on the site with a soft brush to remove the adult

insect thus the insect eggs remained in the rice plant. The rice plants along with

the Green Leafhopper eggs were then carefully transferred to another site in the

rice field covered with a fine net and allowed to reach adulthood after 11 to 13

days upon hatching (Hou, 1987 modified). The number of mortality of Green

Leafhopper was counted after the leaf crude extract has been applied through

spray method every 6 minutes for a total of 30 minutes. The insects were sprayed

with the leaf crude extract between 2:00-3:00 PM.

The study was conducted from December, 2009 to January, 2010.

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Definition of Terms

The following terms are defined for better understanding of the study:

Decoction method It is an extract of herbs produced by boiling the

plant on water.

Extract It is a mean to obtain from a substance by chemical

or mechanical action.

Extraction It is an act of extracting.

Insecticidal It is a chemical substance to kill insects.

Mortality It means death of large numbers.

Mortality rate It is the whole number of death in a given time or

community.

Oviposition It is the process of laying eggs in insects.

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CHAPTER 2

REVIEW OF RELATED LITERATURE

This chapter presents previous studies and information related to the study.

Conceptual Literature

Pansit-pansitan, from the family Piperaceae is an annual herb that is found

in many South American and Asian countries, shallow rooted, that reach 40 cm

high, with succulent stems.

Scientific Classification

Kingdom Plantae – Plants

Class Magnoliopsida – Dicotyledons

Subclass Magnoliidae

Order Piperales

Family Piperaceae – Pepper family

Genus Peperomia Ruiz & Pav. – peperomia

Species Peperomia pellucida (L. HBK)

Common Names: Pansit-pansitan, Ulasimang-bato (Tag.), Shiny bush , silver

bush (Engl.)
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Botanical Description

Pansit-pansitan (Peperomia pellucida (Linn.) (HBK))

It belongs to the family

Piperaceae. It is an herbaceous

annual plant with succulent stems

found in many South American and

Asian countries. The plant grows to

a height of 15 to 45 cm. Its leaves

are alternate, heart-shaped and

turgid, as transparent and smooth as

candle wax. Tiny dot like flowers

Figure 2. Peperomia pellucida (Linn. HBK)
scattered along solitary and leaf-

opposed stalk maturing gradually from the base to the tip and turning brown when

ripe. It propagates by seeds. Numerous tiny seeds drop off

when mature and grow easily in clumps. The species develops during rainy

periods and thrives in loose, humid soils under the shade of trees

(www.drugs.com). The decoction of leaves is used for gout and arthritis

(http://www.stuartxchange.org/Pansit.html). Also the pounded stems are applied

on the face for boils and pimples and have said to have anti bacterial property

against Bacillus subtilis and Staphylococcus aureus. (Lans, Journal of

Ethnobiology and Ethnomedicine, 2007)

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Chemical Content of Botanical Extract

Pansit-pansitan (Peperomia pellucida (Linn.) (HBK))

It was found out that the plant leaf contains ascorbic acid, beta-carotene,

calcium, carbohydrates, fat, fiber, iron, niacin, phosphorus, potassium, protein,

riboflavin, sodium, thiamine, water (http://www.ars-grin.gov/cgi-

bin/duke/farmacy2.pl). Other substances are benzenoid, steroids and apiol found

throughout the plant (Manalo JB, et al. 1983). Alkaloids and tannin are mainly

abundant in leaves (De Padua LS, Pancho JV. 1983).

Alkaloids are plant-derived compound that is toxic or physiologically active.

It has a complex structure and is of limited distribution in the plant kingdom.

Alkaloids have many uses such as poisons which build up nerve impulses

transmitted by the brain which results in restlessness, convulsions, nausea,

vomiting and death (Dales MJ, 1996). Quinine which is a type of alkaloid has

anti-malarial compound. Another form of alkaloids is atropine and is used to

dilate the pupils of the eye. Atropine is also a CNS stimulant and is used as a

treatment for nerve gas poisoning (Murray G, Frank M, 1973).

Apiol (C12H14O4) is an essential oil usually found in some succulent plant

species. It is green, non-viscous, oily liquid with a peculiar odor and a

disagreeable acrid taste (United States National Library of Medicine). In humans

extremely high doses of the chemical may produce effects such as ringing in the

ears, headache and vertigo. In pregnant women it can cause an accumulation of

blood in the pelvic-uterus area and liver damage. High doses of some oils,
 11

especially containing apiol can have narcotic or paralyzing effects

(http://www.juniperandsage.com/oils/toxicity.php).

Tannins are polyphenols that are obtained from various parts of different

plants belonging to multiple species. It is found in abundance in the tree bark,

wood, fruit, leaves, and roots and also in plant gall. Interestingly, tannins are

found almost in all plants and in all climates all over the world however its

percentage varies depending on plant species. Tannins are usually found in large

quantities in the bark of trees where they act as a barrier for micro-organisms like

bacteria and fungi and protect the tree. It is also used in dyeing, photography,

astringent, stops infection, and refining beer and wine

As tannins often lower the absorption of some materials into the body,

tannins are also often known as anti-nutrients. Tannins can be classified into two

broad groups - hydrolysable tannins and condensed tannins. Hydrolysable tannins

are basically derived from simple phenolic acids like gallic acid that when heated

they give away pyrogallol. Pyrogallol is also known as hepatotoxic and has

antiseptic as well as caustic properties. Condensed tannins do not split easily and

hence it is difficult to analyze these. When condensed tannins are heated, catechol

emerges as the final product. Unlike hydrolysable tannins, condensed tannins do

not possess any trace of hepatotoxicity or any adverse side effects and hence are

favorable for use (Harborne Jeffery, H. Baxter. 1983).

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Brief Background about Green Leafhopper (Nephotettix malayanus (Ishihara

et Kawase))

Classification of Green Leafhopper

Domain Eukaryota- eukaryotes

Kingdom Animalia – animals

Subkingdom Bilateria- bilaterians, triploblastic animals

Branch Protostomia- protosomes

Infrakingdom Ecdysozoa- ecdysozomes

Superphylum Panarthropoda

Phylum Arthropoda- arthropods

Subphylum Mandibulata

Class Insecta -insects

Subclass Dicondylia

Order Coleoptera

Suborder Auchenorrhyncha

Superfamily Cicadelloidea

Family Cicadellidae

Genus Nephotettix

Species N. malayanus (Ishihara et Kawase)

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Taxonomy

Green Leafhopper belongs to

the Phylum Arthropoda. It

constitutes the suborder Polyphaga

within the class Insecta. The insect

is said to be dioecious and have

paired reproductive organs. Green

Leafhoppers are serious pests in

rice. They damage the plants

directly by sucking the plant sap Figure 3. Nephotettix malayanus (Ishihara et

Kawase)
resulting in complete plant drying.

Green Leafhoppers are slender, green and may have black markings on the

head or wings. Eggs are white and elongate or cigar-shaped. Individual eggs are

arranged neatly and lie parallel to each other in each egg batch. A single egg

measures 0.9 - 0.8 mm. Upon maturation, the egg turns brownish and develops

red eyes. As the nymph matures, it turns to green. (Pathak MD, 1994).

As the insect matures, blackish markings on the abdomen become more

prominent as well as the blackish band on the last abdominal segment. (Reissig

WH, 1986). They are less than 13mm long and have an average life span of 3

weeks. They disperse rapidly when disturbed. Both adults and nymphs run

sideways and are good jumpers or hoppers. They are generally found in small

numbers at the leaf blade and feed on the upper portion of the rice canopy. A

female hopper can lay as many as 200-300 eggs (Shepard BM, 1995). Green
 14

Leafhoppers are common in rain fed and irrigated wetland environments. They

are not prevalent in upland rice. Both the nymphs and adults feed on the dorsal

surface of the leaf blades rather than the ventral surface. They prefer to feed on

the lateral leaves rather than the leaf sheaths and the middle leaves. They also

prefer rice plants that have been fertilized with large amount of nitrogen

(http://everest.ento.vt.edu/~idlab/vegpests/vegi/LH.jpeg).

Life Cycle

Green Leafhoppers

begin as an egg laid on a

plant leaf. The eggs are

cylindrical, whitish or

pale-yellow and later

become brown with red

eyespots when the

embryos develop. The

Figure 4. Life Cycle of Green Leafhopper
eggs are laid in masses

of 8-16 eggs and hatch within about 4-8 days. Upon hatching the nymph measures

0.9 mm long with whitish and shiny appearance and as it matures, it turns

yellowish. The shape of the nymph is similar to that of the adult except that the

nymph is smaller and is wingless (Estoy A, 2000). After about 10 to 14 days after

hatching the leafhopper turns to adult which has green appearance with black

markings on the head, thorax, and abdomen. A pair of black spots is either present

or absent on the forewings. The adult insect is 4.2-4.3mm long and the total life
 15

cycle from egg to adult is between 22-26 days (Joshi R, 2001). Green

Leafhoppers are most destructive at adult stage due to increased nutrient

consumption from rice plant in preparation for mating and reproduction (Shepard

BM, et al. 1995)

Sevin ® 85 WP

Sevin ® 85 WP is a wide-spectrum insecticide which controls over 100

species of insects on fruit, cotton and other crops as well as on poultry and pets.

Its active ingredient is carbaryl (C12H11NO2) from the chemical group

carbamate (1-naphthyl-N-methyl) (Instant EPA's Pesticide Facts.1996). It kills

insects by disrupting the normal function of their nervous system. Nerve impulses

are transmitted from one nerve to another by a chemical called acetylcholine.

Under normal conditions an enzyme called acetylcholinesterase destroys

acetylcholine so that another nerve impulse may be transmitted. Carbaryl stops

the functioning of the enzyme so acetylcholine builds up resulting in restlessness,

paralysis and death. Carbaryl affects human nerves in the same way as in insect

nerves. However in insects it affects the central nervous system while in humans

it affects the nerve and muscle interaction. Thus carbaryl poisoning symptoms are

different in humans and in insects (Ware G, 2000).

Research Literature

In a study conducted by Dasmariñas, Jover and Pasig (2001) Pansit-

pansitan was used as insecticide for termites. According to the study the plant

contains considerable amounts of monoterpenes, flavonoids, amides and

alkaloids. The chemicals mentioned are known to have insecticidal property in

 16

which it disrupts the nerve and motor functioning of insects thus causing death. In

the experiment mentioned, the researchers prepared four treatments: Treatment A

consisting of 100% Pansit-pansitan extract, Treatment B with 75% Pansit-

pansitan extract, Treatment C with 50% Pansit-pansitan extract, Treatment D with

25% Pansit-pansitan extract and Treatment E as positive control consisting of

unknown commercial pesticide. The treatments were tested on termites randomly

collected and placed in containers with 20 termites in each and three replicates are

made per treatment. Number of mortality was counted every 3min until 15min

was achieved. Results showed that Treatment A (100% Pansit-pansitan extract)

showed the highest mortality rate of 100% compared with the other plant extracts

and is comparable with Treatment E which is the positive control which also

showed 100% mortality rate.

Pansit-pansitan however has lethal activity according to Lim (2006). The

researcher assessed the plant’s lethal activity using the Brine Shrimp (Artemia

salina) Lethality Bioassay. The testing involved the exposure of the organism to

different aqueous extract concentrations of Pansit-pansitan (10%, 30% and 50%)

in three serial dilutions. Mortality among organisms was observed for 48 hours in

experimental and control groups in three replications and LC50 value was

computed using the Spearman-Karber Method. Significant differences among

sample observations were determined by employing One-Way Analysis of

Variance. Lethal activity of the test plant on the invertebrate model was proven

with 9.18 LC50 value and 50 percent extract concentration as the most lethal

treatment.
 17

Another study by Cambe, Delfin and Sales, (2003) is aimed to find out if

oleander (Thevetia peruviana) fruit extract can be used as insecticide against rice

bugs. The experimental design was Completely Randomized Block Design

(CRBD) with the following treatments: Treatment A - (25 ml extract + 25 ml

water), Treatment B - (50 ml extract + 25 ml water), Treatment C - (75 ml extract

+ 25 ml water), Treatment D - (100 ml extract), Controlled set-up (100% water)

and replicated three times. The crude extract was obtained by pounding,

squeezing, and filtering. Phytochemical analysis revealed that extract of the young

fruit contains alkaloid, thevetin, peruvoside and cyanogenic glycosides. The

chemicals have influenced the effectiveness of extract and using greater

concentrations can hasten and increase mortality to 100%. Using the One-way

Analysis of Variance and Duncan’s Test, results led to the conclusion that

oleander can be used as an insecticide against rice bugs.

 18

CHAPTER 3

RESEARCH METHODOLOGY AND PROCEDURE

This chapter presents the research design, methods and adapted procedures in the

study.

Research Design

An experimental method was used in the study to determine the insecticidal

property of different concentrations (50%, 75% and 100%) of Pansit-pansitan leaf

crude extract against Green Leafhopper.

This experiment used three treatments. Different concentrations (50%,

75% and 100%) of the plant’s leaf material have been prepared by dissolving 5g,

7.5g and 10g of Pansit-pansitan leaf crude extract in 10ml of distilled water,

respectively. Sevin ® 85 WP was used as positive control. The positive control

was prepared by dissolving 10g of the insecticide to 10ml of distilled water.

Research Setting

This study along with the collection of plant material and the culturing of the

test organism was conducted at the Dalisay Piggery Farm Compound located in

Lian, Batangas. Phytochemical analysis was done at the IHS Laboratory,

University of Perpetual Help System DALTA located in Las Piñas City. The leaf

crude extract was tested for the following:

 19

Test for Tannin

A few drops of Ferric Chloride T.S. was added to 2ml plant extract.

Test for Alkaloids

The addition of 1% HCL was used to modify the aqueous extract when one

or two drops of Mayer’s reagent was added.

Grease Spot Test

The plant material was pressed on a bond paper. A temporary presence of

grease spot indicates that the plant material contains volatile oil while

permanent spot indicates the presence of fixed oil.

The study was conducted from December, 2009 to January, 2010.

Research Procedure

Collection and Preparation of Plant Specimen

The plant material was collected at the Dalisay Piggery Farm Compound

in Lian, Batangas. The collected plant material was washed with distilled water

to remove dust and insects and air-dried for three days. Afterwards, it was pre-

weighed prior to submission for phytochemical analysis.

Identification of Plant Material

The plant material was initially identified by the researcher and verified

by Mr. Wilfredo F. Vendivil Ph.D, Senior Researcher of Botany Division,

National Museum located at P. Burgos St. Manila.

 20

Identification of Test Organism

The test organism was identified by Mrs. Fe D. Alzona, Researcher of

the College of Agriculture, Crop Protection Cluster, University of the

Philippines Los Baños located at Los Baños, Laguna.

Extraction Procedure

Decoction Method

The researcher used decoction method in order to obtain the leaf crude

extract. Five hundred ml of water was boiled first before adding the leaves. The

chopped leaves which weighed 500g was added. The heat was lowered to 50-

60 °C in order to maintain a slow simmer for 10 to 15 minutes. It was removed

from heat and strained before using.

Preparation of Different Concentrations of Leaf Crude Extract

The preparation of varying concentrations (50%, 75% and 100%) of

Pansit-pansitan leaf crude extract were as follows: for 50%, 5g of leaf extract

was dissolved in 10ml of distilled water, for 75%, 7.5g of leaf extract was

dissolved in 10ml of distilled water and for 100%, 10g of leaf extract was

dissolved in 10ml of distilled water.

Preparation of Sevin® 85 WP (control)

Ten grams (10g) of the powdered form of the control (Sevin 85 WP) was

dissolved in 10ml of distilled water.

 21

Preparation of the Test Organism

Preparation of Green Leafhopper Cage

Sixteen cages were prepared on the natural habitat of the test organism

using a fine net firmly paced on the site by supporting wooden pegs. The first

two cages were used to house the insects in the wild which measured 1m x 1m

x 0.46m. The second two cages were used to house the insect eggs laid by the

previous generation which have the same dimension as the cages that were

used previously. The remaining twelve cages which measured 0.30m x 0.30m

x 0.15m were then used to house the cultured adult forms of Green Leafhopper

in accordance to the number of treatments, trials and control that were used

(Hou, 1987 modified).

Collection of Green Leafhopper

Random wild Green Leafhoppers were collected at Dalisay Piggery

Farm Compound in Lian, Batangas. Insects were collected by constructing

cages on the natural habitat and were cultured to ensure uniformity.

Preparation of Green Leafhopper for Mating

Random Green Leafhopper adults were collected from the wild by

constructing 2 cages which measured 1m x 1m x 0.46m on the natural

environment of the insects. Green Leafhopper eggs were collected after 6 days

by gently brushing the rice plants located on the site with a soft brush to

remove the adult insect thus the insect eggs remained in the rice plant. The rice

plants along with the Green Leafhopper eggs were carefully transferred to
 22

another 2 cages in the natural habitat which measured the same as the other 2

cages that were used previously and was allowed to reach adulthood. After 11

to 13 days mature insects were collected and distributed to the remaining 12

cages which measured 0.30m x 0.30m x 0.15m in accordance with the number

of treatments, trials and control that were used (Hou, 1987 modified).

Application of Leaf Extract to Green Leafhopper

Spray method was done to apply the different treatments using atomizer.

About 10ml of each treatment was sprayed to the respective cages containing

10 cultured Green Leafhoppers. Each treatment had three replications, thus the

total number of cultured Green Leafhoppers used was 120 including the

control. Higher mortality rate of the insect indicates the effectiveness of the

plant extract. The mortality rate of Green Leafhopper was determined after the

extract was applied through spray method at intervals of every 6 minutes for a

total of 30 minutes.

Statistical Treatment

One way analysis of variance (ANOVA) was used to determine the

significant difference in the insecticidal property between Pansit-pansitan

(Peperomia pellucida (Linn.HBK)) leaf crude extract and the positive control

Sevin ®85 WP.

 23

CHAPTER 4

PRESENTATION, ANALYSIS AND INTERPRETATION OF DATA

This chapter presents the data gathered, the analysis of data and the different

findings together with their respective analysis and interpretation of results.

In this study the insecticidal property of Pansi-pansitan leaf crude extract

was tested against Green Leafhopper through spray method. The presentation of

data includes the results of the phytochemical analysis, mortality rate of Green

Leafhopper using different concentrations of Pansit-pansitan leaf crude extract

and Sevin ®85 WP (control) and the result of the ANOVA test.

Table 1. Results of Phytochemical Analysis of Pansit-pansitan (Peperomia

pellucida (Linn.HBK)) Leaf Crude Extract

Tests Theoretical Result Actual Result

Tannins + +

Alkaloids + +

Grease Spot + (volatile oil) + (volatile oil)

Table 1 shows the different constituents present in Pansit-pansitan leaf

extract based on phytochemical analysis. Active components present were tannins

which were indicated by blue to black precipitate, alkaloids which was indicated

by the presence of precipitate and volatile oil which was indicated by a temporary

grease spot. The result justifies the research by De Padua LS and Pancho JV

(1983) on the chemical components of Pansit-pansitan in which the results also

showed the same active components.

 24

Table 2. Mortality Rate of Green Leafhopper Using Different Concentrations

of Pansit-pansitan Leaf Crude Extract and Sevin ® 85 WP (Control)

Treatments Time in Minutes

6 12 18 24 30

Sevin 85 WP 10% 26.67% 33.33% 16.67% 6.67%

(control)

100% 6.67% 13.33% 20% 6.67% 0%

75% 3.33% 10% 16.67% 3.33% 0%

50% 0% 3.33% 10% 6.67% 0%

Table 2 shows the mortality rate of Green Leafhoppers using varying

concentrations of Pansit-pansitan leaf crude extract and Sevin ® 85 WP (control).

Based on the results, the positive control Sevin ® 85 WP showed a mortality

rate of 10% after 6 minutes of application, 26.67% after 12 minutes, 33.33% after

18 minutes, 16.67% after 24 minutes and 6.67% after 30 minutes of application.

The 100% concentration of the leaf crude extract showed a mortality rate of

6.67% after 6 minutes of application, 13.33% after 12 minutes, 20% after 18

minutes, 6.67% after 24 minutes and 0% after 30 minutes of application. The 75%

concentration of the leaf crude extract showed a mortality rate of 3.33% after 6

minutes of application, 10% after 12 minutes, 16.67% after 18 minutes, 3.33%

after 24 minutes and 0% after 30 minutes of application. The 50% concentration

of the leaf crude extract showed a mortality rate of 0% after 6 minutes of

application, 3.33% after 12 minutes, 10% after 18 minutes, 6.67% after 24

minutes and 0% after 30 minutes of application.

 25

It was observed that there is an increasing trend of mortality rate of Green

Leafhopper up to 18 minutes, a decrease in mortality rate after 24 minutes of

application and at 30 minutes, all the different concentrations of Pansit-pansitan

leaf crude extract showed 0% mortality rate. The positive control Sevin ® 85 WP,

showed 6.67% mortality rate from 6minutes onwards to 30 minutes after

application.

It can be inferred from the data that both the positive control, Sevin ® 85

WP and the different concentrations of Pansit-pansitan leaf crude extract provide

optimum mortality rate at 18 minutes after application of Pansit-pansitan leaf

crude extract on Green Leafhoppers.

 26

35

30

25

20
Mortality Sevin ® 85 WP
Rate 100%
15 75%
50%

10

0
6 minutes 12 minutes 18 minutes 24 minutes 30 minutes
Time Interval

Figure 5. Mortality rate of Green Leafhopper using varying

concentrations of Pansit-pansitan leaf crude extract and
Sevin 85 ® WP (control)

Figure 5 shows the mortality rate of Green Leafhoppers using varying

concentrations of Pansit-pansitan leaf crude extract and Sevin ® 85 WP (positive

control). The red column represents Sevin 85 ® WP (positive control), the yellow

column represents 100% concentration of Pansit-pansitan leaf crude extract, the

green column represents 75% concentration of Pansit-pansitan leaf crude extract

and the blue column represents 50% concentration of Pansit-pansitan leaf crude

extract.
 27

Based on the figure, the positive control Sevin 85 ® WP and the varying

concentrations (100%, 75% and 50%) of Pansit-pansitan leaf crude extract

showed the highest mortality rate of 33.33%, 20%, 16.67% and 10% respectively

after 18 minutes of application. The lowest mortality rate of 6.67%, 0%, 0% and

0% was attained for the positive control Sevin 85 WP and the varying

concentrations (100%, 75% and 50%) of Pansit-pansitan leaf crude extract

respectively which occurred after 30 minutes of application.

Table 3. Summary of Computation of ANOVA Test to Determine the

Significant Difference of Pansit-pansitan Leaf Extract and Sevin 85
WP (Control)

Source df SS MS F Critical Decision Conclusion

value
of F

Treatments 3 55 18.333

Error 16 88.8 5.55 3.303303 3.24 Reject Significant

Ho
Total 19 143.8

Table 3 shows the computed value of Pansit-pansitan leaf crude extract and

Sevin 85 ® WP (control). Since the computed F value of 3.303303 is greater than

the critical F value which is 3.24 at 0.05 level of significance, the null hypothesis

is rejected. This means that there is a significant difference in the insecticidal

property between Pansit-pansitan leaf crude extract and the positive control, Sevin

® 85 WP.
 28

CHAPTER 5

SUMMARY, CONCLUSION AND RECOMMENDATION

This chapter discusses the summary, conclusions and recommendations of the

study.

Summary of Findings

This study entitled “The Insecticidal Property of Pansit-pansitan (Peperomia

pellucida Linn. HBK) Leaf Crude Extract against Green Leafhopper (Nephotettix

malayanus Ishihara et Kawase)” was conducted to confirm the active components

present in Pansit-pansitan leaf crude extract using phytochemical analysis,

determine the insecticidal property of Pansit-pansitan based on the mortality rate

of Green Leafhopper, determine the insecticidal property of Pansit-pansitan based

on the mortality rate of Green Leafhopper and lastly to compare the different

concentration of Pansit-pansitan leaf crude extract with Sevin ® 85 WP (positive

control).

Pansit-pansitan leaves were collected at the Dalisay Piggery Farm

Compound in Lian, Batangas. The collected plant material was washed with

distilled water to remove dust and insects and air-dried for three days.

Phytochemical analysis was conducted at the IHS Laboratory, University of

Perpetual Help System DALTA located in Las Piñas City. This experiment has

used three treatments. Different concentrations (50%, 75% and 100%) of the

plant’s leaf material have been prepared by dissolving 5g, 7.5g and 10g of Pansit-

pansitan leaf crude extract in 10ml of distilled water, respectively. Sevin ® 85 WP

 29

had been used as positive control by dissolving 10g of the powdered form of the

insecticide to 10ml of distilled water.

Based on the results of the phytochemical analysis, Pansit-pansitan leaf

crude extract contains alkaloids, tannins and volatile oils.

The effects of the varying concentrations of Pansit-pansitan leaf crude

extract on mortality rate of Green Leafhopper showed that the highest mortality

rate obtained was that of the control (Sevin ® 85 WP) which has 28 deaths or

93.33% followed by the 100% concentration of Pansit-pansitan leaf crude extract

which has 14 deaths or 46.67%. The least mortality rate of Green Leafhoppers has

been observed at 50% concentration of Pansit-pansitan leaf crude extract which

only showed 6 deaths or 20%. The ANOVA test reveals that the null hypothesis

was rejected. There is a significant difference in the insecticidal property between

Pansit-pansitan leaf crude extract and the control, Sevin 85 ® WP.

Conclusions

Based on the above findings, the following conclusions were drawn:

1. active components present in Pansit-pansitan leaf crude extract are

alkaloids, tannins and volatile oil;

2. varying concentrations of Pansit-pansitan leaf crude extract on mortality

rate of Green Leafhoppers showed that the highest mortality rate

obtained was that of the control, Sevin ® 85 WP which showed 28

deaths or 93.33% followed by the 100% concentration of Pansit-pansitan

leaf crude extract which has 14 deaths or 46.67%. The least mortality
 30

rate of Green Leafhoppers has been observed at 50% concentration of

Pansit-pansitan leaf extract which only showed 6 deaths or 20%; and

3. there is a significant difference in the insecticidal property between

Pansit-pansitan leaf crude extract and the positive control, Sevin 85 ®

WP.

Recommendations

Based on the experiment results, the researcher would like to put forward

the following recommendations:

1. a study regarding the insecticidal property of Pansit-pansitan on the

different stages in the life cycle of Green Leafhoppers must be

conducted;

2. test other parts of Pansit-pansitan plant for insecticidal property;

3. test other local plants that may have insecticidal property;

4. try other extensive methods of plant extraction; and

5. a thorough study on the physiological responses of Green

Leafhopper at different time of application.

 1

BIBLIOGRAPHY

Cambe R, Delfin NC,Sales LO. 2003. Insecticidal Effect of Yellow Oleander

Fruit Extract on Rice Bugs. Capiz National High School

Dales MJ. 1996. A Review of Plant Materials Used for Controlling Insect
Pests. U.J Natural Resources Institute, Kent U.K p.84.

Dasmariñas JT, Jover EM, Pasig KC. 2001. Peperomia pellucida for Termite
Control. University of Southern Mindanao Kabacan, Cotabato

De Padua LS, Pancho JV. 1983. Handbook on Philippine medicinal plants,

Volume IV. Los Baños: UPLB Documentation and Information Section,
Office of the Director of Research.

Estoy AB., LB Flor, EM Valdez et al. 2000. Occurance of Major Rice Insect
Pest in PhilRice Stations. Philippine Rice Research Institute (PhilRice).

Harborne Jeffery, H. Baxter. 1983. Phytochemical Dictionary: A Handbook of

Bioactive Compounds from Plants. Taylor & Frost, London. 791 pp.

Hou R. F. 1987. Mass Rearing of Green Rice Hopper (Nephotettix cincticeps).

International Rice Research Institute

Joshi R, Estoy A, Arida G et al. 2001. Rice Technology Bulletin No: 37.
Philippine Rice Research Institute (PhilRice).

Latorre P-Jay D. 2005. Insecticidal Property of Acacia Seeds and Bark

Against Termites. Bureau of Soils Lipa City

Manalo JB, Byung HH, Yong NH, et al. 1983. Studies on Ether-Soluble Neutral
Compounds of Peperomia pellucida. Annual Report of Natural Products
Research Institute, Seoul National University; 22: 16-9.
 1

Manalo JB, Recaña MP, Coronel VQ et al. 1984. Potassium Nitrate from
Peperomia pellucida (L) HBK. NIST J. 1:25-31

Murray G. and Frank Morehouse. 1973. Organic Molecules in Action. Gordon

and Breach.

Pathak MD, Khan ZR. 1994. Insect Pests of Rice. Manila (Philippines):
International Rice Research Institute. 89 p.

Pathak MD, Vea E, John VT. 1967. Control of Insect Vectors to Prevent Virus
Infection of Rice Plants. J. Econ. Entomol. 60:218-225.

Reissig WH, Heinrichs EA, Litsinger JA, Moody K, Fiedler L, Mew TW, Barrion
AT. 1986. Illustrated Guide to Integrated Pest Management in Rice in
Tropical Asia. Manila (Philippines): International Rice Research
Institute. 411 p.

Shepard BM, Barrion AT, Litsinger JA. 1995. Rice-Feeding Insects of Tropical
Asia. Manila (Philippines): International Rice Research Institute. 228 p

Ware, G.W. 2000. The Pesticide Book. Fresno, CA: Thomson Publications. p.
57, 83.

Unpublished sources

Manalo JB. 1983. A Phytochemical Investigation of Pepercmia pellucida (L)

HBK.
UP Manila

Internet sources

Bayer CropScience Chile

(http://www.bayercropscience.cl/soluciones/fichaproducto.asp?id=123)

DOST Investigatory Projects

(http://www.investigatoryprojectexample.com/search/)
Dr. Dukes Phytochemical and Ethnobotanical Databases
 1

(http://www.ars-grin.gov/cgi-bin/duke/farmacy2.pl)
Instant EPA's Pesticide Facts
(http://www.instantref.com/carbaryl.htm)

Juniper and Sage Essential Oils

(http://www.juniperandsage.com/oils/toxicity.php)

Lans. 2007. Journal of Ethnobiology and Ethnomedicine 2007 3:3 doi:

10.1186/1746-4269-3-3
(http://www.ethnobiomed.com/content/3/1/3/table/T4)

Pesticide Management Education Program (PMEP), Cornel University Ithaca, NY

(http://pmep.cce.cornell.edu/profiles/extoxnet/carbaryl-dicrotophos/carbaryl-
ext.html#top)

Sri Lanka Department of Agriculture

(http://www.agridept.gov.lk/institutes_more_sub_links.php?id=184&sMenu=Nati
onal+Botanic+Gardens+(NBG+)

United States Department of Agriculture Natural Resource Conservation Service

(http://plants.usda.gov/java/profile?symbol=PEPE5)

United States National Library of Medicine

(http://chem.sis.nlm.nih.gov/chemidplus/ProxyServlet?objectHandle=DBMaint&
actionHandle=default&nextPage=jsp/chemidheavy/ResultScreen.jsp&ROW_NU
M=0&TXTSUPERLISTID=000523808)

Virginia Tech's Entomology Department. Species of leafhoppers.

(http://everest.ento.vt.edu/~idlab/vegpests/vegi/LH.jpeg)

http://www.herbs2000.com/h_menu/tannins.htm

http://www.oisat.org/pests/insect_pests/hoppers/glh.html
 1

PLATES
 1

Plate 1. Phytochemical Analysis of Plant Material

 1

Plate 2. Materials Used in the Conduct of the Experiment

 1

1. fresh Pansit-pansitan leaves 2. dried Pansit-pansitan leaves

4. filtrate of Pansit-pansitan being 3. dried Pansit-pansitan leaves

evaporated boiled in water

5. Pansit-pansitan leaf crude

extract

Plate 3. Extraction Procedure Done for Pansit-pansitan Leaves

 1

1. cages for containing wild Green 2. cages for containing insect eggs
Leafhoppers

3. cages for containing cultured insects

Plate 4. Green Leafhopper Cages in the Rice Field

 1

Plate 5. Application of Varying Concentrations of Pansit-pansitan Leaf

Crude Extract to Cultured Insects
 1

APPENDICES
 1

Appendix A
Data showing the number of Green Leafhoppers died in varying
concentrations of Pansit-pansitan leaf crude extract and the control (Sevin ® 85
WP) at different time intervals.

Treatments Number of Number of Dead Total Total

Test Leafhoppers Based on Time Number Number
Organisms (minutes) of of Deaths
Deaths in
6 12 18 24 30 Percent

Sevin ® 85
WP
(control) 30 3 8 10 5 2 28 93.33%

50% 30 0 1 3 2 0 6 20%

75% 30 1 3 5 1 0 10 33.33%

100% 30 2 4 6 2 0 14 46.67%
 2

Appendix B
Outline of Research Procedure

Research Procedure Overview

C ollection of Preparation of Different
Plant Material C oncentrations of Leaf C rude Extract

Phytochemical grease Preparation of

Analysis spot test Test Organisms

test for test for preparation of green

tannin alkaloid leafhopper cages

preparation of green
leafhopper for mating

S tatistical Application of L eaf Extract

Treatment to Green Leafhopper
 3

Appendix C
Certification of Borrowing of Laboratory Equipment

University of Perpetual Help – Las Piñas

Pamplona, Las Piñas City

TO: DR. VALERIANO V. TIMBANG JR.

Director, IHS Laboratories
This University
THRU: MRS. CLARISSE ROMULO
Laboratory Coordinator
SUBJECT: As stated
DATE: November 21, 2009
Greetings!
We are fourth year BS Biology students from this University and we are
currently enrolled in Thesis Writing. It is a requirement in this subject to conduct
an experiment/analysis about biological problems. With this regard, may we ask
your good office to allow us to use/borrow some materials and equipments needed
for the conduct of our methodology.
Attached herewith is the list of materials to be used for the different
experiments with the planned schedule.
Hoping for your kind consideration. Thank you so much.

Truly yours

DALISAY, GERARD MAJELL

MOLDEZ, MONIQUE
VALLOSO, JOANNA MARIE
Noted:
MRS. MA.LOURDES C. AGUIRRE
Adviser
 4

DATE LIST OF QUANTITY CHEMICALS

MATERIALS
December 9, Centrifuge 1 Trisodium citrate
2009 (anhydrous)
7:30-10:30 Micropipettor 1
(Class of Mr. Retardo- Test tube 1
Morphoanatomy)

Mortar and pestle 1 Ferric chloride

Water bath 1 Lead acetate
Bunsen burner 1 HCl
Erlenmeyer flask 1 Mayer’s reagent
Filter paper Sodium Carbonate
Beaker(50 ml and 2 10% NaOH
100ml)
Litmus paper (blue
and red)

Blender 1 Ethyl Alcohol (95%)

Funnel 1 Distilled water (100mL)
Filter paper
Water bath 1
Bunsen burner 1
Beaker(100ml) 2
Erlenmeyer flask 2
Evaporating dish 1
Test tubes (big) 5
Pipette 5
aspirator 2
Test tube rack 1
 5

Appendix D
Certification of Statistical Analysis of Data

February 1, 2010

CERTIFICATION

This is to certify that the study entitled “The Insecticidal Property of Pansit-
pansitan (Peperomia pellucida (Linn.) (HBK)) Leaf Crude Extract Against
Green Leafhopper (Nephotettix malayanus (Ishihara et Kawase))” conducted
by Mr. Gerard Majello B. Dalisay was statistically analyzed by the undersigned:

MR. ARTURO TOBIAS CALIZON

Faculty, Mathematics Department

Noted:

MRS. AIDA REGACHO

Chairman, Mathematics Department

 6

Appendix E
Certification of Identification of Plant Material
 7

Appendix F
Certification of Identification of Test Organism
 8

CURRICULUM VITAE

Personal Background

Name: Gerard Majello B. Dalisay

Age: 19 years old

Birthday: April 26, 1990

Address: Herrera Subdivision Lian, Batangas

Name of Father: Honorio L. Dalisay

Name of Mother: Victoria B. Dalisay

Educational Background

School Year

Elementary Nasugbu East Central School 1996-2002

High School Blessed Christ Child Montessori Foundation 2002- 2006

College University of Perpetual Help System DALTA 2006- present