Centella asiatica (Gotu kola) extract as an Insecticide for Mosquitoes

A Capstone Project

Presented to the faculty of

Senior High School Department

PHINMA- Cagayan de Oro College

In Partial Fulfillment

Of the Requirements for the course

STM 009: Research In Science, Technology, Engineering and Mathematics

By: FC1 – G12 – 01 – STEM HEALTH Group 1

Aligsao, Jestoni Jones Bolanio, Thea Teressa Edaño, Myka

Amolata, Michelle Calaurian, Chirstopherson Edio, Van Robert

Asum, Walida Hadji Codilla, Jink Fair Enguito Clive Owen

Bantuas, Abdul Rauf Cuevas, Kim Estoque, Donna Mae

Bantuas, Haneya Atheya Daleon, Venz Gaburno, Claudeth Monica

Bertoldo, Arabela Gift Ann Donglas, Christeah Faith Pascua, Joshua

 Chapter I

INTRODUCTION

The Problem

Centella asiatica (Gotu kola), is a medicinal plant that has been used for

thousands of years in both traditional and modern medicine. Pentacyclic

triterpenes, primarily asiaticoside, madecassoside, asiatic, and madecassic acids,

are among the active substances (Chandrika & Kumara, 2015). The Apiaceae

family includes Centella asiatica, often known as Gotu kola or Tiger Grass. It

thrives in tropical areas of Oceania and Asia, primarily in India, Pakistan,

Madagascar, equatorial Africa, and Central America. Three thousand years ago,

China, India, Africa, the Philippines, Sri Lanka, and Madagascar began using the

Centella asiatica herb as a "panacea" (Vasantharuba et al., 2012). Almost

everyone has endured the uncomfortable sensation of a mosquito bite. The red

mark and itching from mosquito bites are the result of an allergic reaction to the

saliva of the insect. The red mark and itching from mosquito bites are the result of

an allergic reaction to the saliva of the insect. Different forms of standing water are

preferred by various mosquito species for egg laying. In permanent ponds, lakes,

and streams, the presence of beneficial predators like fish and dragonfly nymphs

helps keep these bodies of water largely free of mosquito larvae.

There were 9,482 dengue cases and 16 fatalities as of epidemiological

week of 42 of 2022. In Cambodia, (CFR) 0.2% Were reported each week than the

average cases throughout the same period from 2015 to 2020 in 2022. There were
a total of 413 dengue cases and two fatalities reported during epidemiological week

of 43 of 2022. When compared to the same period in 2021, the number of instances

is 73% lower, but it was reported cases for week 43 in 2022 could change as they

are still being verified and authenticated. As of January 1 there have been 193,010

dengue cases and 629 fatalities (CFR 0.3%) as of 29 October

2022 week 43 compared to the 65,684 instances reported in the same period in

2021 is 194% higher (World Health Organization, 2022). In Singapore, 322 dengue

cases were reported during epidemiological week 44 of 2022, with a total of 29,894

cases so far this year compared to 4,799 dengue cases, this is a 523% increase

cases reported in 2021 (World Health Organization, 2022). Situation in the

Philippines in 2022, country health authorities report 160,594 cumulative cases

through September 10, a 189 percent increase over the number of cases reported

during the same period in 2021 (55,650).Central Luzon accounts for 20 percent of

the cases nationally with 31,730. This is followed by the National Capital Region,

which includes the city of Manila with 16,413 cases, and the Central Visayas, which

includes Cebu, with 13,740 cases. Health officials also report that dengue-related

fatalities have risen to 507 total. The Central Visayas has reported the most deaths

with 84, followed by Central Luzon with 71 (News Desk, 2022).

The researchers purpose of this study is to develop an insecticide with

active toxic agents from a plant extract called Centella asiatica. A non-toxic, easily

accessible, biodegradable substance with wide target-specific activity against

various species of vector mosquitoes. To avoid potential risks, the researchers

intend to look for an alternative insecticide to use.

Conceptual Framework

The unprocessed leaf extracts of Centella asiatica The mosquito Culex

quinquefasciatus has been discovered to be susceptible to asiatica's larvicidal and

adult emergence inhibitory activities. The phenolics, terpenoids, and alkaloids

present in plants may work together or separately to generate larvicidal and adult

emergence inhibitory action against Culex quinquefasciatus may be the cause of

the plant extract's biological activity. Centella asicatica elegans leaf extract

Centella asiatica is better than several neem preparations, which are allegedly

effective at LC50 values between 55 and 65 anti-mosquito larvae ppm (Ascher &

Meisner, 1989). The average fatal concentrations (LC50) of different Melia

azederach parts 30 to 40 ppm of Melia azederach against larva (Culex pipiens)

(Al-Sharook et al., 1991). The effect of different neem extracts and different Melia

azderach's lower portions were slightly lower than for which a report was made leaf

extract from Centella asiatica. The Activity of Centella asiatica that inhibits adult

emergence Asiatica is also comparable to various plant species and different

families of extract (Muthukrishnan et al., 1999). The research's findings at this time

revealed that Centella asiatica elegans leaf extract exemplary larvicidal and adult

efficacy mosquito emergence inhibition activity Quinquefasciatus, the culex.

Additional investigation for a better understanding of this behavior versus a variety

of mosquito species, as well as the extract's active component(s) responsible for

Adult emergence inhibition activity and larvicidal activity Identification of Culex

quinquefasciatus is necessary and, if possible, used in creating a commercial item

or formulation intended for use as mosquitocida.

Centella asiatica leaf extract shows promise as a larvicide and adult

emergence inhibitor against Culex quinquefasciatus and could be used directly in

small-volume aquatic habitats or breeding sites of limited size close to human

habitations. Centella asiatica leaves were evaluated for larvicidal and adult

emergence inhibition activity against mosquitoes (Chinnu et al., 2018).

Schematic Diagram

Independent Variable Dependent Variable

Centella asiatica extract: Numbers of Mosquitoes being

• 5ml killed:

• 10ml • Time duration

• 15ml • Mortality rate
• Intensity

Figure 1. The Centella asiatica extract and the indicators of its effectiveness.
Objectives

General Objective

To determine the effectivity of Centella asiatica as an insecticide.

Specific Objectives

1. To determine the amount of each solution going to be sufficient to eliminate

the mosquitoes:

1.1 5ml

1.2 10ml

1.3 15ml

2. How long does it take for the mosquitoes to die at this exact rate in terms

of these measurements:

2.1 1 hour

2.2 2 hours

2.3 3 hours

3. Is Centella asiatica extract impactful against mosquitoes as an insecticide?

Null Hypothesis

There is no significant effect between centella asiatica and mosquitoes.

Significance of the Study

The outcome would be highly beneficial to anybody, particularly in the field

of health. Furthermore, this research would provide new knowledge about Centella

asiatica. This study discusses Centella asiatica as an Insecticide for Mosquitoes

as well as its benefits are the following areas:

School. This research will be beneficial to our school, PHINMA Cagayan de Oro

College because we can educate them on Centella asiatica as an Insecticide, and

it could preferably benefit PHINMA-COC Senior High School STEM HEALTH

students.

Teacher. The study will be beneficial to the teachers to have a deeper

understanding of this study and for them to promote the use of medicinal plants,

especially Centella asiatica as an insecticide for mosquitoes. The fact that a

teacher is willing to broach this subject in an open and general fashion can be

perceived as a safe invitation for students who want to experiment.

Students. This research will contribute to students of PHINMA-COC. Through this

research, students will have new knowledge about Centella asiatica. With help of
this study, the student will further realize promoting the use of medicinal plants,

especially Centella asiatica as an insecticide for mosquitoes.

Future Researchers. This study will be beneficial to future researchers at PHINMA

Cagayan de Oro College. To conduct an experimental study that would help future

researchers to provide supplements information and analysis, that can help them

as a future reference for further studies in the future. This study covers information

involving Centella asiatica as an Insecticide for Mosquitoes.

Scope and Delimitation of the Study

This experimental study would mainly focus on obtaining valuable

statistical data on the effects of Centella asiatica as an alternative insecticide for

mosquitoes. The samples tested will be randomly captured along selected sites

along the campus premises of PHINMA Cagayan De Oro College without any

target for specific species of mosquitoes. On the other hand, the insecticide used

will be made through pure extracts extracted from the leaves of Centella asiatica

plant without any other chemicals incorporated.

This study would be conducted through experimentation in a home based

setting wherein the researcher would configure each and every set-up trials into

the desired isolated environment. Therefore, it may result to an outcome that may

not generalize well in every condition and every species of mosquito. It would also

not extend to other problems that may possibly arise within the duration of the

experimentation. It won’t cover any other topics aside from the effectiveness of the
extracted Centella asiatica on mosquitoes under trial alone. The research would

be expected to run up until the year of 2023

Definition of Terms

Alternative - There are two possible actions, and alternative is one of them.

Alternatives are always equal from a particular perspective-taking objective.

Alternative is the concept of logical disjunction in both mathematics and logic.

Allergic Reaction – An allergic reaction occurs when the immune system

overreacts to a harmless substance known as an allergen. The immune system

protects the body from infections, viruses and diseases.

Biodegradable Substance – are organic waste matters that can be broken down

into compost or simple organic molecules by biological means. These substances

can decompose due to the action of microorganisms. Examples includes fruits,

vegetables, human waste, etc.

Broach - to initiate a conversation on a certain topic. To broaden and addressed

a given topic thoroughly.

Centella asiatica - Centella asiatica, commonly known as Gotu kola, kodavan,

Indian pennywort and Asiatic pennywort, is a herbaceous, perennial plant in the

flowering plant family Apiaceae. It is indigenous to the tropics of Australia, Asia,

Africa, and the western Pacific Ocean islands. It is consumed as a culinary

vegetable and is used in traditional medicine.

Culex quinquefasciatus - known as the southern house mosquito that is found in

tropical subtropical regions of the world. Use as one of the example for testing a

insecticide.

Conducted - to carry out a particular activity or process, especially in order to get

information or prove facts

Develop – means to build up, grow, or improve gradually over time.

Extracted - to carefully remove a substance from something which contains it,

using a machine, chemical process

Insecticide - Insecticides are substances used to kill insects. They include ovicides

and larvicides used against insect eggs and larvae, respectively. Nearly all

insecticides have the potential to drastically affect ecosystems; many are

hazardous to humans and animals; some grow concentrated as they propagate

down the food chain.

Medical Plants – Medical Plants is any plant that has compounds that can be

utilized therapeutically or that serve as building blocks for the production of

effective pharmaceuticals is considered to be a medicinal plant. Since ancient

times Medical Plants, also known as Medicinal Herbs have been identified and

employed in traditional medical procedures.

Melia azedarach - known as the chinaberry tree. Being experimented as a

insecticide.

Mosquitoes - Mosquitoes are members of a group of almost 3,600 species of

small flies within the family Culicidae. The word "mosquito" is Spanish for "little fly".

Mosquitoes have a slender segmented body, one pair of wings, one pair of halters,

three pairs of long hair-like legs, and elongated mouthparts.

Predator/s – a predator is an animal that kills and eats other animals, and the

threat it poses is relatively clear-cut.

Researcher - an individual who conducts academic or scientific research.

Someone who studies a topic, especially to learn new information or develop new

insights.
 CHAPTER 2

REVIEW OF RELATED LITERATURE

This chapter discusses the review of literature and studies that are closely

connected to the experiment that will be performed.

Centella asiatica (Gotu Kola) is a prized traditional medicine as well as a

nutritionally significant plant. The review discusses the chemistry, nutritional value,

and health advantages of Centella asiatica have been thoroughly covered to

highlight, its use as a traditional food and medicinal. Due to its high concentrations

of useful carotenoids and therapeutically significant triterpenoids, Centella asiatica

is one of the most widely consumed green leafy vegetables (GLVs) in several

nations, including Sri Lanka (Chandrika & Kumara, 2015).

The useful medicinal plant Centella asiatica (L.) is a perennial creeper that

is both native to the Old World and the New World. It can be found all over the

world in tropical and subtropical areas. Centella asiatica has increasingly been

used in foods and beverages over the years, largely because of its advantageous

functional characteristics. The plant's bioactive constituents, particularly the

triterpenic acid (Asiatic acid madecassic acid), triterpenic saponin (madecassoside

and asiaticoside), flavonoids, and other phenolic compounds, have been widely

reported to have potential antioxidant, antimicrobial, cytotoxic, neuroprotective,

and other activities (Vasantharuba Seevaratnam et al., 2012).

 Mosquitoes and other arthropods spread illnesses such as yellow fever,

dengue fever, and other vector-borne illnesses and are well known for their

longterm negative consequences on human health. It is well recognized that

mosquitoes are an essential means of viral transmission. Historically, tropical and

subtropical climates are the ideal locations for mosquitoes to grow, making

mosquito-borne illnesses a serious threat across the world (Mukherjee D, et al,

2019). Aedes albopictus and Aedes aegypti are examples of the two main

mosquito species responsible for transmitting dengue virus in Asia and America

(Anoopkumar, A. N, et al, 2017). When mosquitoes are present in large numbers,

are a nuisance, or spread disease agents, they may be relevant to public health.

As peri-urban suburbs grew into formerly undeveloped natural regions in recent

decades, they provided a wider range of mosquito breeding sites than inner-city

areas, increasing human interaction with mosquitoes. Additionally, mosquitoes that

grow in containers, like Ae. albopictus, are invading urban areas, where they bite

humans constantly throughout the day while the females are looking for blood

meals (Petrić, D, et al, 2014). Virus infection: blood allows the virus to enter the

mosquito's body, where it infects the midgut epithelial cells. It multiplies in the cells

of the midgut, is discharged into the hemocoel, travels via the hemolymph to

various tissues, and eventually reaches the salivary glands to infect new victims

during successive bites. In mosquitos, innate immunity and a local microbiome

protect against the Dengue virus (Mukherjee D, et al, 2019). Because of this, the

control of mosquitoes becomes the need of the hour to prevent widespread

epidemic infections. However, it is difficult to control and prevent the severe

consequences created by mosquito species (Anoopkumar, A. N, et al, 2017).

Foreign Literature

Centella asiatica has been considered as brain tonic due to its wide beneficial

neuroprotective activity. Besides this, various other effects such as

antiinflammatory, ant proliferative, anticancer, antioxidant, antiulcer, wound

healing, etc. have been reported. These effects were tested on animal models and

various phytochemicals such as asiactic acid, asiatosides, polyphenolic

compounds, etc. are proven effective for these responses. The herb is also tested

for its toxicity and interactions with other drugs. Anticonvulsant drugs were found

to interact with Centella asiatica and also toxicological study suggested its use with

caution over a long period of time. In addition, various commercial products are

available throughout the world that has been majorly used for enhancing memory,

antioxidant, skin nourishment, neuroprotective as well as dietary supplements.

Further research work needs to be done more on clinical as well as cultivation

aspects. (Belwal et al., 2019) . Centella asiatica is mostly studied for its medicinal

properties. Very few studies have documented its potential as a biopesticide. A

study has suggested that the leaf extract of Centella asiatica is promising as a

larvicide and adult emergence inhibitor against Culex quinquefasciatus Say

(Culicidae).

The leaf extracts of Eucalyptus globulus and Centella asiatica showed an

overall moderate larvicidal effect when tested against different strains of Ae.
aegypti and An. stephensi. Among the four extracts studied, the hexane extracts

of both plants and the diethyl ether extract of Centella asiatica demonstrated higher

potential for the control of strains of mosquito vectors, followed by

dichloromethane. The methanol extract produced a comparatively less promising

result. The effect of the extracts on larval mortality was dose-dependent, with

increasing rates of larval mortality observed at increasing concentrations. The

highest level of tolerance was observed in the BSN strain of Ae. aegypti towards

the methanol extract of E. globulus. (Nair et al., 2014)

According to Belwal et al., (2019) numerous studies have examined the

variations in larvicidal efficacies of plant extracts tested against different mosquito

species; however, we were not able to find any reports of the larvicidal efficacies

of plant extracts tested against different strains of mosquitoes of the same species.

Susceptibility studies involving strains from different locations could be beneficial

to gauge the efficacy of the drug being studied and as a prelude to investigations

involving the inherent resistance mechanisms of the vectors to the drugs.

Mosquitoes are blood-sucking insects that spread diseases including

Malaria, Dengue, Japanese encephalitis, Yellow fever, etc. Humans must be

protected from mosquito bites in order to avoid mosquito-borne diseases. The

application of repellents such as lotions and liquidators to the skin or clothing

creates a vapor layer with a disagreeable odor or taste that renders a person

undesirable for feeding, therefore, repelling mosquitoes. Plant essential oils, in

general, have been recognized as a major natural resource for repellents in this

area because of their selective characteristics as they are biodegradable, nontoxic,

and eco-friendly. Textile-based mosquito repellents is one the popular method of

protection using cotton, polyester, and other textiles. This method requires no

additional investment in the textile finishing industry, which is a desirable feature

in developing countries. This review details the efforts of technocrats who applied

mosquito repellents to fabrics and the criteria used to determine

repellency.(Anupam et al., 2012)

As stated by Agustina et al., (2022) to prevent the proliferation of mosquito-

borne diseases and to improve the quality of the environment and public health,

mosquito control is essential. The major tool in mosquito control operation is the

application of synthetic insecticides such as organochlorine and organophosphate

compounds. However, this has not been very successful due to human, technical,

operational, ecological, and economic factors. In recent years, the use of many of

the former synthetic insecticides in mosquito control programs has been limited. It

is due to the lack of novel insecticides, high cost of synthetic insecticides, concern

for environmental sustainability, harmful effect on human health, and other non-

target populations, their non-biodegradable nature, higher rate of biological

magnification through the ecosystem, and increasing insecticide resistance on a

global scale2,3. Thus, the Environmental Protection Act of 1969 has framed a

number of rules and regulations to check the application of chemical control agents

in nature4. It has prompted researchers to look for alternative approaches ranging

from the provision of or promoting the adoption of effective and transparent

mosquito management strategies that focus on public education, monitoring and

surveillance, source reduction, and environment-friendly least-toxic larval control.

These factors have resulted in an urge to look for environmentfriendly, cost-

effective, biodegradable, and target-specific insecticides against mosquito

species. Considering these, the application of eco-friendly alternatives such as

biological control of vectors has become the central focus of the control program

in lieu of chemical insecticides.

Mosquitoes can transmit more diseases than any other group of

arthropods and affect millions of people throughout the world. WHO has declared

mosquitoes as “public enemy number one. Mosquito-borne diseases are prevalent

in more than 100 countries across the world, infecting over 700,000,000 people

every year globally and 40,000,000 of the Indian population. They act as a vector

for most life-threatening diseases like malaria, yellow fever, dengue fever,

chikungunya fever, filariasis, encephalitis, West Nile virus infection, etc., in almost

all tropical and subtropical countries and many other parts of the world. (Agustina

et al., 2022)

According to Panneerselvam et al. (2012). Mosquito-borne diseases have

an economic impact, including a loss in commercial and labor outputs, particularly

in countries with tropical and subtropical climates; however, no part of the world is

free from vector-borne diseases. The aim of the present study, to evaluate the

larvicidal, pupicidal, repellent, and adulticidal activities of methanol crude extract

of Artemisia nilagirica were assayed for their toxicity against two important vector

mosquitoes, viz., Anopheles stephensi and Aedes aegypti (Diptera: Culicidae).

In recent years, the use of environment-friendly and biodegradable natural

insecticides of plant origin have received renewed attention as agents for vector

control because they are rich in bioactive chemicals, active against a limited

number of species including specific target insects, and biodegradable. The

effective adult mortality was observed in the methanol extract of A. indica, ethyl

acetate extract of D. biflorus, and ethyl acetate and hexane extract of Z. zerumbet

against C. gelidus and C. quinquefasciatus (Kamaraj et at., 2010).

Synthetic repellents are causing unpleasant effects on humans of bad

aroma, rashes on the skin, eye burning, inhaling, headaches, coughs, sore throats,

nausea, dizziness, asthma, and respiratory irritation. To avoid these adverse

effects chemical repellents may be replaced with botanical agents. Mosquito bites

may be avoided by using mosquito repellents on their skin, housemates, clothing,

and holsters. While repellents are applied to the skin or clothing, they form a vapor

layer with an unpleasant natural scent that protects the user from being bitten. One

of the revolutionary methods is to use a fabric treated with a mosquito-repellent

substance. Human cultures in many regions of the world have employed plant

products against mosquito vectors from the beginning of time. The phytochemical

derived from plant sources may act as larvicidal, adulticidal, insect growth

regulators, and repellents and have different activities. Personal protection for

humans can be provided by repelling and knocking down the mosquito vectors by

creating a textile barrier between the host and the vector. Moisture-sensing pores

of mosquitoes help them to detect live organisms, are closed in olfactory mode,

also known as transpiration repellency and they are unable to locate humans. (
Singh et al., 2018)
A repellent is an insecticide that is repelling and non-killing. Every plant

has a composition of chemical compounds called secondary metabolites. Essential

oils, flavonoids, alkaloids, and aromatic compounds are metabolites in plants that

have the potential to be mosquito repellents (Boate and Abalis, 2020).

Essential oils, also called volatile oils, are secondary metabolites of volatile plants.

This oil exists in fruits, seeds, leaves, flowers, stems, bark, roots, and rhizomes

(Sengül Demirak and Canpolat, 2022). Therefore, plants as a place for mosquito

feeding and resting activities also have a favorable opportunity to become one of

the mosquito control strategies by utilizing secondary metabolite compounds.

Plants around us have the potential insecticides, but it is necessary to identify

which bioactive molecules in repellent plants have a higher effect on

diseasetransmitting mosquitoes (Athuman et al., 2016).

Plants are an integral part of human civilization. Medicinal plants are also

been relied upon by over 80% of the world population for their basic health care

needs. Drugs based on plants are of prime importance for several remedies in

traditional and conventional medicine throughout the world and serve as a

substitute for drug supply in modern medicine. (Mala et al., 2014). Since herbal

medicines play an important role in the treatment of a wide range of diseases, there

is a growing need for quality control and standardization. (Božović, 2012).

Centella asiatica has been considered as brain tonic due to its wide

beneficial neuroprotective activity. Besides this, various other effects such as

antiinflammatory, ant proliferative, anticancer, antioxidant, antiulcer, wound

healing, etc. have been reported. These effects were tested on animal models and

various phytochemicals such as asiactic acid, asiatosides, polyphenolic

compounds, etc. are proven effective for these responses. The herb is also tested

for its toxicity and interactions with other drugs. Anticonvulsant drugs were found

to interact with Centella asiatica and also toxicological study suggested its use with

caution over a long period of time. In addition, various commercial products are

available throughout the world that has been majorly used for enhancing memory,

antioxidant, skin nourishment, neuroprotective as well as dietary supplements.

Further research work needs to be done more on clinical as well as cultivation

aspects. (Belwal et al., 2019) . Centella asiatica is mostly studied for its medicinal

properties. Very few studies have documented its potential as a biopesticide. A

study has suggested that the leaf extract of Centella asiatica is promising as a

larvicide and adult emergence inhibitor against Culex quinquefasciatus Say

(Culicidae).

The leaf extracts of Eucalyptus globulus and Centella asiatica showed an

overall moderate larvicidal effect when tested against different strains of Ae.

aegypti and An. stephensi. Among the four extracts studied, the hexane extracts

of both plants and the diethyl ether extract of Centella asiatica demonstrated higher

potential for the control of strains of mosquito vectors, followed by

dichloromethane. The methanol extract produced a comparatively less promising

result. The effect of the extracts on larval mortality was dose-dependent, with

increasing rates of larval mortality observed at increasing concentrations. The

highest level of tolerance was observed in the BSN strain of Ae. aegypti towards

the methanol extract of E. globulus. (Nair et al., 2014)

Local Literature

According to Sultan,R. A. (2014) Biological activities of Centella asiatica

were determined as cytotoxic activity–LD50 = 1926 μg.mL−1 using Artemia salina;

mild insecticidal activity (40% mortality) against Rhyzopertha dominica; no activity

against Tribolium castaneum and Callosobruchus analis; phytotoxicity (70%

growth regulation at 1,000 μg.mL−1 and 10% growth regulation at 10 μg.mL−1)

against Lemna minor. Mild antibacterial and antifungal activities were noted

against Bacillus subtilis, Pseudomonas aeruginosa, and Candida albicans, while

no activity was noted against Staphylococcus aureus and E. coli. Antioxidant

performance was comparable to ascorbic acid, and analgesic and anti-

inflammatory activities comparable with aspirin at 100 mg.kg−1b.w. (Mahmood, S.

B. Z., Azhar, I., Ahmed, S. W., & Mahmood, Z. A.)

In recent years much interest has been evinced in the use of plant products as

insecticides to controlvector as pests, in view of the environmental and health

hazards by synthetic organic insecticides (2014). In the present study, Centella

asiatica plant extract was prepared using the solvent chloroform. The plant extract

was tested against the various developmental stages of mosquito, Aedes aegypti.

The level of the toxicity of Centella asiatica was expressed in terms of LC50/24
hours values. The LC50 values of chloroform leaf extract for I, II, III & IV instar

larvae of Aedes aegypti were 1.04%, 1.369%, 1.937% and

2.642% respectively. In the present study the preliminary phytochemical analysis

showed the presence of flavanoids, alkaloids, terpenoids, glycosides and steroids

in the chloroform leaf extract of Centella asiatica. The morphogenetic

abnormalities are commonly caused by botanical extracts and the disturbance

from the growth regulating hormones. It is therefore suggested that Centella

asiatica derivatives are considered for vector control operations besides their use

in other fields after exploring field trails. (M.Manimegalai, C.R Hima and D.

Rajmohan).

According to Nair, S. S., (2014) The larvicidal activity of different solvent

leaf extracts (hexane, diethyl ether, dichloromethane, and methanol) of Eucalyptus

globulus and Centella asiatica against two geographically different strains of

Aedes aegypti and Anopheles stephensi was investigated. The extracts were

tested against the late third instar larvae of Aedes aegypti and Anopheles

stephensi, and larval mortality was observed after 24 hours of treatment. LC50 and

LC90 were calculated. The LC50 values of hexane extract of Eucalyptus globulus

against the late third instar larvae of the BSN and JPN strains of Aedes aegypti

and the DLC and KNG strains of Anopheles stephensi were 225.2, 167.7, 118.8,

and 192.8 ppm, while those of the hexane extract of Centella asiatica were 246.5,

268.7, 50.6, and 243.5 ppm, respectively. The LC50 values of diethyl ether extract

of Centella asiatica were 339.6, 134.5, 241, and 14.7 ppm. The hexane extracts of
both plants and the diethyl ether extract of Centella asiatica presented the highest

potential for the control of Aedes aegypti and Anopheles stephensi. The present

findings also reveal the necessity of assaying multiple strains of a species to fully

comprehend the larvicidal efficacy of a compound. (Shetty, V., Shetty, N. J. 2014).

As Stated by Gyawali, R., Aryal, S., Gautam, N., & Manandhar, M. (2015).

Plant extract of Acorus calamus L, Adhatoda vasica L, Artemisia vulgaris L,

Calotropis gigantea (L) W. T. Aiton., Centella asiatica (L) Urb., Curcuma longa L,

Melia azadirach L, Nicotiana tabacum L, Datura stramonium L, Rhus wallichii Hook

F, Punica granatum L, Vitex nugundo L, Wrightia arborea (Dennst.) Mabb. are

studied for insecticidal property against cabbage aphids Brevicoryne brassicae L.

These plants were also evaluated for potentiality against plant pathogens Botrytis

cinerea, Sclerotium rolfsii, Fusarium moniliforme, Fusarium proliferatum, Fusarium

oxysporum and compared with standard fungicide metalaxyl. All screened plants

at concentration of 1.25%, 2.5%, 5%, 10% and 20% exhibited selective response

with different stain of plant fungi. Acorus calamus L, Nicotiana tabacum L,

Adhatoda vasica L, Artemisia vulgaris L shows significant control on cabbage

aphids. Nicotiana tabacum L, showed prominent effect even at lowest

concentration ie 1.25% with 50% for aphids. Diluted extracts of Rhus wallichii Hook

F, Wrightia arborea (Dennst.) Mabb., Curcuma longa L, Calotropis gigantea (L) W.

T. Aiton., at 20% concentration were found most effective against plant pathogens

Sclerotium rolfsii, Fusarium moniliforme, Botrytis cinerea, Fusarium oxysporum

respectively.
Foreign Studies

Among the various control measures for mosquitoes, use of synthetic

insecticides is still the most convenient and indispensable method all over the

world. Chemical measures were first tried but they failed since their overuse

caused irreparable hazards to the environment and humans also developing

resistance among mosquito species and undesirable effect on non - target

organisms, (Thangam and Kathiresan, 1990). The vast number of mosquitoes

present naturally causes the civilians to use mosquito coils and liquidators which

release CFC (chloro fluoro carbon) in a considerable amount that depletes the

ozone which is harmful for the earth and human’s future. These mosquitoes are

vastly present in the developing countries where not much importance is given to

the sanitation. Rain water and sewage can easily get stagnant in the roads and in

the open spaces, these water stagnant and open sewage passages acts as a very

good habitat for the mosquitoes to breed which seriously concerns the civilian’s

day to day life.

Chloroform was used to prepare the plant extract of Centella asiatica. The

Aedes aegypti mosquito, in all of its life stages, was tested using plant extracts.

The level of toxicity of Centella asiatica was expressed in terms of LC50/24 hours

values. For Aedes aegypti larvae in the I, II, III, and IV instars, the LC50 values of

chloroform leaf extract were 1.04%, 1.369%, 1.937%, and 2.642%, respectively.

The chloroform leaf extract of Centella asiatica contained flavonoids, alkaloids,

terpenoids, glycosides, and steroids, according to the preliminary phytochemical

examination of the current study. After investigating field trails, it is advised that
Centella asiatica derivatives be taken into account for vector control activities in

addition to their application in other fields (Devadass et al., 2014).

The hexane extracts of both plants and the diethyl ether extract of Centella

asiatica showed as the highest possible or potential to control the strains of

mosquito vectors. The outcome of the study indicate that the chemicals causing

the insecticidal activity of both plants are likely nonpolar and intermediate in nature.

The present findings also shows that in order to completely understand a

compound's larvicidal activity, numerous strains of a species must be assayed

(Shetty et al., 2014).

Furthermore, another method for getting rid of the mosquitoes and other

vectors that spread diseases like dengue fever, malaria, and yellow fever is to use

synthetic insecticides. However, because of their lack of environmental

friendliness, these synthetic products have the potential to be harmful to human

health as well as the environment. In this case, an effort has been undertaken to

identify the larvicidal qualities of plants that are easily accessible locally, including

Piper nigrum, Centella asiatica, Azadiracta indica, and Leucas aspera. The

mosquito larval death rate was examined at different plant concentrations.

(Extracts of 1%, 1.5%, 2.5%, and 3%). It was determined that the fatality rate would

be 1, 1.5, 2, and 2.5. Three percent of the plant was taken out and compared. In

all the cases, it was observed that the mortality rate increased with an increase in

the concentration of the test solution.

 In addition to, this study which was conducted by Malini et al. (2016) aims

to investigate the antifeedant activity of ethanol extracts from five plants against

the larvae of Spodoptera litura F. The plants that were used were Pterocarpus

indicus, Centella asiatica, Calophyllum inophyllum, Alstonia scholaris, and Lantana

camara. The goals of the study were to find an effective minimum concentration

that has antifeedant properties against third-instar S. litura larvae and to screen

the best extract with antifeedant properties. The result showed that, among the

other plants tested, Centella asiatica had the best extract with antifeedant

properties at a concentration of 2500 ppm and a coefficient of antifeedant activity

in the range of 101–150%, so it is categorized as having good antifeedant

properties. Centella asiatica, with its antifeedant properties, can be used not only

to kill insects but also as an insecticide for mosquitoes.

The common skin condition is mosquito bite reaction. The usual course of

treatment includes topical steroids and oral antihistamines. The treatment's

drawback, however, is the adverse consequences of chronic usage of topical

steroids. Herbal extracts are becoming more popular recently as an alternative to

conventional anti-inflammatory dermatoses treatments. Goal: To evaluate the

efficacy of a 1% hydrocortisone (HC) gel in comparison to a herbal extract gel

including Perilla frutescens, Portulaca oleracea, Ipomoea pescaprae, Aloe vera,

Centella asiatica, and Broussonetia papyrifera. Materials and Methods: A

doubleblinded split randomized control study was carried out on 50 hypersensitive

to mosquito bite participants (15–19 years old). The medications were applied
twice daily after the volunteers received one non-infectious Aedes albopictus bite

on their arms (Udompataikul et al., 2014).

Local Studies

In the Philippines, common mosquito-borne infections include viral

infections such as dengue, Chikungunya, and Japanese encephalitis, and parasitic

infections like malaria and filariasis (DOST 2016). In order to prevent contracting

these diseases, intensive mosquito prevention and control measures, including

personal protective measures against mosquito bites, are of urgent importance.

Among these protective measures, mosquito repellents such as those in lotions,

coils, and liquidators are commonly used for personal mosquito bite prevention

(DOH 2017).

Centella asiatica is a tropical medicinal plant from Apiaceae family native

to Southeast Asian countries Monographs of the plant describing mainly its wound

healing and memory enhancement effects exist in the European Pharmacopoeia,

Commission E of the German Ministry of Health, and World Health Organization

(WHO) .In addition to neuroprotective effect of Centella asiatica, it has been

reported to own a wide range of biological activities desired for human health such

as wound healing, anti-inflammatory, ant psoriatic, antiulcer, hepta- protective,

anticonvulsant, sedative, immune stimulant, cardio protective, antidiabetic,

cytotoxic and antitumor, antiviral, antibacterial ,antifungal, antioxidant,and for lepra

and venous deficiency treatments.

 A perennial herbaceous plant called takip kuhol is widespread in wet

tropical and subtropical areas of South America and India. The aerial portions that

are harvested during flowering are used to make the medication. Centella asiatica

has a long history of usage as a medication, dating back to the late 1800s in France

and as far back as 3000 years in Indian Ayurvedic medicine and 2000 years in

Chinese medicine. It has been dubbed "the fountain of life" because, according to

legend, an ancient Chinese herbalist who consumed gotu kola lived for more than

200 years. This is in contrast to the name's claim that it contains caffeine or cola

(Pacinto, 2014).
 CHAPTER 3

RESEARCH METHODOLOGY

This chapter presents and describes the research design, research setting,

duration and frequency of the study, and data gathering, instrument, and

procedure.

RESEARCH DESIGN

The researcher used a true experimental design because it fits the questions

that the researchers wish to answer. An effective statistical method for determining

the cause and effect of the relationship between various variables is a true

experimental design. True experimental design is one of the most precise types of

research designs, it offers strong evidence to back up the existence of the

relationship. According to Susan Skidmore (2008), there are three types of true

experimental design, first, is the pretest-posttest; this one meets the characteristics

of true experimental design. Volunteers are chosen at random experiments either

the test group or the control group. Second, the post-test only controls group

design. This design differs from the first, this one lacks the pre-intervention

measurement of a pretest. The last is Solomon’s four-group design. This style is

the most powerful of the trio. It does not simply account for any potentially

confusing outcomes of the pre-test but enables you to contrast them with an

experimental as well as a control group that did take the pretest. The experimental

group receives the treatment while the control group does not receive the

treatment. The experimental group should be the one to receive the treatment for
the researcher need to observe if there is any significant effect of the said

experiment. This research design is educational research that revolves around

knowledge, collecting accurate information, and improving analysis.

RESEARCH SETTING

This research takes place at Lower St. Ignatius street Macasandig, Cagayan

de Oro City, this area was chosen because it is a field with greenery and there are

breeding grounds for mosquitoes and reproduction. It is very suitable for this

experiment because it is wide.

Figure 2. Sampling site

Duration and Frequency of the study

In this study, an herbal plant will be tested for its effectiveness against

mosquitos. The herbal plant is the Centella asiatica, it will then be concluded

whether it is a good insecticide or not. The estimated duration for this study is 23
Days. The first week is allocated for the preparation and making of the extract such

as collecting the Gotu kola and preparing the needed instruments for collecting the

extract. The collected extract needed to sit for about 4 weeks according to Asiatic

P, B.B, Brahma, M.B (2010). After 2 days the said extract will be ready to be used

on mosquitos. The mosquitos are caught when they were a larva and then

metamorphose into mosquitoes in just 2-3 days, approximately 8-5 days if we were

to follow the whole process according to the Environmental Protection Agency on

March 8 (2022). In order to test the effectiveness of the extract we will be applying

it to our caught mosquitos and see the result. We will be spraying the extract on

the mosquito to see the result and see how many seconds, minutes, and hours

take for the extract to kick in, we will be conducting multiple trials to make sure how

many mosquitos has affected per ml, 5ml, 10ml, and 15ml. By the end of the day

23rd, it is expected that the researchers to give a good explanation and evaluation

of the study conducted.

Table 1. Gantt Chart of School Activities

Schedule of

Activities Months

November December January February

Class discussion

Title/Topic of the
study approval
Review of Related

Literature and
studies

Methodology

Sampling period

Analysis of data

Final Page

Preparatory

A.Collection of Plant Material

The researchers have collected a total of 200 pieces of Centella asiatica (Gotu

Kola) plant at Zone -10 Zayas, Carmen Cagayan De Oro City, where a large

quantity of Centella asiatica (Gotu Kola) was discovered by the researchers.

B. Preparation of the Insecticide

Following plant identification, 200 pieces of Centella asiatica leaves will be

extracted using a mortar and pestle that have been aseptically cleansed with

running water and air dried. The extraction will begin once the leaves have been

washed under running water by the researchers. The leaves will be prepared by
the researchers so they can be dried for 24 hours. Once the leaves are in the

mortar, the process will begin by uniformly crushing, grinding, and rolling the pestle

across the leaves. This will remain until all of the leaves are crushed but not entirely

crushed. After crushing the leaves with a mortar and pestle, the researchers will

separate the extract from the leaves using a cotton cloth and place it in a single

container. Finally, the Centella asiatica extract was placed in a spray bottle. The

extract is produced from the Centella asiatica plant.

Sampling Procedure

The random selection method was one that the researchers utilized to help

increase the generalizability of the results. We researchers wanted to make sure

that the sample we were using was an effective technique to investigate precisely

and reflects the features of the mosquito group, so it was taken from a bigger

population. The more mosquitoes there are, the better we were able to generalize

our experiment's findings using our sample extract. Researchers might also lessen

the risk of bias affecting the outcomes by randomly choosing test subjects.

Our artificial insecticide is placed on 3 bottles of sprayers and all 3 of the

sprayers are labeled with how much of the extract is inside the bottle. Each bottle

will be sprayed on the box that is assigned to that bottle so we will figure out how

many milliliters of the Centella asiatica extract are needed to kill mosquitoes. There

will be 5ml, 10ml, and 15 ml worth of extract on each bottle and it will be sprayed

on the large box where there are 5 mosquitoes so we will know how effective the
Centella asiatica on killing mosquitoes. To know how fast our insecticide works on

killing mosquitoes we will observe the 3 containers under 1 hour, 2 hours, and 3

hours.

The containment that where we will put the mosquitoes will be a large

transparent box where they can’t escape so we can observe them during the

experiment. There will be 3 large containers where we will put an equal amount of

mosquitoes. The researchers must have 45 caught mosquitoes that will be used in

the experiment and place the mosquitoes in a container. All 3 large containers

should have 15 mosquitoes each so that we will know how effective the insecticide

whether it will be a 5ml worth of spray used or more.

Table 2. The concentration of Centella asiatica dosages in terms of time and

mortality rate towards the variable.
Dosage Time Dead Mosquitos

1hrs

5ml. 2hrs

3hrs

1hrs

10ml. 2hrs
 3hrs

1hrs

15ml.

2hrs

3hrs

DATA ANALYSIS AND PROCEDURE

The researchers used descriptive analysis to analyze the efficacy of Centella

asiatica extract as an effective insecticide against mosquitoes. The research

instrument that was used was the observational method. Researchers observe the

behavior of the mosquitoes after the extract was spray on them. The researchers

determine that the bottle contains extract of Centella asiatica; 5ml, 10ml, and 15ml

are the measurements that show how effective the Centella asiatica extract would

react by the same amount of mosquitoes of each ml. The time duration presented

in objective #2 and the intensity presented in objective #3 could prove how

effective this said extract.

Number of Dead Mosquitoes

× 100 = Mortality Rate
Population of the Mosquitoes
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