THE FEASIBILITY OF OREGANO EXTRACT AS A MOSQUITO

REPELLENT IN ALIDO, MALOLOS, BULACAN

A research paper presented to the faculty of

La Consolacion University Philippines
Senior High School

In partial fulfillment of
the requirements for the subject
Research/Capstone Project

By
Santos, John Enrick
Santos Joshua
Sulleza, Deanne Marise S.
Tahanlangit, Renzo

March 2018
ACKNOWLEDGEMENT
This research would have not been successfully completed without a number of people whose
tireless efforts, support and guidance.

Foremost, I want to offer this endeavour to our God almighty for the wisdom he bestowed upon
the researchers, the strength, peach of mind and good health to finish this research.

The researchers would also like to thank Ms. Maryann Cebricus, a practical research teacher, ,
for giving the researchers the opportunity to make this research ,without whose supervision this
research project would simply not have happened.

To our biology teacher, Sir Kevin Magaoay for providing necessary assistance to complete the
research.

To Prof. Olivia P. Almario, MAED, the principal of senior high school department, for giving us
the oppurtunity to conduct the research.

To all the volunteers participated in this study, for their patience and support for answering the
questions and putting their trust on the research.

Thanks to Deanne Sulleza’s and Renzo Tahanlangit’s parents for accomodating us in their home
to us complete our research. To the researcher's family for their never-ending love and support
for their child and serves as an inspiritation to pursue this undertaking.

To each member of this team, who extend their efforts in finishing this research

To all of them, the researchers extends their deepest gratitude for taking part of this research and
helping them make one step nearer to their dreams.

Godbless us all!

ABSTRACT
The present study was conducted to determine the mosquito repellent activities of selected plant
material which is Oreganum vulgare in order to obtain safe and efficient herbal mosquito
repellent formulations. Direct Heat Infusion process was performed for Oregano leaves for
extraction. The other ingredients such as Isophryl alcohol, Mentha leaves and a little amount of
cloves were purchased from public market. The said product is placed in a 40ml bottled spray
which contain 3.6 % (V/V%) of total active ingredients and 96.4% as other ingredients ( Isophryl
Alcohol, H2O). Mosquito repellency activity testing was carried out using arm-in-cage method.
Control hand (right hand) which is the untreated arm was exposed to the cage where 10- 20
blood-seeking mosquitoes had been placed and the number of mosquitoes that aligned or biting
the arm was recorded in each given time intervals. Same procedure has done to observe the
experimental hand (left hand) which is the treated arm. Three replicates were carried out for each
extract. It showed 100% mosquito repellency in the first replicate. On the second and third
replicate, the product had decreased and has a percentage of 93% to repel mosquitoes. The study
had also found out that an approximate number of 10 times of spray must be applied in the cloth
in order for the repellent to work. Sixteen (16) volunteers from Alido, Malolos Bulacan, have
been asked to participate in this study by wearing black shirt given by the research team. Half of
them anticipated the control group who wore black shirts. Indoor field trial was conducted in
seven days from 8 pm to11 pm by separately applying the mosquito repellent spray on the shirt.
The spray showed 84% mosquito repellency for indoor field trial which were carried out for
three hours each day for a week.

Keywords:
 Thymol-a white crystalline compound present in oil of thyme and used as a flavoring and
preservative.
 Carvacrol-a liquid phenol C10H14O found in essential oils of various mints (as thyme)
and used as a fungicide and disinfectant.
 Monoterpernoid - Monoterpenes are a class of terpenes that consist of two isoprene units
and have the molecular formula C10H16. Monoterpenes may be linear (acyclic) or
contain rings. Biochemical modifications such as oxidation or rearrangement produce the
related monoterpenoids.
 Linalool- a fragrant liquid alcohol C10H18O that occurs both free and in the form of
esters in many essential oils and is used in perfumes, soaps, and flavoring materials.
 P-cymene -. A monoterpene that is toluene substituted by an isopropyl group at position
4.
 Fumigation-is a method of pest control that completely fills an area with gaseous
pesticides—or fumigants—to suffocate or poison the pests within.
 Acaricidal-are pesticides that kill members of the arachnid subclass Acari, which includes
ticks and mites.
 Permethrin- a synthetic insecticide of the pyrethroid class, used chiefly against disease-
carrying insects
I. INTRODUCTION
For the past decade, development and evaluation in the field of plant-based repellent continue to
challenge researchers in terms of dispensing plant- based volatile that implies the viability of
plant-based repellents and duration of repellents in the future.

According to Dr. Marcela (2014) one of the most dangerous insect in the world is Mosquito. It
can harm plants, food and humans. Origanum vulgare, generally known as Oregano,was a
culinary and medical herb from the mint family (Lamiaceae). It was used for cooking and
medicinal purposes, ranging from treating infections to repelling insects. The chemicals that give
the herb its unique and pleasant smell are thymola white crystalline substance of a
pleasant aromatic odor and strong antiseptic properties, pinene gives many compounds
for perfumery, such as artificial odorants most widely encountered terpenoid in nature and was
highly repellant to insects, limonene was a chemical found in the peels of citrus fruits and in
other plants, carvacrol was present in the essential oil oregano, and caryophyllene it boosts
immunity because of the powerful antioxidant compound. (Wagner, 2013)

The said plant was simple, helpful to discover and convenient. The researcher aims to help
diminish the threat of mosquito transmitted sickness. The motivation behind why oregano can
repulse mosquito was that it has the solid compound of thymol and carvacrol which can repulse
mosquito in view of the odor.

The uncontrollable growth of population of the mosquitoes nowadays heightened public health
awareness. Mosquitoes are vital vectors of infections and aggravation pests. It stimulate nuisance
by their bites. A carriers of severe and well-known illnesses such as malaria, arboviral
encephalitis, dengue fever, chikungunya fever, West Nile virus, and yellow fever. Their activity
was affected by climate, light and temperature.(Govindarajan & Sivakumar ,2015)

Repellents limit contact with mosquitoes. Plant-based repellents have been used for generations
in traditional practice as a personal protection measure against host-seeking mosquitoes.
Numerous plants with bug repulsing properties are local to the tropics where they were delivered
for an extensive variety of therapeutic purposes. Some of the plant volatiles were deterrent yet
repellent due to the fact it bear excessive vapour toxicity in conformity with insects.

World Health Organization (WHO) has been revealed that a repellent was recognized as a
valuable infection counteractive action instrument to supplement bug spray based methods for
vector control. It also approves that a plant derived repellent, PMD has been proven to be
suitably efficacious and safe to compete with DEET in the field of disease prevention.

According to Pierre Lutgen (2015), Oregano oils were rich in thymol (38.8%), carvacrol
(32.9%), p-cymene (7,9%). They have solid toxicities against Rhizopertadominica by contact or
by fumigation. Effective, likewise against fruit flies. Thymol adds to the acaricidal properties of
Ocimum gratissimum or known as Clove basil. (I Karpouhtsis et al., J Agriculure and Food
Chem, 1998, 46, 1111-15)
Several studies have been claimed that oregano extract had been proven and tested as an insect
repellent. Researchers have found that biological hazards such as exposure to ticks and
mosquitoes can affect health. They have been suggest to use of permethrin- impregnated clothing
for employers and employees in addition to daily repellent sprays .( Balanay et al, 2017).

Also, a recent evidence such that the phenols carvacrol and thymol that are monoterpenoids
occurring in essential oils of several aromatic and medicinal plants, such as thyme, oregano,
savory, lavender and coriander, can be considered as candidate ingredients for the development
of novel acaricidal formulations to control the populations of I. ricinus and the spread of related
tick-borne diseases (Benelli et al, 2017).

Recent studies have also shown toxicity of these monoterpenoids against

selected mosquito vectors and other arthropod pests. Carvacrol and thymol at all concentrations
tested showed >90% repellency on I. ricinus . Linalool was scarcely effective (50.24%
repellency) only at the concentration of 5% (Benelli et al, 2017).

A study was also conducted by applying mosquito repellent finish on fabric.  Finished cotton
samples were tested against Anopheles stephensi. Marigold (petals)andnirgundi (leaves) extract
finished fabric samples showed 96 and 94 % repellency respectively after 60 minutes of
observation. It remained 56 % and 54 % (after 15 washes) and 54 and 52 % (after exposure in
sun for 3 hours) by the application of marigold and nirgundi extracts respectively. Results was
said to be that microencapsulation technique on selected cotton textile proved effective to repel
mosquitoes up to acceptable level according to WHO.

Studies conducted at the University of Iowa demonstrated the effectiveness of thymol -- one of
the main components of garden thyme (Thymus vulgaris), hardy in zones 5 to 9 -- in controlling
houseflies, according to Pierre Lutgen, writing in "Malaria World." Lutgen also notes the
efficacy of another compound found in thyme -- carvacrol -- for insect control and notes that oil
of oregano contains both of these compounds in concentrations of 38.8 and 32.9 percent
respectively

A study conducted by Licciardelo, Muratore ,Nerin, Russo and Suma (2013) assess its efficacy
of novel functional packaging materials containing three essential oils: citronella, oregano and
rosemary. The results obtained from area tests run in Petri dishes indicated that essential oils that
includes citronella, oregano and rosemary, at concentrations higher than 0.005 μL/cm2 showed
potential in terms of repellent activities against the red flour beetle, Tribolium castaneum.

Previous studies have reported that Carvacrol (75.70%), linalool (9.0%), p-cymene (4.33%) and
thymol (1.9%) were the most abundant compounds in oregano. Against Ae.aegypti, O. onites EO
was repellent at the minimum effective dosage (MED) of 0.011 (±0.00) mg/cm2 in the cloth
patch assay compared to the reference control, N,N-dimethyl-3-methylbenzamide (DEET) with a
MED = 0.007 ± (0.003) mg/cm2. (Agramonte, 2017)
Furthermore, they are easily available, accessible and affordable therefore usage of traditional
repellent plants should be promoted among the local residents in order to reduce vector-borne
disease prevalence. More recently, the Department of Health (DOH,2017) today approaches
nearby officials and families to heighten mosquito anticipation and control measures at home and
in the group, and to shield themselves from being chomped by mosquitoes, especially in high-
hazard zones.

A few plant specialists plant oregano in the garden to repulse nuisances, for example,
mosquitoes, and others assert the fundamental oil was a mosquito repellent. In addition, scientific
proof to help this claim was insufficient, however oregano contains two chemicals with known
anti-agents properties, and in light of the fact that oregano was "generally recognized as safe" by
the U.S Food and Drug Administration (FDA, 2015), it doesn't hurt to test for yourself.

Greenstock and Larrea (2013) also suggest keeping the crops away from traps related with
unsustainable cropping practices. Another belief was that garlic was a powerful anti-agent. It has
a direct anti-agents impact when rubbed on the skin, despite the fact that there are significantly
more compelling anti-agents accessible that likewise have an all the more satisfying scent. The
utilization of garlic nonetheless, has not been appeared to be effective at turning the mosquitoes
away.Therefore, the use of oregano as a mosquito repellent still don’t have a sufficient evidence
to prove that it can repel mosquito.

The researchers aim to discover an alternative solution to avoid various kinds of diseases that
mosquito transports on human subjects such as dengue, malaria, and many more using oregano
extract as mosquito repellent especially in remote areas.

This study seeks to answer the following questions:

1. Is it possible to make a mosquito-repellent using oregano extract? was the oregano

extract components are enough to make a mosquito-repellent?
2. How long would the oregano extract mosquito-repellent last?
3. Is it proven that the oregano extract was safe to use?
4. Does the said plant-based repellent have its advantages or disadvantages?
5. How do mosquitoes behave inside the mosquito box?

This study was eco-friendly for it decreases harmful effect in the environment. This investigation
will be a major help for those people who are having a hard time with mosquitoes. It will end
mosquitoes in this manner it won't cost them much cash. It can decrease terrible symptoms in the
body for it was contains less chemicals than other mosquito-repellants. This examination can
help DOH's battle against mosquito ailments, (for example, dengue and jungle fever). This can
be utilized as apparatus of data in making a material that can battle the said malarias and other
deadly transmitting diseases.As the vast majority of the mosquito repellent items and gadgets
accessible in the market are reported to have harmful effects on human being is the target of the
present investigation was to create viable plant-based mosquito repellent product.
 II. METHODOLOGY

Different authors have measured the efficacy of mosquito repellent in a variety of ways. The
determination of the plant depended on their accessibility as crude materials, logical proof and
traditional used as repellent.

True experimental method was used in this research. The study was to find out the feasibility of
oregano extract as mosquito repellent, the experimental method was suitable for this research
because it characterized by voluntary participation in the study. Since the researchers want to test
the efficacy of the product, this method will help the researchers to answer the hypothesis.

The dependent variable of the study was the number of mosquito that comes to rest on the hand,
and the independent variable was mosquito repellent. The control group was the cloth that hasn’t
sprayed by the oregano extract mosquito repellent while the experimental group was the cloth
that has been sprayed.

For preparation of the oregano extract, crushed and dried oregano leaves were extracted by the
process of Direct Heat Infusion and use solvent like was Isophryl Alcohol. A one cup of water
was boiled for 5-10 minutes. Origanum vulgare (Oregano) leaves and Mentha (Mint) leaves were
added as well as 1g of cloves. Adding such will strengthen the repellent. Also, mint leaves gives
a sweet fragrance compared to chemical insects repellents. After 30 minutes of cooling, the herbs
were strained out and the extractwas mixed with 237 ml of was Isophryl alcohol. The extract was
stored in a cool temperature for about 2-3 hours.

Testing the Mosquito Repellent Activity of Plant Extract Using Arm-In-Cage Method:

This stagewas one of the challenging part for the researchers.The researchers collected
mosquitoes by leaving drum water with an amount of 10-20 percent of water overnight in a
shady area for 3-4 days andwas covered with mosquito net every morning. The investigators also
made an improvised mosquito glass box. Its sidewas made of illustration board and the surface of
the five (5) sideswas covered with a plastic cover that has the dimension of 10 x 6.5 x 6.5 inches.
One side was covered with mosquito net where the arm would be inserted. The guideline used in
this experimentwas based on the article produced by the World Health Organization (WHO).
Arm-in-Cage tests are a common way to explore the effectiveness of repellent formulation. This
technique was suited for the assessment of topical repellents such as crèmes, moisturizers, and
spray formulation and impregnated textiles.

The mosquito box contains 10-20 mosquitoes.Host-seeking behaviour of the mosquitoes was
also tested prior to the experiment. This was done by placing a pre-cleaned hand in the cage and
counting the number of mosquitoes that aligned within ten (10) seconds. If at least five(5)
mosquitoes aligned on the hand, the mosquitoes inside the cage were regarded as host-seeking
and the repellency experiment was continued.This was to ensure that the mosquitoes inside the
mosquito box seeks for its food.
Before insertion of the hand into the cage containing 10-20 mosquitoes, the hands are protected
by stocking socks through which the mosquitoes cannot bite. Movement of the arm was
prohibited while testing. While doing this, the other researchers record the time and take
documentation for evidences. According to World Health Organization (WHO), for the test to
proceed, the biting rate must be ≥10 landings and/or probings in the 30-second period. The right
hand will be considered as the control group while the other hand will identified as the
experimental group.

The covered hand without the mosquito repellent was placed and has been observed for the given
time frames. The data were recorded by the researchers. The control hand was carefully
withdrawn andthe left hand has been sprayed. The hand was desiccated for about 5 minutes.
Consequently, the treated hand was placed in the cage for another 30-second period, testing
should not proceed when the five (5) mosquito landing and/or probing rate on the exposed hand
was. Doing such, the researcher record if how many mosquitoes will land on its arm with respect
to time intervals. These phenomena give the researcher to also monitor the durability of the
product. The experiment was replicated thrice for the validity of the data.

For testing the bio-efficacy of the product, 16 volunteers have been asked to participate in the
study. Suburbs in Alido, Malolos, Bulacan was where the study takes place. Each volunteer were
asked to wear black shirts provided by the research team. To certify the data that will be given by
the participants, the group went to the place everyday and recorded the approximate number of
mosquito bites and aligned that was observed by the volunteers. The shirt must be wear every
night within 3 hours from 8pm -11pm and strictly sprayed five (5) times in front and five (5)
times in back of the shirt. Eight (8) volunteers anticipated the controlled group. These set of
people wore black fabric t-shirt. The remaining participants wore the black fabric t-shirt that
sprayed with oregano extract. The observation last for a week.

STATISTICAL TREATMENT

Statistically, all replicates were calculated and the researchers get its mean value to identify the
mosquito repellency of the plant extract. The mean was the sum of all the values in the data set
divided by the number of values in the data set. This was expressed as:

R 1+ R 2+…+ R n
x̅ =
n

Whereas,

R1 = total no. of mosquitoes aligned in control group and experimental group in Replicate
1
R2 = total no. of mosquitoes aligned in control group and experimental group in Replicate
2
Rn = total no. of mosquitoes aligned in control group and experimental group from the
last replicate
n = the number of replicates done

Percentage repellency (P)was expressed as a proportion of the number of mosquito landings

and/or probings on the treated arm (T) in relation to the number of landings and/or probings on
the control arm (C) of the same individual:

(C−T )
P= x 100
C

Where:

C-= total number of the landings/probings on the untreated arm (arm before repellent
treatment)
T = total number of the landings/ probing on the treated arm ( arm after repellent
treatment)

RESULTS
Table 1.

The constituents in the mosquito repellent spray

Constituents Quantity in spray (420ml)

Active Ingredients
Origanum vulgare (Oregano) leaves 6g
Mentha (Mint) Leaves 6g
Cloves 1g
Other Ingredients
H2O 237 ml
Isophryl Alcohol (70%) 237 ml
Cloves 1g

After analysing the mosquito repellent activity of individual plant extracts, a mosquito repellent
spray which contained active ingredients (3.6% v/v) which includes Oregano leaves, Mint leaves
and Cloves. The inert ingredients (96.4% v/v) of the product that includes water and Isophryl
alcohol having the remaining constituent were prepared as shown in Table 1. The product
mosquito repellent in a 40mL spray bottle includes Origanum vulgare having 6 grams, mentha
having 6 grams, H2O and Isophryl Alcohol having 237 mL each and an amount of 1 gram.

Table 2.

Amount of repellent that will be sprayed

No. of No. of sprays 1 minute 5 minutes 10 minutes Results

trials
First trial 5 sprays 0 2 5 Rejected

Second 10 sprays 0 0 0 Approved

trial
*based on the criteria given by the World Health Organization

The product is sprayed 5 times in the treated hand. According to the results, no mosquito landed
in one minute, two mosquitoes landed in five(5) minutes and six(6) mosquitoes landed within 10
minutes. Therefore, the amount of the repellent is not enough to repel mosquitoes and must be
discontinued. In second trial, the product is sprayed 10 times in the treated hand. In the first
minute no mosquitoes had land, likewise when the product is tested for five (5) minutes,
however, two mosquitoes landed in the treated hand within ten (10) minutes. The applied number
of spray is sufficient for the experiment to continue.
Table 3.1

Replicate no.1

TIME INTERVAL CONTROL EXPERIMENTAL

15 minutes 2 0
25 minutes 3 0
30 minutes 5 0
Total 10 0
% Repellency = 100%
* control = untreated hand
* experimental =treated hand

Table3.1 shows the no. of mosquitoes landed in controlled area and experimental area in the first
replicate. Controlled area was the untreated hand/ the right hand and the other hand was the
treated hand/ the left hand. The number of spray that have been used is ten, five from the back of
the hand and five in palm. In the first trial the untreated hand were inserted. Two mosquitoes
have been land within 5 minutes, three mosquitoes have been land within 25 minutes and five
mosquitoes have been land within 30 minutes in a total of 10 mosquitoes. On the other hand, no
mosquitoes have been land in the said time intervals. Therefore, the mosquito repellency
percentage is 100%.

TABLE 3.2

Replicate no. 2

TIME INTERVAL CONTROLLED EXPERIMENTAL

(UNTREATED) (TREATED)
15 minutes 3 0
25 minutes 5 0
30 minutes 7 1
Total 15 1
% Repellency = 93%
* control = untreated hand
* experimental =treated hand

Table 3.2 signifies the no. of mosquitoes landed in controlled area and experimental area in the
second replicate. The control hand gains three mosquitoes within 15 minutes, five mosquitoes
within five minutes and seven mosquitoes within 30 minutes. Thus, the control hand has the total
number of 15 mosquitoes that were aligned in the hand. In the experimental hand, no mosquitoes
have been observed within 15 minutes and 25 minutes, yet one mosquito have been land when
the product is tested within 30 minutes. As a result, the product has 93% to repel mosquitoes.

Table 3.3

Replicate no. 3

TIME INTERVAL CONTROLLED EXPERIMENTAL

(UNTREATED) (TREATED)
15 minutes 2 0
25 minutes 5 1
30 minutes 5 0
Total 12 1
% Repellency = 93%
* control = untreated hand
* experimental =treated hand

Table 3.3 indicates the no. of mosquitoes landed in controlled area and experimental area in the
third replicate. The untreated hand obtain two mosquitoes within 15 minutes, five mosquitoes
within 20 minutes and five mosquitoes within 30 minutes. This shows that the total number of
mosquitoes landed in the untreated hand is 12. After that, treated hand has been observed. No
mosquitoes have been landed in the said time interval: 15 minutes and 30 minutes. However, one
mosquito have been land within 25 minutes. Hence the product has a percentage of 93%
mosquito repellency.

Table 4

Plant Extract Total no. of Total no. of Total no. of Calculated

Mosquito Mosquito Mosquito Mean Value
bites and bites and bites and of Mosquito
aligned aligned aligned bites and
(Replicate 1) (Replicate 2) (Replicate 3) aligned

Origanum vulgare extract 10 16 14 13.33

Mean values of the mosquitoes controlled area and aligned/bit for plant extract and mosquito
repellency (percentages) of plant extract.

Table 4 shows the efficacy of the Origanum vulgare extract to repel mosquitoes. In replicate 1,
10 mosquitoes is said to be observed in both controlled and experimental hand. In replicate 2,
there is a total number of 16 mosquitoes that have been land in the controlled hand. In Replicate
 3, a total of 14 mosquitoes have been aligned. Computing its mean value, there is an average of
13 mosquitoes observed from the three replicates.

Table 5.1

Bio-efficacy test results of the mosquito repellent spray (controlled group).

Approximate number of mosquito/es bite or aligned

Volunteer # Thursday Friday Saturda Sunday Monday Tuesday Wednesday Total
y
1 5 5 7 6 12 6 7 48
2 8 7 3 4 9 8 4 43
3 4 6 5 2 6 8 4 35
4 6 8 7 4 9 12 6 52
5 7 9 5 6 10 7 5 49
6 4 7 9 9 8 6 8 51
7 7 4 3 5 9 3 5 36
8 6 3 4 7 8 5 10 43

Table 5.1 shows the bio-efficacy test results of the mosquito repellent spray in the controlled
group. Control group are the set of people who wore black shirts in the experiment. The
experiment has been observed within 8pm – 11pm.

Volunteer # 1 assumed that 5 mosquitoes have been land on the first day of the experiment, 5
mosquitoes on the second day, 7 mosquitoes on the third day, 6 mosquitoes on the fourth day, 12
mosquitoes on the fifth day, 6 mosquitoes on the sixth day and 7 mosquitoes on the last day.
Volunteer # 2 observed that 8 mosquitoes have been aligned at the beginning of the experiment.
An approximate number of 7 mosquitoes on the second day, 3 mosquitoes on the third day, 4
mosquitoes on the fourth day, 9 mosquitoes on the fifth day 8 mosquitoes in the sixth day and 4
mosquitoes on the last day. Volunteer # 3 believed that 4 mosquitoes have been landed on the
start of the trial, 6 mosquitoes on the second day, 5 mosquitoes on the third day, 2 mosquitoes on
the fourth day, 6 mosquitoes on the fifth day, 8 mosquitoes on the sixth day and 4 mosquitoes on
the last day. Volunteer # 4 said that 6 mosquitoes have been observed in the first day of the
experiment. 8 mosquitoes have been observed on the second day, 7 mosquitoes on the third day,
4 mosquitoes on the fourth day, 9 mosquitoes on the fifth day, 12 mosquitoes on the sixth day
and 6 mosquitoes on the last day. Volunteer # 5 concluded that 7 mosquitoes were attracted on
the first day of the trial. An estimated number of 9 mosquitoes on the second day, 5 mosquitoes
on the third day, 6 mosquitoes on the fourth day, 10 mosquitoes on the fifth day, 7 mosquitoes on
the sixth day and 5 mosquitoes on the last day of the experiment. Volunteer # 6 believed that 4
 mosquitoes have been landed on the first day of trial. On the second day, 7 mosquitoes were
observed, 9 mosquitoes have been landed on the third and fourth day, 8 mosquitoes on fifth day,
6 mosquitoes on the sixth day and 8 mosquitoes on the last day. Volunteer# 7 said that 7
mosquitoes have been landed on the first day, an approximate number of 4 mosquitoes landed on
the second day, 3 mosquitoes on the third day, 5 mosquitoes on the fourth day, 9 mosquitoes on
the fifth day, 3 mosquitoes on the sixth day and 5 mosquitoes on the seventh day. On the last
volunteer, nearly 6 mosquitoes have been observed on the first night of the experiment, 3
mosquitoes have been observed on the second day, 4 mosquitoes on the third day, 7 mosquitoes
on the fourth day, 8 mosquitoes on the fifth day, 5 mosquitoes on the sixth day, and 10
mosquitoes on the last day.

In approximated number of mosquitoes observed, volunteer # 2 has the highest number of

mosquitoes landed in the first night of the experiment having 8 mosquitoes and volunteer #5 on
the second night having nine mosquitoes. Volunteer # 6 has the highest number of mosquito bite/
landed having a number of 9. On the fifth day, volunteer # 1 has encountered 12 mosquitoes that
has been the largest number of mosquitoes observed in that day. Also, volunteer # 4 observed 12
mosquitoes on the sixth day and volunteer # 8 has the largest number of mosquitoes landed on
the last day of the experiment having an approximate number of 10 mosquitoes. To sum up,
volunteer # 4 has the highest number of mosquitoes that has an approximate number of 52
mosquitoes landed for a week. On the other hand, volunteer# 3 has the lowest number of
mosquitoes that was aligned that has a total number of 35 mosquitoes for a week.

Table 5.2

Bio-efficacy test results of the mosquito repellent spray (experimental group)

Approximate number of mosquito/es bite or aligned

Volunteer # Thursday Friday Saturda Sunday Monday Tuesday Wednesday Total
y
1 0 0 0 0 1 3 5 9
2 0 0 0 0 0 2 2 4
3 0 0 0 0 1 3 4 8
4 0 0 0 0 0 1 5 6
5 0 0 0 0 0 3 4 7
6 0 0 0 0 1 1 2 4
7 0 0 0 0 0 2 5 7
8 0 0 0 0 2 4 5 11

Table 5.2 indicates the results of the bio- efficacy testing of the mosquito repellents in the
experimental group. Experimental group are the set of people who wore black shirts and use the
mosquito repellent spray. The experiment was also observed within 9 pm- 5am.
As expected by the researchers, the mosquito repellency of the product is 100% working from
the beginning of the experiment. Nevertheless, on the fifth day, a minimal number of mosquitoes
have been observed. Volunteer # 1, 3 and 6 said that one mosquito have been landed on the fifth
day, Volunteer # 8 observed two mosquitoes on the same night. On the sixth day, Volunteer # 8
has the largest number of the mosquitoes observed having an approximate number of 4
mosquitoes. Volunteer # 1, 3 and 5 assumed that an approximate number of three mosquitoes
have been aligned. Volunteer # 2 and 7 claimed that two mosquitoes landed on them and
volunteer # 4 and 6 said that only one mosquito have been landed.

The number of mosquitoes landed on the last day were increasing continuously. Volunteer #1
believed that an approximate number of five mosquitoes were aligned while volunteer # 2 and 6
assumed that only two mosquitoes have been landed. Volunteer # 3 and # 5 said that nearly four
mosquitoes have been observed on that day. Also, volunteer # 7 and 8 stated that five mosquitoes
have been landed on the last day of the experiment.

Generally, volunteer # 4 and # 7 has the lowest number of mosquitoes landed that was recorded
for a week having only four mosquitoes. On the contrary volunteer # 8 got the highest number of
mosquitoes aligned having an approximate number of 8 mosquitoes for a week while using the
repellent.

Table 6

Percentage of Mosquito repellency of the repellent

Thursda Friday Saturday Sunday Monday Tuesday Wednesday

y
Control 47 49 43 43 61 55 49 347
group
Experimenta 0 0 0 0 5 19 32 56
l group
% 100% 100% 100% 100% 92% 65% 35% 84%
Repellency

Table 6 shows the percentage of Mosquito repellency of the repellent in Alido, Malolos, Bulacan
that was tested for a week. This indicates the efficacy of the mosquito repellent as the day goes
had passed. As shown above the mosquito repellent out of oregano extract showed 100%
mosquito repellency on the first night up to fourth night of the trial. However, the percentage of
mosquito repellency starts to decrease on the fifth night of the trial. On the fifth night of the trial
the mosquito repellency revealed that there is a percentage of 92% of the product to repel
mosquitoes. However, on the sixth night, the product showed only 65% efficacy of the product.
On the last day, who gives the least percentage in a week , have 35% to repel mosquitoes.
Overall, the mosquito repellency showed 84% mosquito repellency in a week.
 DISCUSSION

Oreganum vulgare also known as oregano leaves contains thymol and carvacrol which can repel
mosquitoes.The 40 ml mosquito repellent spray which contained 10.5% (v/v %) total active
ingredients each, indicated 100% mosquito repellency at the beginning of the experiment. The
mosquito repellent contains oregano leaves which are the main component of the repellent, mint
leaves, water, Isophryl alcohol and a little amount of cloves.

The study conducted three replicates. The first replication of the product also showed 100% of
the efficacy to repel mosquitoes. However, only 93% had been observed on the second and third
replicate

As indicated by the indoor field trial which were participated by 16 volunteers, done for a three-
hour time period every day for seven days, the mosquito repellent spray have demonstrated
100% mosquito repellency in about 4 days. Yet, testing the bio- efficacy of the repellent had
decreased on the fifth trial up to the last which is the fifth day of the week having a total number
of 84% mosquito repellency.

The number of spray was strictly ten (10) times for the repellent to work. These conclude that
there is a relationship between the amount of repellent spray applied in the subject to the duration
and efficacy of the product.

Since herbal plants are non- toxic and natural, the said product was safe to use. Also, no skin
disturbances or rashes have been reported on the skin of the test volunteers with the extract.

The discoveries of this investigation could be fluctuated to numerous variables, for example, the
types of mosquito and the qualities of volunteers and bio-synthetic appeal to mosquitoes. The
aroma that the mosquito repellent is pleasant to the volunteers. However, the mosquito repellent
has the poor life span of the fragrance in the item.

Further investigations are needed to elucidate the efficacy of the herbal mosquito repellent
formulations against a wide range of mosquito species and also to identify active compounds
responsible for mosquito repellent activity in order to utilize them if necessary, in preparing a
commercial product to be used as a mosquito repellent.

Future specialists, in any case, was prescribed to utilize research facilities such as laboratories in
distinguishing the active compounds responsible for repellency for more precise and accurate
results. Likewise, the procedure of extraction of the leaf could be made in various techniques, for
example, steam distillation.
 REFERENCES

Balanay, J.O.G., Banks, V.M., Harris, J.W., &Meshnick, S. (2017, April). Residual effectiveness of
permethrin-treated clothing for prevention of mosquito bites under simulated
conditions.Journal of Environmental Health, 79(8), 8-15.

Benelli, G., Maggi, F., Tabari, M.A., &Youssefi, M.R. (2017, October).Toxic and repellent activity of
selected monoterpenoids (thymol, carvacrol and linalool) against the castor bean tick, ixodesricinus
(acari: ixodidae).Veterinary Parasitology, 245, 86-91. doi: 10.1016/j.vetpar.2017.08.012

Agramonte, N.M., Baser, K.H.C., Bernier, U.R., Carroll, J.F., Demicri, B., Kramaer, M.,…(2017,
February 16). Repellency of the origanumonitesl.essential oil and constituents to the lone star tick and
yellow fever mosquito.Nat Prod Res, 31(18), 2192-2197. doi: 10.1080/14786419.2017.1280485

Mamta, R., Saroj S, J.S., &Saroj, Y. (2017, December).Effect of microencapsulated plant extracts on
mosquito repellency.Journal of applied & natural science, 9(4), 2127-2131.

Govindarajan, M., Sivakumar, R. (2015, October).Laboratory evaluation of Indian medicinal plants as

repellents against malaria, dengue, and filariasis vector mosquitoes.Parasitology Research, 11(2), 601-
612. doi: 10.1007/s00436-014-4222-0

Lutgen, P. (2015, May 6). Mosquitocidal and repellent properties of plant extracts.Malaria world

Mosquito World (2018)Mosquito feeding habitat. Retrieved from http://www.mosquitoworld.net

Licciardello, F., Muratore, G., Nerin, C., Russo, A., & Suma, P. (2013, July) Effectiveness of a novel
insect-repellent food packaging incorporating essential oils against the red flour beetle
(triboliumcastaneum). Innovative food science & emerging technologies, 19, 173-180.

Karunamoorthi, K., Ilango, K., &Endale, A. (2009, September).Ethnobotanical survey of knowledge and
usage custom of traditional insect/mosquito repellent plants among the ethiopianoromo ethnic group.
Journal of ethnopharmacology, 125(2), 224-229.doi: 10.1016/j.jep.2009.07.008

Nordqvist, J. (2017, December 11) “What are the health benefits of oregano?”.Medical news today.
Retrieved from https://www.medicalnewstoday.com/articles/266259.php

Deziel, C. (n.d.). Oil of oregano as a mosquito repellent. Hunker.

World Health Organization.Guidelines for efficacy testing of mosquito repellents for human skin.1-
30.Retrieved from
http://apps.who.int/iris/bitstream/10665/70072/1/WHO_HTM_NTD_WHOPES_2009.4_eng.pdf

Arambewela, L., Ranasinghe, M. S. N., &Samarasighe, S. (2016) Development of herbal mosquito

repellent formulations. International journal of collaborative research on internal medicine & public
health, 8(6), 341-380.
Pradhan, S. S. (N. D.). Preparation of low cost herbal mosquito repellent from Begunia leaf.Department
of chemistry national institute of technology, Rourkela.Retrieved from
http://ethesis.nitrkl.ac.in/2105/1/sushree_thesis.pdf

Morley, J. (2014). Academic phrasebank.The university of Manchester, 1-72. Retrieved from

http://www.kfs.edu.eg/com/pdf/2082015294739.pdf

The epidemic of the mosquito carrier could strike anywhere as long as there is mosquitoes.
Because is the random,