(Group 8) - Sip - Documents (Bronda, Canto, Payas-10-Ste)

Barotac Viejo National High School
Science Technology and Engineering

Barotac Viejo Iloilo
ANTIBACTERIAL EFFECT OF SUMMER GRAPE (Vitis aestivalis)

FRUIT EXTRACT AGAINST E. COLI (Escherichia coli)
An Investigative Project Presented to
the Faculty of Science, Technology and Engineering Department
of Barotac Viejo National High School
Barotac Viejo, Iloilo
In Partial Fulfillment of the Requirements
For Junior High School Completion
Nicole L. Bronda
Razel Ann T. Canto
Christian Andrei B. Payas
March 2020
Approval Sheet
This Science Investigatory Project, entitled “ANTIBACTERIAL EFFECT OF

SUMMER GRAPE(Vitis aestivalis) FRUIT EXTRACT AGAINST E. COLI(Escherichia
coli)” prepared by Nicole L. Bronda, Razel Ann T. Canto and Christian Andrei B. Payas,
in partial fulfilment of the requirements for Junior High School Completion of Science
Technology and Engineering is hereby approved.
IYSA MARIE B. REA JESCEL MAE H. BALDELOVAR

Research IV Teacher English Critic
(Date Signed) (Date Signed)
MARIMAR P. BANABATAC GEROME G. REA

Research Adviser Statistician
LANI JOY O. ESPADA FEROLYN J. BALAJADIA

STE Coordinator Science Coordinator
Accepted as partial fulfillment of the requirements for Junior High School

Completion of Science Technology and Engineering of Barotac Viejo National High
School.
MA. LUISA L. ASONG PhD

Principal IV
(Date Signed)
ii
Acknowledgement
The researchers would like to acknowledge with sincerest gratitude to those
persons who extended professional assistance, shared expertise, gave guidance and
patience behind the success of the study that made them to finish the study for
completion.
First and foremost to the Almighty God for all the blessings and guidance to
pursue their study;
To the Department of Education, for giving them the chance to explore and
develop their knowledge and ideas towards the study;
To Dr. Wellet A. Del Castillo and Dr. Ma. Luisa L. Asong, Principal IV, Barotac
Viejo National High school, for their support and encouragement, in the completion of
the research work;
To Mrs. Grace A. Peñas and Mrs. Ferolyn J. Balajadia, Science Coordinator, for
lending them the laboratory room apparatuses, for the guidance and for giving
suggestions for the improvement of their study;
To Mrs. Lani Joy O. Espada, Science Technology and Engineering Coordinator,
for her encouragement and support throughout the study;
To Ms. Iysa Marie B. Balderas, Research Adviser, for the teachings, provision of
necessary information regarding this research and for sharing her expertise to improve
and help researchers finished the study;
To Ms. Marimar P. Banabatac, Science Critic, for lending her expertise,
suggestions, encouragement and full support in making the study;

To Ms. Jescel Mae H. Baldelovar, English Critic, for editing and checking the
grammatical errors in the completion of the study;
To Mr. Gerome G. Rea, Statistician, for helping the researchers in computing,
solving and analyzing the statistical data of their study;
To Mr. Audie A. Suladay, Head of WVSU-CSL, for guiding the researchers and
giving assistance in conducting their study;
To all Grade 9, Science Technology and Engineering students for helping the
researchers in their studies and for supporting each other in times of difficulties they
encounter during their study; and
Lastly, to Mr. and Mrs. Bronda, Mr. and Mrs. Canto, Mr. and Mrs. Payas, their
supportive parents, for the moral and financial support and for helping them to pursue
their study.
Nicole L. Bronda
Razel Ann T. Canto
Christian Andrei B. Payas
March 2020
Bronda, Nicole L., Canto, Razel Ann T., Payas, Christian, Andrei B., “THE
ANTIBACTERIAL EFFECT OF SUMMER GRAPE (Vitis aestivalis) FRUIT
EXTRACT AGAINST E. coli (Escherichia coli).” Unpublished Science Investigatory
Project. Science Technology and Engineering. Barotac Viejo National High School.
Barotac Viejo, Iloilo, March, 2020.
Abstract
The research project aimed to determine the antibacterial effect of Summer grape fruit
extract that can be used as antibacterial agent against Escherichia coli. More
specifically, the study aimed to determine if Summer grape fruit extract is effective as an
antibacterial agent against Escherichia coli and to determine the best antibacterial
between ripe and raw Summer grape fruit extract. One hundred grams of ripe and raw
Summer grape fruit were weighed and extracted in a slow juicer. Escherichia coli culture
isolates was taken and aseptically transferred into a sterile tube of nutrient broth and
incubated for 24 hours. Three drops of 0.1 mL of the plant extracts were dropped in the
agar wells that were placed in each of the (T1R1, T1R2, T1R3), (T2R1, T2R2, T2R3), the same
amount of the antibiotic was placed into three other control plates. The plates were
examined for Zone of Inhibition formation. The data were recorded and statistically
treated. The researchers found out that ripe Summer grape fruit extract was effective as
antibacterial agent among the three treatments (x= 13.13) but in contrast to
ciprofloxacin (positive control), it is the most effective antibacterial agent. It is therefore
concluded that there is significant difference (p=0.048) between the commercial product
(x=10.00) and the experimental product (x=13.13). As a result, Summer grape fruit
extract is effective and can be used as an antibacterial agent against Escherichia coli.
The researchers recommended that the usage of ROTAVAP method in extraction to
increase the level of concentration against Escherichia coli.
iv
Table of Contents
Page no.
Title Page i
Approval Sheet ii
Acknowledgement iii
Abstract iv
Table of Contents v
List of Tables viii
List of Figures ix
Chapter 1
Introduction of the Study 1
Background of the Study 2
Statement of the Problem 5
Hypotheses 5
Conceptual Framework 6
Significance of the Study 7
Scope and Delimitation of the Study 8
Definition of Terms 8
Page no.
Chapter 2
Review of Related Literature and Studies 10
Review of Related Literature 11
Review of Related Studies 14
Chapter 3
Methodology 18
Purpose of the Study and Research Design 19
Experimental Treatment 21
Materials and Equipment 22
Cost Analysis 24
Data Gathering Procedure 25
Statistical Data Analysis Procedure 27
Chapter 4
Result and Discussions 28
Descriptive Data Analysis and Discussion 29
Inferential Data Analysis and Discussion 31
vii
Page
no.
Chapter 5
Summary, Conclusions, and Recommendations 33
Summary 34
Conclusions 34
Recommendations 35
REFERENCES
vii
List of Tables
Page no.
Table
1 Components of Ripe Summer Grape Fruit Extract in a Treatment 21
2 Components of Raw Summer Grape Fruit Extract in a Treatment 21
3 Components of Ciprofloxacin in a Treatment 22
4 Cost Analysis 24
5 Zone of Inhibition of Escherichia coli using Ripe and Raw Summer Grape 29
Fruit Extract and Ciprofloxacin
6 Mean Average of the Four Treatments in Three 30
Replicates
7 Significant Difference of the Three Treatments in terms of Effectiveness 31
against Escherichia coli
8 One Sample T-test. Significant Difference between Experimental product 32
and Commercial product in terms of their effectiveness as antibacterial agent
against Escherichia coli
viii
List of Figures
Page no.
Figure
1 Input Process Output (IPO) 6
2 Randomized Complete Block Design 20
ix
1
Chapter 1
Introduction of the Study
Chapter 1 consist of (1) Background of the Study (2) Statement of the Problem
and Hypothesis (3) Conceptual Framework (4) Significance of the study (5) Scope and
Delimitations (6) Definition of Terms.
Part One, Background of the Study, presents the introduction of the study, the
reason why was the study conducted and discussed its rationale.
Part Two, Statement of the Problem and Hypothesis, presents the specific and
general problems and enumerates the hypothesis tested
Part Three, Conceptual Framework, shows the Input, Process, Output of the
study.
Significance of the Study, enumerates the different beneficiaries and the benefits that
may obtained from the results of the study.
Part Four, Significance of the Study, enumerates the different beneficiaries and
the benefits that may obtained from the results of the study.
Part Five, Scope and Delimitations, identifies the scope and delimitation
encountered during the conduct of the study. Specifies the coverage of the study.
Part Six, Definition of Terms, presents the conceptual and operational terms. The
terms are arranged alphabetically.

Barotac Viejo Naional High School
Barotac Viejo, iloilo
2
Background of the Study
Nowadays, diseases are rampant in one’s health. Disease is an illness that
affects ones health caused by some microorganisms. Food borne diseases have always
been a major concern in both developing and developed countries. Human struggles
due to the effects of infectious diseases caused by pathogenic microorganism such as
bacteria, viruses, parasites, and fungi. Diseases can be spread directly or indirectly from
one person to another. One of the pathogenic microorganism is the bacteria. Bacteria
can be found everywhere and one of its type that causes different diseases is
Escherichia coli.
Escherichia coli also known as E. coli is a type of bacteria found everywhere
including the environment, foods, intestine of organisms. It is usually found in the lower
intestine of warm-blooded organism. It is one of the most frequent causes of many
common bacterial infections including Cholecystitis, Bactermia, Cholangitis, Urinary
Tract Infection (UTI) and Diarrhea. (Escherichia coli. (E. coli) Infections: Background:
Feb. 11，2019.)
Escherichia coli is the most commonly present bacterium in the human intestine,
which helps in preventing the entry of pathogenic microorganisms. E. coli is non-
pathogenic in normal conditions, but if present in excess, will become causative agent of
various diseases like urinary tract infection, diarrhea, vomiting etc.(Amrita, et.al, 2009)
3
Diarrhea is one of the infectious diseases caused by E. coli. Human having a
bacteria inside the intestine like E. coli can experience diarrhea in which human passes
waste from body very frequently in liquid rather than solid form. Even though E. coli is
helpful to keep the organism’s digestive tract being healthy, it is also harmful that can
cause severe illnesses. Some of its strains such STEC can cause severe food-borne
disease. People eating and drinking contaminated foods might acquire this type of
bacteria that may cause infectious diseases. (Siva, 2011).
Summer grape or scientifically known as Vitis aestivalis is a decidous climber
growing to 20 m. (65ft 7in) at a fast rate. The season for its flower is from the month of
June to July and the seeds ripen from September to October. Because of its compound,
it is used by the native Americans for a variety of purposes. Compuond decoctions are
taken for gynecological, gastrointestrinal, urinary, and pediatric aids. Plants can simply
be taken as a “tonic”. Leaves are taken as a liver aid. Fruits are often made into
beverages, breads, and cakes or simply eaten raw. Summer grape in our local country is
wood vine, shallowly to deeply 3 to 5 lobed leaves 4 to inches long. The bloom season is
in the month of June. In visayan language, it is known as “bika” and are usually found in
decidous woods, thickets, wooded bluffs, etc. Due to its contained property, the
antibacterial property, it is beneficial and helpful on eliminating the effects of phatogenic
microorganism and one is that bacteria arises in the society.(Oliveira, 2012).
According to World Health Organization (WHO) 2019, at least 600 million people
suffer from illness each year and also in the Philippines alone, over 32,000 cases of food
4
and water-borne diseases have been recorded by the Department of Health (DOH) as of
September, 2019.
With this premise, people who cannot afford to buy expensive medicines are
facing financial problems and the researchers conducted this study to determine the
antibacterial effect of summer grape fruit extract against Escherichia coli.

5
Statement of the problem
This study aimed to determine the antibacterial effect of summer grape fruit extract
against Escherichia coli.
Specifically it sought to answer the following questions:
1. Is summer grape fruit extract effective as an antibacterial agent against Escherichia coli?
2. Is there any significant difference in the zone of inhibition of Escherichia coli using ripe
and raw summer grape fruit extract and ciprofloxacin?
3. Is there any significant difference between experimental product and commercial product
in terms of their effectiveness as antibacterial agent against Escherichia coli?
Hypotheses
Based on the preceding questions, the hypotheses below were advanced:
1. There is no significant difference in the zone of inhibition of Escherichia coli using ripe
and raw summer grape fruit extract and ciprofloxacin.
2. There is no significant difference between experimental product and commercial product
in terms of their effectiveness as antibacterial agent against Escherichia coli.

6
Conceptual Framework
Input Process Output
A. Extraction of Summer
grape fruit
B. Preparation of E. coli
culture
C. Preparation of Muller
Hinton Agar (MHA) plates
Summer grape fruit
Summer grape fruit D. Application of Bacteria extract against
Escherichia coli
(ripe and raw) E. Agar Well-Method
F. Application of
Treatments, Positive and
Negative control
G. Incubation and Reading
of the Zone of Inhibition
Figure
1. This figure shows the relationship between the Input, Process, and Output of this study.
Significance of the Study
The study of the antibacterial effect of Summer grape fruit extract against Escherichia
coli has great importance to the advancement of science. This study was beneficial and will
be of great benefit to the following:

7
Community/society.It has a great impact since this will be a natural remedy, people in
the society can lessen the chances of buying expensive medicines. It can be a solution to the
main cause of infectious diseases and it can inform people that using summer grape fruit extract
is effective as antibacterial agent against bacteria.
Entrepreneurs. It will be a great benefit since the study was successful because they
can sell a brand new innovative product. Since it was proven in the study that using summer
grape fruit extract against Escherichia coliis effective, thus summer grape fruit extract were
recommended to examine its chemical components that will be a great idea to produce a new
innovative products using the fruit extract which could benefit the consumers when they have
diarrhea caused by Escherichia coli.
Students. Since this study was proven and tested to fight against Escherichia coli using
summer grape fruit extract, the students could benefit from it because it can can help them to
prevent from diseases caused by Escherichia coli in which they can use the extract as
antibacterial agent during their hand washing program in school.
Future researchers. This study may be one of the basis that a new theory in medicine
and healthcare will arise. This study can also serve as their reference.
8
Scope and Delimitations of the Study
The goal of this study is to determine the antibacterial effect of summer grape fruit
extract with the contrast of negative control against Escherichia coli. This study was only limited
in using the pure extract of summer grape fruit known for its antibacterial effect and inhibitory
effects on Escherichia coliand also it is limited to ciprofloxacin used as positive control. The
extracts were obtained by juicing the summer grape fruits. In addition, the Escherichia coli
bacteria were incubated in a Science Laboratory at West Visayas State University, Iloilo City
and were used as the experimental set-up. This study was conducted at West Visayas State
University, Science Laboratory in October 5-6, 2019.
Definition of terms
Antibacterial - anything that destroys bacteria or suppresses their growth of their ability
to reproduce. Directed of effective against bacteria. (Merriam-WebsterDictionary.2013).
In this study, antibacterial is used to determine how effective are the treatments applied.
Erythromycin - an antibiotic obtained from a strain of bacteria, effective against gram-
positive and some gram-negative bacteria. (AmericanHeritageDictionary, 2012).
In this study, erythromycin is a commercial antibiotic used to compare in summer grape
fruit extract in terms of their effectiveness.Escherichia coli- a type of bacteria that normally
lives in organisms’ intestines. It is also found in the gut of some animals. Human become
infected when he/she swallow even a small amount of Escherichia coli . It is a bacterium
commonly found in the intestines of humans and other animals, some strains of which can
cause severe food poisoning.(LivingScience.2017).

9
In this study, Escherichia coli is the microorganism or bacteria used as a subject in
conducting this study.
Extract- a preparation containing the active ingredient of a substance in concentrated
form. (Merriam-WebsterDictionary.2013).
In this study, extract is used as a substance against Escherichia coli.
Summer grape - a summer grape vine climb nearly all the tree species with which it co-
occurs, scaling heights over 30 ft. (9m). The greenish yellow flowers are produced in panicles
about 2-5 long. It is a vigorous vine climbing to a height of 35 feet by means of tendrils, or
sprawling over low brushes and trees. (LivingScience.2015).
In this study, summer grape were extracted and its extract were used as a substancein
testing the subject of this study.

10
Chapter 2
Review of Related Literature and Studies
This chapter includes the following parts: (1) Review of Related Literature (2) Review of
Related Studies
Part One, Review of Related Literature is composed of:
Antibacterial effect, discusses the uses and purposes of its effect.
Ciprofloxacin, discusses the purpose and nature of Ciprofloxacin.
E.coli (Escherichia coli), discusses the nature of E. coli and the early studies of it.
Distilled Water, discusses the purpose and nature of Distilled Water.
Summer grape fruit, discusses the nature of summer grape fruit.
Distilled Water, discusses the purpose and nature of Distilled Water.
Part Two, Review of Related Studies
Antibacterial effects of herbs and Spices Extract on Escherichia coli
The Antibacterial Effect of Turmeric (Curcurma longa) Leaves and Fire Tree (Delonix
regia) Leaves against Staphylococcus aureus
The Antibacterial Property of Tambis (Syzygiumsamarangense) Leaves against
Staphylococcus aureus
11
Review of Related Literature
Antibacterial effect
Antibacterial activity of nanocarriers was tested in vitro by measuring the zone of
inhibition against suitable microorganisms such as Escherichia coli, Bacillus subtilis,
Pseudomanas aeruginosa. Staphylococcus aureus, etc. (Annur et al. , 2015 ; Yang et al. ,
2016).The microorganism on interest was grown on a compatible sterilized nutrient agar
medium. Test composites on square plastic grids or disks were transferred on the solidified agar
medium and incubated for 24 h at 37 degrees Celsius. After the incubation period, the zones of
inhibition were observed in four directions for each tested sample (Yang et al.,2016).
Previous studies reported the antibacterial activity of prepared nanocarriers such as
silver metal nanoparticles (Arvizo et al., 2012) and chitosan combined silver metal nanoparticles
against Gram negative or Gram positive bacteria (Anisha et al., 2013).
Antibacterial activity
The shelf life of a product can be extended either by adding artificial preservatives or by
taking hygienic measures during the manufacturing process. As the consumer trend today is
towards preservative-free foods with a long shelf-like industry is being forced to rethink its
manufacturing methods. (Shiel, WC., 2018).
Antibacterial activity is the most common type of antimicrobial activity reported for the
acridone alkaloids, although a few cases of antifungal or antiviral activity have been reported.
(Sharma et.al., 2009).

12
Ciprofloxacin
Ciprofloxacin is a fluoroquinolone antibiotic that fights bacteria in the body. It is used to
treat different types of bacterial infections, including skin infections, bone and joint infections,
respiratory or sinus infections, urinary tract infections, and certain types of diarrhea. It is also
used to treat gonorrhea. Ciprofloxacin is also used to treat people who have been exposed to
anthrax or certain types of plague.Ciprofloxacin should be used only for infections that cannot
be treated with a safer antibiotic. Fluoroquinolone antibiotics can cause serious or disabling side
effects that may not be reversible, such as tendon rupture or nerve problems. (Baydar, 2000).
E.coli (Escherichia coli)
Escherichia coli is one of the most frequent causes of many common bacterial
infections, including cholecystitis, bacteremia, cholangitis, urinary tract infection (UTI), and
traveler’s diarrhea, and other clinical infections such as neonatal meningitis and pneumonia.
Most types of E.coli are harmless and even help keep your digestive tract healthy. Some strains
of E.coli infection can include nausea, vomiting, and fever. E.coli comes from human and animal
wastes. During precipitation, E. coli may be washed into creeks, rivers, streams, lakes, or
groundwater. With these are used as sources of drinking water and the water is not treated or
inadequately treated. It may be end up in drinking water. Entero-pathogenic Escherichia coli
(EPEC) are in the family Enterobacteriaceae. The bacteria are gram negative, rod shaped, non-
spore forming,
motile with petritrichous flagella or non-motile, and grow on MacConkey agar (colonies are 2 to
3 mm in diameter and red or colorless). (Ghazali, et. al., 2012).

13
Distilled Water
Distilled water is water that has been boiled into vapor and condensed back into liquid in
separate container. Impurities in the original water that do not boil below or near the boiling
point of water remain in the original container. Thus, distilled water is one of purified water.
Distilled water is safe to use in cooling down engines and cleaning since it is free of
compounds. It is commonly used to top off lead acid batteries often found in cars and trucks.
Lead batteries rely on water to keep cars and trucks running or they will deteriorate. (Salvador,
2013).
Summer grape fruit
Summer grape or scientifically known as Vitis aestivalis or Pigeon grape is a species of
grape native to eastern North America from Southern Ontario east Maine, west to Oklahoma,
and south to Florida and Texas. It is a vigorous vine, growing to 10 meter or more high to
trees.Vitis aestivalis is a deciduous climber growing to 20 m. (65ft 7in) at a fast rate. The season
for its flower is from the month of June to July and the seeds ripen from September to October.
Because of its compound, it is used by the native Americans for a variety of purposes.
Compuond decoctions are taken for gynecological, gastrointestrinal, urinary, and pediatric aids.
Plants can simply be taken as a “tonic”. Leaves are taken as a liver aid. Fruits are often made
into beverages, breads, and cakes or simply eaten raw. Summer grape in our local country is
wood vine, shallowly to deeply 3 to 5 lobed leaves 4 to inches long. The bloom season is in the
month of June. In visayan language, it is known as “bika” and are usually found in decidous
woods, thickets, wooded bluffs, etc.(Oliveira, 2012).

14
Review of Related Studies
Antibacterial effects of Herbs and Spices Extract on E. coli (Escherichia coli)
According to the study of (Amrita, Venogupal et.al, 2009) about the “ Antibacterial effects
of Herbs and Spices Extract on E. coli (Escherichia coli). Escherichia coli are the most
commonly present bacterium in thehuman intestine, which helps in preventing the entry of
pathogenic microorganisms. E. coli are non-pathogenic in normal conditions, but if present in
excess, will become causative agent of various diseases like urinary tract infection, diarrhea,
vomiting etc. With increasing of microorganisms to antibiotics, there is a shift of choice from
allopathic to ayurvedic and naturopathy, where herbs and spices are very common ingredients
of medicines. Herbs and spices are used in Indian recipes as they impart aroma and flavor to it.
Most of the studies performed t check sensitivity of microorganisms for various herbs and spices
involve extraction of the active component (s) with some solvents. However, when these plant
products are used as ingredients of food, the effectiveness of extracts using organic solvents
should not be a criterion. In the present study, frequently used herbs and spices are selected,
their extract are made using distilled water and tested for its antimicrobial effect against E.coli
the most common intestinal non-pathogenic organism. The antibacterial effect of various herbs
and spices were evaluated using various methods. All the herbs and spices tested were able to
inhibit E. coli, but thyme (herb) and Turmeric (spice) were found to be the most effective against
E. coli.
15
The Antibacterial Effect of Turmeric (Curcuma longa) Leaves and Fire Tree (Delonix regia)
Leaves Extract against S. aureus (Staphylococcus aureus)
According to the study of (Barrameda, Laiza D. et, al, 2016) about “The antibacterial
Effect of Turmeric (Curcuma longa) Leaves and Fire Tree (Delonix regia) Leaves Extracts
against S. aureus (Staphylococcus aureus). Fire tree is a large tree with fern like-leaves. Leaves
are oblong, in 18 to 30 pairs, about 1.5 centimeters long. It has antimicrobial, antioxidant, anti-
diarrhea, hepatoprotective and anti-inflammatory properties. In some countries it is used as cure
for skin problems, chest pain, and other ailments. In this study, fire tree and turmeric leaves
were utilized as antibacterial against Staphylococcus aureus. The experiment was conducted at
Microbiology Laboratory of West Visayas State University. Twenty-four MHA (Muller-Hinton
Agar) plates were used in the study. Three plates were used as (+) control were labelled as
SA/F1;SA/F3. The average of the X and Y zone of inhibition from each plate were measured by
means of a ruler and the data obtained was expressed in centimeter. The data taken were
placed in tabulated form and was treated statistically. Results were analyzed and interpreted.
Conclusions and recommendations were expressed. The obtained F-ratio of the differences
were classified according to the different treatments of fire tree and turmeric leaves extracts and
the positive control was 0.375 and the p-value was 0.72. Since the probability value is greater
than 0.05 alpha level, the result shows that the difference in fire tree, turmeric and positive
control were not significant. The results shows that the difference in fire tree and turmeric leaves
extracts could be effective as the positive control. Therefore, it was concluded that fire tree and
turmeric leaves extracts were effective against Staphylococcus aureus.

16
Antibacterial Property of Tambis (Syzygium samaragense) Leaves against S. aureus
(Staphylococcus aureus)
According to the study of (Alanan, Emerald May L. et.al, 2014) entitled “Antibacterial
Property of Tambis (Syzygium samaragense) Leaves against S. aureus (Staphylococcus
aureus). This study aimed to determine if tambis leaves extract can be used as an antibacterial
against Staphylococcus aureus. More specifically, the study aimed to determine if tambis leaves
extract is effective as an antibacterial agent against Staphylococcus aureus and to determine
the best antibacterial between red and green tambis leaves against Staphylococcus aureus.
This is to accomplish further discoveries for plants containing antibacterial property, and which
can help consumers with small budget. One hundred grams each of green and red of fresh
tambis leaves were weighed, pounded, and soaked in ethanol for 48 hours. The ethanol extract
was filtered and was allowed to evaporate. Amber bottles were used to store extracts. A loop full
of the pure culture isolates was taken and aseptically transferred into a sterile tube of nutrient
broth incubated for 15-30 minutes. McFarland’s No. 5 was used to compare the density of the
culture by lacing the tube in a bond paper with rubber bonds. Later, 38 grams of dehydrated
MHA was rehydrated and placed in an Erlenmeyer flask with 1000 mL distilled water.
The rehydrated MHA was heated until it melted. Sterilization in the autoclave at 12
degree for 15 minutes at 15 psi followed. Cooling between 40 degree and 45 degree was
allowed and the MHA was aseptically transferred into each plate at 25 mL each. Plates were
allowed to cool and harden undisturbed. By means of cork borer, holes the agar plate were
punched to make a well. Each plates was labeled as to their level of concentration (100%, 75%,
50% and 25%), the bacteria to be tested the plant part and the number of replicates, same
17
procedure was repeated with the three other control plates. One gram of antibiotic (flucloxacillin
500 mg) was diluted in 70 mL distilled water placed in volumetric flask. Contents were shaken
well in the flask. Into each of the pounded agar plates, a sterile swab moistened with the pure
isolate was used to streak hardened surface in a zigzag manner. Three drops of 0.3 mL of the
plant extracts were placed in the agar wells of each of the plates labeled 100%, 75%, 50% and
25%. The same amount of the standardized antibiotic was placed into the 3 other control
plates. Then, the plates were incubated for 18-24 hours. The plates were examined for Zone of
Inhibition formation. The date were recorded in tabulated form and statistically treated. It was
recorded that 100% concentration of tambis leaves extract was the best antibacterial agent
against Staphylococcus aureus with the mean of 2.42 cm. One-way analysis of variance at 0.05
level of significant confirmed that the 100% concentration of tambis leaves extract has a
significant difference from positive control (Pure Flucloxacillin) and as the most potent
concentration ( the best antibacterial agent against staphylococcus aureus) having an F-value of
118.979> critical value of 3.58. The researchers concluded that the tambis (Syzygium
samaragense) leaves extract can be used as antibacterial agent against staphylococcus
aureus. It was also found out that the best concentration tested was the 100% red tambis leaves
extract. Based on the following findings and conclusions, the researchers recommended that
the red tambis leaves extract is capable to an antibacterial agent against S. aureus.
18
Chapter 3
Methodology
Chapter 3 consist of three parts (1) Purpose of the Study and Research Design (2)
Materials, Data-Gathering Procedure (3) Statistical Data Analysis Tool.
Part One, Purpose of the Study and Research Design, restates the purpose of the study
and describes the research design and the variables used in the research.
Part Two, Materials and Data-Gathering Procedure, shows the materials and equipment
used and discusses the step-by-step process of the research.
Part Three, Statistical Data Analysis Tool, shows the statistical tools used in the
research.
19
Purpose of the Study and Research Design
The purpose of this study was to determine the antibacterial effect of summer grapefruit
extract against Escherichia coli . It was specifically designed to answer the following questions:
1. Is summer grape fruit extract effective as an antibacterial agent against Escherichia
coli?
2. Is there any significant difference in the zone of inhibition of Escherichia coliusing ripe
and raw summer grape fruit extract and ciprofloxacin?
This study used experimental research design wherein, there was ripe, raw,
positive and negative (control group) that consists of three replicates. This was
conducted to make an alternative substance using summer grape fruit extract. The
variables used in the study were gathered, and were experimented to determine the
effect of summer grape fruit extract against Escherichia coli. Ciprofloxacin was used as
the positive control and Distilled water as the negative control in the study against
Escherichia coli.
The researchers also used the randomized complete block design wherein they
used three treatments with three replicates.

20
T1R2
T1R1 T1R2 T1R3

T1R3
T2R1 T2R2
T2R2 T2R3
T2R3
T3R1
T3R1 T3R2
T3R2 T3R3
Figure 2. This figure shows the ripe and raw summer grape fruit extract and ciprofloxacin in
three replicates using Randomized Complete BlockDesign.

21
Experimental Treatment
Table 1. Components of Ripe Summer Grape Fruit Extract in a Treatment.
Treatment Ripe Summer Grape Fruit Extract
T1R1 0.1 mL
T1R2 0.1 mL
T1R3 0.1 mL
Table 1 shows the components of ripe summer grape fruit extract in a treatment with three
replicates and the extract measuresof 0.1 mL.
Table 2. Components of Raw Summer Grape Fruit Extract in a Treatment.
Treatment Raw Summer Grape Fruit Extract
T2R1 0.1 mL
T2R2 0.1 mL
T2R3 0.1 mL
Table 2 shows the components of raw summer grape fruit extract in a treatment with three
replicates and the extract measures 0.1 mL.

22
Table 3. Components of Ciprofloxacin (Positive Control) in a Treatment.
Treatment Ciprofloxacin
T3R1 0.1 mL
T3R2 0.1 mL
T3R3 0.1 mL
Table 3 shows the components of ciprofloxacin in a treatment with three replicates and the
extract measures 0.1 mL.
Materials
The materials used in the study are the following:
Materials Quantity
, Ripe Summer Grape Fruit Extract 50 mL
Raw Summer Grape Fruit Extract 50 mL
Ciprofloxacin 0.3 mL
Distilled Water 0.3 mL

23
Equipment
The equipment used in the study are the following:
a. Alcohol lamp for sterilizing the cork borer.
b. Autoclave for sterilizing the culture media MHA (Muller-Hinton Agar) and other equipment.
c. Beaker for storing the extracts of the summer grape fruit.
d. Cork borer for making wells in agar plates.
e. Electronic weighing scale to weigh summer grape fruit (ripe and raw).
f. Erlenmeyer flask for storing the rehydrated MHA (Muller-Hinton Agar).
g. Glass spreader to spread the bacteria inside the plates.
h. Incubator for incubating the control plates with sample.
i. Micro-pipette for transferring the isolate bacteria and treatments.
j. Petri dish as a plate for agar-making where the sample is placed.
k. Vernier caliper for measuring the zone of inhibition formation of the sample.
24
Cost Analysis
Table 4. The unit Cost of the Variables in Experiment
Materials Amount of Materials Used Unit/Unit Price Price Per Replicate
Ripe Summer Grape Fruit 0.3 mL 50 mL/Php 00.00 Php 00.00
T
Raw Summer Grape Fruit 0.3mL 50 mL/Php 00.00 Php 00.00
Ciprofloxacin 0.3mL 10 mL/Php 100.00 Php 35.50
Distilled Water 0.3 mL 250 mL/Php 10.00 Php 3.33
TOTAL: Php 38.83
The table 4 shows the unit cost of the variables in the treatment. Fifty milliliter (mL) of
ripe summer grape fruit were stored and only 0.3 mL of summer grape fruit extract were used in
the 3 replicates. The same with 50 mL of raw summer grape fruit and only 0.3 mL of summer
grape fruit extract were used in the 3 replicates. One bottle of ciprofloxacin cost Php 100.00 and
only 10 mL of ciprofloxacin were used in the 3 replicates. The amount of distilled water used in 3
replicates is 0.3 mL out of 250 mL that cost Php 10.00. The total price per replicate is Php
38.83.
25
Data Gathering Procedure
A. Extraction of Summer grape fruit
One hundred grams of summer grape fruit (ripe and raw) were collected and it was
weighed using electronic weighing scale. The ripe and raw summer grape fruit were extracted
using Slow juicer. Then, the 50mL ripe summer grape fruit extract were stored at 80mL beaker
and the same with the 50mL raw summer grape fruit extract, but it was separately stored in
another 80 mL beaker. The Erlenmeyer were used to store extracts and were kept in refrigerator
in order for the extracts to avoid contamination.
B. Preparation of Ciprofloxacin
The 10mL liquid form of ciprofloxacin was shaken and measured. Only 0.3mL were used
and were dropped into three control plates.
C. Preparation of E. coli culture
A loop full of pure culture isolate was taken and aseptically transferred into a sterile tube
of nutrient broth (N. broth) and incubated 37 °C for 24 hrs. and were stored in a chiller.
D. Preparation of MHA (Muller-Hinton Agar) plates
The 7.6 g of dehydrated MHA was placed in Erlenmeyer flask and diluted to 200 mL
using distilled water as solvent. In the autoclave, it was sterilized at 121°C for 15 min at 15 psi.
Then, at 45°C it was allowed to cool and aseptically transferred to 6 control plates at 15 mL of
each. The plates were cooled and hardened undisturbed.

26
E. Application of Bacteria
The bacteria inside the culture tube were inoculated in nutrient broth (N. broth) and were
aseptically transferred to 6 control plates using the micro-pipette with the volume 0.1 mL each
plate. Then, in each control plates the bacteria was spread using sterile glass spreader.
F. Agar Well-Method
By means of sterile cork borer, holes of each plate of the agar were punched to make a
well. Each plates was labeled as (T1R1, T1R2, T1R3), (T2R2, T2R2,T2R3), the initial of the bacteria to
be tested, plant part and number of replicates was followed using the ripe summer grape fruit,
raw summer grape fruit, positive control (Ciprofloxacin) and negative control (Distilled water).
G. Application of Treatments, Positive and Negative control
Three drops of 0.1 mL ripe summer grape extract were dropped in the agar wells that
were placed in each of T1R1, T1,R2, T1R3. . Three drops of 0.1 mL raw summer grape extract were
dropped in the agar wells that were placed in each ofT 2R1, T2R2,T2R3. The same amount of the
antibiotic (Ciprofloxacin) as positive control was placed into the 3 control plates. For the
negative control, three drops of 0.1 mL of distilled water were placed in each well in negative
control plate.
H. Incubation and Reading for Zone of Inhibition
Inside the incubator, the plates were placed for 24 hrs. Then, for the zone of inhibition
formation the plates were examined. In each plates, the diameter of zone of inhibition was
27
measured with a Vernier caliper in millimeter (mm) and the average of three replicates of each
treatment were recorded in tabulated form. Then, statistically treated.
Statistical Data Analysis Procedure
The data gathered were used for descriptive and inferential analysis.
Mean. The mean scores gathered from the result were used to determine the mean
difference between summer grape fruit extract and ciprofloxacin in terms of their effectiveness
One-Way Anova Analysis of Variance. The One-way Anova set at 0.05 alpha level of
significance was used to determine the significant difference among treatments in terms of their
effectiveness against Escherichia coli.
One Sample T-test. The One Sample T-test were used to compare the computed
mean of zone of inhibition of Escherichia coli using erythromycin and ripe summer grape fruit
extract.
28
Chapter 4
Results and Discussions
Chapter 4 is consist of Descriptive Data Analysis and Discussion, Inferential Data
Analysis and Discussion.
Part One, Descriptive Data Analysis and Discussion. It discusses and provides results of
the antibacterial effect of summer grape fruit extract against Escherichia coli. In specific time
frame.
Part Two, Inferential Data Analysis and Discussion. It discusses and provides results of
the significance difference between ripe and raw summer grape fruit extract and ciprofloxacin.
29
Descriptive Data Analysis and Discussion

The descriptive findings of the study presented the difference between summer grape fruit
extract (ripe and raw) and ciprofloxacin in terms of their effectiveness against Escherichia coli.
Four treatments were used to determine the most effective against Escherichia coli.
Table 5. Zone of Inhibition of Escherichia coli using Ripe and Raw Summer Grape Fruit Extract
and Ciprofloxacin.
Treatments Zone of Inhibition (mm)

R1 R2 R3
Summer Grape Fruit (Ripe) 11.85 13.25 14.30
Summer Grape Fruit (Raw) 12.30 15.25 11.45
Ciprofloxacin (Positive Control) 33.50 47.20 33.00
This table shows the measurement of zone of inhibition of Escherichia coli using ripe
summer grape fruit, raw summer grape fruit and ciprofloxacin as positive control. In replicate 1,
ripe summer grape fruit has 11.85 mm, raw summer grape fruit has 12.30 mm, and ciprofloxacin
has 33.50 mm. In replicate 2, ripe summer grape fruit has 13.25 mm, raw summer grape fruit
has 15.25 mm, and ciprofloxacin has 47.20 mm. In replicate 3, ripe summer grape fruit has
14.30 mm, raw summer grape fruit has 11.45, and ciprofloxacin has 33.0 mm.
30
Table 6. Mean. Average of the Four Treatments in Three Replicates in terms of Zone of
Inhibition.
Treatments Mean
Treatment 1 (Ripe Summer Grape Fruit Extract) 13.13

Treatment 2 (Raw Summer Grape Fruit Extract) 13.00
Treatment 3 (Ciprofloxacin) 37.90
Treatment 4 (Distilled Water) 6.00
Scale for the Effectiveness
Mean Description
30-40 Most Effective
20-29.99 Effective
10-19.99 Moderately Effective
0-9.99 Least Effective
This table shows the mean average of the four treatments in terms of their zone of
inhibition. Treatment 1 (x= 13.13) and Treatment 2 (x= 13.00) are interpreted as Moderately
Effective, Treatment 3 (x= 37.90) is interpreted as Most Effective and Treatment 4 is the least
effective. The researchers considered that between the four treatments. Treatment 3 had the
best results because it has the highest mean (x= 37.90).

31
Inferential Data Analysis and Discussion
Table 7. ANOVA. Significant Difference of the Three Treatments in terms of Effectiveness
Treatment Mean Sig. Value
Treatment 1 (Ripe Summer Grape Fruit Extract) 13.13

Treatment 2 (Raw Summer Grape Fruit Extract) 13.00 0.001
Treatment 3 (Ciprofloxacin) 37.90
This table shows that there is a significant difference among Treatment 1, Treatment 2
and Treatment 3 in terms of effectiveness (p= 0.001) which is lower than (a= 0.05).
The result shows that in terms of effectiveness, Treatment 3 (ciprofloxacin) was the most
effective among the three treatments (x = 37.90). However, results showed that Treatment 1
and Treatment 2 reached moderately effective. Therefore, treatment 1 and Treatment 2 are
comparable to that Treatment 3 (ciprofloxacin) and in between Treatment 1 and Treatment 2, it
was considered that Treatment 1 (ripe summer grape fruit extract) is more effective against
Esxherichia coli (x = 13.13).

32
Table 8.One Sample T-test.Significant difference between Experimental product and
Commercial product in terms of their effectiveness as antibacterial agent against Escherichia
coli.
Variable Mean Sig. Value
Experimental 13.13
Commercial 10.00 0.048
This table shows the significant difference in the mean zone of inhibition of Escherichia
coli usingexperimental product (x = 13.13) which is better than the commercial product (x = 10).
The mean of the two products has a significant difference (p = 0.048) which is lower than (a =
0.05).
The researcher used 95% confident interval (a = 0.05) that the experimental product (summer
grape fruit extract) (x = 13.13) is more effective than the commercial product (erythromycin). It
shows that there is a significant difference between the mean of the two sample.
33
Chapter 5
Summary, Conclusion, and Recommendations
Chapter 5 consist of three parts; (1) Summary, (2) Conclusions, and (3)
Recommendations.
Part One, Summary, presents the overview of the study.
Part Two, Conclusions, answers the questions presented in the statement of the
problem.
Part Three, Recommendations, presents the recommended things in improving the
study.
34
Summary
This study was conducted to determine antibacterial effect of summer grape fruit extract
(ripe and raw) against Escherichia coli.
It also sought to answer the following:
1. Is summer grape fruit extract effective as an antibacterial agent Escherichia coli?
2. Is there any significant difference in the zone of inhibition of Escherichia coli
using ripe and raw summer grape fruit extractand ciprofloxacin?
The process involves collection and preparation of materials used, 100 grams of
summer grape fruit were gathered and weighed using the electronic weighing scale. Ten MHA
(Muller-Hinton Agar) plates which contains ripe and raw summer grape fruit extract were
incubated for 24 hours. After that, the zone of inhibition were observed and measured by the
researchers and the result showed that the difference in each treatment of variables and
positive control are significant.
Conclusions
In view of aforementioned findings, the following conclusions were drawn:
1. Summer grape fruit extract is effective as an antibacterial agent against Escherichia coli.
35
2. There is a significant difference in the zone of inhibition of Escherichia coli using ripe and raw
summer grape fruit extract and ciprofloxacin.
3.There is a significant difference between experimental product and commercial product in
terms of their effectiveness as antibacterial agent against Escherichia coli.
Recommendations
Based on the following findings and conclusion, the following recommendations are
hereby presented.
1. Utilization of other species of summer grape to be considered in testing the antibacterial
effect.
2. The usage of ROTAVAP method in extraction to increase the level of concentration against
Escherichia coli.
3. Production of antibacterial agent from summer grape fruit in a form of tablet, powder or
solution.
4. Perform phytochemical analysis to determine the antibacterial component present in summer
grape.
5. Find other possible variable that is more concentrated than ripe and raw summer grape fruit
to improve the effect against Escherichia coli.
6. Further study that involves ripe and raw summer grape fruit extract as antibacterial agent

References
Journals:
Alanan, E.M, et. al (2014). Antibacterial Property of Tambis (Syzygium samaragense) Leaves
extracts against S. aureus. Iloilo City. Camacho Printing Press
Barrameda, L, et.al (2016). The Antibacterial Effect of Turmeric (Curcuma longa) Leaves and
Fire Tree (Delonix regia) Leaves extract against S. aureus. Iloilo City. Camacho Printing
Press
Kabir, F, et. al (2015). Antimicrobial Activities of Grape (Vitis vinifera linn.) Pomace
Polyphenols as a Source of Naturally occurring Bioactive Components. 14:2157-61.
Retrieved November 9, 2019, from Retrieved November 9, 2019, from
http://Nurmahani08.blogspot.com/2012/antimicrobial-activities-of-Grape-Pomace-
Polyphenols-as-a-source-of-Naturally-occurring-Bioactive-Components
Nurmahani, MM, et. al (2012). Antibacterial Property of Hylocereus polyrhizus and Hylocereus
undatos Peel Extracts. 19:77-84. Retrieved November 9, 2019, from
http://Nurmahani08.blogspot.com/2012/antibacterial-property-of-Hylocereus-polyrhizus-
and-Hylocereus-undatos-Peel -Extracts
Oliviera, D.A and A.A, Salvador (2012-2013). Antimicrobial Activity and Composition Profile of
Grape (Vitis vinifera) Pomace Extracts obtained by Supersticial Fluids. 164:423-
32. Retrieved November 8, 2019, from http;//www.reconstrue.co.nz/lPCT-J Vol 6
Darley Amarao Oliviera.pdf

Websites:
http://www.news-medical.net/health/What-is-Escherichia-coli.aspx
http://pharmacyinformatics2013-csab.bblogspot.com/2013/07/summer-grape/bika_26.html
http://www.microbiologyonline.og.uk/about-microbiology/introducing-microbes
http://www.ncbi.nih.gov/pmc/articles/PMC3217694/.

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Barotac Viejo National High School

Science Technology and Engineering

ANTIBACTERIAL EFFECT OF SUMMER GRAPE (Vitis aestivalis)

An Investigative Project Presented to

the Faculty of Science, Technology and Engineering Department

of Barotac Viejo National High School

Barotac Viejo, Iloilo

In Partial Fulfillment of the Requirements

For Junior High School Completion

This Science Investigatory Project, entitled “ANTIBACTERIAL EFFECT OF

IYSA MARIE B. REA JESCEL MAE H. BALDELOVAR

(Date Signed) (Date Signed)

MARIMAR P. BANABATAC GEROME G. REA

(Date Signed) (Date Signed)

LANI JOY O. ESPADA FEROLYN J. BALAJADIA

(Date Signed) (Date Signed)

Accepted as partial fulfillment of the requirements for Junior High School

MA. LUISA L. ASONG PhD

The researchers would like to acknowledge with sincerest gratitude to those

pursue their study;

develop their knowledge and ideas towards the study;

the research work;

suggestions for the improvement of their study;

To Mrs. Lani Joy O. Espada, Science Technology and Engineering Coordinator,

for her encouragement and support throughout the study;

and help researchers finished the study;

To Ms. Marimar P. Banabatac, Science Critic, for lending her expertise,

suggestions, encouragement and full support in making the study;

grammatical errors in the completion of the study;

To Mr. Gerome G. Rea, Statistician, for helping the researchers in computing,

solving and analyzing the statistical data of their study;

giving assistance in conducting their study;

encounter during their study; and

Razel Ann T. Canto

Christian Andrei B. Payas

List of Tables viii

Introduction of the Study 1

Background of the Study 2

Statement of the Problem 5

Significance of the Study 7

Scope and Delimitation of the Study 8

Review of Related Literature and Studies 10

Review of Related Literature 11

Review of Related Studies 14

Purpose of the Study and Research Design 19

Materials and Equipment 22

Data Gathering Procedure 25

Statistical Data Analysis Procedure 27

Result and Discussions 28

Descriptive Data Analysis and Discussion 29

Inferential Data Analysis and Discussion 31

Summary, Conclusions, and Recommendations 33

1 Components of Ripe Summer Grape Fruit Extract in a Treatment 21

2 Components of Raw Summer Grape Fruit Extract in a Treatment 21

3 Components of Ciprofloxacin in a Treatment 22

Fruit Extract and Ciprofloxacin

6 Mean Average of the Four Treatments in Three 30

7 Significant Difference of the Three Treatments in terms of Effectiveness 31